Recent progress in in vivo studies and clinical applications of magnesium based biodegradable implants -A review

Prithivirajan Sekar,Narendranath S,Vijay Desai

Corrosion Engineering Lab,Department of Mechanical Engineering,National Institute of Technology Karnataka,Surathkal,Srinivasanagar,Mangalore 575 025,Karnataka,India

Abstract Biodegradable magnesium has regained great attention due to its ability to temporarily offer mechanical strength and degrade completely once the injured pathological tissue is healed.A few clinical applications of Mg-based implants were reported in the last century.However,the knowledge and experience is being gained continuously by studying the host response and degradation behavior of Mg implant in animal models and clinical trials.This led to the development of commercial products emerging from Europe and Asia very recently.The potential of Mg implants in repairing fractures at upper and lower limb of large,small animal models and humans is compared and discussed in detail.In addition the possible future Mg implants that might treat problems concerning to urology and gynecology are reviewed.

Keywords: Biodegradable;Magnesium;Fracture healing;Translational medicine;Clinical trials.

1.Introduction

Magnesium and its alloys are new class of materials with unique capability of degrading inside animal and human body.The density of Mg is comparable to human bone and is also one of the essential elements in the human body [1].The firs use of magnesium as a biodegradable implant dates back to 1878.Magnesium implants have already proved to be successful especially in case of healing certain types of fractures in both in vivo and humans [2].However,rapid degradation in vivo and hydrogen gas formation remained major hindrance to extend their application in various surgical areas.The minimal knowledge and experience in vivo and mediocre development in technology forced the surgeons to choose permanent implants over Mg during the 19th century [3].The permanent implants are now widely used and commercially available.However,they require an additional surgery for explantation and also stress shielding effects are encountered.Currently,Mg regained more attention for treatment of orthopedic implant application.Unlike permanent implants,Mg implants corrode inside animal or human body with simultaneous replacement of native bone.Since,the complete degradation of Mg alloys takes place in vivo the need of secondary surgery is also eliminated [4].In order to develop a successful biodegradable implant,researchers generally carry out in vitro,in vivo evaluation and clinical trials[5].However,each method has its own advantages and disadvantages and is shown in Table 1.In vitro evaluation detects only degradation rate and loss of mechanical integrity.In addition,the hydrodynamic fl w can also be simulated using a peristaltic pump in vitro studies.In contrast,the new bone formation,bone interface strength,gas formation,inflammator reactions can also be detected in vivo.The in vivo method is almost equivalent to clinical trials.However,water content and blood fl w rate of rats,rabbits and humans varies significantl [6].Hence,the clinical trials with more number of patients included in each study is most preferred to evaluate the medical fitnes of implants.From available literature,it is found that most ofreviews on Mg focused on history,alloy composition,in vitro evaluation and comparison of in vitro and in vivo degradation and additive manufacturing [7-15,50-52,64].In contrast,the current review focuses on mechanical integrity of Mg alloys after in vitro,recent developments in vivo studies and use of novel Mg implants in clinical trials.

Table 1 Methods to evaluate the biodegradability of Mg.

Fig.1.Mg implant degradation as a function of fracture healing.

2.Bio mechanical behavior of Mg alloys in vitro

Fig.1 represents the stages of bone remodeling after fracture and degradation of Mg implant.It enlightens the two main requirements of Mg implant (i) maintenance of mechanical integrity until healing(ii)complete degradation or absorption of Mg inside human body[5,8].In vitro assessment is the most basic and easier way of getting insights into the performance of Mg alloys for biomedical applications.From the literature most of the reports tested Mg alloys using immersion,hydrogen evolution and electrochemical methods in simulated body fluid (SBF) [15].In addition,it is well established that corrosion rates obtained in vitro does not match with the in vivo studies.Walker et al.[115] evaluated the performance of fi e Mg based alloys in Earle’s balanced salt solution(EBSS),minimum essential medium (MEM) and MEM with proteins upto 21 days immersion.These fi e Mg alloys were also implanted in the Lewis rats for same duration.They concluded that the corrosion rate obtained using EBSS is similar to that obtained in vivo.Fig.2 depicts the commonly used corrosion medium for evaluating the corrosion rate of biodegradable implants [116].Mei et al.[116,117] comprehensively reviewed the corrosion medium used in evaluating the biodegradable implants.They suggested that cell culture mediums are not suitable for long term immersion studies due to microbial contamination.Based on the review [117] and research of Walker et al.[115],it is reasonable to conclude that EBSS without of any proteins is most applicable medium for corrosion testing of bio absorbable metals.For an implant to be used in clinical applications,in addition to lower corrosion rate sufficien mechanical integrity till healing period is also mandatory.Hence,the recent studies evaluated the mechanical properties of Mg implants before and after immersion in SBF.Yield strength of GZ60K and GZ61K deteriorated upto 9% and 11% respectively after 21 days immersion in SBF [16].In another study Mg-Zn-Ca clip was tested as potential candidate as haemostatic clip for wound closure application.Approximately 80% loss in mechanical properties of clip was observed after 14 days immersion in SBF [17].The mechanical properties of Mg-2Zn-0.8Zr was tested after 28 days immersion in simulated body flui (SBF).The yield strength of Mg-2Zn-0.8Zr was decreased from 222MPa to 181MPa after 28 days [18].Similarly,Hou et al.[19] developed ZX11 Mg for bone plate application.After 28 days of immersion%elongation of both rolled and annealed ZX11 Mg alloy deteriorated to less than 4% elongation.This is probably due to relatively lower thickness of 1.8mm ZX11 plate used.They also reported a non-linear trend in deterioration of mechanical properties with respect to time.This clearly indicates that magnesium corrosion is dynamic in nature.Our research group improved the mechanical properties and enhanced the corrosion resistance of various Mg alloys using equal channel angular pressing (ECAP) [106-112].Our recent study also investigated the mechanical integrity of ZM21 Mg after immersion in Hank’s solution.The % elongation of equal channel angular pressed ZM21 were relatively greater than extruded ZM21 Mg before and after immersion.The mechanical integrity of rolled and ECAPed ZM21 Mg is represented in Fig.3.Even though the above studies evaluated the mechanical integrity after degradation they were not able to mimic the human body environment due to the absence of fl w.The significanc of all the above finding are tabulated in Table 2.

Fig.2.Commonly used corrosive media for testing bio-absorbable metals [116].

3.Flow corrosion to bridge gap between in vitro and in vivo monitoring

In order to bridge the gap between in vitro and in vivo testing,some researchers used a dynamic or fl w corrosion set up using peristaltic pump.This dynamic fl w corrosion set up was firs developed by Mantovani and coworkers [20].They analyzed the potential of AM60 to be used as a potential biodegradable coronary stent.The corrosion rate of AM60 Mg was significantl different in static and fl w conditions.This was mainly because of the shear stress generated during fl w of modifie Hank’s solution.SBF was fl wn over porous Mg in laminar condition by varying three different fl w rates to mimic bone marrow environment.At the end of 3rd day the 95% loss in mechanical integrity was observed at the fl w rate of 0.8ml/min [21].Three different solutions Normal Saline (NS),Phosphate Buffer Solution (PBS) and simulated body flui (SBF) were fl wn over AZ31 Mg alloy at different fl w rates.They also established a relationship between corrosion rate and induced fl w velocity.However,the duration of the study was only 24 h which is insufficien to mimic in vivo corrosion environment [22].Also,the in vitro studies involving fl w cannot assess the bone-implant interaction and functionality of organs.Hence,animal models were chosen to better evaluate the degradation performance,bone implant interface strength,functionality of organs after implantation of Mg.Table 3 depicts the summary of the Mg alloys and its corrosion behavior testing in dynamic condition.

Table 2 Bio mechanical behavior of Mg alloys in vitro.

Table 3 In vitro degradation behavior of Mg alloys under fl w conditions.

4.Corrosion of magnesium alloys in animal models

It is well known that during vivo trials various animal models and implantation site are chosen based on the desired application.Also,there is no standard procedure for deciding the size and implant location.In addition,the water content and blood fl w rate of sheep,miniature pigs,rats and rabbits vary significantl .This is a great hindrance to evaluate and compare the potential of various Mg and its alloys used as implant.Hence,in the present review the studies carried out in each animal model is discussed separately in detail.

Fig.3.Improved mechanical integrity of ZM21 Mg alloy after equal channel angular pressing.

4.1.Mg based implants in large animal models

Fig.4.ZX00 lean Mg alloy implanted in the tibia with and without osteotomy in a growing sheep model [72].

Witte et al.[23] studied the interactions of AZ series(AZ31,AZ91) and rare earth (WE43,LAE442) containing magnesium alloys at bone-implant interface in guinea pigs.The degradation of magnesium alloys was completely dictated by the composition of alloying elements.The fluoro scopic images taken at the bone implant interface revealed the formation of newly formed bone in periosteal and endosteal region.The native bone formation was better in Mg implants than that of degradable polymer.Wang et al.[65] reported that the wound healing patterns in miniature pigs are very close to that of humans.However,fixin cranio-maxillofacial region using biodegradable implants are seldom researched.A new type of hollow screw made of WE42 Mg was coated with PEO technique and inserted in the mandible of minipig.The design of riveted hollow screw eliminated the need of applying torque and the possible failure of implants during insertion [24].Mg-Zn-Sr alloy ring was used a connector in the intestine of bama miniature pigs.14 days post-surgery,the biological functions of intestine restored back to normal without affecting any organs [66].The finding of this study is in accordance with the earlier research in which Payr used Mg as connectors for vessel anastomosis [67,68].The MgYREZr Mg alloy and titanium implants were used for repairing the frontal bone in a mini pig.Osteotomies of both Mg and Ti groups were healed.Lacunas were formed between the implant and bone.Their formation did not affect healing process but their significanc is unknown [69].A rectangular WE43 Mg implant with and without plasma electrolytic coating (PEO) was inserted in the nasal bone of minipig.The PEO coating was successful only in reducing the gas formation and increasing the Ca-P layer formation on WE43.The bending strength of PEO coated and uncoated WE43 decreased from 92N before implantation to 86N after 24 weeks [73].The same research group explored the possibility of using WE43 Mg in repairing the frontal bone of miniature pigs.The PEO coating increased the corrosion resistance,new bone formation and reduced the gas formation in vivo.The plate screw system was not displaced over 24 weeks as observed from micro CT.They reported that the wound healing pattern in miniature pigs is very similar to that of humans.All the osteotomies in orbital rim and zygoma were healed without any dislocation of bone or breakage of implant.In addition the osteotomies performed at the ribs were also healed.However some the Mg screws implanted at ribs were broken due increased mechanical load during breathing.The authors concluded that the degradation rate does not vary significantl in miniature pigs even though WE43 Mg is implanted in different sites [74-77].However,in small animal models such as rats and rabbits it is reported the degradation rate of Mg varies with implantation site.Huang et al.[70] chose a goat model in which they implanted a pure Mg for treating fracture that occurred at femur head.The trend of bone remodeling observed during the period of 48 weeks were similar to that of the human bone.This is the firs study to explore the use of Mg implants in goat model.Only 45% of implant was degraded by the end of 48 weeks.This indicates the relatively higher corrosion resistance of pure Mg which might be due to absence of secondary phases.Magnesium phosphate cements(MPC) were implanted into defects at unloaded femora and weight bearing tibia of sheep.Trabecular new bone formation replaced the defects after complete degradation of MPC at the end of 10 months [71].ZX00 lean Mg alloy was implanted in the tibia with and without osteotomy in a growing sheep model as shown in Fig.4.Fig.5 indicates clearly that fractures consolidated within a period of 12 weeks and there was no significan difference in the healing pattern of tibia with and without osteotomies.They also reported that the screws broke by end of 12 weeks due to lengthening of tibia bone.Fig.6 represents the histology of broken ZX00 Mg implant [72].Weinberg and coworkers [83] firs reported the comparison of degradation performance and new bone formation of ZX00 Mg alloy in small animal (SD rats) and large animal (sheep) and is represented in Fig.7.They observed homogenous degradation and comparable new bone formation of ZX00 Mg both in SD rats and sheep.However,they recommended the use of small animals for initial safety and large animal models for load bearing application.The summary of in vivo testing carried out in large animal models such as sheeps,goats and mini pigs are highlighted in the Table 4.

Table 4 Mg alloys implanted in large animal models.

4.2.Implantation of magnesium in rabbits -a smaller animal model

Mg-6Zn alloy was chosen as a candidate for testing biodegradability in rabbit femora.A gap was observed between the new bone and implant probably due to continuous degradation of Mg alloy.Approximately,87% of Mg-6Zn degraded within 14 weeks [25].New bone formation and bone implant interface of Mg-1Sr alloy was 48% and 16%greater than Mg respectively.The authors concluded that Sr addition increased the corrosion resistance of Mg.The rabbits also metabolized Sr and their release was minimal so that they were not accumulated in the organs [26].ZEK100Mg completely degraded in rabbit tibiae at the end of one year.This complete degradation is mandatory because the secondary surgery for implant removal is no longer required[27].However,the mechanical integrity of implants is also very important to ensure the longevity of magnesium implants.Three point bending test was used to evaluate the biomechanical integrity of three Mg alloys.After 6 months explantation from rabbit tibiae the ranking of bending strength was as follows LAE442>MgCa0.8,WE43[28].The same research group extended the study duration to 12 months.A deterioration of bending force was found to be 86% and 71% in Mg Ca0.8 and LAE442 Mg respectively.Also,the degradation in Mg Ca0.8 Mg was relatively more pronounced than LAE442.Surprisingly,the micro CT scores indicated that the bone implant contact was greater in Mg Ca0.8 than LAE442.The authors concluded that the possible reason could be higher content of Mg in Mg Ca0.8 alloy [29].The LAE442 Mg with fin grains obtained from extrusion exhibited the lowest corrosion rate (0.0134mm/yr) when compared to cast alloy [30].Also,the mechanical stability of extruded LAE442 Mg was always greater that the cast alloy i.e.before and after 27 weeks implantation.The mechanical integrity of yet other novel magnesium alloys ZEK100 and AX30 in rabbits was evaluated by same research group.The results indicated that the biomechanical properties of ZEK100 were comparable to LAE442 alloy.While,AX30 Mg alloy is not suited to heal weight bearing bones.In contrast to conventionally used ex vivo techniques,this study used in vivo micro computed tomography.This technique has the greatest advantages such as (i) quantifying corrosion using three dimension imaging(ii)assessment of implant during any desired time without harming the animal [31].In another study from same research group the biocompatibility of LACer442 (cerium based rare earth alloy) was researched while LAE442 (mixture of rare earths)alloy served as reference.Unfortunately,LACer442 Mg alloy failed within three weeks of implantation due to rapid degradation.Bleeding occurred at periosteal region of tibiae along with gas formation which is evident from micro CT and histology [32].Rare earth Ce,La,Nd exhibited corrosion rate of 1mm/year measured using in vitro for 4 weeks [33].Rare earth containing LAE442 alloy exhibited lower degradation rate in pigs when compared with Ca and Al containing Mg alloys [23].Pitting corrosion is major problem reported in alloy which degraded rapidly in vivo for instance in Mg Ca0.8 and LACer442 Mg.In addition some researchers developed Magnesium based bioceramics by powder metallurgy techniques.New bone formation of MgP-2Sr increased 46%compared to the Mg-P (magnesium phosphate) samples when implanted in a rabbit.The same research group doped various elements Zn,Si and Sr into Mg-P.The new boned formation of all the doped Mg-P were found to be greater than that Mg-P without doping.The new bone formation percentage of MgP,MgP-0.5% Zn,MgP-0.5% Si,MgP-0.5% Sr were 48,55,61 and 73%,respectively [78-80].High purity Mg with negligible amount of secondary phases could relatively exhibit less corrosion rate than Mg with alloy combinations.The rolled HP Mg evinced bone implant contact (BIC) 30%greater than PLLA screws in range of 4 to 24 weeks.The breakage of PLLA screws were observed at 16th week [47].Even though HP Mg is promising candidate for orthopaedic implant application.The mechanical properties can be signifi cantly enhanced by alloying.The effect of alloying may either decrease the corrosion resistance due to formation of microgalvanic cell or increase the corrosion resistance when secondary phase particles inhibit corrosion bioabsorbabale.Mg Ca 0.8 was compared with conventionally used surgical steel SS316L.The inflammatio rate of both implants was equal until 6 weeks in rabbit tibiae.However,at the end of 8th week,inflammatio rate of Mg Ca 0.8 alloy increased comparatively.This is due to the degradation of Mg implants in vivo [48].In all the above discussed studies,magnesium implant in the form of rods is fi ed to rabbit tibiae or femora.In contrast,other studies were done which mimics the more practical situation encountered in human body.These includes fracture at ulna,mandible and tearing of anterior cruciate ligament.

Fig.5.X-ray imaging performed 3,6 and 12 weeks following surgery of the right osteotomized leg and the left intact leg.Inserted screws are displayed on both sides (indicated by arrows).The fracture line in the right leg cannot be depicted in the X-ray image scans [72].

4.2.1.Healing fractures at ulna

Implantation of 99.9% pure Mg (plates and screw combination) in fractured ulna of rabbit healed successfully by 16 weeks.The post mortem analysis revealed that fl xural strength of Mg implanted ulna is comparable to native ulna[45].MAO coatings with 10 and 20 μm were applied to bare Mg-Zn-Ca alloy scaffolds of 15mm length and 10mm length.These scaffolds were used as substitute for healing 15mm bone defect created in the rabbit ulna.The bone defect was completed healed in both bare Mg alloy and MAO coated groups by the end of 18 weeks [82].

4.2.2.Reconstructing anterior cruciate ligament

Anterior cruciate ligament (ACL) bridges thigh to shin bone.i.e.at knee joint.The ligament tear is prone to occur when involved in physical activities especially sports.Cheng et al.[46]carried out an interesting study on reconstruction of ACL in rabbit model.Un-calcifie and calcifie fibro-cartilag tissue is formed between bone and tendon graft after 16 weeks implantation of 99.98% pure Mg.Range of motion (ROM) at knee joint and ultimate load to failure of tendon graft of both Mg and Ti were similar.

4.2.3.Bone formation at mandible

Pull out force of pure Mg,AZ31 Mg and stainless steel(SS) was equivalent to 40N when tested after inserting the screw in synthetic bone.Histology revealed almost complete degradation of pure Mg by 12 weeks at rabbit mandible.In contrast,AZ31 Mg was completely visible and new bone formation was also observed [49]

4.3.Evaluating the safety of Mg implant in the smallest animal models -rats and mice

ZM11 Mg alloy was implanted into rat femora to evaluate the performance of alloy in vivo.The alloy degraded 54% at 18th week post implantation.Calcium phosphate layer is the key element that accelerates the new bone formation.Histology micrographs indicated no significan difference between newly formed and existing cortical bone [54].The same research group extended the present work.In this study ZM11 Mg alloy was implanted in SD rats at cortical bone and bone marrow.The implant at bone marrow degraded completely while only 10% degradation was observed at cortical bone after 6 weeks implantation.This study clearly indicates that the degradation varies significantl with respect to implant location.Histological studies at bone implant interface indicated that with increase in number of weeks new bone tissues were observed.A layer rich in calcium phosphate and membrane comprising of fibroblast were found at Magnesium implant side and new bone tissue side respectively [53].These find ings are in accordance in vivo studies carried out recently[23].Interestingly,Castellani et al.[55] implanted Mg-Y-Nd-HRE and Ti 6Al 7Nb in SD rats in a direction perpendicular to the length of femoral bone.They created in house mechanical set up to determine the strength at bone/implant interface.The contact established between bone/implant interface and the volume ratio of trabecular bone to total tissue volume was relatively higher for Mg implants than Ti 6Al 7Nb.ZX50 Mg degraded completely by 8 weeks,while the residues of WZ21 were found even after 24 weeks inside femur of SD rats.The relatively slower degradation and minimal gas formation of WZ21 resulted in better bone recovery of WZ21 Mg.Though WZ21 Mg exhibited good biocompatibility in vivo,the release of Yttrium in surrounding bone tissues was reported to be toxic.In addition the growth plates are responsible for the longitudinal lengthening of bone.Weinberg and coworkers [84-88] firs explored the interaction of ZX50 and WZ21 Mg implants inserted in femoral growth plate of SD rats.The growth plate at femora of SD rats implanted with WZ21 was comparable to control group due to uniform degradation and less gas formation after 52 weeks.Femoral growth plate did not recover completely due to relatively faster degradation in ZX50 Mg.

Fig.6.Histological depiction of two intact (3 and 6 weeks) and one broken screw (in the 12-weeks group) with transepiphyseal position [72].

Fig.7.ZX00 Mg alloy in small animal (SD rats) and large animal (sheep) [83].

4.3.1.Treatment for fractured mice

Most of the in vivo studies were carried out using drilling a hole in femoral bone and insertion of Mg implants.In contrast,recent studies intentionally fracture the femora to mimic actual fracture condition.One such study compared the degradation of Mg-2Ag with and without fracture to inert steel implants.Mg-2Ag completely degraded rapidly within (19 weeks) in fractured condition to that of unfractured one (30 weeks).The size of femoral bone increased to 40% (unfractured) and 70% (fractured) [56].

4.3.2.Failure of implants due to rare earth accumulation

Fig.8.Gadolinium accumulation in SD rats at different time periods post implantation of Mg-10 Gd alloy [57].

In contrast to uniform degradation observed in vitro studies Mg 10 Gd exhibited peculiar corrosion mechanism called disintegration in vivo due to which the alloy degraded into fin particles.The rapid disintegration of this alloy in Sprague Dawley (SD) rats is due to micro-galvanic corrosion.This is attributed to 10% gadolinium in Mg-10Gd alloy.Also,the rats were unable to excrete the gadolinium that got accumulated in kidney,lung liver and spleen.This Gd accumulation in SD rats upto 36 weeks is represented in Fig.8 [57].

4.3.3.Hydrogen micro-sensor to assess in vivo corrosion

Zhao et al.[58] implanted ZK60,AZ31,Mg8H (a high purity single crystal Mg) Mg subcutaneously into mice and measured the hydrogen gas evolution using micro sensor.The same research group extended the investigation on LAZ611 and WKX41 Mg alloys by using visual hydrogen mapping sensor.This method is simple,cost effective in contrast to micro CT and x-rays.However,it has some of the disadvantages such as less sensitivity,longer time to obtain results and sometimes mice scratch the sensor [62].

4.3.4.Renal vessel occlusion

Biodegradable clip made of Mg-Zn-Ca alloy was used to suture the renal blood vessel.The renal vessel was healed completely by the end of 4th week without any adverse effects [59].The significan results obtained after implantation of Mg in small animal models such as SD rats,mice and rabbits are tabulated in the Table 5.

4.4.Bio-coatings to improve the mechanical integrity of magnesium

It is well established that the coatings on surface of Mg inhibit the corrosion.Hence,various researchers coated Mg with single,double or composite layers [63].Magnesium flu oride coating was applied to LAE442 alloy and tested in rabbits.The degradation rate of coated LAE442 was observed to be relatively lower than uncoated LAE442 Mg for 12 weeks[34].Fluoride coating inhibited the corrosion of AZ31 Mg screw for 3 months.Interestingly,fluorid promoted the regulation of bone morphogenetic protien (BMP -2) type I collagen.Similarly,AZ31 Mg coated with beta tricalcium phosphate enhanced BMP-2 over 12 weeks [35].However,time period of three months is not sufficien to understand the mechanism of ion release and osteogenic action [36].Similarly,Mg-3Zn-0.8Zr Mg coated with MgF2and Ca-P was implanted into femur of Japanese white rabbits.The loss in volume after 3 three months explantation was measured be 37,24 and 23% for uncoated,Mg-F2and Ca-P respectively.The new bone formation and tissue mineral density was relatively higher in Mg-F2than Ca-P and uncoated Mg.The authors reported that Fluoride ions promoted the new bone formation and was also in accordance with the previous study [37].A composite layer with combination of hydroxyapatite (HA),octa calcium phosphate and MgO on top and bottom of Mg-2Zn-Ca alloy were coated.Composite coatings improved the life time of MgZnCa alloy by 12 weeks.In addition,calcium and phosphate ions released from coating aided in formation of new bone [38].Similarly,MAO coating and Fluoride containing HA/MAO composite coating on AZ91 Mg improved the bone formation by 30% and 60%,respectively [39].It can be summarized that,the composite coatings control the release of Mg ion in blood plasma.Mg ions contribute to new bone formation upto certain level.However,the excessive release do not promote new bone formation.The torques were measured during explantation of AZ31 B screws.Bone implant interface strength after 21 weeks were ranked as follows Si coated AZ31>Ti>AZ31>PLLA.Bone interface strength of coated Mg alloy was greater than that of Ti alloys.These coatings degradation was not evaluated until they completely dissolve.Also the effect of these coatings on internal organs is not studied [40,41].Al2O3coatings on AZ91 improved the corrosion resistance.This resulted in controlled release of Mg ions in the surrounding bone.Hence,the new bone formation and the presence of osteoblast like cells were relatively higher when compared to the untreated AZ91[42].MAO coatings on ZX50 inhibited the release of Mg ions in initial 2 weeks post implantation.However,in the later stages i.e.-16 weeks,the degradation rate of MAO coated ZX50 Mg increased rapidly compared to un-treated ZX50[43].In yet another study,ZK60 coated with Mg2SiO4coating using MAO technique was implanted into rabbit femora.Even though the coating inhibited the initial stage of corrosion (2 weeks),in the later stages both coated and bare ZK60 alloy degraded completely by 12 weeks [44].This behavior of coating is due to the following disadvantages (i) it has pores where the corrosion is tend to localize (ii) after degradation over time,the coated and un-coated areas from galvanic cells accelerating corrosion.In addition,both ZK60 and ZX50 has 6% zinc which acts as cathode,forms micro-galvanic cell and accelerate the degradation in vivo.In contrast,some studies reported that the coatings enhances the degradation in vivo.For instance,Ca-Sr-P coated Mg evinced improved new bone formation and corrosion resistance in vivo.However,this study was carried out only for a duration of 4 weeks [81].Hence,it is reasonable to believe that once the coating is partially degraded it tends to form of galvanic cells that will accelerate the corrosion.The summary of coated Mg implants inside animal models are tabulated in Table 6.Possible Future generation Mg implants in urology and gynecology are also reviewed.When urinary duct is blocked by kidney stone or tumor,a stent is implanted to facilitate the fl w of urine from kidney to bladder.Commercially available ureteral stents made of poly urethane (PU) causes infection due bacterial accumulation.Pure Mg and AZ31 and Mg-4Y alloys were tested in vitro using artificia urine (AU) to investigate their degradation behavior.All Mg alloys exhibited more antibacterial nature compared to PU.Hence,Mg possibly has potential to overcome the problems associated with bacterial colony formation and eliminate the need of secondary surgery for implant removal [60].In women,birth control measures are mandatory especially immediately after giving birth to a child i.e.planning the time period between births of their children.This family planning technique is called postpartum contraception.While there are number of techniques available for family planning,intra-uterine device (IUD) is the best recommendation for postpartum contraception according to world health organization (WHO).However,the currently used Dl-lactide-co-glycolide polymer (PDLGA) is not thermally and mechanically stable.To overcome these limitations,Mg alloys were studied in vitro and vivo.Mg alloys were viable to uterine cells in vitro [61].

Table 5 Biodegradation and new bone formation of Mg implants in small animal models.

Table 5 (continued)

Table 6 Performance of coated implants in animal models.

5.Clinical trials and commercialization of magnesium implants

5.1.CE approval of MAGNEZIX implants

In 2013,Syntellix AG,a German based company got Conformite Europeene(CE)approval for Mg implants.This is the greatest achievement in the history of degradable implants and is undoubtedly a milestone.The products developed in the series were named MAGNEZIX.The company also mentioned the list of surgical areas and corresponding fractures where it is implanted.This innovative degradable implant is currently being used in 15 countries where over 5000 surgeries were performed in the recent years [89,90].

5.1.1.Foot and ankle surgery

Windhagen et al.[91,102] used MAGNEZIX screws for correction of Hallux valgus deformity.They compared the performance of Mg group with Titanium group for a period of 6 months.Pain assessment,range of motion at metatarsophalangeal joint and radiological examinations revealed that the healing efficien y of Mg was similar to Ti implants.Hydrogen gas was observed at the firs month during radiographic examination and disappeared after 3 months.In contrast to Chevron osteotomy performed,Choo et al.[103] used scarf technique for the correction of hallux valgus deformity.Short form 36 assessment results of Mg was better than Ti.But,according to the radiology,enhancement in Ti group was greater than Mg after 12 months follow up.Atkinson et al.[104] also used scraf osteotomy technique for correction of hallux valgus deformity using MAGNEZIX implants in 11 patients.The patient reports post-surgery indicated that they can be used in place of titanium implants.The previous studies on correction of hallux valgus deformity reported less number of patients.In contrast,Klauser [105] involved 100 implants made of Mg and Ti and compared their performance.Delayed wound healing was observed in 3 Mg and 4 Ti implants out of 100 implants.In summary,most of degradable Mg was compared with non-degradable Ti group.The reports indicated that the Mg implants were not inferior to Ti in case of performance and therapeutic efficien y.The firs use of MAGNEZIX implants in trauma case was reported by Aktan et al.[97].Metal screws made of MgYREZr were successful in fixin the lateral malleolar ankle fracture in a 43 year old female patient.6 weeks after implantation,a radiolucent area appeared.However,the radiolucency disappeared after 17 months.Another case study involved a 19 year old female with displaced fi ula was treated using MAGNEZIX implants.In,both the case studies range of motion of ankle was restored to normal after a year [98].According to Meyer-Mckeever,the fractures at tibial spines are classifie into types I,II,III and IV based on the lesions observed.Type I and II fractures could be healed by conventional treatment without surgery.However,for the treatment of type III and IV fracture surgery is mandatory because it directly affects anterior cruciate ligament.Hence,MAGNEZIX compression screws were employed in treatment of three such patients below 20 years of age.International knee documentation (IKDC) scale and lysholm values after 12 months indicated that the functionality of knee recovered significantl .The Mg screws were absorbed after 6 months and replaced by native bone at the end of one year [99].Trauma at foot and ankle region generally during sports results in osteochondral lesions of talus(OLT).The OLTs are usually classifie based on plain radiography,computed tomography (CT) and magnetic resonance imaging (MRI).Accordingly,based on the clinical finding the lesions greater than 15 mm2should be treated with osteotomy.First trauma surgery of MAGNEZIX implants were carried out by Kose et al.[101].Secondly,the same research group compared bio-absorbable Mg versus permanent Ti implants which were employed in healing medial malleolus fracture.While there are several osteotomy techniques available Acar et al.[100] chose biplane chevron osteotomy.The find ings implied that both Mg and Ti implants were successful in treating OLT.This is mainly because of the advantages such built in locking property and the larger surface area that supports healing.

5.1.2.Repairing fractures at mandible,wrist and elbow

Biber et al.[92] reported a case where they used MAGNEZIX for the fixin intra-articular fracture occurred atelbow.It is interesting note that the patient was 73 years old.Another case study reported fixatio of humerus fractures in a 50 year old person [96].After insertion of MAGNEZIX implants the range of motion was reversed to normal and Mayo score was 100.Also,it is also noteworthy to mention that such type of fractures were already been cured with help of Mg in the past.These studies were successful only in children.However,MAGNEZIX implant exhibited good fracture union in 73 year old patient as fore mentioned.This indicates that the current technology and experience could possibly revolutionize implant application.The treatment for mandible fracture surgery is complicated and hence it remains debatable.MAGNEZIX screws were employed to fi displaced fracture that occurred at the head of mandible in six patients.This is firs ever report to use an innovative Mg bio-absorbable implants in the treatment of oral and maxillofacial surgery.The lower jaw moments were recovered back to normal after three months [95].Wichelhaus et al.[93] tried to reconstruct the wrist fracture occurred due to trauma using MAGENZIX implants.The patient was 40 y.o and it was a case study reporting only single patient.The corrosion in vivo deteriorated the mechanical integrity within 6 months which led to formation of cysts.Hence,the surgeons concluded that MAGNEZIX implants are not recommended for repairing wrist fractures.Another case study also did not recommend MAGNEZIX implants,because of cyst formation in three out of fi e patients[94].Table 7 summarizes the detailed data of MAGNEZIX implants used in various surgical areas.

Table 7 MAGNEZIX implants employed in repairing fractures at various surgical areas.

5.2.Clinical trials in Asia

Lack of blood supply results in necrosis of bone tissue.Zhao et al.[113] used a pure Mg screws for the treatment of one such necrosis that occurred in femora of a 17 year old boy.Two years post-operation,the CT images and radiographs clearly shows that the Pure Mg screw was almost completely degraded.In addition,the functionality of hip was assessed using Harris scores.The Harris hip scores were found to be 37,74,83 and 86 for pre-operation,post operation,one year and two years respectively.The same research group extended the study in which 19 cases with displaced femora.They used the combination of Pure Mg implants with vascularized iliac grafting.Compared to the previous studies that used internal fixation combination of internal fixatio with osteotomy or vascularized iliac grafting,the success rate was relatively higher upto 99.4%.In this study the combination of magnesium screws and vascularized iliac grafting is reported to achieve hip reunion in all 19 patients [114].It is worth mentioning that the MAGNEZIX implants were not yet employed in the treatment of hip joint.In contrast to them,pure Mg implants were successful in recovering the hip joint motion.RESOMET alloy comprising Mg-5Wt% Ca-1 Wt% Zn got approval from Korea Food and drug administration for carrying out clinical trials.They were used as fixat ve device to heal distal radius fracture in 53 patients.The fracture union was achieved in all patients by the end of 6 months and degraded almost completely by the end of 12 months [118].Magmaris,an Mg based scaffolds are currently used for cardiology related treatments.They have proved themselves to be better than the conventionally used PLLA stents.In addition,Magmaris scaffolds having strut size of 150μm and an expandable diameter of 0.6mm are absorbed upto～95%.Fig.9 represents the current state of art biodegradable implants [119,120].Generally,the addition of Ca and Zn elements into Mg are recommended since they help in bone formation.In contrast,Al and Rare earths are not suggested to be used in alloying Mg because of the potentials risks involved.For instance,when Mg -10 Wt% Gd was implanted in SD rats Gd accumulated in internal organs.However,the MAGNEZIX implants are similar to a WE43 Mg alloy system consists of 3 Wt% of rare earths.These implants were certifie and also proved to be successful in treating fracture.Hence,it is reasonable to believe that the limited use of rare earths in Mg is acceptable.In contrast,RESOMET alloy does not contain any rare earths and was also successful in clinical trials.

Fig.9.Current state-of-the-art applications of biodegradable metals.Devices at developmental stages -(a) Mg operative clip mounted on the clip applier,(b) MgF2 coated Mg-2wt%Nd alloy nasal stent,(c) Loaded Mg microclip in microlaryngeal forcep,(d) Mg stent for neurological application,(e) Mg-based intramedullary nail.Devices at clinical stages -(f) Resomet orthopaedic devices (Courtesy of U&I Corporation),(g) DREAMS 2nd generation stent,(h)Magnezix screw and pin (Courtesy of Syntellix),(i) Iron-based suture anchor (Courtesy of Fraunhofer IFAM),(j) Velox CD vascular closure device (Courtesy of Transluminal Technologies),(k) Pure Mg Screws [119].

5.3.Comparison of surgical areas implanted with Mg in animal models and clinical trials

The comparison of surgical areas implanted with Mg in animal models such as rats,mice,rabbits,pigs,sheep with humans are tabulated in Table 8.From the Table 8 it is apparent that commercially available MAGNEZIX implants were successful in treating hallux valgus deformity and malleolus factures.In addition,Pure Mg implants produced by Zhao et al.[113,114] research group successfully treated the fracture at hip joint.The possibility of using Mg implants in other surgical areas remains unexplored.Mg implantation resulted in new bone formation equivalent to native bone in surgical areas such as ulna,anterior cruciate ligament,femora and tibia of rabbits.However,some authors claim that the in vivo studies carried out on small animal models are only useful in evaluating the safety of Mg since they do not exactly mimic the human body conditions.It is noteworthy to mention that the miniature pigs were able to mimic humans because their bone healing pattern was found to be similar to humans.The frontal bone,orbital rim,nasal bone and ribs implanted with Mg alloy were tested pre clinically in miniature pigs.In summary,the future research should focus on exploring application of Mg implants in the above mentioned surgical areas in large animal model and then implementing the same in clinical trials.

Table 8 Comparison of surgical areas implanted with Mg in animal models and clinical trials.

6.Recommendations and future directions

The longstanding utilization and commercialization of conventional bio-inert implants exist till date.However,biodegradable Mg implants,especially MAGNEZIX and Magmaris evince better performance than Ti and PLLA implants respectively.This led to commercialization of MAGNEZIX using which 5000 surgeries were performed in over 15 countries.While,70% of surgeons prefer Magmaris implants over PLLA.This indicates visibly that the Mg implants has immense potential to replace age-old bio-inert and polymer based implants.The above mentioned success of Mg implants is possible only because of the intensive research carried out by team of various experts such as surgeons and materials scientists.However,several mysteries remain in the degradation process of Mg and it is in the early stages of commercialization.This review compares the performance of Mg implants in animal models and clinical trials.It also give insights into surgical areas treated with Mg implants.The review also suggest the future directions in order to broaden the application of Magnesium alloys.

1.Since there is no gold standard procedure for evaluating biocompatibility of Mg alloys.The comparison of the performance in in vivo becomes very difficult Therefore when a novel Mg alloy is designed and fabricated the head to head comparison with performance of known Mg alloy becomes mandatory.For this purpose the size of the implant,location and animal model of the novel Mg alloys should be maintained same as that of the reference (for instance LAE442 Mg alloy).

2.In order to verify the safety of Mg implants and their applications in non-load bearing conditions testing inside small animals is sufficient However,in order to mimic the load bearing conditions similar to that of humans,large animals such as miniature pigs and sheep models are recommended.

3.The performance of Mg implants were comparable to that of Titanium in case of treating fractures that occurred in upper limb i.e.oral,maxillofacial and elbow regions and some parts of lower limb i.e.foot and ankle region.Hence,the future studies should focus on extending their application in other surgical areas.