Effect of total Panax notoginseng saponins on the activity of human endothelial nitric oxide synthase gene promoter

Abstract

Abstract: Objective To study the mechanism of total Panax notoginseng saponin (tPNS) in regulating the transcription activity of human endothelial nitric oxide synthase (heNOS) gene promoter.Methods With gene recombination technique,we subcloned the heNOS gene promoter sequence (from -1 to -1 600 bp) into the BglⅡ/HindⅢ sites of the firefly luciferase reporter gene vector,pGL2-Basic (Promega),to yield the recombinant plasmid designated as peNOS-Luc. With lipofectamine- mediated co-transfection technique,peNOS-Luc,pGL2-Basic and pCMV-β were cotransfected into NIH3T3 cells,which were treated with lipopolysaccharide (LPS),tPNS and transforming growth factor β1 (TGFβ1) respectively. The relative activities (Luc/β-gal) were subsequently determined in the cell lysates to evaluate the effects of these 3 factors on the activity of heNOS gene promoter. Results Double restriction enzyme digestion and sequencing both confirmed that the recombinant plasmid,peNOS-Luc,was constructed correctly,which could be effectively expressed in NIH3T3 cells. Upon LPS stimulation,the luciferase activity was obviously decreased,contrary to the results of tPNS and TGFβ1 treatment,and between the latter two agents,TGFβ1 produced higher transcription activity.Conclusions A firefly luciferase reporter gene vector containing heNOS gene promoter sequence has been constructed correctly. tPNS can up-regulate the activity of heNOS gene promoter in NIH3T3 cells.

CLC Number:

R329.
R285.5

ZHANG Li-hua¹, JIA Yu-hua¹, SUN Xue-gang¹, XING Fei-yue², LIU Zhi-feng², SONG Ge², JIANG Yong². Effect of total Panax notoginseng saponins on the activity of human endothelial nitric oxide synthase gene promoter[J]. Journal of Southern Medical University, 2004, 24(10): 1113-1116.

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