Transplantation Of PLGA In Combination With Glial-derived Neural Cell As A Neuroprotective Therapy For Spinal Cord Injury In Rats

Research background:Spinal cord injury(SCI),as a central nervous system injury that seriously endangers human health,usually causes persistent paralysis,sensory loss,intractable pain and other neurological dysfunction,and also leads to dysfunctions of multiple organ systems such as urinary,respiratory,and circulatory systems.SCI usually has the characteristics of high incidence,high disability rate and high treatment cost.Also,there is no effective treatment strategy,which seriously threatens human health and affects the quality of life of patients,and imposes a huge burden on individuals,families and society.Cell transplantation therapy is one of the most promising methods for the treatment of SCI.Astrocyte is one of the potential cell sources for cell transplantation because of its abundant sources and low immunogenicity.Some studies used the Neurogenin2(Ngn2)gene to directly reprogram it to differentiate into neural cells and showed certain therapeutic effect for the SCI animal after transplantation.However,like other types of cell used for transplantation,its further application is limited due to low survival rates in SCI lesion.The rise of biological tissue engineering materials has brought new hopes for the treatment of SCI.Biological scaffolds not only have good biocompatibility,degradability,but also can maintain cell survival,regulate their growth,proliferation,differentiation,and other biological behaviors.The application of tissue engineered nerve scaffolds provides a new approach for treating SCI.The present study sought to use PLGA(Poly(lactic-co-glycolic acid))scaffold in combination with glial-derived neural cells to construct tissue-engineered spinal cord,and then implant it into the rat model with SCI,which be used for exploring the role and mechanism of promoting SCI neurological recovery.Methods:1 The cerebral cortex of neonatal rats was achieved and mechanically disrupting,and astrocytes were obtained by digestion,isolation,and culture.The purity of extracted astrocytes was identified by the expression of GFAP utilizing immunofluorescence technique.2 After lentivirus-mediated Ngn2 gene being transduced into rat astrocyte(Ngn2-astrocyte),using immunofluorescence technique to detect the expression of neuron-related markers to prove whether astrocyte could differentiate into neural cells.3 In vitro the tissue engineered spinal cord was constructed by PLGA scaffold in combination with Ngn2-astrocyte,and the binding of cell and scaffold were observed under electron microscope.4 Adult female SD rats were used to establish the SCI model by Allen’s method.They were randomly divided into three groups:PLGA+Ngn2-astrocyte group,simple Ngn2-astrocyte group,and PB S control group.Transplantation were carried out after 1 week.5 The BBB locomotor rating scale was used to evaluate the recovery of motor function after SCI in rats.Meantime,the histological condition of spinal cord injury and the expression level of related proteins were respectively detected by HE staining and immunofluorescence technique,which were used to observe the recovery of spinal nerve function in experimental rats furtherResults:1 The cerebral cortex of neonatal rats was successfully obtained,and the third generation of rat astrocyte was further cultured,isolated,and passaged.The purity of astrocyte was determined to be over 97%.2 Ngn2 gene can be efficiently transduced into the third generation of rat astrocyte by lentivirus and stably expressed.In vitro,the morphology of astrocyte gradually transformed into neural cell and expressed neuron-related markers.3 Ngn2-astrocyte can adhere,grow,and migrate on the PLGA scaffold.4 PLGA+Ngn2-astrocyte or Ngn2-astrocyte was successfully transplanted into SCI lesion.The survival and differentiation of transplanted cells in rats were observed by immunofluorescence technique.5 PLGA+Ngn2-astrocyte transplantation group was significantly better than the other two groups in terms of BBB motor function score and pathological changes after spinal cord inj ury.Conclusions:The transplanted cells in PLGA+Ngn2-astrocyte group can effectively survive and differentiate into neural cells in SCI lesion,fill the SCI cavity,and promote the recovery of motor function in rats.It can better promote SCI repair than the simple Ngn2-astrocyte transplantation.