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The Osteogenic Differentiation Effect of the FN Type 10-Peptide Amphiphile on PCL Fiber

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The Osteogenic Differentiation Effect of the FN Type 10-Peptide Amphiphile on PCL Fiber International Journal of Molecular Sciences Article The Osteogenic Differentiation Effect of the FN Type 10-Peptide Amphiphile on PCL Fiber Ye-Rang Yun 1, Hae-Won Kim 2,3,4,* and Jun-Hyeog Jang 5,* ID 1 Industrial Technology Research Group, Research and Development Division, World Institute of Kimchi, Nam-Gu, Gwangju 61755, Korea; [email protected] 2 Institute of Tissue Regeneration Engineering (ITREN), Dankook University, Cheonan 330-714, Korea 3 Department of Nanobiomedical Science & BK21 PLUS NBM Global Research Center for Regenerative Medicine, Dankook University, Cheonan 330-714, Korea 4 Department of Biomaterials Science, School of Dentistry, Dankook University, Cheonan 330-714, Korea 5 Department of Biochemistry, Inha University School of Medicine, Incheon 22212, Korea * Correspondence: [email protected] (H.-W.K.); [email protected] (J.-H.J.); Tel.: +82-41-550-3081 (H.-W.K.); +82-32-890-0930 (J.-H.J.); Fax: +82-41-550-3085 (H.-W.K.); +82-32-882-1877 (J.-H.J.) Received: 1 November 2017; Accepted: 3 January 2018; Published: 4 January 2018 Abstract: The fibronectin type 10-peptide amphiphile (FNIII10-PA) was previously genetically engineered and showed osteogenic differentiation activity on rat bone marrow stem cells (rBMSCs). In this study, we investigated whether FNIII10-PA demonstrated cellular activity on polycaprolactone (PCL) fibers. FNIII10-PA significantly increased protein production and cell adhesion activity on PCL fibers in a dose-dependent manner. In cell proliferation results, there was no effect on cell proliferation activity by FNIII10-PA; however, FNIII10-PA induced the osteogenic differentiation of MC3T3-E1 cells via upregulation of bone sialoprotein (BSP), collagen type I (Col I), osteocalcin (OC), osteopontin (OPN), and runt-related transcription factor 2 (Runx2) mitochondrial RNA (mRNA) levels; it did not increase the alkaline phosphatase (ALP) mRNA level. These results indicate that FNIII10-PA has potential as a new biomaterial for bone tissue engineering applications. Keywords: FNIII10; peptide amphiphile; PCL fiber; osteogenic differentiation activity 1. Introduction Recently, various attempts have been made to find new biomaterial for tissue engineering. Among the attempts, research on the peptide amphiphile (PA) has been widely reported. PA has hydrophilic and hydrophobic components and a self-assembly ability as a peptide-based molecule. There is increased interest in the extracellular matrix (ECM) molecule mimetic PA because it can promote certain cellular activities including adhesion, proliferation, and differentiation [1–5]. Based on these characteristics, biomimetic PA is extensively utilized for tissue engineering as a new biomaterial. For example, the Arg-Gly-Asp (RGD) peptide, composed of L-arginine, glycine, and L-aspartic acid, participates in cellular attachment via integrin [6]. For this reason, numerous studies on PA-RGD have been reported in the pharmaceutical field [7–9] as well as in the tissue engineering field [10–12]. For instance, the combination of hydroxyapatite (HA) and PA-RGD enhances the osteoinductive and osteoconductive activities of the scaffold [13]. Moreover, studies on amphiphile containing fibronectin (FN) are commonly demonstrated and reported. We also previously demonstrated that the engineered fibronectin type 10-peptide amphiphile sequence (FNIII10-PA) potentiates utilization for bone tissue engineering by enhancing the adhesion, proliferation, and differentiation of rat bone marrow stem cells (rBMSCs) [14]. In another study, the tenascin-C mimetic PA (TN-C PA) showed self-assembly activity. Nanofiber gels containing TN-C Int. J. Mol. Sci. 2018, 19, 153; doi:10.3390/ijms19010153 www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2018, 19, 153 2 of 10 PA were shown to promote neurite outgrowth, which could potentially be used for artificial matrix therapy in neuronal regeneration [15]. PolycaprolactoneInt. J. Mol. Sci. 2018, 19, 153 (PCL) is a biomaterial that is used mostly for bone tissue engineering2 of 10 [16,17]. PCL is a biodegradable polyester with a low melting point (60 ◦C) and a low glass transition containing TN-C PA were shown to promote neurite outgrowth, which could potentially be used for temperature (−60 ◦C). Because of its biodegradation properties, PCL is regarded as a suitable artificial matrix therapy in neuronal regeneration [15]. biomaterialPolycaprolactone for long-term implantable (PCL) is a biomaterial devices [that18]. is Above used mostly all, PCL for hasbone advantages tissue engineering in tissue [16,17]. engineering applicationsPCL is fora biodegradable drug delivery polyester devices, with sutures, a low ormelting adhesion point barriers(60 °C) and without a low safety glass transition problems for the Food andtemperature Drug Administration (−60 °C). Because (FDA) of its approvalbiodegradation [19]. pr However,operties, PCL it isis difficultregarded foras PCLa suitable to influence cellularbiomaterial activity becausefor long-term it does implantable not provide devices an [18]. ECM Above type all, structure PCL has foradvantages the cells in [tissue20]. Recently, engineering applications for drug delivery devices, sutures, or adhesion barriers without safety nanofibrous PCL has been the preferred choice for bone tissue engineering applications by improving problems for the Food and Drug Administration (FDA) approval [19]. However, it is difficult for cellularPCL activities to influence and cellular mechanical activity properties. because it does PCL not isprovide indisputably an ECM type the structure most used for biomaterialthe cells [20]. in bone tissue engineeringRecently, nanofibrous research. PCL has been the preferred choice for bone tissue engineering applications by Previously,improving cellular we revealed activitiesthat and geneticallymechanical pr engineeredoperties. PCL FNIII10-PA is indisputably promoted the most cell used adhesion, proliferation,biomaterial and in differentiationbone tissue engineering of rBMSCs. research. In the present study, we investigated whether or not FNIII10-PAPreviously, with PCL we fiber revealed promoted that genetically the MC3T3-E1’s engineered cellular FNIII10-PA activities. promoted FNIII10 cell adhesion, was used as the proliferation, and differentiation of rBMSCs. In the present study, we investigated whether or not negative control in all experiments. Initially, the protein adhesion activity of FNIII10-PA was measured. FNIII10-PA with PCL fiber promoted the MC3T3-E1’s cellular activities. FNIII10 was used as the Then, thenegative effects control of FNIII10-PA in all experiments. with PCL Initially, fiber onthe MC3T3-E1protein adhesion cells activity adhesion of andFNIII10-PA proliferation was were investigated.measured. Furthermore, Then, the effects the osteogenicof FNIII10-PA differentiation with PCL fiber activity on MC3T3-E1 of FNIII10-PA cells adhesion with PCL and fibers on MC3T3-E1proliferation cells was were investigated investigated. by Furthermore, measuring the the osteogenic osteogenic differentiation differentiation activity marker of FNIII10-PA mRNA level. with PCL fibers on MC3T3-E1 cells was investigated by measuring the osteogenic differentiation 2. Resultsmarker and mRNA Discussion level. 2.1. Morphology2. Results and of PCL Discussion Fibers PCL2.1. Morphology has been of commonly PCL Fibers used as a biomaterial in the tissue engineering field because of its non-toxic andPCL biodegradable has been commonly characteristics. used as a biomaterial On the other in the hand, tissue PCL engineering demonstrated field because a slow of degradation its rate, poornon-toxic mechanical and biodegradable properties, characteristics. and low cell On adhesion. the other Particularly, hand, PCL PCLdemonstrated shows low a slow affinity and stiffnessdegradation in cells due rate, to poor poor mechanical hydrophilicity, properties, leading and low to cell the adhesion. reduction Particularly, in cell proliferation, PCL shows low migration, differentiation,affinity and and stiffness regeneration in cells due [20 to]. poor However, hydrophilicity, nanofibrous leading PCL to the was reduction reported in cell to proliferation, improve the cellular migration, differentiation, and regeneration [20]. However, nanofibrous PCL was reported to activities in recent studies [21,22]. FNIII10 was connected to the PA sequence LLLLCCCGGDSDS improve the cellular activities in recent studies [21,22]. FNIII10 was connected to the PA sequence (L, leucine;LLLLCCCGGDSDS C, cysteine; G, (L, glycine; leucine; C, D, cysteine aspartic; G, acid; glycine; S, serine) D, aspartic to create acid; S, FNIII10-A serine) to create sequence FNIII10-A as illustrated in Figuresequence1A. PCL as illustrated fiber was in Figure fabricated 1A. PCL by fiber the electrospinningwas fabricated by technique.the electrospinning Figure technique.1B shows that the electrospunFigure 1B shows PCL that fiber the was electrospun uniform PCL with fiber straight was uniform fibers. with Average straight fibers. diameter Average of diameter PCL fibers was approximatelyof PCL fibers 1 µ Mwas (Figure approximately1C). Hence, 1 μM PCL (Figure fiber 1C). was Hence, suitable PCL fiber to investigate was suitable the to cellinvestigate adhesion the activity of FNIII10cell adhesion in combination activity of with FNIII10 biomaterial in combinat inion the with present biomaterial study. in the present study. FigureFigure 1. Structure 1. Structure offibronectin of fibronectin type type 10-peptide10-peptide amphiphile amphiphile (FNIII10-PA) (FNIII10-PA) (A) and (A
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