The Effects of Spinal Implant Wear Debris Particles

ISRN UTH-INGUTB-EX-MTI-2021/004-SE Examensarbete 15 hp Juni 2021

The effects of spinal implant wear debris particles

Eslam El Ammarin Cecilia Thomas

Högskoleingenjörsprogrammet i medicinsk teknik The effects of spinal implant wear debris particles

Eslam El Ammarin and Cecilia Thomas

Abstract

The goal of this literature study was to study the effects of spinal implant wear debris particles on the body in general, and on microglia cells in particular. The method of the literature study was searching for scientific peer-reviewed papers on the topic. Spinal implants are used to fix spinal problems such as deformities or injuries. All implants wear down in the body. This produces wear debris particles. The body’s immune system reacts to the particles, triggering inflammation, osteolysis and implant loosening. The reaction depends on particle type and the location of the particles. Cobalt chrome particles are more toxic than stainless steel particles. Metal particles are more inflammatory than ceramics and most polymers. Microglia are immune cells specific to the brain and spinal cord. These cells would be one of the cells reacting to wear debris from spinal implants. Not many studies have been made on the interaction between microglia and wear particles. Some cells react differently to wear particles on their own, compared to when they are combined with other cell types. It is important to study the body as a whole system, and not just one cell type, as the results may differ. Several studies have concluded that wear particles induce an inflammatory response, and that the resulting inflammation is mild and does not have any severe negative effects. How much inflammation is required for a severe negative effect such as osteolysis is unclear. In conclusion, the perfect spinal implant does not exist. All spinal implants generate wear debris, and the body reacts to every type of debris. Maybe science will one day find the perfect implant material that does not induce a biological reaction.

Faculty of Science and Technology Uppsala University, Uppsala

Supervisor: Estefanía Echeverri Correa Subject reader: Gry Hulsart Billström Examiner: Caroline Öhman Mägi

Svensk populärvetenskaplig sammanfattning

Ryggimplantat används för att åtgärda missbildningar, skador och problem i ryggen. Implantat nöts ner i kroppen och då bildas partiklar, som sprids till omkringliggande vävnader. Nötningspartiklar kan orsaka problem, då kroppens immunförsvar reagerar på dem. Detta kan leda till inflammation och infektion, och även att implantatet lossnar. Det kan krävas ytterligare operationer för att ta bort eller byta ut implantat som orsakar problem. Ett bra implantatmaterial slits mindre, vilket skapar färre partiklar som kan orsaka inflammation. Det är också fördelaktigt om partiklarna i sig orsakar mindre inflammation. Mindre inflammation leder till färre biverkningar av implantaten.

Makrofager är en typ av immuncell som reagerar på nötningspartiklar. Deras jobb är att skydda kroppen mot hot, till exempel sjukdomar, skadade celler eller främmande föremål såsom nötningspartiklar. Mikrogliaceller är en speciell typ av makrofager som finns i ryggraden och hjärnan. Det är relevant att studera mikrogliaceller, då de kommer reagerar på och påverkas av nötningspartiklar från ryggimplantat.

Det finns flera olika typer av implantat, som kan se olika ut, och tillverkas av olika material beroende på användningsområde. Implantat kan vara gjorda av polymerer, keramer, och metaller. Metallerna kan vara till exempel kobolt krom, rostfritt stål eller titan, samt legeringar som kombinerar metaller med ett eller flera andra element.

Implantaten kan vara skruvar, plattor eller stavar som används för att stabilisera delar av ryggraden, eller för att laga en skadad ryggrad. Samma resultat kan fås med hjälp av en så kallad bur (cage på engelska). Det är ett implantat som placeras mellan två kotor och hjälper kotorna att växa ihop. En steloperation tar bort rörligheten mellan två eller flera ryggkotor. Det finns även andra typer av implantat som behåller rörligheten, såsom diskproteser.

För att kunna använda implantatet i människokroppen behöver det genomgå olika typer av tester. Först görs in vitro-test, där experiment utförs på celler i till exempel provrör eller petriskålar. In vitro-test studerar cellens reaktion på ett visst material, till exempel om cellerna överlever.

Om materialet ger bra resultat i in vitro-testerna, görs in vivo-test, vilket betyder djurförsök. Ett levande djur består av många olika organsystem och celltyper som

interagerar med varandra. En celltyp kanske inte påverkas alls av materialet som testas, samtidigt som en annan celltyp påverkas kraftigt. Därför är det viktigt att testa ett materials effekt på en komplicerad organism, såsom ett helt djur, då det kan ge olika resultat jämfört med in vitro-tester (celltester). Om både in vitro och in vivo- testerna är godkända kan materialet testas kliniskt, på människor.

Syftet med detta examensarbete var att göra en litteraturstudie av vetenskapliga artiklar om ryggimplantat och deras nötningspartiklars effekt på kroppen i allmänhet, och mikrogliaceller i synnerhet.

Litteraturstudiens metod bestod av att söka efter vetenskapliga artiklar relaterade till projektets syfte. Artiklarna skulle handla om ryggimplantat och dess nötningspartiklar. Även den biologiska effekten av implantatet eller partiklarna skulle nämnas i artiklarna.

Studier visar att kroppen påverkas av alla typer av ryggimplantat. Kroppen reagerar olika beroende på var implantaten är placerade, och vilka material de är gjorda av. Några exempel är att kobolt krom har mer negativ effekt än rostfritt stål. Metaller skapar mer inflammation än keramer och de flesta polymerer. Flera studier har kommit fram till att implantatpartiklar skapar en mild inflammation som inte leder till några större skador. Hur mycket inflammation som krävs för att orsaka skador i kroppen, som att implantat lossnar, är oklart.

En studie visade att olika kombinationer av celler reagerar olika på implantatpartiklar. Försök på en celltyp gav olika resultat jämfört med försök på en kombination av celltyper. Det visar att det är viktigt att se kroppen som ett komplicerat system, där olika delar av kroppen påverkar varandra.

Många studier handlar om hur makrofager reagerar på nötningspartiklar från implantat. Det finns få studier om hur mikrogliaceller påverkas av implantatpartiklar. Vi skulle vilja se framtida studier om mikrogliacellers reaktion på implantatpartiklar av olika material.

Sammanfattningsvis har alla material har för- och nackdelar. Inget implantatmaterial är perfekt. Förhoppningsvis leder framtida forskning till ännu bättre implantatmaterial.

Preface

This report is our degree project for our Bachelor’s in Biomedical Engineering at Uppsala university.

Our original plan was to study the effects of silicon nitride particles on microglia cells. This was going to be done using microglia cell cultures in a lab. Because of the COVID-19 pandemic, we did a remote literature study instead.

We would like to thank our supervisor, Estefanía Echeverri Correa, for her help and guidance through our degree project. Thanks to her, we stayed on the right path, and the task seemed a little less difficult.

We are also grateful to our subject reader, Gry Hulstart Billström, for her ideas and expertise.

Table of Contents

1 Introduction ...... 1

1.1 Goal ...... 2

1.2 Research question ...... 2

1.3 Scope and delimitations ...... 2

1.4 Outline ...... 3

1.5 Method ...... 3

1.6 Discussion regarding sources ...... 4

1.7 Author contribution ...... 5

2 Theory ...... 6

2.1 Anatomy of the spine and spinal cord ...... 6

2.2 Cells of the central nervous system ...... 8

2.3 Microglia ...... 9

2.4 Spinal implants ...... 11

2.5 Materials used in spinal implants ...... 13

2.6 Implant wear debris ...... 13

2.7 Morphological properties of wear debris ...... 14

2.8 The biological response to spinal implant debris ...... 15

2.9 In vitro, in vivo and clinical studies ...... 16

3 Literature study ...... 17

3.1 Bryan cervical disc ...... 17

3.2 In vitro studies...... 18

3.2.1 CoCr particles on the dura mater ...... 18

3.2.2 SS and CoCr particles with astrocyte and microglia cells ...... 20

3.3 In vivo studies ...... 20

3.3.1 Particles applied to spinal cord ...... 20

3.3.2 Titanium particles applied to retroperitoneal and epidural space ...... 21

3.3.3 PEEK particles applied to epidural and intervertebral disc spaces ...... 22

3.3.4 Cobalt chromium particles in epidural space ...... 23

3.4 Clinical and retrieval studies ...... 24

3.4.1 Metallosis caused by spinal implants ...... 24

3.4.2 M6-C artificial cervical disc failure ...... 25

3.4.3 Dynesys spinal implants ...... 28

4 Discussion ...... 31

4.1 Bryan cervical disc study ...... 31

4.2 Dynesys article ...... 31

4.3 Animal studies might not accurately predict results in humans ...... 32

4.4 Use of different animals to study spinal implants ...... 33

4.5 Ethical aspects of animal testing ...... 34

4.6 The perfect implant material does not exist ...... 34

5 Conclusion ...... 35

5.1 Further studies ...... 35

References ...... 37

Abbreviations

Below is a list of abbreviations used in this report

CNS - Central nervous system

CoCr - Cobalt chromium

CoCrMo - Cobalt chrome molybdenum

IVD - Intervertebral disc

MoM - Metal on metal

MoP - Metal on polymer

Ni – Nickel

PCU - Polycarbonate urethane

PEEK - Polyetheretherketone

Si3N4 - Silicon nitride

SS - Stainless steel

TDR - Total disc replacement

Ti - Titanium

UHMWPE - Ultra high molecular weight polyethylene

1 Introduction

Spinal implants are used to fix problems with the spine. These problems include congenital deformities, traumatic injuries, and degeneration caused by old age or disease.

Scoliosis is an abnormal sideways curvature of the spine. People with mild cases might not have any symptoms at all, and do not require treatment [1]. More severe cases of scoliosis can lead to back pain and the inability to stand up straight. It can also cause deformation of the rib cage, pressing against the lungs and heart, restricting breathing and the heart’s ability to pump blood [2,3].

As a person ages, the joints of the body degenerate. This is also true for the intervertebral discs of the spine, where the phenomenon is called degenerative disc disease. Wear and tear of the spine increases with age, which also affects the intervertebral discs. This leads to shrinking of the discs, and they cannot function as normal. The intervertebral discs lose their shock absorbing properties, leading to pain with every movement of the spine [4,5].

Traumatic injuries can also cause spinal fractures. Broken vertebrae not only hurt, but can also cause further problems if bone fragments push against the spinal cord. Spinal dislocations can also damage the spinal cord. Damage to the spinal cord can be catastrophic, causing pain, loss of sensation and muscle control, and paralysis [6,7].

While spinal implants can solve the above mentioned problems, they can also create new problems. Implants in the body wear down over time [8]. This results in particles releasing from the implant into the surrounding tissue. These particles can trigger biological responses such as inflammation, cytotoxicity (cell death) and hypersensitivity to the implant material [9]. These responses can lead to pain, infection and loosening of the implant [8]. If that happens, the implant would not help at all, or even make the problems worse for the patient. There is a need for better implant materials that wear less and trigger a milder biological response, in order to mitigate these negative effects. Before a new material is implanted into a living body, the reaction of relevant cells to the implant material needs to be studied.

There are many materials that can be used for different types of implants, including spinal implants [8,10,11]. Although spinal implants are never directly in contact with the spinal cord and central nervous system (CNS), wear particles released from spinal implants can migrate into the surrounding tissue [12]. That surrounding tissue includes the spinal cord and CNS. The CNS, like other organ systems in the body, contains specialised cells that behave differently compared to other cell types. To know how the body would react to a spinal implant, we need to investigate how neural cells react to the implant material.

Microglia, a type of neural cell, are the macrophages of the central nervous system [13]. Part of their job is to defend the CNS against pathogens or foreign materials. Microglia would be one of the main cell types reacting to the spinal implants and their wear particles, therefore it is important to know how microglia react to different implant materials.

1.1 Goal

The goal of this project is to do a literature study of different materials used in spinal implants and their effects on the living body, including microglia cells. While there are few studies combining spinal implants and microglia, separate studies about spinal implants and microglia should give useful information about how the two will react together.

1.2 Research question

How do wear particles from spinal implants made of different materials affect the body in general, and microglia cells in particular?

1.3 Scope and delimitations

This literature study covers spinal implants, wear debris, wear particles and microglia cells. As we have not done any experiments, we are only reporting the results of previous studies.

There are far more studies related to hip and knee implants compared to spinal implants, and more about macrophages than microglia. When possible, we have focused on spinal implants and microglia, and excluded papers on hip and knee implants and macrophages.

This report does not cover the mechanical properties of spinal implant materials.

1.4 Outline

This section describes the outline of the report.

The report begins with an introduction, describing what spinal implants are, what they are used for, and problems they might cause. The introduction also includes the goal of this project, our research question, and the scope and delimitations of our literature study.

The method describes our search for information about the topic, and our process deciding what papers to include or exclude. We also discuss our sources, and explain the specific contributions of each author.

The theory explains the terminology, anatomy, physiology and processes mentioned in the literature review. This section is meant to help the reader understand what is going on, without having to be an expert in the field.

The literature study section describes various peer-reviewed papers we have read. The case descriptions are sorted according to study type: in vitro, in vivo, or clinical studies.

In the discussion section we discuss our own thoughts about the papers included in the literature review.

The paper ends with a conclusion, and recommendations about further studies.

1.5 Method

The main source of information was peer-reviewed scientific papers. Our supervisor sent us a few papers on the topic. We then continued our own search using keywords from those papers. The Uppsala university library and PubMed databases

were used, as well as the Google search engine. Search terms included microglia, spinal implants, wear debris, wear particles, in vitro, in vivo and clinical studies.

After reading the abstract, we determined if the paper fulfilled our goal of studying the biological response to spinal implant wear particles. If so, the paper was included in the report. Some general information about implants were taken from papers related to knee and hip implants. Studies about spinal implants that did not discuss wear debris or wear particles were excluded. Of all the studies we read about, ten are discussed in chapter 3 of this report.

We used the software Zotero to manage our references.

1.6 Discussion regarding sources

Our main source of information were peer-reviewed studies, which should be reliable. As scientific knowledge moves forward, some information may prove to be incorrect, or become obsolete. Our sources are fairly recent, most are from the 2010’s, so they should still be relevant.

In the theory section, veterinary anatomy and physiology textbooks have been used. The information taken from the textbooks is of a general nature, true for all mammals, including humans.

There are internet websites listed in the reference list. These have mostly been used to find pictures, or explain terminology. We deemed information from hospital websites, such as the Mayo Clinic, to be reliable.

Some information about spinal implant wear debris was hard to find, but existed for knee and hip implants. In those cases, we used the information available about hip and knee implants.

It was more difficult to find articles that describe the in vitro studies than finding articles about in vivo and clinical studies. There was also not as much information about microglia cells as other cell types.

1.7 Author contribution

Cecilia wrote the abstract, introduction, and the theory about anatomy, spinal implants, and the morphological properties of wear debris. She also wrote about the in vivo studies, and the clinical studies with the M6-C and Dynesys implants, and the conclusion. Since Cecilia is fluent in English, she has helped with the grammatical structure of Eslam’s work.

Eslam wrote the Swedish summary, and the theory of the cells of the central nervous system, including microglia, and about the biological response to spinal implant debris. In the studies section, she was responsible for the Bryan cervical disc study, the in vitro studies, and the clinical study about metallosis. She also wrote about future studies.

2 Theory

This section explains theory elements that help the reader understand the scientific and technical concepts in the literature study section.

2.1 Anatomy of the spine and spinal cord

The spine, also known as the vertebral column, is made up of many bones called vertebrae. The spine is similar in all mammals, but the number of vertebrae varies depending on species. This report will focus on the human spine. The vertebral column is divided into regions: cervical, thoracic, lumbar, sacrum and coccyx. A diagram of the human vertebral column is shown in Figure 1. The cervical spine has seven vertebrae that connect the head to the rest of the body. Below the cervical vertebrae are the 12 thoracic vertebrae, which connect to the ribs. Below the thoracic vertebrae are the five lumbar vertebrae, followed by the sacrum and coccyx. The sacrum and coccyx bones are made of fused vertebrae [14]. Vertebrate animals with tails have caudal Figure 1: The spinal column [14] vertebrae instead of a fused coccyx [15].

The individual vertebrae vary in size and shape depending on where in the spine they are located, but their general structure is the same. All vertebrae have a hole in the middle, the vertebral foramen, which form the spinal canal that contains the spinal cord. Vertebrae also have processes sticking out laterally and dorsally (towards the sides and back), where muscles and ligaments attach. The vertebral body is somewhat cylindrical, and supports the weight of the upper body. Between the vertebral bodies, there are intervertebral discs that allow for movement of the spine and function as shock absorbers [14]. Vertebral discs have a gel-like center called the nucleus pulposus, with a fibrous ring called annulus fibrosus around it. There are also cartilaginous end-plates where the disc meets the vertebral bodies [16]. An illustration of a vertebra is shown in Figure 2.

Figure 2: The anatomy of a vertebra seen from above (left) and three vertebrae seen from the side (right) [17].

Figure 3: Cross section of the spinal cord inside a vertebra [18]. The spinal cord is encased by connective tissue known as the meninges. It consists of three layers: the dura mater on the outside, the arachnoid in the middle, and the pia mater closest to the spinal cord. The spinal cord itself consists of grey matter in the middle shaped like the letter H or a butterfly, and white matter on the outside. Spinal nerves leave the spinal cord through the intervertebral foramina (holes between vertebrae). The spinal nerves then go out to the rest of the body, including the arms and legs [14,19,20]. A cross section of the spinal cord can be seen in Figure 3.

2.2 Cells of the central nervous system

The brain and the spinal cord make up the central nervous system (CNS). The main cell type of the nervous system are neurons, the nerve cells. Neurons are responsible for receiving, transmitting and sending nerve signals through the body [21]. Different types of neurons perform different functions. Sensory neurons, also known as afferent neurons, convert stimuli such as touch or sound to nerve signals. These signals are sent through the spinal cord, to the brain, where they can be processed. This allows the individual to “feel” the touch, or “hear” the sound. Motor neurons, or efferent neurons, receive signals from the brain, sending them to various parts of the body such as muscles. These signals function as instructions to move your hand, for example [19].

Glial cells have a supporting function. The word glia means glue, and has been used to describe the cells holding neurons together [22]. The number of glial cells are the same as neurons, and microglial cells are 10-20 percent of glial cells [23].

Different types of glial cells include oligodendrocytes, astrocytes and microglia. An illustration of these is shown in Figure 4. Oligodendrocytes are cells that help produce the myelin sheaths that facilitate signal transmission in the neurons’ axons [24]. Astrocytes are shaped like stars and are the largest neuroglial cells. Astrocytes help the neurons move during development, and are part of the blood-brain barrier. Some sources also count ependymal cells as glial cells. Ependymal cells line the walls of the brain’s ventricular system and the spinal cord [21].

Figure 4: Illustration of microglia, astrocytes, oligodendrocytes, and neurons [24].

2.3 Microglia

Microglia cells are immune cells located in the entire central nervous system. Figure 5 shows different types of microglia cells. Microglia cells' responsibility is to respond to both injuries and diseases. One way to achieve that is by phagocytising foreign material and diseased cells. Phagocytosis is when a cell neutralises injured cells, pathogens, or foreign particles by engulfing or ingesting them [22].

Under normal circumstances, microglia are in a non-activated state. They have a ramified phenotype, meaning they have long thin processes sticking out. The microglia are constantly moving around, surveying the environment [21]. If the non- active microglia come into contact with something abnormal, such as foreign material or injured cells, they activate. When activated, microglia transform into an amoeboid shape, with fewer thicker processes [25]. These activated microglia cells proliferate,

increasing the number of microglia in the region. They also phagocytose any possible threats.

Figure 5: Different types of microglia cells. Picture A shows a type of microglia cells called ramified, which have radially thin projecting processes together with developed ramifications. Picture B shows a type called hypertrophied microglia which have short thick processes. Picture C shows a type of microglia cells called monopolarised and picture D shows bipolarised microglia cells. The difference between monopolarised and bipolarised cells is that monopolarised cells have just one process, and bipolarised cells have two processes pointing in different directions [25]. Microglia can also release a variety of substances, such as cytokines. Cytokines are cell signalling molecules. Some cytokines are pro-inflammatory, meaning they promote inflammation. Other cytokines can have an anti-inflammatory function, lowering inflammation. One of the functions of cytokines is to signal other microglia cells to activate them, so more cells can respond to a threat [26]. Microglia can also release cytotoxic substances to kill injured or diseased cells, or invading organisms [27].

There are different types of microglia cell lines available for use in cellular experiments. Microglia cells have been taken from mice, rats, macaques and humans. Most of these cells have been taken from the brain or spinal cord. Studies have revealed differences between rodent and human microglial cell lines, both regarding function and aging. This means that rodent and human cells may react very differently, and studies on rodent microglia may not accurately predict how human microglia will react to the same treatment. Microglia cells from non-human primates, such as rhesus macaques, are meant to bridge the gap between rodents and humans [13].

There have been attempts to create microglia from stem cells. The cells created resembled human foetal microglia, but also resembled other cells such as macrophages. These cells should be used with caution, as they are not exactly like microglia cells [13].

2.4 Spinal implants

Spinal implants can be used to correct deformities or repair structures in the spine. Implants are available in different sizes, depending on the size of the spinal structures in the patients receiving the implants. Broken intervertebral discs can be replaced with an artificial intervertebral disc (IVD) in a procedure called a total disc replacement (TDR). An IVD is meant to mimic the function of the intervertebral disc, retaining motion [28]. Figure 6 shows an artificial intervertebral disc. An alternative to TDR is to fuse the vertebrae Figure 6: An artificial intervertebral disc [30]. on both sides of the affected disc. When fusing vertebrae, they are fixed in place, and all motion between those vertebrae is lost. Vertebral fusion can be done using cages or fixation using a combination of screws, rods, and plates. Cages are porous, allowing the bone to grow through them, and in turn fusing two vertebrae together [11,29]. Figure 7 shows cage implants in the lumbar spine.

Figure 7: Lumbar spine cage implants. The darker implant is made of titanium, the lighter is made of PEEK [31].

Scoliosis is when a person has an abnormal curvature of the spine. Most cases are mild and do not require treatment. An example of a mild case of scoliosis is Cecilia, one of the authors of this report. Her spine can be seen in Figure 8. More severe cases of scoliosis require surgery to straighten the spine. This can be done with rods and screws holding the spine straight [1,11,29]. An example of scoliosis corrected with surgery is shown in Figure 9. These types of surgeries fuse the vertebrae of the spine. When the spine is fused, it is held in place and cannot move. In order to preserve vertebral motion, dynamic stabilisation devices can be used instead [29].

Figure 8:One of the authors of this report, Cecilia, has mild scoliosis.

Figure 9: Scoliosis before and after corrective surgery [32]. 12

2.5 Materials used in spinal implants

Common metals used in spinal implants include cobalt chrome (CoCr), stainless steel (SS) and titanium (Ti). SS is stronger, stiffer and cheaper than CoCr and Ti. Ti is more biocompatible than SS and CoCr. A biocompatible material is not harmful to living tissue. Metals can lead to artifacts in medical imaging, with CoCr and SS giving more artifacts than Ti [11].

Polyetheretherketone (PEEK), a thermoplastic polymer, is also used in spinal implants. It is hydrophobic and bioinert, and does not easily bond to bone. PEEK can be coated or doped to facilitate osseointegration (bone integration). PEEK is radiolucent, and gives fewer imaging artifacts than metals [11].

Silicon nitride (Si3N4) is a ceramic spinal implant material. It can be made porous or with a rough surface, facilitating osseointegration. Silicon nitride is partly radiolucent, so both implant and surrounding bone can be seen in X-rays. It is also non-magnetic, and does not give artifacts in magnetic resonance imaging (MRI) [10]. Si3N4 has good mechanical properties, and it is biocompatible and osteoconductive, which means bone can grow on the material [33].

2.6 Implant wear debris

All implants degrade to some extent in the body. This can happen through wear, corrosion, or a combination of both. This leads to wear debris particles being released from the implant into the surrounding tissue. These particles range in size from nanometres to micrometres [8,9]. Metals also corrode, releasing by-products such as ions into the tissue surrounding the implant. Wear and corrosion weaken the structural integrity of implants, causing implants to break and fail. There is also a biological response to implant debris, which can cause inflammation, osteolysis (breakdown of bone) and hypersensitivity to the implant material. In turn, this can lead to pain and loosening of the implant [8].

Metal on metal (MoM) implants, such as a metal plate attached to bone with a metal screw, generate metallic wear debris. Metal on polymer (MoP) implants, such as

artificial intervertebral discs made of metal and polymer parts, generate mostly polymer wear debris, and very little metal debris [8].

The ideal spinal implant material is biologically inert and biocompatible, to avoid any negative consequences of biological reactions to the implant [11]. Some types of implants should degrade in the body, to allow the body’s own tissues to replace it. Other types of implants should degrade as little as possible, so their function does not deteriorate over time. Less degradation also leads to fewer wear particles, which means fewer foreign particles that can trigger an immune response to the implant and its wear debris particles.

To measure how much debris has been generated by an implant, the implant can be weighed before it is implanted in the body, and after it has been taken out of the body. The difference in weight indicates the amount of wear debris generated from the implant. Another method of assessing the amount of wear particles is to look at a tissue sample with a microscope, and count the particles [8].

2.7 Morphological properties of wear debris

Not all wear particles are created equal. The morphology (shape) of wear particles depends on several factors: material, load and if the implant is fixed or moving [34]. There is not much information about spinal implant debris, but debris from hip and knee implants has been studied extensively. The assumption is that hip and knee implants made of the same materials will produce wear debris similar to that of spinal implants. Particles resulting from metal-on-metal wear are generally smaller and rounder, while particles from metal-on-polymer are larger and more elongated [8].

The same implant can also generate wear debris of different shapes. The debris can be spherical, or shaped like discs, blocks, strips, sheets, etc. Figure 10 shows different morphologies of UHMWPE (ultra high molecular weight polyethylene) debris from a simulation test of an artificial hip joint consisting of UHMWPE and a CoCrMo (cobalt chrome molybdenum) alloy [35].

Figure 10: UHMWPE wear debris from an artificial joint simulator [35].

2.8 The biological response to spinal implant debris

Different materials affect the spine in different ways and the biological response to different materials also varies. Osteolysis is when bone breaks down due to the immune response to wear debris particles [36]. When bone breaks down around an implant, the implant no longer has any tissue to adhere to. This will cause the implant to loosen.

Immune cells may identify wear particles as something foreign, a threat that needs to be defeated. The cells responsible for this immune response are often macrophages. As microglia are the macrophage equivalent of the central nervous system, they are assumed to have a similar function when they encounter wear particles. Macrophages respond to wear particles by initiating an inflammatory response. During the inflammatory response, macrophages release cytokines. These cytokines

have pro-inflammatory and pro-osteoclastic effects, meaning they increase inflammation and the breakdown of bone [37].

2.9 In vitro, in vivo and clinical studies

Before any new type of implant can be used in a patient, it has to be studied as much as possible, to make sure it is safe to use, and that it will not harm the patient. The first step is in vitro studies. Vitro means glass in latin. In vitro studies, “studies in glass”, are done on cells inside containers such as petri dishes or test tubes [38]. These studies give the first pieces of information about how the body’s cells, and by extension the entire body, react to the implant.

One of the types of tests done in in vitro studies is cytotoxicity testing. Cytotoxic means toxic to cells [39]. The test determines if the implant material causes cell death, and the effects on cell growth, reproduction, and the morphology of the cell [40,41].

If the cells are not negatively affected in extensive in vitro tests, it is deemed safe to test the material on an entire living organism. A substance can have an entirely different effect on a complex system, such as a whole living animal, than on cells in a test tube. These tests are known as in vivo, or “within the living”. Testing is first done on animals, often on mice [42].

If both extensive in vitro studies and in vivo studies on animals show promising results, it is safe to move on to human test subjects. Technically, this is also in vivo testing, because humans are living animals, but it is often called clinical studies. If any of the testing phases result in harmful effects, the testing is stopped, and does not move on to the next phase. This is an important step in ensuring the safety of test subjects, and reducing potential suffering.

3 Literature study

This section describes in vitro, in vivo and clinical studies that have been done on how spinal implants and their wear debris affect cells, animals and humans. The cases are sorted in three categories: in vitro, in vivo and clinical studies. The study about the Bryan cervical disc includes elements from every category, and has been placed on its own.

3.1 Bryan cervical disc

An article written by Anderson et al. discusses both in vivo and clinical studies [43]. They describe Bryan cervical discs, their wear properties, and the early clinical results. The Bryan cervical disc is an artificial intervertebral disc implant made of a titanium alloy and polyurethane. Figure 11 shows an image of the implant. Figure 11: The Bryan cervical disc [43].

In the in vivo study they tested the inflammatory response in goats. The goats had a discectomy (disc removal) between C4 and C5 in the cervical spine. The Bryan cervical disc was placed in 11 goats. On four goats, the vertebrae were fused with a titanium plate instead. The goats were sacrificed (euthanised) after three, six or 12 months. After euthanasia, tissue samples were collected. There were no signs of abnormalities related to the Bryan cervical disc implants in any of the treatment groups. The goats whose vertebrae were fused had more wear debris than the ones who received disc implants. The disc implants generated wear debris made of polyurethane, while the titanium implants of the control group generated titanium wear debris.

The clinical study included patients with problems related to radiculopathy and myelopathy, pain or neurological deficits because of pinched or damaged nerves.

These problems were caused by herniation or stenosis of intervertebral discs. An MRI image of a herniated disc, which has caused compression of the spinal cord, is shown in Figure 12. The patients were divided into two groups: one-level patients received one Bryan disc, and two-level patients received two Bryan discs, at different levels of the cervical spine.

Two years after receiving the implant, the one- level patients were evaluated using a Figure 12: Herniation of the disc questionnaire about experienced pain and between C6 and C7. Note how it is compressing the spinal cord [43]. impacts on daily activities such as movement. There was also a radiographic assessment of motion and implant stability. The results were that 45 out of 73 level-one patients were rated as excellent, seven good, 13 fair and eight patients were rated poor. The two-level patients were evaluated one year after receiving implants. 21 out of 30 patients were rated excellent, three good, five fair and one poor.

The conclusion of this study was that the amount of wear in in vivo studies was good compared to the expected age of the prosthesis. The animal study showed no signs of inflammatory response. The short-term clinical results were satisfactory, and equivalent to spinal fusion treatments [43].

3.2 In vitro studies

3.2.1 CoCr particles on the dura mater

An article written by Papageorgiou et al. [44] discusses the biological effects of CoCr particles on the dura mater, the outermost layer of the tissue that encases the spinal cord. Dura mater tissue taken from pigs was used as an organ culture model. This was meant to study the structure of the dura mater in its entirety, as opposed to cells taken from the dura mater. This allowed the structural changes of the dura mater to be studied, not just cell viability.

Dura mater tissues were exposed to two different doses of CoCr particles. There were also negative controls, where tissues were not exposed to any particles. The tissues were studied immediately after particle exposure (day zero), and after seven days. Cell viability was measured using a so-called MTT assay, where the colour changes depending on the number of viable cells.

There was no significant effect on cell viability at day zero or day seven. After seven days, there was structural damage to the dura mater tissues exposed to both doses of CoCr particles. The outer epithelial layer had detached from the rest of the tissue. The remainder of the tissue appeared similar to tissues from the control samples. Images of the detached layers are shown in Figure 13.

There were significant levels of inflammatory cytokines in the tissues exposed to CoCr particles, suggesting that CoCr stimulates an inflammatory response in the tissue.

The authors comment that the study did Figure 13: Dura mater tissue. D: Not exposed to not investigate if the structural damage to CoCr particles. E: Exposed to a lower dose of CoCr. F: Exposed to a higher dose of CoCr. Note the outer layer of the tissue affected the how the outer epithelial layer at the top of images E and F have detached from the rest of the tissue barrier function or mechanical integrity of [44]. the dura mater as a whole. Further studies are required to investigate this, as well as if other implant materials such as PEEK, stainless steel or titanium have similar effects [44].

3.2.2 SS and CoCr particles with astrocyte and microglia cells

Lee et al. have studied the effects of stainless steel (SS) and cobalt chrome (CoCr) wear particles and ions on astrocyte and microglia cells taken from rats [9]. They tested astrocytes and microglia together in a co-culture, and a cell culture with only astrocytes. Cell viability and DNA integrity were investigated in both 2D and 3D cell cultures. Cell viability was measured using an ATP-lite assay, and DNA integrity was measured using a Comet assay that measures DNA damage. Varying doses of particles were tested, between 0.05 and 50 μm3 per cell.

The results of the study showed that CoCr is more harmful to cells than SS. This affected both cell viability and DNA integrity. When CoCr was added to a combination of astrocytes and microglia, the microglia were damaged, but not astrocytes. Astrocytes sustained more damage when exposed to CoCr without microglia. This suggests that microglia protect astrocytes from damage [9].

3.3 In vivo studies

3.3.1 Particles applied to spinal cord

Cunningham et al [12] applied particles of 11 different implant materials to the spinal cord dura mater of New Zealand White rabbits. The materials included metals, polymers and ceramics. Images of particles being applied to the spinal cord can be seen in Figure 14. There was also a control group with rabbits that had sham surgeries. This means the rabbits had the same surgery done, but no particles were applied to the dura mater. There were a total of 120 rabbits used in the study, 10 in each group. Half the rabbits were euthanised and studied three months after the particles were applied, the other half after six months.

Figure 14: Posterior view of the spinal cord at the L5-6 level, after removing the spinous process and surrounding ligaments. A: without any particles added. B: with PCU (polycarbonate urethane) particles added. C: With PET particles added [12]. 20

All experimental groups had developed more epidural fibrosis than the control group, showing that the particles cause scar tissue. The rabbits who received metallic particles developed more epidural fibrosis than the ones that received polymeric particles. There were no visual signs of infection on any of the test subjects.

The levels of cytokines and macrophages in the spinal cord and surrounding fibrous tissue were measured. The control group had increased levels of cytokines and macrophages, showing that the surgery itself also triggers a biological response. All metallic implants and ultra-high molecular weight polyethylene (UHMWPE) had significantly elevated levels of IL-6 cytokines after three months compared to the other implant types and the control group. At six months, the IL-6 levels of metal and UHMWPE groups had gone down to match the levels of the other test groups, including the control group.

Macrophage levels were highest in the stainless steel and UHMWPE groups after three months. After six months, macrophage levels were still high in stainless steel, but the macrophage levels in the UHMWPE group had gone down to similar levels as the control group. At the six month mark, macrophage levels in the cobalt-chrome group had risen to high levels similar to the stainless steel group.

The particles made of polymeric and ceramic materials remained localised to the area of application, and had not spread further. On the other hand, metallic particles had spread along the spinal cord. The authors of the study concluded that there was a systemic reaction to implant debris, but the reaction was mild and not acute [12].

3.3.2 Titanium particles applied to retroperitoneal and epidural space

Chang et al. [45] compared the effects of titanium particles applied to the retroperitoneal and epidural spaces. The retroperitoneal space is the space between the peritoneum, which encloses most of the intestines, and the lumbar spine [46]. The epidural space is the space around the spinal cord, see Figure 3.

23 New Zealand white rabbits weighing 2.5 - 3 kg were used as test subjects. Titanium particles were placed in the retroperitoneal space of ten rabbits, and into the epidural space of 13 rabbits. Two rabbits were paralysed because of injuries sustained during the surgical procedure. These were euthanised immediately, and

not included in the study. After four or 12 weeks, the remaining animals were euthanised, and various tissues from the animals were evaluated.

All animals were healthy at the time of euthanisation. Particles were found in the tissues surrounding the surgical sites, as well as in other tissues studies, such as the kidneys, lymph nodes, and in some cases, the liver and the brain. There were no adverse biological effects of the particles, no inflammation, and minimal cellular response.

The conclusion of the study was that the titanium particles had travelled from the sites they had been deposited in, but there was a minimal biological response to these particles [45].

3.3.3 PEEK particles applied to epidural and intervertebral disc spaces

Hallab et al. [47] compared the application of PEEK particles to the epidural versus the intervertebral disc spaces. The hypothesis was that there would be a smaller response to debris particles in the intervertebral disc space, as it lacks blood vessels.

30 New Zealand white rabbits were used in the study, divided into three groups of ten. Group 1 had PEEK particles placed epidurally, onto the dura of the spinal cord. Group 2 served as the epidural control group. They were opened up in the same way as group 1, but did not receive any particles. Group 3 had PEEK particles placed intradiscally; inside the discs between the L4 and L5 vertebrae. The discs between the L2 and L3 vertebrae were perforated, but received no particles, to serve as the intradiscal control. Half the animals were euthanised after three months, and the other half after six months. The tissues around the placement of the PEEK particles were studied, as well as systemic tissue samples from organs and lymph nodes.

There were no signs of infection or neurological damage in any of the test subjects. PEEK particles were visible around the areas they were placed, in all individuals that received particles. There was no evidence of particles in the systemic tissues.

There was an increase in inflammatory cytokines in both the epidural and intradiscal groups. In the intradiscal group, the cytokine levels were only 10-20% higher compared to the controls. In the epidural group, there was a 50-100% increase in

cytokine levels compared to the control group. The authors concluded that the inflammatory response was much greater when the PEEK particles were placed epidurally, compared to intradiscally. Although there was an inflammatory response, the response was mild. There were no pathological changes as a result of the PEEK particles [47].

3.3.4 Cobalt chromium particles in epidural space

A study by Hallab et al. [48] measured the inflammation caused by cobalt chromium and nickel wear debris. Three different dose levels of CoCr-alloy particles were injected into the epidural space of 30 New Zealand white rabbits. A negative control group had contrast agent injected instead of particles, and a positive control group received nickel (Ni) particles. The negative control group was supposed to have less inflammation compared to the other groups, and the positive control group was supposed to have more severe inflammation compared to the other groups. Half the rabbits were euthanised and studied after 12 weeks, the other half after 24 weeks.

By the second day after surgery, all rabbits treated with nickel had neurological deficits such as paralysis, and were euthanised. The other animals appeared normal after the procedure. After 12 or 24 weeks, wear debris was still present at the injection site. Almost all animals had wear debris above and below the injection site, along the spinal cord.

The animals who received the highest dose of CoCr particles had epidural fibrosis, formation of fibrous tissue around the injection site. One animal with the highest dose had been euthanised on day eight. It had signs of necrosis, infection, and excessive inflammation. The other animals in the same treatment group did not have any signs of adverse tissue effects. No animals had any particles observed in distant organs, such as the brain, heart or lungs. These organs were also normal.

The individuals treated with nickel showed signs of necrosis (dead tissue), degeneration (breakdown) and haemorrhage (bleeding) of the spinal cord and surrounding tissue.

Animals treated with CoCr and Ni particles had increased amounts of cytokines compared to control animals without particles. The rabbits with Ni particles demonstrated the highest amounts of cytokines, even though they were only alive for 23

two days after receiving particles. The response to CoCr particles increased with higher dosages, and was higher at 24 weeks than at 12 weeks.

The authors concluded that spinal implant debris leads to an inflammatory response, and that the response to CoCr particles is milder than the response to Ni particles. The authors would like to see further studies to discern how much inflammation is required for osteolysis and implant loosening to occur [48].

3.4 Clinical and retrieval studies

3.4.1 Metallosis caused by spinal implants

An article by Ayers et al. describes three cases of metallosis caused by spinal implants [49]. Metallosis is the discoloration of tissue associated with metallic implants. The first case is about a patient who had many spinal surgeries which ended with a lumbar fusion. There was also inflammation in the muscles. Tissues were coloured black because of the consecutive operations. The patient’s explanted rods can be seen in Figure 15.

Figure 15: Spinal rods with surface damage due to fretting (black arrows) and galling (white arrows). Left image is a CoCrMoC alloy, the right image is a Ti6Al4V alloy. These rods were explanted from the patient in case 1, but similar damage was seen in the other cases [49]. The second case is about a patient who had multiple lumbar spine procedures, complicated by infection. He ended up with recurrent lung infections, as well as back and leg pain. Upon revision surgery, tissues were grey and black coloured. He also had a big amount of fluid in the area. The discoloured tissue was removed. Analysis revealed it consisted of necrotic fat tissue and fibrous connective tissue.

The third and last case is about a patient who had many symptoms in the back and legs. The patient had an operation in the lower part of the spine to stabilize the lumbar spine. The operation did not relieve her pain, and ended with a deformation of the spine. The soft tissues around the implant had also been coloured black. Revision surgeries consisted of removing the implants and surrounding tissue. New implants replaced the old implants. There were no signs of infection in the removed tissue, but the tissue was not sent for analysis. The patient did well after the revision surgeries.

The paper concluded that all explanted rods showed evidence of corrosive damage and fretting wear. This suggests that all metal implants will degrade to some extent. The discoloration of the surrounding tissue is further evidence of metallic wear particles from the implants. There was no evidence of one single mechanism degrading the implants, suggesting several processes contribute. Some patients showed signs of infection and inflammation in the tissues surrounding the implants [49].

3.4.2 M6-C artificial cervical disc failure

A case report by Clark et al. describes failure of a cervical disc replacement due to osteolysis caused by wear debris [50]. The 20-year-old patient presented with symptoms of paresthesia and cervical myelopathy. Paresthesia is the sensation of tingling or burning, or “pins and needles”. It is caused by pressure on nerves, which in turn can be caused by nerve damage or an underlying neurological disease [51]. Cervical myelopathy is caused by the compression of the cervical spinal cord. It can lead to pain and loss of fine motor skills [52]. The patient was diagnosed with degeneration of the cervical spine and disc herniation.

Treatment involved the placement of artificial cervical discs at C4-5 and C5-6. The placement of the artificial discs can be seen in Figure 16. The implants used were of the type M6-C, see Figure 17. The implant has titanium endplates, a viscoelastic PCU (polycarbonate urethane) nucleus, and an UHMWPE annulus around the nucleus. Around the annulus is a PCU sheath, designed to minimise debris migration [53].

Figure 16: Immediate postoperative images of the disc replacement implants [50]. The cervical vertebrae have been numbered. The titanium endplates are radiopaque, and show up on the X-ray images. The PCU and UHMWPE parts of the implants are radiolucent, and can not be seen on X-ray images.

Figure 17: M6-C implant. From bottom left to right: sheath, endplate, artificial nucleus and artificial annulus [54].

Initially, the patient's symptoms resolved after receiving the artificial cervical discs. Five years after the surgery, the patient developed paresthesias and decreased fine motor dexterity. Six years after surgery, the patient sought treatment for his symptoms. CT imaging revealed osteolysis of the C5 and C6 vertebral bodies (Figure 18). The implants had migrated into the vertebral bodies. MRI revealed stenosis (narrowing of the spinal cord).

Figure 18: Osteolysis of vertebral bodies [50]. CT image taken six years after the initial surgery. Note the change compared to Figure 16. The patient had a revision surgery to remove the failed implants. During surgery, a tear along the annular sheath of one of the implants was noted. A C5 corpectomy was performed, or the removal of the C5 vertebral body, a carbon-fiber corpectomy cage placed in its stead. The cervical spine was then fused from C3 to C7. The patient had no symptoms 16 months after the revision surgery. An X-ray image taken after the revision surgery is shown in Figure 19.

Figure 19: X-ray images taken after revision surgery [50]. Tissue samples were taken around the implants. Polyethylene was found in the tissue, suggesting osteolysis caused by polyethylene particles in the tissue. The broken sheath around the implant allowed it to move more than it should, increasing wear. The PCU nuclear core and annular fibers of the implant had degraded, releasing polyethylene into the surrounding tissue. The author concludes that the wear debris, and the osteolysis it caused, was the reason for the implant failure [50].

3.4.3 Dynesys spinal implants

Neukamp et al. describe the retrieval of Dynesys spinal implants from five patients [55]. The Dynesys implant was designed as a dynamic stabilisation system, to stabilise the spine without fusing vertebrae. Figure 20 shows a spine model with Dynesys implants. The point of this is to preserve the biomechanical functions of the spine, allowing normal movement of the spine. The rate of complications concerning Dynesys implants range from 19 to 34%, reported in other studies.

Figure 20: Dynesys implants on a spinal model [56]. The system consists of three parts: titanium alloy pedicle screws, polycarbonate urethane (PCU) spacers, and a cord made of polyethylene terephthalate (PET). The pedicle screws are screwed into the vertebra, through the pedicle into the vertebral body. Several pedicle screws are connected by the cord, which runs through the spacers. The cord bears tensile loads and limits flexion of the spine, while the spacer bears compressive loads and limits extension of the spine. Figure 21 shows the screws, spacer and cord, while Figure 22 shows the placement of the screws in the vertebra.

Figure 22: An illustration of the placement of the pedicle screws in the vertebra [58]. Figure 21: An image showing screws, spacer and cord of a Dynesys spinal implant system [57].

All five patients in the study had their implants removed during revision surgeries. The reason for the revision surgeries was that the patients experienced pain. Four patients had the implants for about two years, one patient had their implants for about five years. All cords showed signs of fraying, and one of the cords that had been implanted for five years had snapped. All spacers had bent, the deformation ranged between 0.2° to 13.6°. Three out of five cases showed signs of abrasive wear on the spacers. There was also deformation where the cord exited the spacer. Screws had loosened in all patients, but none of the screws showed signs of damage.

Tissues from around the implants were collected during the revision surgeries. Tissue from three of the patients contained wear debris and showed signs of macrophage infiltration and inflammation. The wear particles had a round shape, and most were in the 1-10 µm size range. The largest wear particles were in the tissues from the patient with the snapped cord. All wear particles consisted of PCU or PET, as only the cords and spacers had worn. All cases had necrotic tissue, or tissues with signs of cells dying. Some tissues developed more fibrocartilage, in response to stress.

The authors of the study assumed that the Dynesys implant is stable, as long as the implant does not fail. They acknowledge that this was only a short-term study, with few cases. The authors suggest that further long-term studies about the implant’s long-term implications are needed [55].

4 Discussion

This section contains our own thoughts and opinions of the papers included in the literature review.

4.1 Bryan cervical disc study

The clinical study rated patient outcomes based on radiological images and a patient questionnaire [43]. The paper does not explain why some patient outcomes were rated better or worse. Were there any specific factors influencing the outcome? The results of a tissue analysis would have been interesting to see. Obtaining tissue samples would be an invasive procedure, wanting to avoid that is understandable.

Other clinical studies included in our literature study, such as the papers by Ayers et al., Clark et al., and Neukamp et al. [49,50,55] have included retrieval surgeries where the implants were taken out and analysed along with tissue samples. This may give useful information about why patient outcomes differed with the Bryan cervical disc implant.

4.2 Dynesys article

The authors of the paper about the Dynesys implant concluded that the implant system is biocompatible if it does not undergo a major failure [55]. We think this is a strange conclusion to make, as the implant did break in one patient, and we consider that a major failure.

Under conflicts of interest, it is stated that one of the authors holds shares in the company making the implants. Studies have shown that conflicts of interest are associated with biases in research. These biases include only publishing certain outcomes, choosing comparators that produce favourable results, and drawing conclusions that are inconsistent with the study results [59]. There is also evidence of industry-funded studies showing more favourable results compared to studies without industry funding [60]. There is a risk that the authors of the Dynesys study exaggerated the positive results of the study, and downplayed the negative results, because they wanted the implant to be successful. The conflicts of interest, and the

biases associated with that, could explain the strangle conclusion made by the authors.

The article mentions other studies reporting complications requiring revision surgeries in 19 to 34% of patients with Dynesys implants. Other types of dynamic stabilisation systems seem to have similar revision rates. A follow up study by Reyes-Sanchez et al. regarding the Accuflex rod system reports 22.22% of patients requiring hardware removal after two years [61]. This allows us to assume the Dynesys implant is neither better nor worse than similar implants.

4.3 Animal studies might not accurately predict results in humans

Animals may not react the same way as humans to an implant or its wear debris. Even with positive results from animal studies, it is important to remember that the result may not be the same in humans. In an article, Bracken argues that while some animal studies accurately predicted the outcome in humans, there are cases where the results of animal and human studies were very different [62]. One of the main reasons for the differing results is that the animal studies are poorly designed. In another article, Pound et al. list animal study design problems [63]. Some of the problems include that the test animals have not received the illness or injury in the same way that humans do. This means that the animals tested do not have the same medical conditions as the humans receiving treatment for the same ailment. Another issue is that doses tested on animals may be irrelevant to humans. Lastly, small experimental groups are insufficient for statistical significance.

Similar problems can be seen in the animal studies included in our literature review. All implants and particles were tested in healthy animals, without underlying spinal problems. There were about 10 animals in each group in almost all studies, which may not be enough for statistical significance. Cunningham et al. placed a large, one-time dose of particles onto the dura mater of rabbits [12]. In the human body, wear particles are released from implants over time, and not all at once.

4.4 Use of different animals to study spinal implants

Goats are used to study spinal implants, partly because they weigh about 60 to 80 kg, similar to humans. Goat and human vertebrae are also of similar shape and size [64]. This means that the vertebrae of goats and humans carry similar loads. Of course, humans are bipeds (walk on two legs), and goats are quadrupeds (walk on four legs), so the loads on the spines are not in the same direction. The loads on goat and human spines are still more similar than in rabbits weighing only a few kilograms [65], or in rats or mice that are even smaller.

An article by Smit discusses the relevance of using quadruped animals as in vivo models of human spines [66]. The vertebrae of quadrupeds are stronger and denser than human vertebrae, which means a screw will have a stronger hold in a quadruped. This means that implants may loosen more easily in humans than quadrupeds. The author of the article also states that the load on quadruped and human spines are still quite similar, which justifies the use of those animals when testing spinal implants. The article concludes that although there are some differences between the spines of quadruped animals and humans, they are similar enough to be useful in studies.

Another article by Drespe et al discusses the use of different animals to study spinal fusion [67]. Primates (such as monkeys and apes) are most like humans genetically, and they also walk upright, unlike most other mammals. Primates are not widely used in animal studies. Many laboratory facilities are designed to house smaller animals, such as rabbits, rats, and mice. Few facilities can accommodate primates. Primates are also more expensive to house, and difficult to purchase. Public opinion is also a factor, as testing on primates is seen as less acceptable compared to other animals.

Our opinion is that primates are a better choice of animal for in vivo tests from a scientific viewpoint, as they are most like humans. Meanwhile, we understand the difficulties involved with using primates. In our literature study, goats and rabbits were used for in vivo testing [12,43,45,47,48]. It may have been difficult or even impossible to perform tests on primates, so they might have chosen other mammals instead. We believe it is better to have results from studies with animals that are less like humans, than no in vivo results at all. 33

4.5 Ethical aspects of animal testing

In one of the studies described in the literature study rabbits were injected with nickel particles as a positive control, to give a more severe response than the CoCr particles that was the main focus of the study [48]. These particles caused major damage to the rabbits’ spinal cords, and by day two all had been euthanised. This was probably because they were in such bad condition that their suffering had to be ended for animal welfare reasons. We think the positive control groups given nickel particles resulted in unnecessary suffering of animals, and should have been avoided. Could the study not have been done, and reached the same results, without the positive control group? The study would have shown a significant difference between the negative control group and the individuals who received CoCr particles anyway.

The Three Rs is a set of guidelines for more ethical animal testing, and is also part of Swedish and European law regarding animal testing [68]. The guidelines include reducing the pain and suffering of laboratory animals as much as possible [69]. The positive control group of the study mentioned above was perhaps not in accordance with these guidelines.

4.6 The perfect implant material does not exist

This literature study has covered many different implants made from many different materials. Some materials seem to be better than others. For example, stainless steel implants have been shown to be less irritant than cobalt chrome [9]. Metals and UHMWPE particles cause greater inflammation than particles from ceramics and other polymers [12]. Titanium is more biocompatible than stainless steel and cobalt chrome [11]. Implants can also break because of material fatigue [58].

All implant material types have advantages and disadvantages [11]. No implant is perfect, without any negative effects or properties. We hope further scientific research may one day lead to the perfect implant material.

5 Conclusion

All spinal implants degrade to some extent in the body. This can happen due to wear or corrosion. This generates wear debris particles. The particles vary in shape and size depending on the material, and the type of wear the implant is subjected to. The particles can spread from the area surrounding the implant to nearby tissues. The body reacts to the particles, triggering an immune response. This immune response can lead to inflammation, pain, the breakdown of bone, and implant loosening. The biological response to particles varies in type and intensity depending on the amount and type of particles, as well as the location of the particles.

A study has found cobalt chrome particles to be more toxic than stainless steel particles. Another study found that metal and UHMWPE particles cause more inflammation than particles made of other polymers and ceramics.

Different cell types can have different reactions to the same type of wear particle. Microglia seem to protect astroglia from the damaging effects of wear particles. It is important to study the reaction of the body as a whole system, and not just one or two types of cells, as the results may differ.

Several studies conclude that particles cause an inflammatory response, and that the response is mild. The level of inflammation required for the response to be severe, and lead to negative effects such as osteolysis, is unclear.

5.1 Further studies

We would like to see the effect of wear particles on microglia studied, as it appears to not have been done before. This can be done through in vitro studies, where wear particles are added to microglia cell cultures. The effect on the microglia cells can be measured by cytotoxicity tests [40,41]. It would be interesting to compare the effects of wear particles made of different materials, as well as the amount of particles needed to cause a significant microglial cell response.

One study showed that cobalt chromium wear particles lead to the loosening of the outermost layer of the dura mater [44]. It would be interesting to study if that affects the function or integrity of the dura mater, and if particles made from other materials have the same effect.

Several studies concluded that wear particles induced a mild biological reaction, with inflammation that was not severe enough to have any significant effects [12,45,47,48]. The level of inflammation needed to cause significant negative effects such as osteolysis is unclear. We would like to see studies investigating where this threshold is.

The in vivo studies mentioned in this literature study were much shorter than the intended lifespan of spinal implants [12,43,45,47,48]. We would like to see studies on whether short term tests accurately predict the long term results.

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