Regionspecific Survival,Migration, Differentiation Of Induced Neural Stem Cells Into The Mouse Brain And The Induced Local Immunological Rejection

Objective: Induced neural stem cells(iNSCs)can be obtained from rodent and human somatic cells through the forced expression of defined factors.It has the potential clinical application values in the treatment of central nervous system diseases.Studies have confirmed that iNSCs can differentiated into mature astrocytes,neurons and oligodendrocytes in vitro,the purpose of this paper is to explore the survival,migration and differentiation of implanted iNSCs in different mouse brain regions,and to investigate the immunological rejection in the brain followed neural stem cell transplantation.Methods: iNSCs were plated into serum-free DMEM /F12(1?1)medium for subculture,and gave them the nestin immunofluorescence identification;Preparation of iNSCs suspension;We grouped 36 adult male C57 mice randomly according to the different time of transplantation and brain regions,Each group contains four mice.The grouping is as follows: 2 weeks transplantation group(hippocampus,Striatum,cortical motor area,cortical sensory area)and 6 weeks transplantation group(hippocampus,Striatum,cortical motor area,cortical sensory area),According to the group,and with the help of a mouse brain stereotaxic instrument,we transplanted the iNSCs suspension into corresponding C57 mice brain regions;After corresponding period of time,we removed the brain tissue from the infused mice and obtain the cryostat slice.With the help of green fluorescent protein,we can observed the survival and migration of iNSCs under fluorescence microscope;With the aid of cell specificity antibodies,for example,Tuj1,nestin,MAP2,GFAP,we can explore the differentiation of iNSCs,and with the aid of immue cell specificity antibodies,CD8,CD3,CD45 and so on,we can investigate the lymphocyte infiltration induced by the transplantation of iNSCs.Results: Under the condition of in vitro culture,iNSCs are capable of stable clone proliferation,and remain nestin positive.Two time points,we all found iNSCs survival in different brain regions,but the survival percentage differ greatly amongdifferent brain regions iNSCs(P<0.05),survival ratio in Striatum,hippocampus is relatively high;The transplanted iNSCs can migration into the surrounding brain tissue in 4 weeks,and the migration shows regional specificity;GFP positive cells were found to migrate along subgranular layer zone;2 weeks and 4 weeks after iNSCs transplantation,living cells in each brain region showed Tuj1 negative,Part of live cells are nestin positive in the 4 weeks groups,but the nestin positive cells rate differ greatly among different brain regions(P < 0.05),rate in the hippocampus is higher;In 4 weeks groups of striatum and cerebral cortex,some live cells showed GFAP positive,in some hippocampus regions near the lateral ventricle,some MAP2 positive living cells were found;At 4 weeks,we found the lymphocyte infiltration in all transplant sites,including CD8 positive cells,CD3 positive cells,CD45 positive cells,In the hippocampus,for example,the number of immune cells exist significant differences compared with control group(P<0.05).Conclusion: iNSCs show stable clonal proliferation in vitro culture condition,and keep its neural stem cell characteristics;iNSCs can survive and gradually integrated into local brain tissue,but the survival shows regional specificity;iNSCs has the ability to migrate in the adult brain,in hippocampus area,the iNSCs can migrate along the subgranular layer zone;In the short term,after transplantation,the differentiation efficiency of iNSCs is low,but the differentiation trend shows regional specificity;iNSCs also have certain immunogenicity,they can induce the occurrence of local immune rejection in the process of brain transpla.