HTERT Gene Mediated Schwann Cells Transplantation On Spinal Cord Injury Damage Area Of Nerve Repair Related Proteins In Rats

Objective: To understand the liposome transfection method will human telomerase reverse transcriptase(h TERT) gene transfection on in vitro cultured Schwann cells proliferation and cell cycle.Further explore the h TERT gene modified Schwann cell transplantation for spinal cord injury nerve damage zone detection and repair effects on neurological recovery related to the expression of related Protein.Methods: Schwann cells by recombinant adeno-associated virus as a vector mediated telomerase reverse transcriptase(h TERT) gene transfection, divided into three groups: control group, no-load virus group, h TERT transfection group. By rt-pcr and Western blot detection Schwann cells in the expression of h TERT gene and protein before and after transfection. Application of cell growth curve, MTT observation of cell growth is optimized; The variation of the flow cytometry to determine the cell cycle distribution.71 adult female Wistar rats, successful model 66, were randomly divided into control group, SCs group, h TERT-SCs group, 22 / group, established in accordance with acute spinal cord injury model modified Allen method. Respectively, before modeling, one day after modeling, three days, one week, two weeks, three weeks, four weeks through the BBB rating, ramp test, a modified Tarlov score assessment of motor function. 72 h after modeling by RT-PCR, Western blot detection of spinal cord injury around AQP4 / 9, MMP9 / 2 gene transcription and protein expression, TUNEL assay apoptosis. 4 weeks after modeling, HE staining and fluorescence microscopy PKH-26 labeled SCs survival and distribution of GFAP detection by immunohistochemistry, Nogo and expression of NF200, the row HRP tracing of nerve fiber regeneration by SEP MEP observed in rats and physiological nerve recovery.Results:After the h TERT gene by retrovirus-mediated transfection PLXSN Schwann cells of rats after transfection h TERT, h TERT transfected with the control group, negative group were transfected h TERT gene expression and protein levels compared to the growth of cells speed was significantly faster, G0 / G1 cell cycle phase reduction, an increase in the number of cells in S phase. There were significant differences(p<0.05) between the groups. RT-PCR, Western blot analysis showed that h TERT gene transfected rat Schwann cells expressing h TERT; lower extremity motor function evaluation h TERT-SCs group rats than SCs, SCs group than the control group. 72 hours after modeling h TERT-SCs group apoptosis index was significantly lower than the control group(p<0.05). 72 hours after modeling, compared with the control group significantly compared AQP4 / 9 gene and proteinexpression in h TERT-SCs group lower(p <0.05). 4 weeks after modeling, HE staining seen in the control group and spinal cord tissue loss cavity formation, no axonal through. SCs group shows a small amount of axonal damage zone-like structure, syringomyelia smaller, h TERT-SCs group showed more nerve axon-like structure, no syringomyelia. PKH-26 positive cells labeled: Up h TERT-SCs group, SCs group followed, at least in the control group, and there were significant differences between the groups(p<0.05). 4 weeks after modeling, h TERT-SCs group of glial fibrillary acidic protein(GFAP), the expression of neurofilament protein(NF-200) was significantly higher than the control group, the difference was statistically significant(p<0.05), h TERT- Nogo protein expression SCs group significantly lower than the control group, the difference was statistically significant(p<0.05), HRP-positive nerve fibers number: h TERT-SCs group the most, SCs group followed, at least in the control group, there are differences between the groups statistically significant(p <0.05). SEP and MEP latency period: h TERT-SCs group <SCs group <control group, and the difference between the groups was significant(p<0.05); amplitude: h TERT-SCs group> SCs group> control group, and each group of The difference between significant(p<0.05).Conclusion: Mediated telomerase reverse transcriptase(h TERT) gene transfected Schwann cell telomerase activity was significantly increased, to promote in vitro rat Schwann cell proliferation by snow retroviral PLXSN carrier. h TERT gene modified Schwann cell transplantation regeneration can promote synaptic spinal cord injury, spinal cord injury AQP reduce surrounding area / 9, MMP9 / 2 gene transcription and protein expression and neuronal apoptosis and promote glial fibrillary acidic protein(GFAP), neurofilament(NF-200) expression increased, decreased the expression of Nogo protein, improve motor function and electrophysiological function in rats.