| Objective To investigate the effectiveness of heat shock protein 70(HSP70) gene transfection in rats following cerebral hypoxia-ischemia.Methods①293 cells were infected with recombinant adenovirus (vAd-HSP70) then collect the virus after infected successfully which could be proved by the inverted fluorescence microscope。②The HSP70 using adenovirus-mediated gene therapy was transferred into the mouse through caudal vein. The expression level of HSP70 mRNA in the brain tissue of the mouse was detected by reverse-transcription polymerase chain reaction (RT-PCR) after injecting 24h,48h,72h respectively.③The rats were randomly divided into three groups:(1) HI (hypoxia-ischemia, n=25)group(2) vAd-HSP70 (subjected to hypoxia-ischemia brain injury after transferring HSP70 48h, n=25) group (3)normal control group(NC, n=25). RT-PCR and immunohistochemistry were used to analyze the expression of HSP70 mRNA and HSP70 in the brain tissue at 2h,24h,48h,72h and 7d respectively. The changes of brain tissue were observed by pathological section.Results①the recombinant adenovirus (vAd-HSP70) could infect the 293 cells effectively, and collect the virus successfully.②The expression of HSP70 gene could be tested in the mouse after administration of AdHSP through the caudal vein and reach its peak at 48h (0.81181±0.23416), and there was statistics significance compared with 24h and 72h groups (P<0.05).②RT-PCR shows there was an increase in the expression of HSP70 2h after HI (0.7762±0.4855) and reach its peak at 24h (1.8087±0.4142), then declined at 72h (0.9700±0.0391). The HI 7d group (0.3458±0.0904) was still higher than the NORM group. Also, the expression of HSP70 in vAd-HSP70 48h HI group was higher than HI group at the corresponding times, at 2h,48,72h were significantly higher than HI group(P<0.05). There were not HSP70 positive cells in NC group, after hypoxic ischemia the HSP70 positive cells began to increase. The HSP70 positive cells could be found at 2h after HI, and reach the peak at 24h and 48h, after 72h they began to decrease, till 7d were still higher than the NC group. The HSP70 positive cells in the vAd-HSP70 groups were more than HI group at the corresponding times, and there was statistics significance at 2h,48h,72h (P<0.05), especially the 48h group. The brain tissue of NORM group is normal, endochylema and nucleus are clear. In HI group, there are only a few neurons edema at 2h, some neurons edema at 24h and the intercellular substance edema lightly. At 48h the intercellular substance edema severely. At 72h the neurons edema obviously. the brain blood vessel vasodilatation, congestion. Tissue sample from the brain revealed significantly less evidence of hypoxia-ischemia induced tissue damage in vAd-HSP70 groups compared with the HI group.Conclusions①The HSP70 using adenovirus-mediated could be tested in the rats②The vAd-HSP70 has the effect of brain protection against hypoxia-ischemia。... |
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