A Plasmid PLXSN-bcl-2 Intraventrically Injected Into Transient Focal Ischemic Rat Brain And The Influence To The Expression Of The Hsp70 Protein

At present, many researchers are interested in the bcl - 2 gene transfer therapy of the cerebral ischemic diseases. There are two critical problems in all the transgenetic therapy: the pathway and the vector. Most of the study adopted viruses as vector and injected at the ischemic focal cortex. In this study , to find a more convenient and more effective method, we took plasmid as vector , injected directly into the transient focal ischemic rat lateral ventricle, and observed if this new method could protect the ischemic rat brain more effectively. Hsp70 protein family, who are redundant in the stress cell, have been proved of the protective effect to the ischemic neurons. Some studies have found that the protective effect of the hsp70 protein family to the ischemic tissue in much extend were due to their function of antagonizing apoptosis and the influence to bcl - 2 protein. So this time we want to find if there is some relationship between the bcl - 2 protein and hsp70 protein family, which will be the bases to the further research .MethodsConstructing the rat model of transient focal cerebral ischemia with the modified Longa method. Then, immediately injected plasmid pLXSN - bcl - 2 (100ng/ul) , vector pLXSN( 100ng/ul) and normal saline 20ul slowly into the rat lateral ventricle of the three groups,respectively. At the same time, 1 hour after occlusion we withdrew the nilon string. All the survived rats were decapitated at 3h 6h 24h 48h 72h after reperfusion . Immunohistochemistry methodwas performed to confirmed the expression of the bcl - 2 protein and the hsp70 protein, and the infarct area was delineated by triphenyltetrazolium chloride stain. We measure the infarct volume of the rat brain directly at different time points after the ischemic insult (6h ~72h).Results1. The expression of bcl - 2 protein and hsp70 protein could be observed from 3 h time point after reperfusion, and reached peak at 24 h time point, since when the expression decreased. The expression of bcl - 2 protein and hsp70 protein in rats injected with pLXSN - bcl - 2 was greater than that of the control groups at each time point, and much of the expression were found at the " penumbra field "and the hippocampus caudate nucleus thalamus ,where were much sensitive to ischemic insult. Further more, the expression of bcl - 2 protein in rats injected with pLXSN - bcl - 2 was global while that in the control group was limited at the infarct focal site.2. The infarct volume of the rats injected with pLXSN - bcl - 2 is smaller than the control groups at 6h 24h 48h 72h time point, and there was no significant difference between the two control groups.Conclusion1. Intraventrically injected plasmid pLXSN - bcl -2 could increase the expression of the bcl - 2 protein in the ischemic rat brain, and the production could reduce the infarct volume, so that it could protect the cerebral tissue a-gainst ischemic insult. Accordingly, this method of transgenic therapy was feasible for rat transient focal cerebral ischemia.2. The overexpression of bcl - 2 protein could promote the expression of hsp70 protein. We presumed that maybe there were cooperative relationship between the bcl - 2 protein and the hsp70 protein .