Study Of Analysis And Comparison Of The Components Of AECM And Histocompatibility In Different Species

ObjectiveIt is our goal in the field of neuroscience to explore suitable nerve graft to bridge the nerve gap between the two stumps of the damaged nerve trunk.With the development of tissue engineering,homogeneous acellular nerve transplant gained well development,it was used as bioactive surrogate of nerve,having better bionic property and exhibiting excellent histocompatibility,may offer fitting microenvironment for axonal regeneration,so as to facilitate and induce effective regeneration of the injured nerve.However,the source of donator will be restricted in clinical application.Whether heterogeneous nervous acellular transplant could be used as eligible graft or not must be solved urgently.In our experiment,by comparing the composition and immunogenicity of acellular extracellular matrix(AECM)in different species of mammalian,we aim at supplying experimental evidence for the solution of the source of graft in clinical peripheral nerve repair.Materials and Methods1.Experimental animals30 Wistar rats and 8 SD rats weighed 200～250g,and 8 hybrid dogs weighed 16kg.The animals were supplied by China Medical University.2.Preparing of acellular transplantsThe optimized chemical extracted nerve transplants were obtained from dogs and rats,and the transplants were also prepared via hypotonic-enzymatic method from rats. 3.Observed indices and detected methods(1)Observing the morphology and construction of AECM by employing the light microscope and electron microscope.(2)Observing and comparing the residual myelin in AECM by chromotropic acid 2R-brilliant green staining,then analyzing the content of myelin fragment semiquantitatively.(3)Observing and comparing the positive expression of laminin and S-100 in AECM by immunohistochemistry,then analyzing the remained content semiquantitatively.(4)Observing and comparing the bands of nerve proteins separated by SDS-PAGE.(5)Observing and comparing the immunoreaction of AECM embedded under cutis by HE dyeing.Results1.Both AECM obtained by the optimized chemical protocol and AECM of rat obtained via hypotonic-enzymatic method reserved structural integrity of vessel of basilar membrane.2.There was no significant difference in the staining intensity of residual myelin in AECM between dog and rat,and no significant difference between different decellularization protocols.3.There was no significant difference in the integrated optical density value of laminin and S-100 positive product in AECM between dog and rat,and no significant difference between different decellularization protocols.4.There were obvious growth associated protein bands,and myelin protein P0 bands vanished in both AECM obtained by the optimized chemical protocol and AECM of rat obtained via hypotonic-enzymatic method.5.The homogeneous and heterogeneous AECM embedded under cutis caused weak immunological rejection. Conclusions1.AECM of different species of mammalian have better bioactivity and excellent histocompatibility.2.AECM obtained via optimized chemical extracted protocol and hypotonicenzymatic method appear to be bionic,thus could be used as suitable transplant in efficacious peripheral nerve repair.