materials

Review Biocompatibility of Polyimides: A Mini-Review

Catalin P. Constantin 1 , Magdalena Aflori 1 , Radu F. Damian 2 and Radu D. Rusu 1,*

1 “Petru Poni” Institute of Macromolecular Chemistry, Romanian Academy, Aleea Grigore Ghica Voda 41A, Iasi-700487, Romania; [email protected] (C.P.C.); mafl[email protected] (M.A.) 2 SC Intelectro Iasi SRL, Str. Iancu Bacalu, nr.3, Iasi-700029, Romania; [email protected] * Correspondence: [email protected]; Tel.: +40-232-217454

 Received: 14 August 2019; Accepted: 25 September 2019; Published: 27 September 2019 

Abstract: Polyimides (PIs) represent a benchmark for high-performance on the basis of a remarkable collection of valuable traits and accessible production pathways and therefore have incited serious attention from the ever-demanding medical field. Their characteristics make them suitable for service in hostile environments and purification or sterilization by robust methods, as requested by most biomedical applications. Even if PIs are generally regarded as “biocompatible”, proper analysis and understanding of their biocompatibility and safe use in biological systems deeply needed. This mini-review is designed to encompass some of the most robust available research on the biocompatibility of various commercial or noncommercial PIs and to comprehend their potential in the biomedical area. Therefore, it considers (i) the newest concepts in the field, (ii) the chemical, (iii) physical, or (iv) manufacturing elements of PIs that could affect the subsequent biocompatibility, and, last but not least, (v) in vitro and in vivo biocompatibility assessment and (vi) reachable clinical trials involving defined polyimide structures. The main conclusion is that various PIs have the capacity to accommodate in vivo conditions in which they are able to function for a long time and can be judiciously certified as biocompatible.

Keywords: biocompatibility; polyimides; polyimide films; biostability; structural biocompatibility; in vitro testing; in vivo screening

1. Introduction Six decades after their entry into the market of commercial polymeric materials for aerospace, defense, and optoelectronics, polyimides (PIs) still maintain their status as a benchmark for high-performance polymers on the basis of a remarkable collection of thermoxidative resistance, outstanding mechanical characteristics, long-term reliance, and accessible production pathways [1–7]. The reputation of this engineering family among advanced-technology industries is backed by a handful of traits: excellent dielectric features, strength and flexibility, chemical and radiation inertness, interesting optical attributes, durability, prolonged shelf life, and others [8–10]. As a result, PIs easily established a plethora of applications as ( or thermosetting) dielectric films, wire insulation coatings, advanced composite matrices, specialty , and molding resins, photoimageable layers, flexible circuits, non-woven materials, advanced fabrics, medical tubing, etc. [7–10]. As expected, all these commercially-relevant features come with a premium cost. Nevertheless, the certified versatility and performance of this polymeric family render PIs as the materials of choice for high-performance, advanced uses in which breakdown, malfunction, or replacement costs are even higher [2,7,11,12]. Due to this remarkable combination of features, PIs incited serious attention from the exponentially growing medical field. This was doubled by the already accumulated knowledge regarding their functionalization, tuning, and processing which can be used to satisfy manifold complex requirements of present biomedical topics [12]. Therefore, several polyimide-based materials have been explored

Materials 2019, 12, 3166; doi:10.3390/ma12193166 www.mdpi.com/journal/materials Materials 2019, 12, 3166 2 of 27 in the last two and a half decades in various biomedical applications, from micropatterned cell substrates to implantable neuronal sensors. The main drive of research in this area is mainly focused on novel, processable, tailorable PI materials of superior performance to satisfy the innovation of the future. However, special interest must be directed towards their ability to make it in the ever-demanding biomedical field, biocompatibility being the first item on a long checklist. Biocompatibility represents a complex endeavor for producers of and medical devices. It is influenced by the entire lifecycle of a product, from the initial stages of design and materials development up to the market clearance and entry, and is the subject of tight regulations, guidance, and supervision. Hence, biocompatibility assessment embodies a composite enterprise of many variables, from the chemical nature or physical characteristics of the envisaged material to the tissue contact span or the nature of that tissue. Therefore, some pointers and perspectives are necessary when a (an already intricate and expensive to manufacture, high value) material arrives at the point of selecting the proper (also elaborate and high-priced) in vitro or in vivo biocompatibility evaluation technique [13,14]. PIs are sometimes described as “biocompatible” based on the seminal work presented in [15], which is dated 1993 and deals with the in vitro evaluation of five commercial polyimidic products. Even so, up-to-date, proper analysis of their biocompatibility and safe use in biological systems is deeply needed. Many times, a PI-based material designed for, e.g., further implanting is reported as biocompatible only by (in the best case scenario) citing the above-mentioned research or on the basis of some scarcely described in vitro cytotoxicity tests. On other occasions, their reputation can be at risk due to research performed on generally-described “polyimide-based” biomedical devices with poor results [16]. PIs are engineered starting from a rather small collection of building blocks which nevertheless provide a broad range of PI formulations. This wide PI variety demands more data, interpretations, and correlations to achieve the “biocompatible” label since even a small change in the macromolecular architecture can generate noticeable differences in terms of cytotoxicity, thrombogenicity, interaction with various cells and tissues, biostability and many others [17,18]. Ultimately, each polyimidic material intended to be employed in any biomedical application needs to be severely tested. However, a starting point on the topic is imperative and, in the absence of any solid known review dedicated to the matter, the present work aims to take the first steps in this regard. This mini-review is designed to encompass some of the most robust available research on the biocompatibility of various commercial or in-house synthesized PIs and to comprehend their potential in the biomedical area. It follows all reachable articles related to the biocompatibility of polyimides published in the last two decades, as present in the Web of Science database (topic: “biocompatibility” and “polyimide”, timespan: 2000–2019, all databases). The research was performed by considering (i) the newest concepts in the field, (ii) the chemical, (iii) physical, or (iv) manufacturing elements of PIs that could affect the subsequent biocompatibility, and, last but not least, (v) in vitro and in vivo biocompatibility assessment and (vi) reachable clinical trials involving defined polyimide structures.

2. Polyimides: Synthesis, General Features, and Biomedical Applications PIs represent a family of polymeric architectures acknowledged first and foremost for their high-temperature resistance, which covers all their material forms: films, coatings, varnishes, binders, tapes, fibers, composites, and foams. The chemistry of PIs is by nature a broad domain of materials science and covers a wide assortment of macromolecular backbones, starting materials, and synthetic pathways. Therefore, they are able to deliver manifold characteristics covering quite some large boundaries, which can be easily amended by relatively inconsequential structural modifications. There are many excellent monographs, reviews, and books dedicated to this class of polymers and the present work does not intend to reproduce their content, but to resume the essentials and to build the basics for the upcoming chapters [1–3,7,8,11]. This section is dedicated to aromatic polyimides, which are planar imidic macromolecules containing rigid aromatic rings and various flexible, heteroaliphatic, unsymmetrical, bulky or pendant structural motifs. We chose to focus Materials 2019, 12, x FOR PEER REVIEW 3 of 27 Materials 2019, 12, 3166 3 of 27 various flexible, heteroaliphatic, unsymmetrical, bulky or pendant structural motifs. We chose to focuson the on aromatic the aromatic polymers polymers since since the majority the majority of polyimidic of polyimidic materials materials tested tested for biomedical for biomedical uses usessubscribe subscribe to this to structural this structural architecture, architecture, even if some even commercial if some orcommercial in-house aliphatic or in-house backbones aliphatic have backbonesfound their have way found into the their field way [19 ,into20]. the Moreover, field [19, the20]. review Moreover, is mainly the review centered is onmainly polyimidic centered films on polyimidic(also encompassing films (also foils encompassing and coatings) since, foils withand somecoatings) noticeable since, exceptions with some which noticeable will be underlinedexceptions whichat their will rightful be underlined place, this isat the their format rightful chosen place, by most this bio-relatedis the format applications chosen by and most devices. bio-related applicationsIn general and terms, devices. polyimides are attained by a one- or two-stage reaction between a diamine and aIn dianhydride general terms, in apolyimides polar aprotic are solvent attained (N,N-dimethylacetamide by a one- or two-stage andreaction N-methyl between pyrrolidinone a diamine andbeing a dianhydride the most used in ones) a polar which aprotic follows solvent a polycondensation (N,N-dimethylacetamide mechanism and (although N-methyl apyrrolidinone polyaddition beingprocess the is most also used available). ones) which The two-stage follows a pathwaypolycondensation (Scheme1 mechanism) implies the (although formation a polyaddition of a highly processsoluble, intermediateis also available). polyamic The acidtwo-stage (or polyamidic pathway acid,(Scheme PAA) 1) through implies a mildlythe formation exothermic of a reaction highly soluble,between theintermediate starting compounds polyamic atacid modest (or temperaturespolyamidic acid, (room PAA) temperature through usually a mildly suffi ces).exothermic This is reactiona stepwise between process the that starting takes placecompounds at a rate at depending modest temperatures on the ’ (room reactivity.temperature Afterward, usually suffices).the PAA intermediateThis is a stepwise is transformed process intothat thetakes corresponding place at a rate polyimidic depending structure on the via monomers’ thermal or reactivity. Afterward, the PAA intermediate is transformed into the corresponding polyimidic chemical imidization and concurrent H2O release. This procedure is usually employed in laboratories structureor by industry via thermal in the or preparation chemical imidization of polyimidic andmaterials concurrent which H2O arerelease. rather This insoluble procedure in theiris usually final employedform [2,21 ,in22 ].laboratories or by industry in the preparation of polyimidic materials which are rather insoluble in their final form [2,21,22].

Scheme 1. SchematicSchematic representation representation of of the two-step synthesis of a conventional polyimide.

The one-stage pathway eliminates the PAA precursor from the process and involves the stepwise The one-stage pathway eliminates the PAA precursor from the process and involves the of the (at least) two monomers at high temperatures and concomitant thermal stepwise polymerization of the (at least) two monomers at high temperatures and concomitant (sometimes chemical) imidization of the macromolecular backbone. This procedure is mainly followed thermal (sometimes chemical) imidization of the macromolecular backbone. This procedure is in-house in the synthesis of soluble, readily processable polyimides [1,23,24]. mainly followed in-house in the synthesis of soluble, readily processable polyimides [1,23,24]. The PAA route is the most used technique to produce commercial PI films. Usually, the PAA The PAA route is the most used technique to produce commercial PI films. Usually, the PAA precursor is stable and displays a long shelf life, which makes it easy to handle, deposit, transport, precursor is stable and displays a long shelf life, which makes it easy to handle, deposit, transport, or shape it into films, coatings, or fibers. PAA films are cast or spin-coated, and afterward thermally or shape it into films, coatings, or fibers. PAA films are cast or spin-coated, and afterward thermally dehydrated to generate the final imide film. The main drawback of this process is inefficient imidization dehydrated to generate the final imide film. The main drawback of this process is inefficient (cyclization) along the polymer chain, which usually does not render major flaws in the final material. imidization (cyclization) along the polymer chain, which usually does not render major flaws in the

Materials 2019, 12, 3166 4 of 27 Materials 2019, 12, x FOR PEER REVIEW 4 of 28 finalThe imidizationmaterial. The reaction imidization implies reaction only the implies release ofonly water, the whichrelease can of bewater, easily which removed can nowadaysbe easily removedby both small- nowadays or large-scale by both small- manufacturing, or large-scale and manufacturing, the obtained polyimidic and the obtained films, foils polyimidic or coatings films, are foilsmostly or freecoatings of voids are mostly or defects. free of voids or defects. Based onon these these simple simple and and straightforward straightforward synthetic synthetic methods, methods, most PIsmost can PIs be relativelycan be relatively amended amendedand tailored and to tailored supply to various supply features. various Thisfeatures. is usually This is performed usually performed based on judiciousbased on monomerjudicious monomermodifications modifications through properthrough chemical proper chemical tools or, tool to as lessor, to extent, a less byextent, small by interventions small interventions within withinthe manufacturing the manufacturing process, process, with little with sacrifice little sacrific of the desirede of the propertiesdesired properties of the final of the material. final material. The first Theapproach first approach is used on is aused large on scale a large to resolve scale to PIs’ reso biggestlve PIs’ bottleneck: biggest bottleneck: reduced solubility,reduced solubility, high softening high softeningtemperatures, temperatures, and troublesome and troublesome or costly processing or costly coming processing from highlycoming symmetrical, from highly polar, symmetrical, and rigid polar,macromolecular and rigid macromolecular backbones with strongbackbones intra- with and strong interchain intra- interactions. and interchain interactions. The same same structural structural elements elements mentioned mentioned above above unlock unlock the high-performance the high-performance character character of PIs: thermoxidativeof PIs: thermoxidative stamina,stamina, chemical chemical (organic (organicsolvents included) solvents included) and mechanical and mechanical durability, durability, excellent insulatingexcellent insulating properties, properties, and long-term and long-term reliance. These reliance. traits These make traits them make suitable them for suitable service for in servicehostile environmentsin hostile environments and purification/sterilization and purification/sterilization by robust methods, by robust as methods, requested as by requested most biomedical by most applications.biomedical applications. Moreover, as Moreover, it will be asdetailed it will belater, detailed PIs have later, the PIs capacity have the to capacity accommodate to accommodate to in vivo conditionsto in vivo conditions in which they in which are able they to are function able to for function a long fortime a longwithout time a withoutharming a response. harming response. All of the above and and the the positive positive experience experience of of heavily heavily employing employing PIs PIs in in microelectronics microelectronics lead to the successful successful use of polyimidic materials in high end, complex complex biomedical biomedical devices like bio micro-electromechanical-systems (bio-MEMS),(bio-MEMS), e.g., re retinaltinal and neural neural implants implants [16,25]. [16,25]. PIs PIs fulfill fulfill a three-folda three-fold function function inin these these devices: devices: a flexible a flexible (and unstretchable)(and unstretchable) mechanic mechanic substrate substrate for the embedded for the embeddedmetallic elements, metallic an elements, efficient an insulating efficient substrateinsulating between substrate adjacent between conductor adjacent conductor paths and anpaths overall and an(chemical) overall encapsulant(chemical) encapsulant to protect the to device protect against the device the adverse against liquid the adverse surroundings. liquid Dependingsurroundings. on Dependingthe final application, on the final PIs enable application, the production PIs enable of bio-MEMS the production devices ofof small-bio-MEMS or micro-sizes devices andof small- thickness, or micro-sizesvarious shapes, and high thickness, density of electrodes,various shapes, a large numberhigh density of stimulating of electrodes,/recording elements,a large andnumber variable of stimulating/recordingdegrees of flexibility or elements, stiffness. Asand a consequence,variable degrees the useof flexibility of PIs ensures or stiffness. the smallest As a possibleconsequence, intrusion the usein the of biologicalPIs ensures frame, the ansmallest additional possible key feature intrusion for implantablein the biological devices. frame, Moreover, an additional PIs are already key feature easily forprocessed implantable by well-established devices. Moreover, micromachining PIs techniquesare already in conventionaleasily processed clean room by facilitieswell-established which are micromachiningalso employed in thetechniques production in conventional of bio-MEMS, clean thus ensuringroom facilities the desired which repeatability, are also employed pattern accuracy, in the productionand low costs of [ 7bio-MEMS,,8,16]. The complexity thus ensuring of such the a desired device is repeatability, depicted in Figure pattern1, showing accuracy, a flexible, and low stacked, costs [7,8,16].polymer-metal-polymer The complexity microsystem of such a placeddevice onis thedepi visualcted cortexin Figure of a cat.1, showing a flexible, stacked, polymer-metal-polymer microsystem placed on the visual cortex of a cat.

Figure 1. A high-density array of electrodes for mapping neural activity: flexible electrode array Figure 1. A high-density array of electrodes for mapping neural activity: flexible electrode array insulated and encapsulated by polyimide (unknown trademark) layers placed on the visual cortex of a insulated and encapsulated by polyimide (unknown trademark) layers placed on the visual cortex of

Materials 2019, 12, x FOR PEER REVIEW 5 of 28

Materialsa cat; 2019 (inset), 12, x FORthe folded PEER REVIEW electrode array inserted into the interhemispheric fissure. Adapted from [26],5 of 28 with permission from © 2011 Springer Nature. a cat; (inset) the folded electrode array inserted into the interhemispheric fissure. Adapted from [26], Materialswith 2019 permission, 12, x FOR fromPEER ©REVIEW 2011 Springer Nature. 5 of 28 MaterialsIn 2019a broad, 12, 3166 sense, there are two main PI versions employed in biomedical research: 5 of 27 conventional, non-photosensitive PI materials (cPIs), and photosensitive polyimides (psPIs). The a cat; (inset) the folded electrode array inserted into the interhemispheric fissure. Adapted from [26], latterMaterialsIn contain 2019a broad, 12 ,photoreactive x FOR sense, PEER REVIEWthere side- are or end-groups,two main PIusually versions of acrylic employed nature. in Representatives biomedical research: of5 bothof 28 with permission from © 2011 Springer Nature. conventional,versions are commercially non-photosensitive available PI mostlymaterials as aromatic(cPIs), and polymers. photosensitive cPIs are polyimides typically used (psPIs). starting The lattera containcat; (inset) photoreactive the folded electrode side- orarray end-groups, inserted in tousually the interhemispheric of acrylic nature. fissure. Representatives Adapted from [26], of both fromIn their a PAAbroad precursors, sense, there while are psPIs two are main mostly PI basedversions on PAA employed variations, in namelybiomedical esters research: or salts versionswith arepermission commercially from © 2011available Springer mostly Nature. as aromatic polymers. cPIs are typically used starting conventional,of theircat; polyamic (inset) non-photosensitive the acids folded [27,28]. electrode Both PI formsmaterials array areinserted (cPIavailables), and into as photosensitive liquids the interhemispheric or varnishes, polyimides which fissure. (psPIs). are cast AdaptedThe or from [26], fromspin-coated their PAA on variousprecursors, substrates while psPIs and thermallyare mostly cured based untilon PAA an imidizedvariations, film namely is obtained. esters or Some salts latterwithIn contain a permissionbroad photoreactive sense, from there side-© 2011are or end-groups,two Springer mainNature. PIusually versions of acrylic employed nature. in Representatives biomedical research: of both ofcommercial their polyamic cPIs areacids directly [27,28]. available Both forms and are employed available as as preimidized liquids or varnishes, films, like which the well-known are cast or versionsconventional, are commercially non-photosensitive available PI mostlymaterials as aromatic(cPIs), and polymers. photosensitive cPIs are polyimides typically used (psPIs). starting The spin-coatedKaptonIn aor broad Ultem on various sense,polyimidic substrates there materials are and two (for thermally mainthe actual PI cured versionsstructures until an employedof imidizedthe most usedfilm in biomedical commercialis obtained. PIs Some research: see conventional, fromlatter theircontain PAA photoreactive precursors, whileside- orpsPIs end-groups, are mostly usually based ofon acrylicPAA variations, nature. Representatives namely esters ofor bothsalts commercialTable 1). cPI cPIs films are aredirectly processed available by andphotolithography employed as preimidizedand dry/wet films, etching like thewhich well-known imply a non-photosensitiveofversions their polyamic are commercially acids PI[27,28]. materialsavailable Both mostlyforms (cPIs), areas aromaticavailable and photosensitive polymers.as liquids cPIsor varnishes, are polyimides typically which used (psPIs).are starting cast or The latter contain Kaptonphotoresist or Ultem or hard polyimidic mask asmaterials an etch (for template the actual. The structures psPI counterparts of the most used are commercialmanufactured PIs seeby photoreactivespin-coatedfrom their PAA on various side-precursors,or substrates end-groups, while psPIsand thermallyare usually mostly cured based of acrylicuntil on PAA an nature. imidizedvariations, film Representatives namely is obtained. esters or Some salts of both versions are Tablestraightforward 1). cPI films patterning are processed through UVby exposurephotolithography and developer and dry/wetchemicals, etching which whicheliminate imply some a commercialof their polyamic cPIs areacids directly [27,28]. available Both forms and areemployed available as as preimidized liquids or varnishes, films, like which the well-known are cast or commerciallyphotoresist or availablehard mask mostlyas an etch as aromatictemplate. polymers.The psPI counterparts cPIs are typically are manufactured used starting by from their PAA Kaptonprocessing or Ultemsteps. polyimidicAs a drawback, materials the photo-defina (for the actualble structures versions areof the moisture most used sensitive commercial and prone PIs see to straightforwardspin-coated on variouspatterning substrates through and UV thermallyexposure andcured developer until an chemicals,imidized film which is obtained.eliminate Somesome precursors,waterTable sorption1). cPI while filmsand uptake, psPIsare processed arewhich mostly needs by photolithography basedadditional on stabilizers PAA variations,and or dry/wet restricts namely etchingtheir application which esters imply orin saltsvivo a of their polyamic processingcommercial steps. cPIs Asare adirectly drawback, available the photo-defina and employedble versions as preimidized are moisture films, sensitive like the and well-known prone to acids[25].photoresist [27,28 or]. hard Both mask forms as arean etch available template as. The liquids psPI orcounterparts varnishes, are whichmanufactured are cast by or spin-coated on waterKapton sorption or Ultem and polyimidic uptake, which materials needs (for additional the actual stabilizers structures orof restrictsthe most their used applicationcommercial in PIs vivo see variousstraightforwardTable 1). substrates cPI films patterning andare processedthermally through UVby cured exposurephotolithography until and an developer imidized and dry/wetchemicals, film etching is which obtained. whicheliminate Someimply some commerciala cPIs are [25]. Table 1. Structure, trade name, and producer of commercial polyimides (or precursors) tested for processingphotoresist steps.or hard As amask drawback, as an the etch photo-defina template. bleThe versions psPI counterpartsare moisture sensitiveare manufactured and prone byto directlytheir available biocompatibility. and employed as preimidized films, like the well-known or Ultem polyimidic water sorption and uptake, which needs additional stabilizers or restricts their application in vivo straightforwardTable 1. Structure, patterning trade throughname, and UV producer exposure of comme and developerrcial polyimides chemicals, (or precursors) which eliminate tested for some materials[25]. (for the actual structures of the most used commercial PIs see Table1). cPI films are processed processingtheir biocompatibility. steps. As a drawback, the photo-definable versions are moisture sensitive and prone to bywater photolithography sorption and uptake, and which dry/ wetneeds etching additional which stabilizers imply or arestricts photoresistTrade their applicationName or hard 2; Producer maskin vivo as an etch template. The[25]. psPI Table counterparts1. Structure, trade are name, manufactured and producer of comme by straightforwardrcial polyimides (or precursors) patterning tested through for UV exposure and their biocompatibility.Structure of Polyimide (or Precursor 1) BiocompatibilityTrade Name 2; Producer Studies developer chemicals, which eliminate some processing steps. As a drawback, the photo-definable Table 1. Structure, trade name, and producer of commercial polyimides (or precursors) tested for versions are moistureStructure of sensitive Polyimide and (or Precursor prone to1) water sorptionBiocompatibility and[References] uptake, Studies which needs additional their biocompatibility. Trade Name 2; Producer stabilizers or restricts their application in vivo [25]. [References] 1 1 Structure of Polyimide (or Precursor ) BiocompatibilityTradePI2611 Name ; Du 2; Producer Pont Studies Table 1. Structure, trade name, and producer of commercial polyimides (or precursors) tested PI2611 1; Du Pont for their biocompatibility.Structure of Polyimide (or Precursor 1) Biocompatibilityin[References] vitro; in vivo Studies

in[7,16,27,29–40] vitro; in vivo 2 PI2611[References] 1; TradeDu Pont Name ; Producer 1 Structure of Polyimide (or Precursor ) Biocompatibility Studies [7,16,27,29–40][References] 1 DurimidePI2611in vitro; 7000 1; Duin vivo ;Pont Fujifilm 1 Materials 2019, 12, x FOR PEER REVIEW PI26116 of; Du28 Pont 1 Durimidein[7,16,27,29–40] vitro; 7000 in vivo; inFujifilm vitro; in vivo [7,16,27,29–40]

Materials 2019, 12, x FOR PEER REVIEW [27,29,36,41–43]in vitro;in vitro in vivo 6 of 28 Durimide[7,16,27,29–40] 7000 1; Fujifilm

1 [27,29,36,41–43][15]Durimide 7000 ; Fujifilm Materials 2019, 12, x FOR PEER REVIEW in vitro 6 of 28 DurimideKapton;in vitro; 7000 Du in 1 vivo;Pontin Fujifilm vitro; in vivo [27,29,36,41–43]

Thermid[15] EL 5512; theoretical;Kapton;[27,29,36,41–43]in vitro;in vitro Du inin vitro; vivo Pont in Materials 2019, 12, x FOR PEER REVIEW Nationalvivo Starch and6 of 28 Kapton; Du Pont

theoretical;Chemical in vitro; Co in ThermidKapton;[27,29,36,41–43][15]theoretical; DuEL 5512;Pont in vitro; in vivo [15,20,27,30,44–46]vivo Nationalin vitroStarch[15 , 20and,27 ,30,44–46] in vitro theoretical;Chemical in vitro;Co in [15,20,27,30,44–46]ThermidKapton; DuEL 5512;Pont [15] ThermidNationalThermid ELvivo[15] 5010;Starch EL 5010; National and Starch and Chemical Co

in vitro theoretical;NationalChemical Starch in vitro;Co and in in vitro [15,20,27,30,44–46]ThermidChemicalvivo EL Co5010; [15]

Thermid EL 5512; Ultem 1010;[15] GE Nationalin vitroStarch and National Starch and Chemical Co

[15,20,27,30,44–46] ThermidThermidChemical ELin 5512;vitro EL 5010;Co National Starch and Chemical Co [15] UltemNational 1010; Starch GE Plastics andin vitro in vitro [15] Chemical[15] Co Thermid EL 5010; in vitro UltemNational 1010; Starch GE Plastics and

[15] ChemicalUltem Co 1010; GE Plastics Silbem;[15] GE Plastics in vitro in vitro [15] Ultem 1010;in vitro GE Plastics Silbem; [15]GE Plastics

in [15]vitro Silbem; GE Plastics in vitro in vitro

Silbem; GE Plastics [15]

[15] PI2525[15] 1; Du Pont in vitro PI2525 1; Du Pont Silbem;in vitro; GE in Plastics vivoin vitro; in vivo [15]1 PI2525 ; Du Pont[28 ,47,48]

[28,47,48]in vitro in vitro;1 in vivo PI2525 ; Du Pont

[15] Pyralin [28,47,48]PI 2700 1; Du Pont in vitro; in vivo

1 PI2525in vitro; Du Pont Pyralin PI[28,47,48] 2700 1; Du Pont

in vitro;[28] in vivo in vitro1 Pyralin PI 2700 ; Du Pont [28,47,48] Probimide [28]7500 1; Fujifilm in vitro Pyralin PI 2700 1; Du Pont in vitro; in vivo Probimide [28]7500 1; Fujifilm

in [49]vitro in vitro; in vivo Probimide 7500 1; Fujifilm [28] Neopulim;[49] Mitsubishi in vitro; in vivo Probimide 7500 1; Fujifilm in vitro Neopulim;[49] Mitsubishi in vitro; in vivo [19] in vitro Neopulim; Mitsubishi 1 structure is represented as the commercial polyimide precursor, but all biocompatibility[49] tests were

2 [19] performed on the imidized form of the polymeric material; common tradenames or inabbreviations vitro for the monomers: PMDA (pyromellitic dianhydride), BPDA (biphenyl-tetracarboxylic acid 1 structure is represented as the commercial polyimide precursor, but all biocompatibilityNeopulim; tests Mitsubishi were performed on the imidized form of the polymeric material; 2 common tradenames or abbreviations[19]

for the monomers: PMDA (pyromellitic dianhydride), BPDA (biphenyl-tetracarboxylic acid 1 structure is represented as the commercial polyimide precursor, but all biocompatibilityin vitrotests were performed on the imidized form of the polymeric material; 2 common tradenames or abbreviations

for the monomers: PMDA (pyromellitic dianhydride), BPDA (biphenyl-tetracarboxylic[19] acid 1 structure is represented as the commercial polyimide precursor, but all biocompatibility tests were performed on the imidized form of the polymeric material; 2 common tradenames or abbreviations for the monomers: PMDA (pyromellitic dianhydride), BPDA (biphenyl-tetracarboxylic acid

Materials 2019, 12, x FOR PEER REVIEW 6 of 28

Materials 2019, 12, x FOR PEER REVIEW 6 of 28 in vitro Materials 2019, 12, x FOR PEER REVIEW 6 of 28 in vitro [15] in[15] vitro Thermid EL 5512; [15] NationalThermid Starch EL 5512; and NationalChemical Starch Co and ThermidChemical EL Co5512; in vitro National Starch and Chemicalin vitro Co [15] in[15] vitro

Ultem 1010; GE Plastics [15] Ultem 1010; GE Plastics in vitro

Ultem 1010;in vitro GE Plastics [15]

in[15] vitro

Silbem; GE Plastics [15] Silbem; GE Plastics in vitro

Silbem;in GEvitro Plastics [15]

in[15] vitro 1

PI2525 ; Du Pont [15] PI2525 1; Du Pont Materials 2019, 12, 3166 in vitro; in vivo 6 of 27

PI2525in vitro; 1; Duin vivo Pont [28,47,48]

Table 1. Cont. in vitro;[28,47,48] in vivo 1 Pyralin PI 2700 ; Du Pont [28,47,48]Trade Name 2; Producer Structure of Polyimide (or Precursor 1) Pyralin PI 2700Biocompatibility 1; Du Pont Studies in vitro [References] Pyralin PI 2700 1; Du Pont

in vitro [28] Materials 2019, 12, x FOR PEER REVIEW Pyralin PI 2700 1; Du Pont 7 of 28 in[28] vitro in vitro dianhydride), PPD (p-Phenylenediamine), 6FDA (4,4′-(hexafluoroisopropylidene)diphthalicProbimide 7500 1; Fujifilm[28] Materials 2019, 12, x FOR PEER REVIEW [28] 7 of 28 anhydride), ODPA (4,4'-oxydiphthalic anhydride), ODA (4,4’-oxydianiline).Probimide 7500 1; Fujifilm in vitro; in vivo Probimide 7500 1; Fujifilm dianhydride), PPD (p-Phenylenediamine), 6FDA (4,4′-(hexafluoroisopropylidene)diphthalicProbimide 7500 1; Fujifilm Materials 2019, 12, x FOR PEER REVIEW in vitro; in vivoin vitro; in vivo 7 of 28 anhydride),The properties ODPA of (4,4'-oxydiphthaliceach commercial anhydride), polyimidic ODA film (4,4’-oxydianiline).can fluctuate broadly [49] even[49 within] the same PI version (cPI or psPI), due to structural features, manufacturingin vitro;[49] in technique, vivo imidization dianhydride), PPD (p-Phenylenediamine), 6FDA (4,4′-(hexafluoroisopropylidene)diphthalic temperature, and final dimensions of the material, which can Neopulim;inflict modifications Mitsubishi in terms of The properties of each commercial polyimidic film can fluctuate broadly even within the same anhydride), ODPA (4,4'-oxydiphthalic anhydride), ODA (4,4’-oxydianiline). [49] PIdensity, version polymer (cPI chainor psPI), orientation, due to mechanicalstructural features,features, andmanufacturing moistureNeopulim; uptake. MitsubishiNeopulim;technique, The Mitsubishi issue imidization becomes in vitro in vitro temperature,even more complicated and final dimensionswhen it comes of theto the material, evaluation which of PIcan materials inflict modifications of noncommercial[19] in terms nature of The properties of each commercial polyimidic film can fluctuateNeopulim; broadlyin vitro Mitsubishi even within the same density,(synthesized polymer in the chain laboratory). orientation, The mechanical plethora of features, aliphatic, and semialiphatic, moisture uptake.[19] or aromatic The issue PI structures becomes PI version1 (cPI or psPI), due to structural features, manufacturing technique, imidization evenmakes more it 1simply structurestructure complicated impossibleis represented is represented when as to the itinclude ascommercialcomes the commercialtheir to polyimide the proper eval precursor, polyimideuationties into but of precursor, allclearly PI biocompatibility materials defined butin all[19] vitrotestsof biocompatibilityboundaries. noncommercial were Fortunately tests nature were temperature, and final dimensions of the material, which can2 inflict modifications in terms of performedperformed on on the the imidized imidized form of form the polymeric of the polymericmaterial; 2 common material; tradenamescommon or abbreviations tradenames or abbreviations (synthesizedor not, the1 structure number in the is represented oflaboratory). PIs which as the commercialThe were plethora tested polyimide forof precursor, altheiriphatic, biocompatibility but allsemialiphatic, biocompatibility is tests orrelatively aromaticwere small, PI structures some of density, forpolymer the monomers: chain orientation, PMDA (pyromellitic mechanical dianhydride), features, BPDA and (biphenyl-tetracarboxylicmoisture uptake.[19] The acid issue dianhydride), becomes for the monomers: PMDA (pyromellitic dianhydride), 2BPDA (biphenyl-tetracarboxylic acid makestheir structures it PPDperformedsimply (p-Phenylenediamine), being impossibleon the presentedimidized to form 6FDAinclude in of Tablethe (4,4 polymeric0 -(hexafluoroisopropylidene)diphthalictheir 2. proper material;ties common into clearly tradenames defined anhydride),or abbreviations boundaries. ODPA (4,40-oxydiphthalic Fortunately even more1 structure complicated is represented when as the it commercial comes to polyimide the eval precursor,uation but of all PI biocompatibility materials testsof noncommercial were nature or not, theanhydride),for numberthe monomers: ODA of PIs (4,4 PMDA0 -oxydianiline).which (pyromellitic were tested dianhydride), for their BPDA biocompatibility (biphenyl-tetracarboxylic is relatively acid small, some of (synthesized performed in the on thelaboratory). imidized form The of the plethora polymeric material;of aliphatic, 2 common semialiphatic, tradenames or abbreviations or aromatic PI structures Table 2. Structure and abbreviation of synthetic polyimides (or precursors) tested for their their structuresfor the beingmonomers: presented PMDA (pyromellitic in Table 2. dianhydride), BPDA (biphenyl-tetracarboxylic acid makesbiocompatibility. itThe simply properties impossible of each to commercialinclude their polyimidic properties film into can clearly fluctuate defined broadly boundaries. even within Fortunately the same PI or not,version the number (cPI or psPI), of PIs due which to structural were tested features, for their manufacturing biocompatibility technique, is relatively imidization small, temperature, some of Table 2. Structure and abbreviation of synthetic polyimides (or precursors) tested for their theirand structures final dimensions being presented of the material, in Table which 2. can inflict modifications in terms of density, polymer chain biocompatibility. orientation, mechanical features, and moisture uptake. The issue becomesAbbreviation even more complicated whenTable it comes2. Structure to the and evaluation abbreviation of PI materials of synthetic of noncommercial polyimides (or nature precursors) (synthesized tested infor the their laboratory). Structure of Polyimide (or Precursor 1) Biocompatibility Studies Thebiocompatibility. plethora of aliphatic, semialiphatic, or aromatic PI structures makes itAbbreviation simply impossible to include their properties into clearly defined boundaries. Fortunately or not, the number of PIs which were Structure of Polyimide (or Precursor 1) Biocompatibility[References] Studies tested for their biocompatibility is relatively small, some of their structuresAbbreviation being presented in Table2.

Table 2. Structure and abbreviation of synthetic polyimides[References] (or precursors) tested Structure of Polyimide (or Precursor 1) BiocompatibilityDOCDA-PPD Studies 1 for their biocompatibility.O O

N N H H [References] Abbreviationtheoretical 1 O O DOCDA-PPD StructureHOOC of PolyimideCOOH (or Precursor 1) Biocompatibility Studies CH3 n N N H H [References][50] theoretical 1 HOOCO COOHO DOCDA-PPD CH3 n 1 DOCDA-PPD N N [50] H H DOCDA-DDEtheoreticaltheoretical 1 HOOC COOH [50] CH3 n [50] DOCDA-DDEtheoretical 1 1 DOCDA-DDE theoreticaltheoretical[50] DOCDA-DDE 1 [50] [50] EPICOLN-PPDtheoretical EPICOLN-PPD [50] theoreticaltheoretical;EPICOLN-PPD; inin vitro vitro [20,51]

theoretical[20,51]; in vitro EPICOLN-PPD

[20,51] theoreticalEPICOLN-APB; in vitro

[20,51] theoreticalEPICOLN-APB; in vitro

theoretical[51]; in vitro EPICOLN-APB

[51] theoretical; in vitro

[51]

Materials 2019, 12, x FOR PEER REVIEW 7 of 28

dianhydride), PPD (p-Phenylenediamine), 6FDA (4,4′-(hexafluoroisopropylidene)diphthalic anhydride), ODPA (4,4'-oxydiphthalic anhydride), ODA (4,4’-oxydianiline).

The properties of each commercial polyimidic film can fluctuate broadly even within the same PI version (cPI or psPI), due to structural features, manufacturing technique, imidization temperature, and final dimensions of the material, which can inflict modifications in terms of density, polymer chain orientation, mechanical features, and moisture uptake. The issue becomes even more complicated when it comes to the evaluation of PI materials of noncommercial nature (synthesized in the laboratory). The plethora of aliphatic, semialiphatic, or aromatic PI structures makes it simply impossible to include their properties into clearly defined boundaries. Fortunately or not, the number of PIs which were tested for their biocompatibility is relatively small, some of their structures being presented in Table 2.

Table 2. Structure and abbreviation of synthetic polyimides (or precursors) tested for their biocompatibility.

Abbreviation

Structure of Polyimide (or Precursor 1) Biocompatibility Studies

[References]

1 O O DOCDA-PPD

N N H H theoretical HOOC COOH CH3 n [50]

DOCDA-DDE 1

theoretical

[50]

Materials 2019, 12, 3166 EPICOLN-PPD 7 of 27

theoretical; in vitro Table 2. Cont.

Abbreviation[20,51] Structure of Polyimide (or Precursor 1) Biocompatibility Studies [References] Materials 2019, 12, x FOR PEER REVIEW EPICOLN-APB 8 of 28 EPICOLN-APB Materials 2019, 12, x FOR PEER REVIEW theoreticaltheoretical;; inin vitro vitro 8 of 28 [6FDA-6FAP51] [51] 6FDA-6FAPin vitro 6FDA-6FAP in vitrovitro[52–54] [52–54] [52–54] O O O O N Ar N O O SPAB-xxDA HN S NH O O O O SPAB-xxDA O N Ar N SPAB-xxDA O n in vitroin vitro HN S NH [55] O O O CF3 O On in vitro[55] Ar = ;; ; CF3 CF3 O [55] Ar =1 ;; ; 2 biocompatibility tests were performed onCF the polymeric precursor (polyamidic acid); common tradenames 1 3 2 biocompatibilityor abbreviations tests for were the performed monomers: on the DOCDA polymeric (5-(2,5-dioxotetrahydrofuryl)-3-methyl-3-cyclohexene-1,2- precursor (polyamidic acid); common tradenames or abbreviationsdicarboxylic for anhydride), the monomers: DDE (4,40 -diaminodiphenylDOCDA (5-(2,5-di ether),oxotetrahydrofuryl)-3-methyl-3-cyclohexene-1,2- APB (1,3-bis(3-aminophenoxy) benzene), 1 6FAP (2,2-bis(3-amino-4-hydroxyphenyl) hexafluoropropane). 2 biocompatibilitydicarboxylic testsanhydride), were performed DDE (4,4 on′-diaminodiphenyl the polymeric precursor ether), (polyamidicAPB (1,3-bis(3-aminophenoxy) acid); common tradenames benzene), or 6FAP abbreviations(2,2-bis(3-amino-4-hydroxyphenyl) for the monomers: hexafluoropropane).DOCDA (5-(2,5-di oxotetrahydrofuryl)-3-methyl-3-cyclohexene-1,2- dicarboxylic3. Biocompatibility: anhydride), DDE Concepts, (4,4′-diaminodiphenyl Standards, and ether), Assessment APB (1,3-bis(3-aminophenoxy) benzene), 6FAP (2,2-bis(3-amino-4-hydroxyphenyl)3. Biocompatibility:The term “biocompatibility”, Concepts, hexafluoropropane). Standards, the “holy and grail” Assessment of any researcher working in the or medical domains, has reached such a common and careless usage in countless areas and contexts 3. Biocompatibility:The term “biocompatibility”, Concepts, Standards, the and“holy Assessment grail” of any researcher working in the biomaterial or thatmedical its sense domains, and gravity has reached have beensuch attenuateda common andand blurredcareless [usage17]. On in onecountless side, thereareas isand a constant contexts fluctuationthatThe termits sense in“biocompatibility”, its and operation gravity and have significance.the been “holy attenuated grail” On the of and other,any blurredresearcher the modern [17]. working exponentialOn one in side, the evolution biomaterialthere is ofa constant biology or medicalandfluctuation medicinedomains, in generates hasits operationreached several such and mutationsa significance. common inandthe On careless concepts the other, usage and the in understanding moderncountless exponential areas of biocompatibility.and contextsevolution of thatThe biologyits presentsense andand section gravitymedicine is designed have generates been to touchattenuated several some keyandmutati notionsblurredons and in[17]. the attributes On concepts one relatedside, andthere to theunderstanding is fielda constant and place of fluctuationthembiocompatibility. in ain contemporary its operation The present context.and significance. section is designed On the to other, touch the some modern key notions exponential and attributes evolution related of to biologythe fieldInand its and beginnings,medicine place them generates biocompatibility in a contemporary several was mutati regarded context.ons asin thethe property concepts of aand material understanding to be neutral of from biocompatibility.the physical,In its beginnings, chemical,The present biocompatibility and section physiological is designed was point regarded to touch of view. someas the Nowadays, key property notions theof anda term material attributes has progressedto be related neutral to to from an the intricatefieldthe physical,and conceptplace chemical, them describing in a and contemporary thephysiological interaction context. point between of view. a material Nowadays, and a the living term biological has progressed framework. to an TheintricateIn its definition beginnings, concept accepted describingbiocompatibility by the the academic interaction was regarded majority between as is the “thea propertymaterial ability ofand of a a materiala material living tobiologicalto be perform neutral framework. from with an the appropriatephysical,The definition chemical, host accepted response and physiological by in athe specific academic point application”. majority of view. Thereis Nowadays, “the is ability also anthe of expanded terma material has versionprogressed to perform that to includes with an an intricatetheappropriate concept concept of hostdescribing “bioactivity” response the in (aninteraction a specific action stimulated application”between a bymaterial. There the material) is and also a an living and expanded thus biological becomes version framework. muchthat includes wider Thethan thedefinition concept simple accepted inertia of “bioactivity” [17 by,18 the]. This academic (an comes action asmajority stimulated a necessary is “the by complement abilitythe material) of a of material theand IUPAC thus to becomesperform (International withmuch Unionan wider appropriateofthan Pure simple andhost Applied responseinertia [17,18]. Chemistry) in a specific This definition comesapplication” as whicha necessary. There describes is alsocomplement an biocompatibility expanded of the version IUPAC as the that “ability(International includes to be the inconceptUnion contact of of withPure “bioactivity” aand living Applied system (an Chemistry)action without stimulated producing definition by an thewh adverse ichmaterial) describes effect” and [biocompatib56 thus]. For becomes a detailedility much as and the intriguing wider“ability to thandiscussion besimple in contact inertia regarding with [17,18]. a theliving This meaning system comes of withoutas “biocompatibility” a necessary producing complement an and adverse its often of effect”the improper IUPAC [56]. uses,For(International a thedetailed reader and is Uniondirectedintriguing of Pure towards and discussion Applied reference regarding Chemistry) [13], in the which definition meaning is judiciously whof “bichiocompatibility” describes argued biocompatib that “a and biocompatible itsility often as improperthe material“ “ability uses, to is anthe be inabusereader contact and is with thedirected proper a living towards descriptive system reference without expression [13], producing isin “intrinsically which an isadverse judiciously biocompatible effect” argued [56]. system”. For that a “adetailed biocompatible and intriguingmaterial“Biocompatibility discussion is an abuse regarding assessments and the the proper meaning pursue descriptive the of potential“biocompatibility” expression risk brought is “intrinsically and by its amaterial often biocompatible improper for a living uses, organism, system”. the readere.g., is patient, Biocompatibilitydirected by employingtowards assessments reference certain conditions[13], pursue in wh the andich potentia environmentsis judiciouslyl risk brought which argued are bythat close a “ama or biocompatibleterial similar for to a clinical living material“circumstances.organism, is an e.g.,abuse Naturally, patient, and the by the proper employing multiplicity descriptive certain which ex condit governspressionions biomaterials, is and “intrinsically environments their biocompatible applications which are and close system”. interactions or similar withtoBiocompatibility clinical biological circumstances. systems assessments makes Naturally, it difficultpursue the or the evenmultiplici potentia impossibletyl whichrisk to providebrought governs aby harmonized,biomaterials, a material generally theirfor aapplications living accepted, organism,orand practical interactions e.g., patient, evaluation withby ofemploying biocompatibilitybiological certain systems condit [13 ,14makes,ions17]. Theandit difficult assessmentenvironments or ofeven (implantable)which impossible are close medical orto similarprovide devices, a to clinicalharmonized, circumstances. generally Naturally, accepted, the multiplicior practicalty which evaluation governs of biomaterials, biocompati bilitytheir applications[13,14,17]. The and assessmentinteractions ofwith (implantable) biological systemsmedical makesdevices, it thedifficult main orbio-related even impossible application to provideof polyimidic a harmonized,materials, generally is established accepted, by internationally or practical acknowledged evaluation of standards. biocompati Thebility ISO-10993 [13,14,17]. (International The assessmentOrganization of (implantable) for Standardization) medical guidelinesdevices, theare themain most bio-related important application and used standards of polyimidic in Europe, materials,Asia (mostis established countries) by and internationally the US (even acknowledged if the FDA (U.S. standards. Food and The Drug ISO-10993 Administration) (International provides Organizationsome more for severeStandardization) requisites guidelineson certain are topics). the most These important guidelines and areused not standards to be considered in Europe, as a Asia mandatory(most countries) checklist; and they the USbuild (even a recommended if the FDA (U.S. gene Foodral framework and Drug of Administration) suitable, efficient provides tests for the someanalysis more severe of biocompatibility, requisites on andcertain judicious topics). proofs These of equivalenceguidelines are are notaccepted to be [18]. considered as a mandatory checklist; they build a recommended general framework of suitable, efficient tests for the analysis of biocompatibility, and judicious proofs of equivalence are accepted [18].

Materials 2019, 12, 3166 8 of 27 the main bio-related application of polyimidic materials, is established by internationally acknowledged standards. The ISO-10993 (International Organization for Standardization) guidelines are the most important and used standards in Europe, Asia (most countries) and the US (even if the FDA (U.S. Food andMaterials Drug 2019 Administration), 12, x FOR PEER REVIEW provides some more severe requisites on certain topics). These guidelines9 of are 28 not to be considered as a mandatory checklist; they build a recommended general framework of suitable, efficientAt teststhis forpoint, the analysiswe must of biocompatibility,underline that, andcurren judicioustly, there proofs is no of equivalencePI-based material are accepted certified [18]. accordingAt this to point, the weabove-mentioned must underline that,standards. currently, According there is no to PI-based our assessment, material certified the BPDA-PPD according topolyimidic the above-mentioned architecture standards. (abbreviated According accordingly to our to assessment, the starting the monomers: BPDA-PPD diphenyl polyimidic dianhydride, architecture (abbreviatedBPDA, and accordinglyp-phenilene to diamine, the starting PPD) monomers: is the most diphenyl used dianhydride, polyimidic BPDA,material and inp bio-related-phenilene diamine, studies PPD)and applications, is the most used being polyimidic the main materialsubject of in roughly bio-related 24% studies of the and articles applications, analyzed being for this the review. main subject Even ofthis roughly material, 24% commercially of the articlesanalyzed available for as this PI2611 review. (DuPont; Even this HD material, Microsystems) commercially or U-Varnish-S available as (UBE), PI2611 (DuPont;fails in this HD regard. Microsystems) Nevertheless, or U-Varnish-S several research (UBE), teams fails in explored this regard. its application Nevertheless, in neural several interfaces research teamsand demonstrated explored its application its overall in biocompatibility neural interfaces andas demonstrateda bulk material its overallor encapsulant biocompatibility of a long-term as a bulk materialimplantable or encapsulant device, based of a on long-term in vitro implantableand in vivo device,assays, based as it will on in be vitro discussedand in later vivo [16,25].assays, as it will be discussedBiocompatibility later [16,25 tests]. can be partitioned on three levels: in vitro (effects of materials on culturedBiocompatibility cells), in vivo tests (tissue can reactions be partitioned after onimplantation, three levels: usuallyin vitro in(e animals),ffects of materials and preclinical on cultured tests cells),(performancein vivo (tissue and reactions reactions afterof biomaterials implantation, in usually normal in animals),clinical conditions and preclinical after tests implantation (performance in andhumans) reactions [57]. of biomaterialsThey are assessing in normal “the clinical big conditions three” afterbiological implantation effects: incytotoxicity humans) [57 ].(in They vitro), are assessingsensitization “the (in big vivo), three” and biological irritation effects: (in cytotoxicityvivo), which (in are vitro common), sensitization to biomaterials (in vivo), anddesigned irritation for (medicalin vivo), devices. which are Despite common the to absence biomaterials of the designed complex forbiological medical and devices. physiological Despite the environment, absence of the in complexvitro studies biological are employed and physiological on a large environment, scale and arein able vitro tostudies provide are valuable employed information on a large regarding scale and arethe ablebiological to provide toxicity valuable of a given information material regarding and to direct the biological further toxicitystudies of(Figure a given 2). material They involve and to lower direct furthercosts and, studies most (Figure important,2). They a involvesmaller lowerextent costs of and,ethical most controversies important, aas smaller compared extent to of in ethical vivo controversiesexperiments, aswhich compared are nevertheless to in vivo experiments, closer to an whichideal biological are nevertheless research closer methodology to an ideal [14,18,57]. biological researchBoth types methodology of screening [ 14imply,18,57 a ].robust Both research types of screeningmethodology imply and a robustjustification research which methodology can be attained and justificationby a thorough, which sequential can be attainedchemical by and a thorough, physical investigation. sequential chemical and physical investigation.

Figure 2. SequentialSequential chemical, chemical, physical, physical, and and biological biological investigation investigation steps steps of polymeric of polymeric materials materials for forbiocompatibility biocompatibility assessment. assessment. Adapted Adapted from from [18], [ 18with], with permission permission from from © 2018© 2018 Royal Royal Society Society of ofChemistry. Chemistry.

Therefore, biocompatibilitybiocompatibility reviewingreviewing isis aa complexcomplex processprocess thatthat followsfollows aa gradualgradual investigativeinvestigative systemsystem that is influenced influenced by by several several parameters parameters co comingming from from the the material. material. In Inthe the particular particular case case of ofpolymers, polymers, polyimides polyimides included, included, these these parameters parameters are are bulk bulk characteristics characteristics (overall composition, degree of crystallinity, morphology),morphology), surface features (surface chemistry and and energy, energy, hydrophilicity, morphology), degradation kinetics and products, leachables and extractablesextractables and theirtheir potentialpotential toxicity. An ideal pattern for comprehending and assessing all these features does not exist, but the systematic approach provided in [17,18] is a good starting point. However, the employment of standardized analyses should become the generally accepted rule, since (especially in a scientific environment inclined towards open access) it enables judicious comparisons between various studies and empowers knowledge related to the biological behavior of materials and their biosafety.

3.1. Biostability of Polyimides

Materials 2019, 12, 3166 9 of 27 toxicity. An ideal pattern for comprehending and assessing all these features does not exist, but the systematic approach provided in [17,18] is a good starting point. However, the employment of standardized analyses should become the generally accepted rule, since (especially in a scientific environment inclined towards open access) it enables judicious comparisons between various studies and empowers knowledge related to the biological behavior of materials and their biosafety.

Biostability of Polyimides When it comes to biomaterials designed for implantable medical devices, biostability is a concept which needs a separate discussion. This is also the case of PIs since most of their bio-related applications deal with long-term implantable devices, e.g., neural prostheses. In a broad sense, biostability implies various aspects, mainly of chemical and sometimes mechanical nature, related to the stability and integrity of the investigated polymeric material or the system containing it. For example, the mechanical support and electrical or chemical insulating prepared from one or several PI films should not degrade or delaminate in the harsh environment provided by the human body. The good adhesion between metals and PIs is certified by their long common use in microelectronics. However, the delamination of metals (Ti, Au, Pt) from PIs is a serious issue for the long-term durability of bio-MEMS and a threat for their host [16,25]. The aqueous surroundings of an presume interactions between the PI material and different ions and molecules of various sizes (salts and proteins) and a certain amount of absorbed water. While the former does not pose a threat due to the general chemical inertness of PIs, water uptake can be a two-fold problem even for these generally hydrophobic polymers. Moisture adsorption causes stretching and has a plasticizing effect, which becomes a serious issue for the long-term use of PIs, especially for the more moisture-sensitive psPI type. Moreover, any hydrolytic-induced alteration or degradation can be exacerbated by pH changes of the environment and, in the case of bio-MEMS, by voltages coming from the microelectrode arrays. A common and accessible approximation of a polymeric material’s biostability can be performed by in vitro soaking tests in various media (saline physiologic solutions, saline buffer salts, cell culture media, etc.) at body temperature [25]. Sometimes, these biostability assays are accomplished in accelerated aging conditions (at higher temperatures in the same media) in order to boost diffusion and moisture uptake and asses the influence of water and aging and to estimate a failure period. One of the least polar PIs, BPDA-PPD, excels in this regard since it displays a very low water uptake (0.045%) and a subsequently reduced coefficient of expansion induced by the wet environment [16,29,44]. This is one of the main reasons for the extensive study of its commercial forms (liquid varnish precursors PI2611 and U-Varnish-S, thermally cured in-house until imidization) as the polymeric support and encapsulant of many neuroprosthetics. This polymeric material raises above other frequently employed PIs, like Kapton, several psPIs, fluorinated PIs, and other BPDA-based PIs, which have a much higher water uptake or, in some extreme cases, prove slow degradation in the wet environment [1,11,16,25,58]. Both BPDA-PPD-based commercial PIs are quite stable to the plasticization effect of water. More importantly, they are able to sustain excellent mechanical features after long-term storage in simulated body conditions (20 months at 37 ◦C in water or saline phosphate buffer, PBS) or after accelerated aging (60 and 85 ◦C in water) [29]. These observations are supported by several studies regarding the in vivo assessment of electrodes based on the aforementioned PIs, which showed them to properly operate even at 12 months post-implantation [59,60]. Subjected to the same simulated body conditions as PI2611 and U-Varnish-S, a commercial psPI material (Durimide 7510; Fujifilm) also showed surprising long-term stability in saline buffer and deionized water environments, despite a small, yet tolerable decrease of the mechanical characteristics [29]. None of the three investigated PIs showed any visible changes in the chemical composition of the macromolecular backbone, as evaluated by infrared spectroscopy. Materials 2019, 12, 3166 10 of 27

However, all of them displayed a decrease of the mechanical characteristics after being stored for a long period (20 months) in PBS at 85 ◦C, indicating an undeniable sensitivity to alkali metal salts and ions at this temperature. The Upilex 25S foil, a commercial preimidized version of BPDA-PPD liquid precursors, provided resembling stability at body temperature in water and PBS. When exposed to more extreme experimental conditions, it showed a mass loss and some mutations in the overall macroscopic behavior. For example, after being incubated in PBS at 85 ◦C for 18 months, this PI foil displayed a heterogeneous area with changedMaterials 2019 rugosity, 12, x FOR and PEER revealed REVIEW several creases on its surface (Figure3). Moreover, it started to11 tearof 28 apart while carefully being handled or rinsing. All these observations corroborate to the reasoning thatobservations at least somecorroborate parts of to the the materials, reasoning most that likelyat least uncyclized some parts polyamic of the acid,materials, dissolved most in likely PBS atuncyclized high temperatures. polyamic acid, dissolved in PBS at high temperatures.

Figure 3.3.SEM SEM micrographs micrographs of theof the surface surface of Upilex25S of Upilex25S film after film (left) after and (left) before and (right) before being (right) incubated being

inincubated PBS at 85 in◦ CPBS for 18at months.85 °C for Reproduced 18 months. from Reprod [29],uced with from permission [29], with from permission© 2010 Elsevier. from © 2010 Elsevier. Another conclusion coming from these studies is that proper attention must be directed towards the experimentalAnother conclusion conditions coming in which from accelerated these studies aging is assaysthat proper are performed. attention must The mechanismsbe directed bytowards which the polyimidic experimental materials conditions are failing in whichin vitro acceleratedare not always aging dependent assays are on performed. the diffusion The or uptakemechanisms of water by andwhich temperature-subordinated polyimidic materials are processes failing in must vitro be are taken not into always consideration, dependent alone on the or indiffusion combination or uptake with theof former.water and For temperature- example, thesubordinated aforementioned processes incubation must inbe PBS taken at 85 into◦C forconsideration, various months alone doesor in notcombination prove to bewith a proper the former. method For toexample, evaluate the the aforementioned long-term conduct incubation of PIs atin bodyPBS at temperature. 85 °C for various months does not prove to be a proper method to evaluate the long-term conductMoreover, of PIs at any body of these temperature. results require validation by in vivo testing, and the ISO standards provide someMoreover, guidance for any finding of these the rightresults experiments require validation in this regard. by in These vivo evaluations testing, and also the need ISO to standards approach anyprovide structural some guidance changes takingfor finding place the in theright material experiments after implantation. in this regard. For These instance, evaluations one brief also paper need onto approach the topic any reports structural delamination changes and taking a 30% place drop in the in the material tensile after strength implantation. of PI2611 For after instance, 11 months one inbrief vivo paper[61]. on This the di topicfference reports underlines delamination that in vitro and modelsa 30% drop and experimentsin the tensile are strength not able of to PI2611 encompass after the11 months potential in evivoffect [61]. of enzymes This difference on the PI underlines material. that Therefore, in vitro even models if PBS and is experiments much closer are to the not body able environmentto encompass as the compared potential to watereffect inof termsenzymes of salt on or th ionse PI content material. and Therefore, pH, in vitro evensoaking if PBS tests is inmuch this mediumcloser to canthe body give resultsenvironment quite far asfrom compared the in to vivo waterones. in Atterms first of glance, salt orit ions seems content futile and to particularize pH, in vitro thesoaking differences tests in between this mediumin vitro canand givein results vivo assays, quite fa evenr from if strictlythe in vivo referring ones. toAt polyimidic first glance, materials. it seems Nonetheless,futile to particularize there are the several differences studies between which try in to vitr drawo and a direct in vivo line assays, between even the if two, strictly an at referring least risky, to ifpolyimidic not erroneous materials. endeavor. Nonetheless, there are several studies which try to draw a direct line between the two,Taking an at into least consideration risky, if not all erroneous of the above, endeavor. it seems rational to accede to the necessity of a paradigm changeTaking regarding into consideration implants proposed all of inthe [17 above,], and replaceit seems “biocompatibility” rational to accede with to the “biotolerability”, necessity of a whichparadigm is defined change as “theregarding ability ofimplants materials proposed to reside inin the [17], body and for longreplace periods “biocompatibility” of time with only with low degrees“biotolerability”, of inflammatory which is reaction”. defined as “the ability of materials to reside in the body for long periods of time with only low degrees of inflammatory reaction”.

4. The Influence of Chemical Composition, Physical Features, and Manufacturing of Polyimides on Their Biocompatibility Outcome

Despite the intensive research in the field, the basic elements which control polymers’ biocompatibility are not yet fully comprehended. This is especially the case of the chemical building blocks or the overall chemical blueprint that govern the interaction of an implanted polymeric biomaterial with the host system. This is available for polyimides as well and a straight line between the chemical formulation of a polyimidic material and its biocompatibility is inaccessible. On the other side, any correlation between important factors like interfacial free energy or molecular weight and biocompatibility is unreachable due to disparate or missing information in the available literature. However, based on several biocompatibility assays related to polyimides, we identified

Materials 2019, 12, 3166 11 of 27

4. The Influence of Chemical Composition, Physical Features, and Manufacturing of Polyimides on Their Biocompatibility Outcome Despite the intensive research in the field, the basic elements which control polymers’ biocompatibility are not yet fully comprehended. This is especially the case of the chemical building blocks or the overall chemical blueprint that govern the interaction of an implanted polymeric biomaterial with the host system. This is available for polyimides as well and a straight line between the chemical formulation of a polyimidic material and its biocompatibility is inaccessible. On the other side, any correlation between important factors like interfacial free energy or molecular weight and biocompatibility is unreachable due to disparate or missing information in the available literature. However, based on several biocompatibility assays related to polyimides, we identified some key points which connect the chemical structure or surface of polyimides and the biocompatibility outcome of the resulting materials. These are addressed in the following subsections and are related to (i) the chemical nature of the secondary components of the polyimidic material (and a particular case related to functional groups’ influence), (ii) the surface wettability (balance between the hydrophilicity and the hydrophobicity), (iii) mechanical features of the polymeric film, (iv) surface topography or roughness.

4.1. Chemical Composition The assessment of biocompatibility also covers the evaluation of any biochemical reaction determined by the interaction between the biological framework and the overall material or its secondary components. The latter are generally called leachables or extractables and display a variety of sources: processing aids, unremoved solvents, antioxidants, stabilizers, , catalysts, oligomers, monomers, contaminants, degradation products or debris, etc. [17,18]. As detailed earlier, PI films and foils come from straightforward synthetic and manufacturing pathways and display overall chemical resistance and tolerance to conventional sterilization (with a soft spot for alkali or aqueous solutions). These features drastically reduce the long list of leachable sources and increase the importance of sample preparation and investigation in the overall biocompatibility outcome According to the ISO guidelines, an extraction ratio (sample surface area/extractant volume) of 6 cm2/mL is recommended for polymeric films with a thickness below 0.5 mm. The extraction media needs to properly represent the environment in which the material or device is intended for use. Likewise, the extraction conditions must come close to the circumstances in which the material is going to be used. However, in many cases, the available studies on polyimides’ biocompatibility deviate from these guidelines and a comparison between their outcomes must be done with caution. Of course, these guidelines should be applied only in the case of properly purified polyimidic materials, an attribute which, unfortunately, is not found in all scientific papers dedicated to PIs. There are many factors that can lead to a large variation in the main features of PI materials and affect their biocompatibility and their post-implant behavior. Alongside structural diversity, commercial availability of two forms of the same material (PAA precursor and preimidized film), small variations in the processing methods (different curing temperatures included) or even different sizes of the same sample are some the most common reasons for different biocompatibility outcomes of polyimidic materials. For example, the relatively high water uptake and moisture sensitivity of Kapton, perhaps the most known and generally used PI, hampers its long-term application in bio-MEMS. In comparison, the PI2611 precursor or its Upilex 25S cyclized counterpart do not suffer from such a disadvantage [29]. As detailed in the previous section, the PI’s stability reaches its maximum in the fully cured state, that is when the PAA amount is reduced to a minimum by its proper treatment at elevated temperatures [27]. One of the few in-depth studies treating the interaction between cells and commercial PI leachables investigates the effect of extractables coming from three commonly employed PIs (preimidized Kapton foil and in-house thermally imidized PI 2610 and HD 3007 precursors) on mitochondria and cell viability [30]. The research follows ISO 10993 protocols and is focused on some of the most expected leachables Materials 2019, 12, 3166 12 of 27 like PAA, incompletely cyclized products and residual solvents (in this case, N-methyl-2-pyrrolidone and gamma butyrolactone). The study does not cover degradation products since they are mostly expected after long-term in vivo employment. It was found that the incubation of human capillary and microvascular endothelial cells with extracts coming from the three PIs did not generate any significant stress or cytotoxicity, since no relevant mutations in the marker of mitochondrial stress (membrane potential) or the more severe markers of cell viability (apoptotic and necrotic cell death) were detectedMaterials 2019 (Figure, 12, x 4FOR). PEER REVIEW 13 of 28

Figure 4. CellCell viability viability of of SV-HCEC SV-HCEC human endothelial cells cells after 24 h exposure (Polyimide (PI) 2 samples in cell culture media at 37 ◦°CC for 48 h, 1 cm /mL/mL media ratio) to extracts coming from various common PIs:PIs: PMDA-ODA PMDA-ODA (Kapton), (Kapton), BPDA-PPD BPDA-PPD (thermally (thermally imidized imidized PI2611 precursor),PI2611 precursor), and polyimidic and adhesivepolyimidic (thermally imidized(thermally HD3007 imidized precursor). HD3007 Negativeprecursor). controls: Negati untreatedve controls: cells, untreated high-density cells, polyethylenehigh-density (HDPE). Positive (HDPE) control:. Positive natural control: latex rubber. natural Reproduced latex rubber. from Reproduced [30], with permissionfrom [30], fromwith permission© 2015 John from Wiley © and2015 Sons.John Wiley and Sons.

The successsuccess of thisof this assay assay has a manifoldhas a explanationmanifold explanation that mostly residesthat mostly in the high-temperatureresides in the (300high-temperature◦C) gradual treatment (300 °C) whichgradual is treatment used to imidize which the is used films. to On imidize one side, the the films. completely On one imidizedside, the PIcompletely chains are imidized highly inert,PI chains while are the highly unavoidable inert, while small the portion unavoidable of uncyclized small portion backbones of uncyclized is reduced tobackbones a minimum. is reduced At the sameto a time,minimum. the ineluctable At the same residual time, solvent the ineluctable content, already residual regarded solvent ascontent, of low toxicity,already regarded is too low as to of interfere low toxicity, with the is finaltoo low result. to interfere Moreover, with the the two final psPIs result. are heavily Moreover, cross-linked the two andpsPIs therefore are heavily any macromolecularcross-linked and mobility therefore or solventany macromolecular and small components mobility elutionor solvent is very and limited. small Althoughcomponents promising, elution theseis very results limited. need Although to be followed promising, by further these research results dealing need to with be anyfollowed potential by cytotoxicityfurther research of aged dealing PIs. with any potential cytotoxicity of aged PIs. Another study follows the cytotoxicity of four noncommercial PIs filmsfilms cast from N,N-dimethylacetamide andand containing containing the samethe sa diamineme diamine and several and commonlyseveral commonly employed anhydrideemployed fragmentsanhydride fragments (SPAB-xxDA, (SPAB-xxDA, their chemical their structurechemical structure is shown is at shown the end at ofthe Table end 2of)[ Table55]. 2) All [55]. of theAll investigatedof the investigated samples samples did notdid providenot provide any any leachables leachables that that could could prove prove toxic toxic for for humanhuman dermal fibroblasts,fibroblasts, as determined by indirect andand directdirect (days(days 1,1, 3,3, and 6) contact assays. The latter was used to build anan anhydride-dependentanhydride-dependent bi biocompatibilityocompatibility scale which, at this point, cannot be completely comprehended: 6FDA 6FDA >> BPDABPDA >> PMDAPMDA >> ODPA.ODPA. Nevertheless, Nevertheless, the the study study concludes that the 6F (hexafluoroisopropylidine)(hexafluoroisopropylidine) structuralstructural motif motif limits limits the absorptionthe absorption of human of fibroblasthuman andfibroblast contributes and tocontributes improved to biocompatibility. improved biocompatibility. It would be It really would helpful be really to see helpful if the to biocompatibility see if the biocompatibility differences betweendifferences PIs between or the above PIs or order the wouldabove beorder kept would when changingbe kept when the starting changing amine. the Thisstarting work amine. represents This anwork interesting represents starting an interesting point for more starting detailed point research for more focused detailed on the research role of various focused functional on the groupsrole of uponvarious the functional biocompatible groups characteristics upon the biocompatible of a polyimidic characterist material.ics of a polyimidic material.

4.2. Surface As for any given polymeric material [17,18], the wettability of a PI surface, commonly expressed by contact angle values, has a major influence on its biological response. Enhanced protein adsorption and cell adhesion are usually obtained in the case of more hydrophilic surfaces. However, as detailed previously, any increase in water uptake can have a negative impact on the long-term PIs’ mechanical features and reliance. Nevertheless, if, for example, a PI film is intended for cell or tissues culture usage, then it needs a surface of higher hydrophilicity, which, in the case of this type of polymers, can be attained by laser or plasma treatment [45,50,62,63] or chemical modifications [64–68]. The field of surface modifications of PI materials is so vast, that it would actually require a review of its own and therefore is not treated herein.

Materials 2019, 12, 3166 13 of 27

4.2. Surface As for any given polymeric material [17,18], the wettability of a PI surface, commonly expressed by contact angle values, has a major influence on its biological response. Enhanced protein adsorption and cell adhesion are usually obtained in the case of more hydrophilic surfaces. However, as detailed previously, any increase in water uptake can have a negative impact on the long-term PIs’ mechanical features and reliance. Nevertheless, if, for example, a PI film is intended for cell or tissues culture usage, then it needs a surface of higher hydrophilicity, which, in the case of this type of polymers, can be attained by laser or plasma treatment [45,50,62,63] or chemical modifications [64–68]. The field Materialsof surface 2019 modifications, 12, x FOR PEER ofREVIEW PI materials is so vast, that it would actually require a review of its14 ownof 28 and therefore is not treated herein. These observations observations were were confirmed confirmed by by cytotoxicity cytotoxicity assays assays of ofcommercial commercial PIs. PIs. For Forexample, example, the conventionalthe conventional PI2525 PI2525 film film has has a higher a higher hydrophilicity hydrophilicity (roughly (roughly 25° 25 ◦smallersmaller contact contact angles) angles) and surface free energy (a surplus of ca 22 mJ/m mJ/m2)) as as compared compared to to the the homologous homologous photosensitive photosensitive PI2771, PI2771, which results in a small increase in the adhesion and proliferation of BHK-21 (baby hamster kidney cells) fibroblasts,fibroblasts, as it can be observed inin FigureFigure5 5..

Figure 5. SEMSEM micrographs micrographs (three (three different different magnificat magnifications)ions) showing showing the the prevalence, prevalence, morphology, morphology, and adherence adherence of of BHK-21 BHK-21 cells cells at 24 at h 24 after h cultured after cultured on different on di polyimidefferent polyimide surfaces: surfaces:(A,D,G)

(conventional,A,D,G) conventional, non-photosensitive non-photosensitive PI2525 cured PI2525 at 200 cured °C; ( atB,E 200,H) ◦photosensitiveC; (B,E,H) photosensitive PI2771 cured PI2771 at 200 °C;cured (C, atF,I 200) photosensitive◦C; (C,F,I) photosensitive PI2771 cured PI2771at 350 °C. cured Adap at 350ted ◦fromC. Adapted [28], with from permission [28], with from permission © 2010 Elsevier.from © 2010 Elsevier.

The same rule of thumb (higher hydrophilicity for better compatibility) is available for PI biomaterials thatthat come come in contactin contact with with blood blood since surfacesince surface energy andenergy wettability and wettability drive the adsorptiondrive the adsorptionof plasma proteinsof plasma and proteins ulterior and adhesion ulterior ofadhesion platelets of toplatelets their surface. to their Bloodsurface. compatibility Blood compatibility implies impliesrestricted, restricted, if any, platelet if any, adhesionplatelet adhesion and activation, and activation, and thrombogenicity and thrombogenicity [18]. Even [18]. if theEven ultimate if the ultimateevaluation evaluation of hemocompatibility of hemocompatibility needs to beneeds based to onbe basedin vivo onassays, in vivoin assays, vitro investigations in vitro investigations and even andtheoretical even theoretical calculations calculations can be valuable can be in valuable this regard. in this regard. For example, example, the the theoretical theoretical assessment assessment of of blood blood compatibility compatibility can can be bedone done by computing by computing the spreadingthe spreading work workof some of basic some blood basic cells blood and cellsprotei andns on proteins the surface on theof the surface polymeric of the material. polymeric This wasmaterial. applied This wasto appliedtwo PI tomaterials: two PI materials: a noncommercial a noncommercial film filmcontaining containing alicyclic alicyclic sequences, sequences, EPICLON-PPD (for its structure, see Table 2) and the conventional, aromatic Kapton film [20]. The theoretical study judiciously estimated a stronger non-adsorbent (of blood components) character and a subsequently more hemocompatible behavior for the aliphatic material as compared to the aromatic one. Even if further experimental confirmation of these results is needed, they are already sustained by the higher contact angle and lower polarizability values of the noncommercial film, which, according to the aforementioned observations, support the theoretical outcome. Other studies confirmed experimentally that various commercial PIs are hemocompatible, also underlying certain variability across formulations [30]. The discussion about surface biocompatibility needs more detail since the main bio-application of PI materials envisages implanting, and therefore they need to show a minimal effect on the body. Any implantation of a biomaterial (which has already been certified as leachables-free and overall non-toxic by standardized in vitro assays) triggers a mild physiological inflammatory response. This

Materials 2019, 12, 3166 14 of 27

EPICLON-PPD (for its structure, see Table2) and the conventional, aromatic Kapton film [ 20]. The theoretical study judiciously estimated a stronger non-adsorbent (of blood components) character and a subsequently more hemocompatible behavior for the aliphatic material as compared to the aromatic one. Even if further experimental confirmation of these results is needed, they are already sustained by the higher contact angle and lower polarizability values of the noncommercial film, which, according to the aforementioned observations, support the theoretical outcome. Other studies confirmed experimentally that various commercial PIs are hemocompatible, also underlying certain variability across formulations [30]. The discussion about surface biocompatibility needs more detail since the main bio-application of PI materials envisages implanting, and therefore they need to show a minimal effect on the body. MaterialsAny implantation 2019, 12, x FOR of PEER a biomaterial REVIEW (which has already been certified as leachables-free and overall15 of 28 non-toxic by standardized in vitro assays) triggers a mild physiological inflammatory response. reactionThis reaction starts starts with with immediate immediate protein protein adsorpti adsorptionon and and evolves evolves after after ro roughlyughly three three weeks weeks with macrophages and giant cells walling the implant with a fibrousfibrous capsule (Figure6 6))[ 17[17].].

Figure 6.6.The Thein vivo incomposite vivo composite reaction to syntheticreaction biomaterials:to synthetic timeline biomaterials: and monocyte timeline/macrophage and monocyte/macrophagephenotypes. Adapted from phenotypes. [17], with Adapted permission from from [17],© with2015 permission Elsevier. from © 2015 Elsevier.

This aggregate, chemical, chemical, physical, physical, and and biological biological process process is is summed summed up up as as the foreign-body responseresponse (FBR)(FBR) and and depends depends on manyon many factors, factors, from leachablesfrom leachables and endotoxins and endotoxins to size and to mechanical size and mechanicalfeatures of thefeatures implanted of the material implanted or itsmaterial biospecific or its interactions biospecific withinteractions its surroundings. with its surroundings. The ultimate “biocompatible”The ultimate “biocompatible” attestation of a attestation biomaterial of is givena biomaterial by an unsophisticated is given by an FBR unsophisticated resulting in a stable, FBR resultingnonadherent, in a stable, well-defined nonadherent, fibrotic well-defined tissue enclosing fibrotic the tissue implant. enclosing Of course, the implant. this capsule Of course, should this not capsuleharm the should implant not in harm any way,the implant and it could in any actually way, and prove it could a valuable actually asset, prove by supplying a valuable additional asset, by supplyingspatial fixation. additional Several spatial studies fixation. regarding Several the stud implantationies regarding (mostly the implantation focused on (mostly the peripheral focused and on thecentral peripheral nervous and system) central of nervous various system) PI-encapsulated of various devices PI-encapsulated evidenced device thats this evidenced type of that material this typeexerts of amaterial mild, quiescent exerts a mild, FBR quiescent (confined FBR to the (confi immediatened to the vicinity immediate of the vicinity material) of the and material) qualifies and as qualifiesbiocompatible as biocompatible [16,25,35–39 [16,25,35–39,42,43].,42,43].

4.3. Mechanical Mechanical Effects Effects As indicatedindicated earlier,earlier, there there are are several several mechanically-driven mechanically-driven interactions interactions between between the implant the implant and its andenvironment its environment which a whichffect the affect success the ofsuccess an implant, of an implant, and they and can bethey summed can be undersummed the under term structural the term structuralbiocompatibility biocompatibili [25]. Anyty mechanical [25]. Any mismatchmechanical between mismatch the hardbetween biomaterial the hard and biomaterial the soft target and tissue the softin terms target of tissue flexibility in terms/rigidity, of shapeflexibility/rigidity, (sometimes even shape texture) (sometimes or weight even leads texture) to micromovements, or weight leads cell, to micromovements,tissue, or material cell, irritation tissue, and or mate damage,rial irritation and, in the and end, damage, to implant and, in failure. the end, There to implant are three failure. types Thereof factors are responsiblethree types for of suchfactors a drawback. responsible First, for theresuch isa thedrawback. implant First, manufacturer, there is whichthe implant needs manufacturer,to make sure that which the needs implant to ismake not rubbingsure that or the irritating implant the is tissuenot rubbing by, e.g., or replacing irritating athe sharp-edged tissue by, e.g.,design replacing with a more a sharp-edged rounded one. design Second, with there a ismore the surgeonrounded performing one. Second, the operation,there is the which surgeon needs performing the operation, which needs to minimize such a rubbing-inflicted irritation by proper placement and anchoring [17]. Still, most of the responsibility for mechanical mismatch belongs to the polymeric material, through its mechanical and biological functionalities. Polyimidic materials are the prime polymeric materials of many implant-related applications (especially the demanding neural ones) since they provide a wide range of mechanical features, from easy flexibility to straight stiffness, and enable the facile construction of multilayered implantable devices. However, they are still prone to mechanical mismatch, by means of manufacturing, incompatible mechanical features, and deterioration induced by the biological media. For instance, mechanically-inflicted surface modification can alter the interaction of a PI material with living cells and plasma proteins. The rubbed surface of a noncommercial fluorinated PI (6FDA-6FAP, Table 2) displayed dissimilar hydrophobicity and roughness as compared to the starting material, which resulted in different biological behavior in terms of plasma protein adsorption, platelet adhesion, and fibroblast proliferation [52]. Rubbing proved an advantage in this particular case since the generated nano-ordered domains determined selective protein adsorption

Materials 2019, 12, 3166 15 of 27 to minimize such a rubbing-inflicted irritation by proper placement and anchoring [17]. Still, most of the responsibility for mechanical mismatch belongs to the polymeric material, through its mechanical and biological functionalities. Polyimidic materials are the prime polymeric materials of many implant-related applications (especially the demanding neural ones) since they provide a wide range of mechanical features, from easy flexibility to straight stiffness, and enable the facile construction of multilayered implantable devices. However, they are still prone to mechanical mismatch, by means of manufacturing, incompatible mechanical features, and deterioration induced by the biological media. For instance, mechanically-inflicted surface modification can alter the interaction of a PI material with living cells and plasma proteins. The rubbed surface of a noncommercial fluorinated PI (6FDA-6FAP, Table2) displayed dissimilar hydrophobicity and roughness as compared to the starting material, which resulted in different biological behavior in terms of plasma protein adsorption,

Materialsplatelet 2019 adhesion,, 12, x FOR and PEER fibroblast REVIEW proliferation [52]. Rubbing proved an advantage in this particular16 of case 28 since the generated nano-ordered domains determined selective protein adsorption and spheroidal andaggregation spheroidal of cells aggregation (Figure7). of The cells key (Figure point of 7). the Th studye key is thatpoint this of type the ofstudy mechanical is that modificationthis type of mechanicalhas a decisive modification impact on thehas propera decisive functioning impact on of the an implant.proper functioning of an implant.

Figure 7.7. Phase-contrastPhase-contrast images images of of fibroblast fibroblast cells cells at 48at h48 after h after cultured cultured on unrubbed on unrubbed (left) and (left) rubbed and rubbed(right) surface (right) ofsurface a synthetic of a synthetic fluorinated fluorinated polyimide polyimide (6FDA-6FAP). (6FDA-6FAP). Adapted from Adapted [53], withfrom permission [53], with permissionfrom © 2002 from Elsevier. © 2002 Elsevier.

In anotheranother case,case, the the inherent inherent mechanical mechanical characteristics characteristics of the of PI the material PI material do not do fully not comply fully comply with its within vivo itsintended in vivo use.intended For example, use. For the example, photosensitive the photosensitive Probimide 7520 Probim was testedide for7520 intracortical was tested neural for intracorticalimplants due neural to its proven implants biocompatibility due to its proven and flexibility.biocompatibility The latter and mitigates flexibility. tissue The irritation latter mitigates coming tissuefrom the irritation micromotion coming of afrom hard the PI materialmicromotion on the of brain’s a hard soft PI tissue material [69]. on However, the brain’s this flexibilitysoft tissue can [69]. be However,a disadvantage: this flexibility the PI-based can electrodebe a disadvantage: bends and overarchesthe PI-based during electrode implantation bends and and, overarches as a consequence, during implantationit cannot pass and, the brain’sas a consequence, pia mater. The it cannot structural pass stiffnessthe brain’s can pia be resolved,mater. The for structural example, stiffness by using can an beextra resolved, layer of for silicon, example, but this by willusing complicate an extra layer manufacturing, of silicon, but reinitiate this will biocompatibility complicate manufacturing, screening and reinitiateadd a new biocompatibility source of delamination screening or and degradation add a new [49 source]. of delamination or degradation [49]. In some cases, this behavior is just a case of a mechanicalmechanical mismatch between the implanted PI and the target tissue, which could be ruled out by selecting another PI material. As As a a counterexample counterexample to the above, another flexible flexible psPI, namely Durimide 7510, did not induce any negative mechanical effecteffect after 6 6 months months of of epiretinal epiretinal implantation implantation in in rabb rabbitit eyes eyes [41]. [41 ].In Inthis this case case,, the the flexibility flexibility of the of the PI (alsoPI (also doubled doubled by bya round-edged a round-edged design) design) helped helped to eliminate to eliminate any anymechanical mechanical compression compression on the on retinathe retina and andallowed allowed relatively relatively simple simple and convenient and convenient implantation, implantation, as it will as be it willdetailed be detailed in the section in the dedicatedsection dedicated to in vivo to experiments. in vivo experiments.

4.4. Manufacturing Manufacturing The construction process of any given implant, even even in in the early production stages of raw materials cancan alsoalso inflict inflict a diaff differenterent biocompatible biocompatible behavior. behavior. In the In case the of case PI films, of PI the films, most importantthe most important preparation step is the imidization of the material. This is usually performed by curing the film at temperatures ranging between 150 and 400 °C, depending on the structure or on the manufacturers’ indications for the commercial materials [1,58]. The cyclization temperature can have a major influence on the adhesion, proliferation, and morphology of certain cells and, by extension, on the FBR behavior, since PI reach their top chemical stability only if completely cured (the maximum possible cyclization eliminates or minimizes PAA residues and solvent traces). One experimental argument of this hypothesis was provided by the treatment of spin-coated conventional PI2611 precursor at two different temperatures, followed by autoclave sterilization and fibroblast populating experiments. The film cured at 200 °C showed a weak cellular adhesion and proliferation, while its counterpart cured at 400 °C displayed superior fibroblast adhesion and viability (Figure 8) coming from a more efficient elimination of chemical residues and/or solvent traces [27].

Materials 2019, 12, 3166 16 of 27 preparation step is the imidization of the material. This is usually performed by curing the film at temperatures ranging between 150 and 400 ◦C, depending on the structure or on the manufacturers’ indications for the commercial materials [1,58]. The cyclization temperature can have a major influence on the adhesion, proliferation, and morphology of certain cells and, by extension, on the FBR behavior, since PI reach their top chemical stability only if completely cured (the maximum possible cyclization eliminates or minimizes PAA residues and solvent traces). One experimental argument of this hypothesis was provided by the treatment of spin-coated conventional PI2611 precursor at two different temperatures, followed by autoclave sterilization and fibroblast populating experiments. The film cured at 200 ◦C showed a weak cellular adhesion and proliferation, while its counterpart cured at 400 ◦C displayed superior fibroblast adhesion and viability Materials(Figure 82019) coming, 12, x FOR from PEER a moreREVIEW e ffi cient elimination of chemical residues and/or solvent traces17 [27 of]. 28

Figure 8. Scanning electron microscopy images (500 , 2500 ) showing the viability and adherence Figure 8. Scanning electron microscopy images (500×,× 2500×)× showing the viability and adherence of L929of L929 fibroblast fibroblast cells cells at at24 24h after h after cultured cultured on on the the su surfacesrfaces of of conventional conventional PI2611 PI2611 cured cured at at 200 200 °C◦C (top) (top) and at 400 °C◦C (bottom). Reproduced Reproduced from from [27], [27], with permission from © 20082008 John John Wiley Wiley and and Sons. Sons. The same conclusion was drawn by curing a commercial psPI, PI2711, at two different temperatures, The same conclusion was drawn by curing a commercial psPI, PI2711, at two different 200 C (Figure5B,E,H) and 350 C (Figure5C,F,I), respectively, and observing some variations in the temperatures,◦ 200 °C (Figure 5B,E,H)◦ and 350 °C (Figure 5C,F,I), respectively, and observing some adhesion and proliferation of BHK-21 fibroblasts, along small changes in the surface free energy and variations in the adhesion and proliferation of BHK-21 fibroblasts, along small changes in the surface zeta potential (and surprisingly constant contact angle values). free energy and zeta potential (and surprisingly constant contact angle values). The straightforward dependence between imidization temperatures and biological behavior The straightforward dependence between imidization temperatures and biological behavior was also observed in the case of a fluorinated PI (6FDA-6FAP; Figure9). In this particular instance, was also observed in the case of a fluorinated PI (6FDA-6FAP; Figure 9). In this particular instance, protein adsorption, neutrophil adhesion, and complement activation are inversely proportional protein adsorption, neutrophil adhesion, and complement activation are inversely proportional to to temperature, showing better values at lower cyclization temperatures. This behavior is determined temperature, showing better values at lower cyclization temperatures. This behavior is determined by a high temperature-induced rearrangement of molecular fragments at the outermost surface, by a high temperature-induced rearrangement of molecular fragments at the outermost surface, which results in low surface energy and high hydrophobicity [53]. which results in low surface energy and high hydrophobicity [53]. Further studies employing other PI materials are needed in order to fully grasp the effect of imidization temperature on their micro- and macro-molecular structure, surface, hydrophobicity, and subsequent biocompatible behavior. From another point of view, it must be underlined that many of the results discussed in the last two subsections were obtained in static conditions. The broad range of flow conditions implied by tissue- and blood-contact devices brings additional variables and unknowns to the issue and protein

Materials 2019, 12, x FOR PEER REVIEW 17 of 28

Figure 8. Scanning electron microscopy images (500×, 2500×) showing the viability and adherence of L929 fibroblast cells at 24 h after cultured on the surfaces of conventional PI2611 cured at 200 °C (top) and at 400 °C (bottom). Reproduced from [27], with permission from © 2008 John Wiley and Sons.

The same conclusion was drawn by curing a commercial psPI, PI2711, at two different temperatures, 200 °C (Figure 5B,E,H) and 350 °C (Figure 5C,F,I), respectively, and observing some variations in the adhesion and proliferation of BHK-21 fibroblasts, along small changes in the surface free energy and zeta potential (and surprisingly constant contact angle values). The straightforward dependence between imidization temperatures and biological behavior was also observed in the case of a fluorinated PI (6FDA-6FAP; Figure 9). In this particular instance, proteinMaterials 2019adsorption,, 12, 3166 neutrophil adhesion, and complement activation are inversely proportional17 of to 27 temperature, showing better values at lower cyclization temperatures. This behavior is determined byadsorption, a high temperature-induced cellular adhesion, or rearrangement complement activationof molecular may fragments present a at significant the outermost variation surface, from whichthe static results experiments. in low surface energy and high hydrophobicity [53].

Figure 9. The amount of three adsorbed proteins on the surfaces of the same 6FDA-6FAP fluorinated polyimide membrane cured at three different temperatures (pH 7.4; (PSt) control). Reproduced from [53], with permission from © 2002 John Wiley and Sons.

Further, the manufacturing stages of implantable bio-MEMS, for example, need to reduce any source of leachables and minimize their toxicology. Once again, PIs are quite resistant to micromachining techniques, so the toxicity risk in this regard is minor but still noteworthy. However, these well-known general features of PIs do not exclude the in-depth physicochemical characterization of any given PI-based material and of its final products (a biocompatible PI does not guarantee a biocompatible PI-based device). Special attention must be dedicated to any modification in the composition and properties of the PI material which could arise from hydrolytic or enzymatic degradation during its in vivo employment, especially since it is the case of devices conceived for long-term implantation.

5. Biocompatibility Screening of Polyimides by In Vitro Methods Cytotoxicity evaluations represent the first stage of the biocompatibility assessment of any given biomaterial and they enable qualitative and quantitative evaluations of the possible biological risk of their usage. Despite their specificity shortage, these tests represent the most suitable, inexpensive, reliable and, very important, reproducible means to review the biological response induced at the cellular level and to simulate, with some limitations, in vivo circumstances. Besides the available standards and regulations, the selection of cells and cellular models employed in these studies is (or should be) determined by the envisaged outcome. Usually, the basic cytotoxicity screening is performed on common cultures of isolated, immortalized cells (like L929, HeLa, 3T3). The more thorough cytocompatibility investigations use specific cell lines (like animal or human endothelial, epithelial cells), according to the foreseen biological interactions and medical usage. Combined assays of both types, either in direct or indirect contact mode, can deliver a more accurate image of a material’s cytotoxicity and should be applied to complement its physicochemical evaluation. Even if PIs are not yet certified by ISO standards, there are several research papers in which their low, if any, cytotoxicity is demonstrated. Some of the most solid ones are being cited in this section, while an example of a successful cell viability assay is presented in Figure 10. However, the available scientific literature on the topic provides a wide range of methods and cell lines, and an Materials 2019, 12, 3166 18 of 27 explicit definition of assessment conditions is needed in order to perform any reliable interpretation or comparison of the assay’s results [17,18,57]. For instance, the first systematic in vitro evaluation of commercial polyimidic products dates back to 1993 and does not follow the currently available ISO guidelines, but another set of prior, nevertheless trustworthy standards [15]. Even if this is deservedly one of the most cited papers when PIs are described as “biocompatible”, any judgment based on it or Materialscomparison 2019, 12 with, x FOR the PEER current REVIEW guidelines is not reasonable. 19 of 28

Figure 10. Cell viability assay examination of BHK-21 fibroblast fibroblast cells at 24 h post-culture on PI2525 polyimide: ( (AA)) live live (viable: (viable: green green color)/dead color)/dead (non-viable: (non-viable: red red color) color) staining staining of ofcells cells on on the the surface surface of PI2525;of PI2525; (B) ( Bpositive) positive control control (latex (latex rubber). rubber). Scale Scale bar: bar: 200 200 µm.µm. Reproduced Reproduced from from [28], [28], with with permission permission from © 20102010 Elsevier. Elsevier.

Various commercial and noncommercial PI materials were proven as non-toxic by a multitude of in vitrovitro,, direct, oror indirect,indirect, cytotoxicitycytotoxicity assays. Most of them specificallyspecifically declares to comply with the ISO guidelines, and investigate the the viability, proliferation, degeneration, and and lysis lysis of of different different common cell types: • L929 mouse fibroblast cells (Kapton, PI2611, Durimide 7020, Durimide 7510, PI2525, L929 mouse fibroblast cells (Kapton, PI2611, Durimide 7020, Durimide 7510, PI2525, 6FDA-6FAP, • 6FDA-6FAP, EPICOLN-PPD) [27,41,44,48,51,52], EPICOLN-PPD) [27,41,44,48,51,52], • 3T3 mouse fibroblasts cells (Probimide 7520) [49], 3T3 mouse fibroblasts cells (Probimide 7520) [49], • OP9 mouse stromal cells (Neopulim) [19], OP9 mouse stromal cells (Neopulim) [19], • rat primary Schawnn cells (Kapton, PI2611, Durimide 7020) [27], rat primary Schawnn cells (Kapton, PI2611, Durimide 7020) [27], • rat astroglial cells (unknown PI trade name) [70], • rat astroglial cells (unknown PI trade name) [70], • bovine endothelial cells (unknown PI trade name) [70], • bovine endothelial cells (unknown PI trade name) [70], • BHK-21 baby hamster kidney fibroblasts (PI2525, PI2771) [28], BHK-21or more babyspecific hamster cell lines: kidney fibroblasts (PI2525, PI2771) [28], • • human retinal pigment cells (PI2525) [47], or more specific cell lines: • AGSE human gastric adenocarcinoma cells (unknown PI trade name) [70], • human retinalmesenchymal pigment stem cells cells (PI2525) (Kapton) [47], [20], • • AGSEHeLa, human gastricepithelial adenocarcinoma cells (6FDA-6FAP) cells (unknown[54], PI trade name) [70], • • humanSV-HCEC mesenchymal human endothelial stem cells cells (Kapton) (Kapton, [20 PI2610,], HD 3007) [30], • HeLa,human human dermal epithelial fibroblasts cells (SPAB-xxDA) (6FDA-6FAP) [55]. [54 ], • SV-HCEC human endothelial cells (Kapton, PI2610, HD 3007) [30], • The low cytotoxic outcome of these tests and the well-established PIs’ surface treatment methodshuman were dermal further fibroblasts combined (SPAB-xxDA) to attain an [interesting55]. controlled positioning of living cells along • within a given pattern [70]. A KrF laser-based technique was employed to garnish the surface of a The low cytotoxic outcome of these tests and the well-established PIs’ surface treatment methods commercial psPI with an extra carbon layer and to manipulate its interaction with various cell lines were further combined to attain an interesting controlled positioning of living cells along within (Figure 11). For example, bovine endothelial cells proved quite immune to this surface modification, a given pattern [70]. A KrF laser-based technique was employed to garnish the surface of a commercial while rat astroglial cell lines showed guided adherence and proliferation by avoiding the psPI with an extra carbon layer and to manipulate its interaction with various cell lines (Figure 11). nonadherent carbon layer, an observation which can prove very helpful for further cellular For example, bovine endothelial cells proved quite immune to this surface modification, while rat culture-related applications. astroglial cell lines showed guided adherence and proliferation by avoiding the nonadherent carbon layer, an observation which can prove very helpful for further cellular culture-related applications.

Materials 2019, 12, x FOR PEER REVIEW 19 of 28

Figure 10. Cell viability assay examination of BHK-21 fibroblast cells at 24 h post-culture on PI2525 polyimide: (A) live (viable: green color)/dead (non-viable: red color) staining of cells on the surface of PI2525; (B) positive control (latex rubber). Scale bar: 200 µm. Reproduced from [28], with permission from © 2010 Elsevier.

Various commercial and noncommercial PI materials were proven as non-toxic by a multitude of in vitro, direct, or indirect, cytotoxicity assays. Most of them specifically declares to comply with the ISO guidelines, and investigate the viability, proliferation, degeneration, and lysis of different common cell types: • L929 mouse fibroblast cells (Kapton, PI2611, Durimide 7020, Durimide 7510, PI2525, 6FDA-6FAP, EPICOLN-PPD) [27,41,44,48,51,52], • 3T3 mouse fibroblasts cells (Probimide 7520) [49], • OP9 mouse stromal cells (Neopulim) [19], • rat primary Schawnn cells (Kapton, PI2611, Durimide 7020) [27], • rat astroglial cells (unknown PI trade name) [70], • bovine endothelial cells (unknown PI trade name) [70], • BHK-21 baby hamster kidney fibroblasts (PI2525, PI2771) [28], or more specific cell lines: • human retinal pigment cells (PI2525) [47], • AGSE human gastric adenocarcinoma cells (unknown PI trade name) [70], • human mesenchymal stem cells (Kapton) [20], • HeLa, human epithelial cells (6FDA-6FAP) [54], • SV-HCEC human endothelial cells (Kapton, PI2610, HD 3007) [30], • human dermal fibroblasts (SPAB-xxDA) [55]. The low cytotoxic outcome of these tests and the well-established PIs’ surface treatment methods were further combined to attain an interesting controlled positioning of living cells along within a given pattern [70]. A KrF laser-based technique was employed to garnish the surface of a commercial psPI with an extra carbon layer and to manipulate its interaction with various cell lines (Figure 11). For example, bovine endothelial cells proved quite immune to this surface modification, while rat astroglial cell lines showed guided adherence and proliferation by avoiding the nonadherentMaterials 2019, 12 ,carbon 3166 layer, an observation which can prove very helpful for further cellular19 of 27 culture-related applications.

Materials 2019, 12, x FOR PEER REVIEW 20 of 28

Figure 11. Optical micrographs of two cell lines on the surface of a polyimide sample with carbon Figurepatterns: 11. (aOptical) carbon micrographs layer pattern; of two(b) rat cell astroglial lines on thecells; surface (c) bovine of a polyimideendothelial sample cells. withReproduced carbon patterns:from [70], (a with) carbon permission layer pattern; from ( b©) 2008 rat astroglial Springer cells; Nature. (c) bovine endothelial cells. Reproduced from [70], with permission from © 2008 Springer Nature. 6. Biocompatibility Screening of Polyimides by in vivo Methods 6. Biocompatibility Screening of Polyimides by In Vivo Methods In vivo screening is the next level of biocompatibility testing, the critical step in the evaluation of theIn biological vivo screening response is the generated next level by of biocompatibilitya material. While testing, well developed the critical and step comprehensive, in the evaluation the of the in biologicalvitro methodology response generatedis not able by to a material.accurately While model well many developed of the and aspects comprehensive, implied by the a inclinical vitro methodologybiological reaction. is not ableThe tolatter accurately brings modeladditional many hurdles of the aspectslike high implied controversy, by a clinical ethical biological issues, reaction.premium Thecosts, latter cumbersome brings additional bureaucratic hurdles challenges, like high etc. controversy, [57]. ethical issues, premium costs, cumbersomeSeveral commercial bureaucratic PIs challenges, have been etc. the [ 57subject]. of in-depth in vivo studies that authenticated their compatibilitySeveral commercial with living PIsbiological have beenorganisms. the subject Most ofof in-depththe reachablein vivo clinicalstudies trials that involved authenticated defined theirpolyimide compatibility structures with listed living in biologicalTables 1 and organisms. 2 (Kapton, Most Durimide of the reachable 7050, Durimide clinical trials 7510, involved PI2611, definedPI2525, polyimideU-Varnish structuresS) but also listed a few in unknown Tables1 and co2mmercial (Kapton, DurimidePIs which 7050,are simply Durimide and 7510, frustratingly PI2611, PI2525,named “polyimides”. U-Varnish S) butThis also review a few is unknownnot intended commercial to reproduce PIs whichor castigate are simply the conclusions and frustratingly of these namedarticles, “polyimides”. but to underline This some review of the is notmost intended spectacula tor reproduce and meaningful or castigate cases theof PI’s conclusions in vivo testing of these as articles,the ultimate butto and underline undeniable some proof of the ofmost their spectacularbiocompatibility. and meaningful cases of PI’s in vivo testing as the ultimateThe first and example undeniable in this proof regard of theirdeals biocompatibility.with a flexible commercial cPI, PI252, which was used to buildThe the first components example in of this an regard artificial deals retina with awith flexible the commercialaim to stimulate cPI, PI252, retinal which neurons was used in cases to build of thedegenerated components photoreceptors of an artificial retina[47]. withIts thein aimvivo to stimulatebiocompatibility retinal neurons was intested cases ofat degenerated12 weeks photoreceptorspost-implantation [47]. in Its rabbitin vivo eyes,biocompatibility as a free-floati wasng tested and a at fixed 12 weeks implant post-implantation (Figure 12). Both in rabbit of them eyes, asproved a free-floating good andbiological a fixed implantand mechanical (Figure 12). Bothstability of them and proved no goodinflammatory biological andresponse, mechanical the stabilityelectroretinogram and no inflammatory and microscopic response, investigations the electroretinogram not being able and to find microscopic any disparities investigations between notthe beinghealthy able and to the find transplanted any disparities eye. between the healthy and the transplanted eye.

Figure 12. InIn vivo vivo biocompatibilitybiocompatibility tests tests performed performed on on a polyimide a polyimide (PI2525) (PI2525) microelectrode microelectrode array array (3 × (33 mm,3 mm 18 ,µm 18 µthickm thick polyimide polyimide support) support) for retinal for retinal stimulation: stimulation: (a) fixed (a) fixedpolyimide-based polyimide-based array arrayat 12 × atweeks 12 weeks after afterimplantation implantation in rabbit in rabbit eyes; eyes; (b) ( bfree-floating) free-floating polyimide-based polyimide-based array array at at 12 12 weeks weeks afterafter implantationimplantation inin rabbitrabbit eyes.eyes. Adapted from [[47],47], with permissionpermission fromfrom ©© 2004 Elsevier.

As a complement of the former, thethe secondsecond exampleexample ofof successfulsuccessful PIPI implantationimplantation is related to a commercialcommercial psPIpsPI (Durimide (Durimide 7510) 7510) used used in the in preparation the preparation of MEMS of for MEMS epiretinal for electricalepiretinal stimulation. electrical Itsstimulation. reliability andIts reliability stability were and assessed stability by were chronic assessed 6-month by implantationchronic 6-month in rabbit implantation eyes, which in showed rabbit aeyes, nonirritant, which showed biologically a nonirritant, safe PI-based biologically device with safe high PI-based biocompatibility device with and high no biocompatibility mechanical pressure and orno mismatch.mechanical The pressure study or details mismatch. the lack The of cytotoxicitystudy details of the the lack thin of PI cytotoxicity sheet, which of didthe notthin bring PI sheet, any injurywhich ordid modifications not bring any to injury the inner or modifications retina or its layers, to the as inner it can retina be observed or its layers, in Figure as it 13cana–c. be Moreover,observed in Figure 13a–c. Moreover, the PI material was immune to sterilization and the harsh biological environment, and, also important, no delamination was noticed 6 months after the implantation (Figure 13d) [41].

Materials 2019, 12, 3166 20 of 27 the PI material was immune to sterilization and the harsh biological environment, and, also important, noMaterials delamination 2019, 12, x FOR was PEER noticed REVIEW 6 months after the implantation (Figure 13d) [41]. 21 of 28

Figure 13. LongLong term term inin vivo biocompatibility evaluation of a microelectrode array based on photosensitive Durimide 7510 polyimide layer implanted withinwithin aa rabbitrabbit eye:eye: (a) phase-contrastphase-contrast image (400(400×)) of of the inner retinal morphology at 6 weeks post-implantationpost-implantation (RGC: retinal ganglion × cell layer; ILM: inner limiting membrane); ( b) light microscopemicroscope image (400(400×)) of the inner retinal × morphology at 6 months post-implantation; (c) light microscopemicroscope imageimage (400(400×)) of the control rabbit × retina; ( d) SEM micrograph of the undamaged implanted electrode electrode at at 6 months post-operation. Reproduced from [[41],41], with permissionpermission fromfrom ©© 2013 BioMed Central, under CC BY 2.0.

The mechanical mismatch issue encountered in ne neuroprostheticsuroprosthetics may find find a solution by using electrospinned PIPI nanofibers nanofibers instead instead of castedof casted PI films.PI films. A neural A neural electrode electrode based based on such on highly such flexiblehighly andflexible permeable and permeable polyimidic polyimidic nanofibers nanofibers provides aprovides smaller contact a smaller area contact and reduced area and neural reduced compression neural (Figurecompression 14). In(Figure exchange, 14). In these exchange, features these enable features a decrease enable in a decrease the shrinkage in the of shrinkage the nerve of tissue the nerve and long-termtissue and (12 long-term weeks), steady(12 weeks), recordings steady of neuralrecordings signals of [neural36]. Moreover, signals [36]. biologically Moreover, active biologically principles canactive be principles loaded on can such be nanofibers loaded on to such enhance nanofibers its affinity to enhance with the its sciatic affinity nerve with tissue the sciatic or to pave nerve the tissue way foror to very pave interesting the way for neural very drug-delivery interesting neural applications. drug-delivery applications.

Materials 2019, 12, 3166 21 of 27 Materials 2019, 12, x FOR PEER REVIEW 22 of 28

FigureFigure 14.14. HistologicalHistological imagesimages ofof nanofiber,nanofiber, polyimidepolyimide (PI1338)-based(PI1338)-based electrodeselectrodes afterafter implantationimplantation inin thethe sciaticsciatic nerve of of a a rat: rat: (a (a) )pristine pristine electrode electrode straight straight after after implantation implantation (A (,AB,)B magnifications) magnifications of ofthe the contact contact area area between between tissue tissue and and electrode); electrode); (b) ( bpristine) pristine electrode electrode at 4 at days 4 days post-implantation; post-implantation; (c) (electrodec) electrode with with small small holes holes at at4 4days days post-implantation; post-implantation; (d (d) )electrode electrode with with large large holes holes at 44 daysdays post-implantation.post-implantation. ReproducedReproduced fromfrom [[36],36], withwith permissionpermission fromfrom ©© 2017 AmericanAmerican ChemicalChemical Society.Society. At this point, we must note that the same cPI material steps forward from the handful of in vivo At this point, we must note that the same cPI material steps forward from the handful of in vivo tested polyimidic materials: the BPDA-PPD macromolecular architecture. Most likely known tested polyimidic materials: the BPDA-PPD macromolecular architecture. Most likely known by the by the trade names of its liquid precursors, PI2611 and U-Varnish-S, this film was heavily tested as trade names of its liquid precursors, PI2611 and U-Varnish-S, this film was heavily tested as a a passive component in a broad range of applications. Its uses span from retina stimulation and blood passive component in a broad range of applications. Its uses span from retina stimulation and blood sensing to cortical recordings, phantom pain relief, and intra-neural stimulating electrodes for the sensing to cortical recordings, phantom pain relief, and intra-neural stimulating electrodes for the peripheral and central nervous systems [25]. peripheral and central nervous systems [25]. Perhaps the most tremendous example of in vivo biocompatibility of a PI-based material comes Perhaps the most tremendous example of in vivo biocompatibility of a PI-based material comes from an intraneural flexible electrode built on this commercial PI, tf-LIFE4, which was implanted in a from an intraneural flexible electrode built on this commercial PI, tf-LIFE4, which was implanted in human amputee with the aim of controlling a robotic hand (Figure 15)[33]. After a 4-week in vivo a human amputee with the aim of controlling a robotic hand (Figure 15) [33]. After a 4-week in vivo test period (as imposed by European Authorities), the implant proved biocompatible and stable even test period (as imposed by European Authorities), the implant proved biocompatible and stable when the patient was performing everyday life activities. However, a fibrotic tissue reaction was even when the patient was performing everyday life activities. However, a fibrotic tissue reaction detected by visual inspection during the removal of the device, but ethical reasons did not allow its was detected by visual inspection during the removal of the device, but ethical reasons did not allow histological confirmation. The device itself and the cited study signify a real breakthrough in the field its histological confirmation. The device itself and the cited study signify a real breakthrough in the of implantable robotics for amputees and also open great opportunities for polyimidic materials. field of implantable robotics for amputees and also open great opportunities for polyimidic There are also other available examples of intraneural flexible electrodes based on PI materials, materials. with TIME and SELINE flexible electrodes being the main competitors of tf-LIFE. The main difference between the three is the direction in which they are implanted and the three-dimensional geometry of the device. In all cases, the PI2611 polyimide is the material of choice for insulation and mechanical support attainable through standard photolithographic techniques. The available studies concur that this polyimidic material provides a good compromise between biocompatibility, mechanical features, and insulating efficacy [27,39,46]. All these electrodes showed promising results in functionality, selectivity, invasiveness, and biocompatibility studies performed on animal models and human volunteers. The main application limits are related to the overall device design, mechanical anchorage to the nerve, and post-implantation stability. Further research is needed to improve the stability of the electrodes for acute and chronic studies and to comparative asses the devices’ chronic functionality. This type of study will also contribute greatly to the currently reduced body of knowledge related to the long-term stability of polyimides in vivo conditions. Materials 2019, 12, 3166 22 of 27 Materials 2019, 12, x FOR PEER REVIEW 23 of 28

Figure 15. Surgical implantation of of a t tf-LIFE4f-LIFE4 intraneural electrode implant based on a polyimide (PI2611) substrate substrate (10 (10 µmµm in in thic thicknesskness and and 5 5cm cm in in length). length). (A ()A A) Amagnified magnified photographic photographic view view of implantof implant components components during during surgical surgical implantation; implantation; (B (B) )schematic schematic illustration illustration of of the the overall overall tf-LIFE4 implant system; ( C) photographic view median nerve, tf-LIF tf-LIFE4E4 electrode and cable during surgical implantation; ( (D)) external transit cables after implantation.implantation. Reproduced Reproduced from from [33], [33], with permission from © 20102010 Elsevier. Elsevier. 7. Conclusions and Future Directions There are also other available examples of intraneural flexible electrodes based on PI materials, with BiocompatibilityTIME and SELINE assessment flexible of electrodes a polyimidic bein materialg the designedmain competitors for the medical of tf-LIFE. field must The comply main withdifference a composite between enterprise the three of is many the direction variables, in from which the chemicalthey are implanted nature or physical and the characteristics three-dimensional of the geometryenvisaged of PI the to thedevice. tissue In contactall cases, span the orPI2611 the nature polyimide of that is giventhe material tissue. Aof judiciouschoice for screening insulation order and mechanicalshould start support with an in-depthattainable evaluation through ofstandard all available photolithographic data, followed techniques. by the integration The available of chemical studies and concurphysical that investigations, this polyimidic basic andmaterial specific provides in vitro screening,a good compromise and legitimate between in vivo characterization.biocompatibility, mechanicalBiomedical features, applications and insulating are more demandingefficacy [27,39,46]. than ever All for novel,these biocompatible,electrodes showed skillful promising materials whichresults canin functionality, simultaneously selectivity, provide invasiveness, lightweight, and superior biocompatibility mechanical studies features performed and uncomplicated on animal modelsmanufacturing. and human PIs resolvevolunteers. many The of main these application requirements limits by anare unparalleled related to the combination overall device of inherent design, mechanicalcharacteristics, anchorage ease of to tuning, the nerve, facile and production, post-implantation and manifold stability. utility. Further Several research commercial is needed and to improvenoncommercial the stability materials of the developed electrodes from for polyimidicacute and chronic building studies blocks and have to been comparative severely testedasses the by devices’both in vitro chronicand infunctionality. vivo methods Th inis the type last of two study decades will which also leadcontribute to a quite greatly impressive, to the yetcurrently mostly reduceddisconnected body bodyof knowledge of knowledge. related Evento the if long-term PIs are not stability yet certified of polyimides by ISO standards,in vivo conditions. a respectable amount of research authenticate their non-toxic nature in vitro and demonstrate cell viability and 7.proliferation Conclusions of and diff erentFuture common Directions cell types on various PI materials. Moreover, a small number of commercially-available PIs were labeled as “biocompatible” by in-depth, acute, and chronical in vivo Biocompatibility assessment of a polyimidic material designed for the medical field must studies on animal models and, in few cases, even on human volunteers. comply with a composite enterprise of many variables, from the chemical nature or physical The positive results provided by these scrupulous investigations enable a wide exploration characteristics of the envisaged PI to the tissue contact span or the nature of that given tissue. A of polyimides in various biomedical applications, from micropatterned cell substrates and blood judicious screening order should start with an in-depth evaluation of all available data, followed by sensing to cortical recordings, retina stimulation, and intra-neural stimulating electrodes for the the integration of chemical and physical investigations, basic and specific in vitro screening, and peripheral and central nervous systems. legitimate in vivo characterization. It is safe to reason that commercial or synthetic PI materials are far from reaching their peak usage Biomedical applications are more demanding than ever for novel, biocompatible, skillful in medical devices and implants. On one side, many studies are dedicated to providing tailored PIs materials which can simultaneously provide lightweight, superior mechanical features and uncomplicated manufacturing. PIs resolve many of these requirements by an unparalleled

Materials 2019, 12, 3166 23 of 27 specially designed for the biomedical area since the demand for unique implants can be also met through polyimidic materials. Moreover, recent studies report physically or chemically modified and composite PIs for the same type of applications. On the other hand, intense research is focused on improving the functionality and stability of medical devices based on the “classic”, commercial PIs described herein. Despite all this, it is still difficult to discern which PI-based material works best for different biomedical usages. An outlook into the future of PIs in the biomedical area can follow multiple directions. First, more studies are needed to access a straight line between the chemical blueprint of a polyimidic material and its biocompatibility. The data presented here come close to a basis for tailoring PIs to achieve in vitro biocompatibility. Even so, correlations between important factors like curing temperature, interfacial free energy, or molecular weight and biocompatibility are still unreachable. Additionally, further research dealing with any potential cytotoxicity of aged PIs is needed. Second, PI-based medical devices are far from clinical controlled performances. Further in vivo assays of recognized or new PI materials and their outcomes’ in-depth comparisons could be a first step in solving this issue. The in vivo studies should also address the long-term chemical and biological inertness of PIs as the main prerequisite of medical devices’ biocompatibility. There is a lack of data related to changes in the chemical compositions, mass loss, and macroscopic behavior of PIs after long-term in vivo employment. At the same time, the in-depth analysis of specific tissue responses and FBRs triggered by PIs is required. These missing pieces in the body of knowledge related to PIs are a first limit to their expansion in the biomedical market. A rule of thumb for any of these biocompatibility assays should be the use of validated and standardized test methods. Otherwise, any applicable, comparable, and conclusive results would be out of reach. In another direction, polyimidic materials alone or intraneural electrodes based on them are still prone to mechanical mismatch, a defect which hampers the development of neuroprosthetics. While the latter is a matter of device design, new PIs must be tested to mitigate the material-related difficulties. Another issue to be addressed in this area is the water uptake of some PIs, especially of the psPI type. Moisture adsorption causes stretching and has a plasticizing effect which limits their application. Finally, the delamination of metals from PIs remains a topic of concern for the long-term durability of bio-MEMS. The main drive of research in the field of polyimides will logically remain focused on novel, polyimidic materials of superior performance. However, special interest must be directed towards their ability to make it even further in the biomedical area, with proper (especially in vivo) biocompatibility being the first item on a long checklist. The ability of polyimides to tolerate or even adapt to the circumstances created by a living environment is the pivotal element in the production of stable, reliable medical devices destined to long-term, nontoxic usage. A couple of success stories cited herein pave the way for more intensive research focused on the biocompatibility of old or new polyimides as to further improve their bio-performance and safety and to unlock unmet biomedical possibilities and applications. Nevertheless, an undeniable change has occurred in the way polyimides are regarded by the biomedical field and the concepts of “biocompatibility” and “polyimides” are more congruent than ever.

Author Contributions: Conceptualization, R.D.R. and M.A.; methodology, R.D.R. and M.A.; investigation, R.D.R., C.P.C. and R.F.D.; resources, C.P.C. and R.F.D.; Writing—Original draft preparation, R.D.R., C.P.C. and R.F.D.; Writing—Review and Editing, R.D.R., C.P.C. and M.A.; visualization, R.F.D.; project administration, R.D.R.; funding acquisition, M.A. Funding: The authors acknowledge the financial support of this research through the Project “Partnerships for knowledge transfer in the field of polymer materials used in ”, ID P_40_443, Contract No. 86/8.09.2016, SMIS 105689, co-financed by the European Regional Development Fund by the Competitiveness Operational Programme 2014–2020, Axis 1. Research, Technological Development and Innovation in support of economic competitiveness and business development, Action 1.2.3. Knowledge Transfer Partnerships. Conflicts of Interest: The authors declare no conflict of interest. Materials 2019, 12, 3166 24 of 27

References

1. Liaw, D.-J.; Wang, K.-L.; Huang, Y.-C.; Lee, K.-R.; Lai, J.-Y.; Ha, C.-S. Advanced polyimide materials: Syntheses, physical properties and applications. Prog. Polym. Sci. 2012, 37, 907–974. [CrossRef] 2. Bryant, R.G. Polyimides. Ullmann’s Encycl. Ind. Chem. 2014, 1–27. [CrossRef] 3. Teo, A.J.T.; Mishra, A.; Park, I.; Kim, Y.-J.; Park, W.-T.; Yoon, Y.-J. Polymeric Biomaterials for Medical Implants and Devices. ACS Biomater. Sci. Eng. 2016, 2, 454–472. [CrossRef] 4. Wang, Y.; Yang, T.; Hasebe, Y.; Zhang, Z.; Tao, D. Carbon Black-Carbon Nanotube Co-Doped Polyimide Sensors for Simultaneous Determination of Ascorbic Acid, Uric Acid, and Dopamine. Materials (Basel) 2018, 11, 1691. [CrossRef][PubMed] 5. Shao, D.; Xu, C.; Wang, H.; Du, J. Enhancing the Dyeability of Polyimide Fibers with the Assistance of Swelling Agents. Materials (Basel) 2019, 12, 347. [CrossRef][PubMed] 6. Bruma, M.; Damaceanu, M.-D.; Rusu, R.-D. Study of thin films made from aromatic polymers containing six-member imide rings. High Perform. Polym. 2012, 24, 31–39. [CrossRef] 7. Georgiev, A.; Dimov, D.; Spassova, E.; Assa, J.; Dineff, P.; Danev, G. Chemical and Physical Properties of Polyimides: Biomedical and Engineering Applications. In High Performance Polymers—Polyimides Based—From Chemistry to Applications; Abadie, M., Ed.; InTechOpen: London, UK, 2012. [CrossRef] 8. Zhao, Q.; Whittaker, A.K.; Zhao, X.S. Polymer Electrode Materials for Sodium-ion Batteries. Materials (Basel) 2018, 11, 2567. [CrossRef][PubMed] 9. Damaceanu, M.-D.; Rusu, R.-D.; Musteata, V.-E.; Bruma, M. Dielectric Behavior of Thin Films made from poly(oxadiazole-naphthylimide)s. Soft Mater. 2010, 9, 44–63. [CrossRef] 10. Rusu, R.D.; Damaceanu, M.D.; Bruma, M.; Ronova, I.A. Effect of conformational parameters on thermal properties of some poly (oxadiazole-naphthylimide) s. Iran. Polym. J. 2011, 20, 29–40. 11. Hergenrother, P.M. The Use, Design, Synthesis, and Properties of High Performance/High Temperature Polymers: An Overview. High Perform. Polym. 2003, 15, 3–45. [CrossRef] 12. Rusu, R.D.; Damaceanu, M.D.; Constantin, C.P. Intelligent Amide- and Imide-Based Polymeric Materials for Biomedical Applications; Aflori, M., Ed.; CRC Pres: Boca Raton, FL, USA, 2018; pp. 167–190. 13. Williams, D.F. There is no such thing as a biocompatible material. Biomaterials 2014, 35, 10009–10014. [CrossRef][PubMed] 14. Asghari, F.; Samiei, M.; Adibkia, K.; Akbarzadeh, A.; Davaran, S. Biodegradable and biocompatible polymers for tissue engineering application: A review. Artif. Cells Nanomed. Biotechnol. 2017, 45, 185–192. [CrossRef] [PubMed] 15. Richardson, R.R.; Miller, J.A.; Reichert, W.M. Polyimides as biomaterials: Preliminary biocompatibility testing. Biomaterials 1993, 14, 627–635. [CrossRef] 16. Ordonez, J.; Schuettler, M.; Boehler, C.; Boretius, T.; Stieglitz, T. Thin films and microelectrode arrays for neuroprosthetics. MRS Bull. 2012, 37, 590–598. [CrossRef] 17. Ratner, B.D. Chapter 3—The Biocompatibility of Implant Materials. In Host Response to Biomaterials; Badylak, S.F., Ed.; Academic Press: Oxford, UK, 2015; pp. 37–51. [CrossRef] 18. Bernard, M.; Jubeli, E.; Pungente, M.D.; Yagoubi, N. Biocompatibility of polymer-based biomaterials and medical devices—Regulations, in vitro screening and risk-management. Biomater. Sci. 2018, 6, 2025–2053. [CrossRef][PubMed] 19. Maenosono, H.; Saito, H.; Nishioka, Y. A Transparent Polyimide Film as a Biological Cell Culture Sheet with Microstructures. J. Biomater. Nanobiotechnology 2014, 5, 17–23. [CrossRef] 20. Buruiana, L.I.; Barzic, A.I.; Stoica, I.; Hulubei, C. Evaluation of blood cells and proteins spreading on imidic polymers containing alicyclic sequences. J. Polym. Res. 2016, 23.[CrossRef] 21. Damaceanu, M.D.; Rusu, R.D.; Bruma, M.; Rusanov, A. Fluorinated Heterocyclic Polyperyleneimides. Rev. Roum. Chim. 2010, 55, 953–961. 22. Sava, I.; Damaceanu, M.-D.; Constantin, C.-P.; Asandulesa, M.; Woli´nska-Grabczyk,A.; Jankowski, A. Structure—Promoted high performance properties of triphenylmethane-containing polyimides and copolyimides. Eur. Polym. J. 2018, 108, 554–569. [CrossRef] 23. Damaceanu, M.-D.; Rusu, R.-D.; Bruma, M. Copolyimides containing perylene and hexafluoroisopropylidene moieties. High Perform. Polym. 2012, 24, 50–57. [CrossRef] Materials 2019, 12, 3166 25 of 27

24. Damaceanu, M.-D.; Constantin, C.-P.; Bruma, M.; Belomoina, N.M. Highly fluorinated polyimide blends—Insights into physico-chemical characterization. Polymer 2014, 55, 4488–4497. [CrossRef] 25. Hassler, C.; Boretius, T.; Stieglitz, T. Polymers for neural implants. J. Polym. Sci. Part B Polym. Physics 2011, 49, 18–33. [CrossRef] 26. Viventi, J.; Kim, D.H.; Vigeland, L.; Frechette, E.S.; Blanco, J.A.; Kim, Y.S.; Avrin, A.E.; Tiruvadi, V.R.; Hwang, S.W.; Vanleer, A.C.; et al. Flexible, foldable, actively multiplexed, high-density electrode array for mapping brain activity in vivo. Nat. Neurosci. 2011, 14, 1599–1605. [CrossRef] 27. Sun, Y.; Lacour, S.P.; Brooks, R.A.; Rushton, N.; Fawcett, J.; Cameron, R.E. Assessment of the biocompatibility of photosensitive polyimide for implantable use. J. Biomed. Mater. Res. A 2009, 90, 648–655. [CrossRef][PubMed] 28. Myllymaa, S.; Myllymaa, K.; Korhonen, H.; Lammi, M.J.; Tiitu, V.; Lappalainen, R. Surface characterization and in vitro biocompatibility assessment of photosensitive polyimide films. Colloids Surf. B Biointerfaces 2010, 76, 505–511. [CrossRef] 29. Rubehn, B.; Stieglitz, T. In vitro evaluation of the long-term stability of polyimide as a material for neural implants. Biomaterials 2010, 31, 3449–3458. [CrossRef] 30. Starr, P.; Agrawal, C.M.; Bailey, S. Biocompatibility of common polyimides with human endothelial cells for a cardiovascular microsensor. J. Biomed. Mater. Res. Part A 2016, 104, 406–412. [CrossRef] 31. Del Valle, J.; de la Oliva, N.; Müller, M.; Stieglitz, T.; Navarro, X. Biocompatibility evaluation of parylene C and polyimide as substrates for peripheral nerve interfaces. In Proceedings of the 7th International IEEE/EMBS Conference on Neural Engineering (NER), Montpellier, France, 22–24 April 2015; pp. 442–445. 32. Lago, N.; Yoshida, K.; Koch, K.P.; Navarro, X. Assessment of biocompatibility of chronically implanted polyimide and platinum intrafascicular electrodes. IEEE Trans. Biomed. Eng. 2007, 54, 281–290. [CrossRef] 33. Rossini, P.M.; Micera, S.; Benvenuto, A.; Carpaneto, J.; Cavallo, G.; Citi, L.; Cipriani, C.; Denaro, L.; Denaro, V.; Di Pino, G.; et al. Double nerve intraneural interface implant on a human amputee for robotic hand control. Clin. Neurophysiol. 2010, 121, 777–783. [CrossRef] 34. Xue, N.; Sun, T.; Tsang, W.M.; Delgado-Martinez, I.; Lee, S.-H.; Sheshadri, S.; Xiang, Z.; Merugu, S.; Gu, Y.; Yen, S.-C.; et al. Polymeric C-shaped cuff electrode for recording of peripheral nerve signal. Sens. Actuators B Chem. 2015, 210, 640–648. [CrossRef] 35. Ceyssens, F.; Puers, R. Insulation lifetime improvement of polyimide thin film neural implants. J. Neural Eng. 2015, 12, 054001. [CrossRef][PubMed] 36. Heo, D.N.; Kim, H.J.; Lee, Y.J.; Heo, M.; Lee, S.J.; Lee, D.; Do, S.H.; Lee, S.H.; Kwon, I.K. Flexible and Highly Biocompatible Nanofiber-Based Electrodes for Neural Surface Interfacing. ACS Nano 2017, 11, 2961–2971. [CrossRef][PubMed] 37. Hernandez-Sebastian, N.; Diaz-Alonso, D.; Renero-Carrillo, F.J.; Villa-Villasenor, N.; Calleja-Arriaga, W. Design and Simulation of an Integrated Wireless Capacitive Sensors Array for Measuring Ventricular Pressure. Sensors (Basel) 2018, 18, 2781. [CrossRef][PubMed] 38. Wurth, S.; Capogrosso, M.; Raspopovic, S.; Gandar, J.; Federici, G.; Kinany, N.; Cutrone, A.; Piersigilli, A.; Pavlova, N.; Guiet, R.; et al. Long-term usability and bio-integration of polyimide-based intra-neural stimulating electrodes. Biomaterials 2017, 122, 114–129. [CrossRef][PubMed] 39. Cutrone, A.; Del Valle, J.; Santos, D.; Badia, J.; Filippeschi, C.; Micera, S.; Navarro, X.; Bossi, S. A three-dimensional self-opening intraneural peripheral interface (SELINE). J. Neural Eng. 2015, 12, 016016. [CrossRef][PubMed] 40. Fekete, Z.; Pongrácz, A. Multifunctional soft implants to monitor and control neural activity in the central and peripheral nervous system: A review. Sens. Actuators B Chem. 2017, 243, 1214–1223. [CrossRef] 41. Jiang, X.; Sui, X.; Lu, Y.; Yan, Y.; Zhou, C.; Li, L.; Ren, Q.; Chai, X. In vitro and in vivo evaluation of a photosensitive polyimide thin-film microelectrode array suitable for epiretinal stimulation. J. Neuroeng. Rehabil. 2013, 10, 48. [CrossRef][PubMed] 42. Lee, H.C.; Ejserholm, F.; Gaire, J.; Currlin, S.; Schouenborg, J.; Wallman, L.; Bengtsson, M.; Park, K.; Otto, K.J. Histological evaluation of flexible neural implants; flexibility limit for reducing the tissue response? J. Neural Eng. 2017, 14, 036026. [CrossRef] 43. Starr, P.; Bartels, K.; Agrawal, C.M.; Bailey,S. A thin-film pressure transducer for implantable and intravascular blood pressure sensing. Sens. Actuators A Phys. 2016, 248, 38–45. [CrossRef] Materials 2019, 12, 3166 26 of 27

44. Park, G.; Chung, H.-J.; Kim, K.; Lim, S.A.; Kim, J.; Kim, Y.-S.; Liu, Y.; Yeo, W.-H.; Kim, R.-H.; Kim, S.S.; et al. Immunologic and Tissue Biocompatibility of Flexible/Stretchable Electronics and Optoelectronics. Adv. Healthc. Mater. 2014, 3, 515–525. [CrossRef] 45. Antanaviciute, I.; Simatonis, L.; Ulcinas, O.; Gadeikyte, A.; Abakeviciene, B.; Tamulevicius, S.; Mikalayeva, V.; Skeberdis, V.A.; Stankevicius, E.; Tamulevicius, T. Femtosecond laser micro-machined polyimide films for cell scaffold applications. J. Tissue Eng. Regen. Med. 2018, 12, e760–e773. [CrossRef][PubMed] 46. Patil, A.C.; Thakor, N.V. Implantable neurotechnologies: A review of micro- and nanoelectrodes for neural recording. Med. Biol. Eng. Comput. 2016, 54, 23–44. [CrossRef][PubMed] 47. Seo, J.-M.; Kim, S.J.; Chung, H.; Kim, E.T.; Yu, H.G.; Yu, Y.S. Biocompatibility of polyimide microelectrode array for retinal stimulation. Mater. Sci. Eng. C 2004, 24, 185–189. [CrossRef] 48. Bae, S.H.; Che, J.-H.; Seo, J.-M.; Jeong, J.; Kim, E.T.; Lee, S.W.; Koo, K.; Suaning, G.J.; Lovell, N.H.; Cho, D.-I.D.; et al. In Vitro Biocompatibility of Various Polymer-Based Microelectrode Arrays for Retinal ProsthesisMicroelectrode Arrays for Retinal Prosthesis. Investig. Ophthalmol. Vis. Sci. 2012, 53, 2653–2657. [CrossRef][PubMed] 49. Lee, K.; Singh, A.; He, J.; Massia, S.; Kim, B.; Raupp, G. Polyimide based neural implants with stiffness improvement. Sens. Actuators B Chem. 2004, 102, 67–72. [CrossRef] 50. Popovici, D.; Barzic, A.I.; Barzic, R.F.; Vasilescu, D.S.; Hulubei, C. Semi-alicyclic polyimide precursors: Structural, optical and biointerface evaluations. Polym. Bull. 2015, 73, 331–344. [CrossRef] 51. Stoica, I.; Barzic, A.I.; Butnaru, M.; Doroftei, F.; Hulubei, C. Surface topography effect on fibroblasts population on epiclon-based polyimide films. J. Adhes. Sci. Technol. 2015, 29, 2190–2207. [CrossRef] 52. Nagaoka, S.; Ashiba, K.; Kawakami, H. Interaction between biocomponents and surface modified fluorinated polyimide. Mater. Sci. Eng. C 2002, 20, 181–185. [CrossRef] 53. Kanno, M.; Kawakami, H.; Nagaoka, S.; Kubota, S. Biocompatibility of fluorinated polyimide. J. Biomed. Mater. Res. 2002, 60, 53–60. [CrossRef] 54. Nagaoka, S.; Ashiba, K.; Kawakami, H. Biomedical Properties of Nanofabricated Fluorinated Polyimide Surface. Artif. Organs 2002, 26, 670–675. [CrossRef] 55. Waris, G.; Siddiqi, H.M.; Bolte, M.; Hussain, R.; Akhtar, Z. Synthesis and characterization of processable aromatic polyimides and their initial evaluation as promising biomaterials. Colloid Polym. Sci. 2013, 291, 1581–1593. [CrossRef] 56. Vert, M.; Doi, Y.; Hellwich, K.-H.; Hess, M.; Hodge, P.; Kubisa, P.; Rinaudo, M.; Schué, F. Terminology for biorelated polymers and applications (IUPAC Recommendations 2012). Pure Appl. Chem. 2012, 84, 377–410. [CrossRef] 57. De Moraes Porto, I.C.C. Polymer Biocompatibility. In Polymerization; De Souza Gomes, A., Ed.; InTechOpen: London, UK, 2012. [CrossRef] 58. Ree, M.; Shin, T.J.; Lee, S.W. Fully Rod-like Aromatic Polyimides: Structure, Properties, and Chemical Modifications. Korea Polym. J. 2001, 9, 1–17. 59. Ceballos, D.; Valero-Cabré, A.; Valderrama, E.; Schüttler, M.; Stieglitz, T.; Navarro, X. Morphologic and functional evaluation of peripheral nerve fibers regenerated through polyimide sieve electrodes over long-term implantation. J. Biomed. Mater. Res. 2002, 60, 517–528. [CrossRef] 60. Lago, N.; Ceballos, D.; Rodríguez, F.J.; Stieglitz, T.; Navarro, X. Long term assessment of axonal regeneration through polyimide regenerative electrodes to interface the peripheral nerve. Biomaterials 2005, 26, 2021–2031. [CrossRef] 61. Schuettler, M.; Koch, K.P.; Stieglitz, T. Investigations on explanted micromachined nerve electrodes. In Proceedings of the 8th Annual International Conference of the IFESS, Maroochydore, Australia, 1–5 July 2003; pp. 306–310. 62. Stoica, I.; Barzic, A.I.; Aflori, M.; Hulubei, C.; Harabagiu, V.;Vasilescu, D.S. Three-Dimensional Nanostructures with Biocidal Activity Created on a Siloxane-Containing Copolyimide Film. Key Eng. Mater. 2015, 638, 98–103. [CrossRef] 63. Chen, C.C.; Wang, F.H.; Chang, S.C.; Yang, C.F. Using Oxygen Plasma Pretreatment to Enhance the Properties of F-Doped ZnO Films Prepared on Polyimide Substrates. Materials (Basel) 2018, 11, 1501. [CrossRef] 64. Teymouri, S.; Calejo, M.T.; Hiltunen, M.; Sorkio, A.; Juuti-Uusitalo, K.; Skottman, H.; Kellomäki, M.; Vullum-Bruer, F. Collagen-immobilized polyimide membranes for retinal pigment epithelial cell adherence and proliferation. Cogent Chem. 2017, 3.[CrossRef] Materials 2019, 12, 3166 27 of 27

65. Lin, S.P.; Chen, J.J.; Liao, J.D.; Tzeng, S.F. Characterization of surface modification on microelectrode arrays for in vitro cell culture. Biomed. Microdevices 2008, 10, 99–111. [CrossRef] 66. Heo, D.N.; Ko, W.-K.; Lee, W.J.; Lee, S.J.; Lee, D.; Heo, M.; Rim, H.; Bae, M.S.; Lee, J.B.; Ahn, B.-S.; et al. Enhanced Biocompatibility of Polyimide Film by Anti-Inflammatory Drug Loading. J. Nanosci. Nanotechnol. 2016, 16, 8800–8804. [CrossRef] 67. Julien, S.; Peters, T.; Ziemssen, F.; Arango-Gonzalez, B.; Beck, S.; Thielecke, H.; Buth, H.; Van Vlierberghe, S.; Sirova, M.; Rossmann, P.; et al. Implantation of ultrathin, biofunctionalized polyimide membranes into the subretinal space of rats. Biomaterials 2011, 32, 3890–3898. [CrossRef] 68. Sridar, S.; Churchward, M.A.; Mushahwar, V.K.; Todd, K.G.; Elias, A.L. Peptide modification of polyimide-insulated microwires: Towards improved biocompatibility through reduced glial scarring. Acta Biomater. 2017, 60, 154–166. [CrossRef] 69. Rousche, P.J.; Pellinen, D.S.; Pivin, D.P.; Williams, J.C.; Vetter, R.J.; Kipke, D.R. Flexible polyimide-based intracortical electrode arrays with bioactive capability. IEEE Trans. Biomed. Eng. 2001, 48, 361–371. [CrossRef] 70. Hopp, B.; Smausz, T.; Papdi, B.; Bor, Z.; Szabó, A.; Kolozsvári, L.; Fotakis, C.; Nógrádi, A. Laser-based techniques for living cell pattern formation. Appl. Phys. A 2008, 93, 45–49. [CrossRef]

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).