Construction of multi-epitope gene of periodic Brugia malayi and its expression in eukaryotic cells / 中华地方病学杂志

Objective:To construct a eukaryotic expression plasmid pcDNA3.1(+)-BmCPI502/BmGAPDH720 of periodic Brugia malayi cysteine protease inhibitor/glyceraldehyde 3-phosphate dehydrogenase (CPI502/GAPDH720) multi-epitope gene and observe its protein expression in Hela cells. Methods:Primers were designed according to the predicted gene sequences of T cell epitopes. The target gene fragment was amplified by reverse transcription (RT)-PCR using plasmid pGEM-T-CPI621 as template. The fragment was cloned into prokaryotic expression plasmid pET-28a(+) to construct prokaryotic expression plasmid pET28a(+)-BmCPI502. The BmCPI502 gene and BmGAPDH720 gene were amplified by RT-PCR, respectively. The gene fragments of pcDNA3.1(+), BmCPI502 and BmGAPDH720 were digested by double enzyme digestion and ligated with the target gene. The eukaryotic expression plasmid pcDNA3.1(+)-BmCPI502/BmGAPDH720 was constructed. The recombinant plasmid was transfected into Hela cells and verified by RT-PCR to obtain the desired target bands. The expression product was detected by sodium dodecyl sulfate polyacrylamide gelelectrophoresis (SDS-PAGE).Results:The recombinant plasmid pET28a(+)-BmCPI502 was obtained, and the 502 bp specific fragment was identified by enzyme digestion, which was in line with the expected value; the eukaryotic expression plasmid pcDNA3.1(+)-BmCPI502/BmGAPDH720 of the multi-epitope gene was successfully constructed, and the fragment size was in line with the expected value. The eukaryotic expression plasmid pcDNA3.1(+)-BmCPI502/BmGAPDH720 was transfected into Hela cells and the recombinant protein was stable expressed. SDS-PAGE analysis showed that the relative molecular weight ( Mr × 10 3) of the recombinant protein was about 50. Conclusions:The eukaryotic expression plasmid of pcDNA3.1(+)-BmCPI502/BmGAPDH720 of periodic Brugia malayi multi-epitope gene is successfully constructed, and the corresponding recombinant protein is obtained in eukaryotic cells. This study has laid a foundation for further study of the purification and biological activity of the recombinant protein.

Effects of hydroxysafflor yellow A on iNOS expression in rats of focal cerebral ischemic injury / 中国药理学通报

Aim This was to study the effects of hydroxysafflor yellow A(HYSA)on iNOS expression in rats following focal cerebral ischemia. Methods Rat cerebral ischemia-reperfusion injury was induced by middle cerebral artery occlusion. The expression levels of iNOS were examined using immunohistochemical method. The neurological outcome and the cerebral infarct area were evaluated. Results The expression levels of iNOS in HYSA groups were significantly lower than those in ischemic model group. Treatment with HYSA also decreased the cerebral infarct area and the neurological deficit score. Conclusion HYSA induced down regulation of iNOS expression, which may mediate the protective effect of HYSA on cerebral ischemia.

Pharmacological Effects of Zhongfengkang on Model Mice with Cerebral lschemia / 中成药

Objective: To study the pharmacological effects of Zhong Feng Kang on model mice with cerebral ischemia and after restoration of blood flow. Methods: To make the model of cerebral ischemia and after restoration of blood flow with thread embolism, and measure the area of cerebral infarction and observe cerebral pathologic change. Results: Zhong Feng Kang could reduce the area of cerebral infarction, improve cerebral pathologic change on model mice. Conclusion: The results confirmed that the clinical data of Zhongfengkang would provide a basis of pharmaceutical application