Experimental Study On The Repair Of Facial Nerve Injury In Rabbits With Adipose Derived Stem Cells Combined With Platelet Rich Plasma

ObjectiveObjective to investigate the effect of combined allogeneic adipose tissue derived stem cells(ADSCs)and acellular xenogeneic nerve(AXN)combined with autologous platelet rich plasma(PRP)on facial nerve injury in early stage.MethodsThe adult healthy 300~350g SD female rats were harvested and the bilateral sciatic nerve 15 mm was cut.The acellular xenogeneic nerve(AXN)was prepared by chemical combined freezing and thawing,and was stored at 4 degrees fridge.One subcutaneous fat pad of the neck of adult healthy New Zealand rabbits were harvested,and adipose-derived stem cells(ADSCs)were cultured in vitro by single enzyme digestion.Meanwhile,platelet rich plasma(PRP)was extracted simultaneously.After the ADSCs was labeled with CM-Dil,it was implanted into the xenograft after treatment..Another female New Zealand white rabbits 32,randomly divided into A group,B group and C group as the experimental group,group D as control group,8 rats in each group.The model of four groups of animals was built and the 1cm defect model of the left lateral buccal branch of the nerve was established.Group A: ADSCs combined AXN combined with PRP to repair nerve defect group;B group: ADSCs compound AXN repair nerve defect group;C group: simple AXN repair nerve injury group;D group: Auto nerve graft group.After transplantation,the experimental animals were divided into cage rearing.At the fourth and eighth weeks after operation,the animals in the experimental group and the control group were observed for gross morphology.The angle between the upper lip and the midline of the affected side was measured,and the electrophysiological detection of the regenerating nerve was performed.At the eighth week after operation,the regeneration nerves in each repair group were sliced,histologically observed,Western-blot protein detected,transmission electron microscope observed,and then analyzed and evaluated the recovery of early neurological function in the experimental group and control group after regeneration.ResultsAfter the operation,all the experimental animals survived well and no serious complications occurred.The incision of the left side of all the different groups was healed well and no infection was found.After fourth weeks,eighth weeks were found in each experimental group implanted xenogenic nerve transplantation and injury of nerve stump goodbiocompatibility and cover the surface with thin vascular fibrous membrane,although there are some absorbed phenomenon,but not obvious detachment,and no formation of neuroma.Through the general observation,the symmetry of the upper lip angle and the midline angle of the affected side were compared.The angulation of the autograft group was most close to the healthy side.However,the recovery effect of ADSCs combined with AXN and PRP injection group was similar to that of the autograft group.Light and transmission electron microscopy were used to observe the distal and proximal nerve segments of regenerated nerve segments.Myelinated nerve fibers were seen in the anastomotic ends of each group.CM-Dil labeled stem cells were observed in group A and B combined with adipose tissue derived mesenchymal stem cells,and the passing rate of A group was higher.ADSCs composite AXN combined with PRP repair nerve defect group and autologous nerve graft group had more regeneration myelinated nerve fibers,arranged in a regular and orderly manner,larger myelin sheath,larger diameter,lamellar structure and B group.In the simple AXN group,there was no myelinated nerve fiber angle in the regenerated nerve,but the myelinated nerve fibers were irregular in arrangement,and the myelin sheath was thinner.The regeneration of nerve specific NF-200,NGF,and BDNF was detected by immunohistochemistry and Western-blot protein.In the autograft group,the protein content was the highest.ADSCs combined with AXN and PRP group was higher than that of ADSCs composite AXN group.The AXN group had the least difference(P<0.05),(P>0.05)and there was no significant difference between A group and D group.ConclusionsIn this experiment,adipose-derived stem cells were cultured and identified in vitro.It is proved that adipose derived stem cells have multiple differentiation abilities and can form Schwann cells(SCs)under the induction of autologous platelet rich plasma.Schwann cells are myelin forming cells in the peripheral nervous system and play an important role in regeneration after injury.The sciatic nerve of SD rats was removed by chemical combined freeze-thaw treatment,which had good immunity and good scaffold function,and a matching xenogeneic nerve graft was obtained.In the process of nerve repair,the xenogenic nerve transplantation of acellular stent as a non immune support guide axonal regeneration,carrying a number of cavity of the differentiation of adipose derived stem cells(ADSCs)effect of early repair as seed cells combined with injection of autologous platelet rich plasma autograft.This lays the experimental foundation for the further research of animal nerve graft instead of human peripheral nerve to repair peripheral nerve injury,and opens up new research directions for nerve regeneration and repair.