| PrefaceThe disease incidence rate and the case fatality rate of the ischemic cerebrovascular disease increase year by year , the ischemic cerebrovascular disease assumes the young tendency and is more and more seriously threating people's health and life. Therefore,the effective treatment of this disease is always an important question.The therapeutic efficacy of the bone marrow mensenchymal stem cells(MSC) to ischemic cerebrovascular disease was already confirmed in many ways.But,the apoptosis of MSC is a problem.How to decrease the apoptosis and increase the contribution of MSC need to investigate.This research coadministration MSC with the bcl-2 gene which is an identified anti-apoptosis gene,observing modified Neurological Severity Scores,the expression of brain-derived neurotrophic factor and the apoptosis of the cells, to discuss the therapeutic effect and the mechanism of the grafting MSC and Plasmid pLXSN mediated transfer of bcl-2 gene to cerebral ischemia rats.MethodsForty ischemia-reperfusion rat models were established by transient occlusion of the left middle cerebral artery(MCAo)to induce focal cerebral ischemia for 2h, followed by reperfusion,and were randomly divided into the physiological saline group, the pure MSC group, the pure bcl-2 gene group and the experimental group (MSC+bcl-2 group), separately graft pLXSN-bcl-2 gene and MSC in 3h and 24h into the carotid artery and the tail vein after 2 hours of MCA o. Each group of rats were divide into 3d and 14d two time spots (each time spots 5 rats). Neurological Deficits Score(NDS) were observed at3d,14d.Their cerebral tissues were obtained at 3 or 14 day after 2 hours of MCAo.Brain sections were stained with hematoxylin and eosin(HE) for surveying the volume of infarction. Double-staining immunohistochemistry of brain sections was used to identify brain-derived neurotrophic factor (BDNF),Bcl-2 protein expression level and the distribution and quantity in the brain of the BrdU marked MSC. Taken two sections of each sample,examined and photographed using computerized video imaging microscopy.Date were acquired from different fields in the margin of the infarction focus, hippocampus and corpus striatum region by analysing the positive area percentage or counting the number of positive cells at high power magnification(400×).SPSS 13.0 statistical soft was used to analy the date .All data were expressed as x±s.A value P<0.05 was considered statistically significant.Results1,MSC cultivation:the cell morphous from 3 generation to 6generation were stable, the Fusiform shape and applanation shape were the typical examples.After 6 generation ,there were a lot of grana and vacuolus in the cells ,then the cells fell off and died;2,pLXSN-bcl-2 which was cut after the EcoRI,XhoI double enzyme has two belts, one is 850 bp bcl-2 pieces, another is 5863 bp pLXSN split-ring pieces;3,The score of neurologic impairment in the experimental group at 3d(5.94±0.27) was significantly lower than that in the physiological saline group, and that at 14d (2.02±0.62)was significantly lower than that in the control group(P<0.05);.4,The expression of BDNF significantly increased in the experimenta group compared with the control groups;5,Many BrdU positive cells could be seen at the infarction hemisphere in the experimental group and pure MSC group.The number of engrafted MSC in the experimental group was significantly higher than that in the pure MSC group;6,The expression of Bcl-2 at 3d,14d significantly increased in the experimental group compared with the control groups;7,The cell apoptosis significantly decreased in the experimental group compared with the control groups.Conclusion These results suggest that MSC and bcl-2 gene combined transplantation has the obvious therapeutical effect in rats with cerebral ischemia. Its mechanism possibly is that bcl-2 gene can reduce the apoptosis of grafted MSC while reduce cellular apoptosis in the brain.
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