| Background and objective:In recent decades,with the rapid progress of society and continuous improvement of medical technology,people’s living standards and life expectancy have been greatly improved,and the structure of population is slowly turning into an aging society.For this reason,the mobidity of degenerative diseases of spine such as cervical,lumbar intervertebral disc protrusion and vertebral canal stenosis that cause incidence of neck,shoulder and lumbocrural pain increase,which resultsin an increase of the socioeconomic burden by years.At present,various routes of laminectomy,discectomy and spinal fusion are used in the treatment of degenerative diseases of the spine.More than 60 years ago,Robinson et al.pioneered the anterior cervical discectomy and fusion(ACDF)surgical technique.Cloward and Hodson reported surgical techniques for posterior lumbar interbody fusion(PLIF)surgery.Since then,with continuous improvement of clinical application,spinal fusion has achieved good clinical efficacy in the treatment of spinal degenerative diseases.So far,Cervical ACDF and lumbar PLIF surgery are still the common surgical methods in clinical practice.At present,polyether ether ketone resin(PEEK)is the most commonly used fusion cage in clinical interbody fusion surgery.After more than ten years of clinical application,it has been proved that PEEK material fusion cage has the advantages of good biocompatibility,high strength,X-ray penetration,which is a good candidate for make interbody fusion cage.However,it is inert material,and the hydrophobic property of its surface goes against cell adhesion and growth,and its osteogenic activity is poor,which makes it difficult to fuse with bone interface and eventually form pseudojoint.Moreover,its elastic modulus is still significantly higher than that of vertebral endplate,which is close to cortical bone of human body,and there is still a high incidence of fusion cage sedimentation.Therefore,it is particularly important to explore a new interbody fusion cage with good biocompatibility,low elastic modulus,osteogenic activity and absorbability and degradability by human body in order to improve the success rate of interbody fusion surgery.Based on previous studies on biodegradable absorbable materials,we found that the biodegradable material polycaprolactone(PCL)combined with bioceramicsβ-tricalcium phosphate(β-TCP)had good strength and osteogenic effect as bone defect replacement materials.Through our preliminary designs and in vivo animal experiments,it has been verified that this material as an interbody fusion cage has good biocompatibility and osteoinductive effect,and can be degraded and absorbed.Therefore,the purpose of this study is to manufacture a new PCL/β-TCP(mass ratio are 50:50)cervical interbody fusion cage using 3D bio-printing technology and computer-aided design to optimize the structure of the cages.In this way the cage has enough mechanical strength to meet the needs of clinical applications.Then we conduct degradation experiments and cervical ACDF experiments on sheep to observe the biomechanical properties,degradation characteristics and the fusion effect in vivo and in vitro.On the basis of confirming its safety and effectiveness,pilot small-sample clinical trials were conducted to analyze the safety,fusion effect and adverse reactions of the new fusion cages used in human ACDF surgery,in order to provide theoretical basis and technical guidance for the early application of biodegradable PCL/β-TCP fusion cages in clinical practice.Methods:Part One:Design,manufacture and physical and chemical properties analysis of the biodegradable PCL/β-TCP cervical fusion cages:Selecting mass ratio for the 50:50PCL andβ-TCP powder,blending them homogenously.Using the double screw extrusion mechanism to extrude 5mm diameter rod after melting and mixing in 75℃,then putting the rod with biological 3D printers to print the biodegradable PCL/β-TCP cervical fusion cages according to the fusion cage model is designed by CAD in advance.The fusion cages were analyzed by scanning electron microscope to observe its apparent structure,micro-CT scanning to analyze its structure such as volume,porosity and pore size etc.INSTRON universal testing machine to test the mechanical strength of cage,and gel penetration chromatographic analysis of PCL molecular weight in the fusion cage.Part Two:In vitro degradation experiment of biodegradable PCL/β-TCP cervical fusion cage:Immerging the biodegradable PCL/β-TCP cervical fusion cage of experimental group in 37℃simulated body fluids and replace the fluids every two weeks.After soaking 2 weeks,4 weeks,12 weeks,26 weeks and 52 weeks,taking out the cage to study the cage degradation characteristics in vitro through detecting the weight loss rate,mechanical strength,the molecular weight,analysis of SEM and Micro-CT scanning.Blank control group was placed in a closed container in room temperature for 52 weeks for the same detection.Part Three:ACDF experiments on sheep:Choosing 12 adult female small tail cold sheep,operating anterior C3/4 and C4/5 intervertebral disc resection,and implanting biodegradable PCL/β-TCP fusion cage using plate and screws of anterior cervical vertebral body to fix the vertebral body.Establishing sheep anterior cervical decompression and fusion experimental model,and using the PEEK fusion cage as the control group.X-ray examination were performed in 1 week,6 months and 12 months after surgery,evaluating the cervical interbody fusion effects through the X-ray image.And the sheep were killed in 6 months and 12 months after surgery,the fusion cages and the surrounding cervical vertebral bodies were collected.Histopathological examination was performed to observe the osteogenic fusion of the tissue around the fusion cage,and to analyze the cervical interbody fusion effect and fusion mode of PCL/β-TCP fusion cages after ACDF operation on sheep.Part Four:The biodegradable PCL/β-TCP fusion cage in small sample exploratory clinical trial of human ACDF:The above experiments verified that the biodegradable PCL/β-TCP fusion cage had good mechanical properties,There were good fusion effect and no adverse reactions in sheep ACDF experiment.Therefore,after the review and approval of the ethics committee of the hospital,a total of 12 patients who required ACDF surgery were selected.Among them,6 patients underwent ACDF with biodegradable PCL/β-TCP cervical interbody fusion cage,and 6 patients received ACDF with PEEK fusion cage as control group.Follow-up functional examination,VAS and JOA score were performed in 3 days and 1,3,6 and 12 months after surgery to evaluate the clinical efficacy.X-ray and CT examinations were performed to analyze and evaluate the fusion effect and the presence of intervertebral space collapse.Blood routine,erythrocyte sedimentation rate,C-reactive protein(CRP),liver and kidney function were monitored for evaluation inflammatory reaction and hepatorenal toxicity.Results:Part One:Design,manufacture and physical and chemical properties analysis of the biodegradable PCL/β-TCP cervical fusion cages.1.PCL andβ-TCP with a mass ratio of 50:50 can be well mixed,and the rod produced has a smooth surface,uniform structure and no cracks.The cervical interbody fusion cage for experiment was successfully manufactured by the 3D printer,and the appearance was intact and flawless.2.Scanning electron microscopy(SEM)showed that the surface of biodegradable PCL/β-TCP fusion cage was coarse and had micropores.3.Micro-CT scannig results showed that the pore diameter(trabecular gap)of the fusion cage was about 211μm,the porosity was about 18.7%,and the volume was about 431mm~3.4.INSTRON universal testing machine was used to test the compressive strength of the cervical fusion cage.The maximum compressive strength was 49MPa and the work compressive strength was 23MPa..5.The mass ratio ofβ-TCP was about 48.77%in the cage,and the molecular weight of PCL was about 100,000.Part Two:In vitro degradation experiment of biodegradable PCL/β-TCP cervical fusion cage.1.The biodegradable PCL/β-TCP interbody fusion cages were degraded in vitro for 52weeks.The appearance of the cage was little changed,the overall structure was intact,and the surface and porous area had slight structural loss..2.After 52 weeks in vitro degradation,the cage weight loss rate reached 9.23%(P<0.05),and the degradation curve was relatively straight.3.After 26 weeks of in vitro degradation,the mechanical strength of cage decreased about14.5%and there was no significant difference compared with the compressive strength of cage before degradation in vitro.(P>0.05).After 52 weeks in vitro degradation,the compressive strength of the cage decreased about 18.2%(p<0.05).4.After 52 weeks in vitro degradation,the molecular weight of PCL in cage decreased from about 100,000 to about 70,000(P<0.05).Part Three:ACDF experiments on Sheep.1.The mechanical strength of the biodegradable PCL/β-TCP fusion cage was good,and the height of the intervertebral space was not significantly reduced during the experimental period.X-ray examination showed that the fusion cage had no deformation and collapse,and the fusion rate of PEEK group and biodegradable PCL/β-TCP group reached 100%in 12 months after surgery.2.The X-ray examination in both groups showed the sheep cervical vertebral sequence was stable in 6 and 12 months after surgery.And there was obvious bone growth in the central bone graft area of the PEEK fusion cage that connected the upper and lower endplates.In biodegradable PCL/β-TCP fusion cage,the bone growth and osteogenesis around the cage were between in the cage surface and endplates.There was bone growth in the porous area of the cage too,and the bone mass increased from 6months to 12months after surgery.3.The interbody fusion site of the PEEK cage was located at the bone graft site in the center of the cage,while the fusion site of the biodegradable PCL/β-TCP fusion cage was located at the bony interface between the cage surface,the endplate,and the bony bridge around the cage.Part Four:The biodegradable PCL/β-TCP fusion cage in small sample exploratory clinical trial of human ACDF.1.In both groups,there was no increase in monitoring inflammation and liver and kidney function indexes at 1,3,6 and 12 months after surgery,and no local or systemic discomfort symptoms such as fever and pain were found.2.In both groups,the overall symptoms of the patients were relieved,the VAS,JOA score and limb function were improved after surgery,and the clinical treatment rate was 100%.3.Within 1 year after surgery,X-ray and CT examination showed that the biodegradable PCL/β-TCP cage had no collapse or deformation,the height of the intervertebral space was well maintained,and the stability of the spinal sequence was good.At 12 months after surgery,CT showed that the cervical interbody fusion was good,the fusion rate of biodegradable PCL/β-TCP cage reached 100%,and the fusion effect was not significantly different from that of the PEEK cage.Conclusion:This project used 3D printer to manufacture the biodegradable PCL/β-TCP cervical interbody fusion cages with a PCL andβ-TCP mass ratio of 50:50.Through initial analysis of physical and chemical properties,degradation experiment in vitro,anterior cervical interbody fusion experiment on sheep and small sample clinical trial of human anterior cervical discectomy and fusion,we could conclude that:1.The biodegradable PCL/β-TCP cervical interbody fusion cages were manufactured by computer aided design and 3D printing technique.It was a porous structure in the center of the surrounding entity.SEM showed that the surface microstructure was rough with micropores,which was conducive to tissue and cell attachment and osteogenesis induction.2.The compressive strength of the PCL/β-TCP cage was good,and the working strength reached 23MPa.After 52 weeks of degradation in vitro,the mechanical strength attenuation was less,reaching more than 80%of the initial strength,which still had a good supporting effect.3.The degradation of the PCL/β-TCP cage in vitro was slow and the degradation curve was straight.The degradation started from the surface and porous area of the cage,and the degradation process was related to the continuous decrease of molecular weight of PCL.4.The anterior cervical interbody fusion experiments on sheep showed that the properties of the PCL/β-TCP cages were stable during the degradation process,and there was no degradation product stimulation and local inflammatory reaction,and the biocompatibility was good.5.The anterior cervical interbody fusion experiments on sheep showed that the PCL/β-TCP cages had good mechanical strength and could maintain the stability of vertebral body sequence without cervical external fixation after surgery,and the cervical vertebral body and the cage did not collapse and the intervertebral space has not decreased.6.Biodegradable PCL/β-TCP cage in the anterior cervical interbody fusion experiments on sheep showed good fusion results,achieving the total integration in 12months after surgery.In the porous area of cage bone ingrowth was observed.The fusion mode of PEEK cage was in the center of bone graft,while the fusion mode of PCL/β-TCP cage was different from the PEEK cage,it fused by means of the combination of the interface between the cage and bone,and the bone bridge around the cage.7.The biodegradable PCL/β-TCP fusion cages have shown good fusion results and clinical efficacy in patients undergoing ACDF 1 year after surgery,and no specific adverse reactions were observed. |
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