Effects Of Valsartan And Angiotensin Ⅱ On NOS And NO In Endothelial Cells Of Cultured Human Coronary Artery

Objective To Build a model of adult coronary artery endothelial cells culture in vitro, and in order to study the effects of valsartan and angiotensin Ⅱ (AngⅡ) on nitric oxide synthase(NOS) activity and nitric oxide (NO) level in cultured human coronary artery endothelial cells.Methods Endothelial cells were separated by collagenase digesting and scraping from adult coronary artery of sudden death, they were cultured with Dulbecco's modified Eagle medium (DMEM).The second generation cells were divided into two parts, one was freezed, and the other was identified by phase contrast microscope, transmission electron microscope and immunohistochemical staining;The freezed endothelial cells were resurrected and planted into culture bottles of 5 millilitres. They were divided into contrast group, AngⅡ group (10 μ mol/L), valsartan group(10μmol/L), Anglland different concentrations valsartan (0.1μmol/L, 1μmol/L , 10μmol/L) groups. The drugs which included valsartan and AngⅡ were put in when the third generation cells had reached 60 percent of culture bottle. We examined the NOS activity, NO level of cultured human coronary artery endothelial cells after the cells with Anglland valsartan had been incubated for 36 hours.Results lhere were 30-40% adherent rate of cells in primary cultures within 24 hours, the endothelial cells were growing at second day;the quantity of cells had increased obviously at 5lh day and the bodies of cell connected tightly;the borderline between cell and cell was indistinct at 7"1 day;the cultured cells became confluent monolayer and arranged closely at 10'" day. The cells in secondary culture showed polygons, all of them had obvious borders and grew actively, according with the biological characteristic of vascular endothelial cells. By transmission electron microscope, the specific cytoplasmic organelle (Weibel-Palade bodys, WPBs) of endothelial cells had been found in their cytoplasm. The immunohistochemical staining of the rabbit anti-factor VIII related antigen showed the cultured endothelial cells contain factor VIH related antigen (Vffl-R Ag);The difference of NOS activity between 6 groups had statistical significance(F=407. 747, P=0. 000);The difference of NO level between 6 groups had statistical significance(F=258. 918, P=0. 000);NOS activity and NO level were lower in Angll group than in control group, there were statistical significance between the two groups(P=0. 000);but there were not statistical significance between valsartan group and control group in NOS activity and NO level (P >0. 05);NOS activity and NO level were lower in Angll group than in Ang II +0. 1 u mol/L valsartan group, there were statistical significance between the two groups (P=0. 000);NOS activity and NO level were lower in Angll group than in AngII +1 V- mol/L valsartan group, there were statistical significance between the two groups (P=0.000);NOS activity and NO level were lower in Angll group than in AngII+10umol/L valsartan group, there were statistical significance between the two groups (P=0. 000);NOS activity and NO level were lower in AngII+0. 1 V- mol/L valsartan group than in Ang II +1 u mol/L valsartan group, there were statistical significance between the two groups (P=0.000);NOS activity and NO level were lower in Ang II +1 n mol/L valsartan group than in AngII+10 u mol/L valsartan group, there were statistical significance between the two groups (P=0. 000).Conclusion Lit was proved that the cultured cells were endothelial cells.2. Using alone AngII could decrease NOS activity and NO level of human coronary artery endothelial cells3. Using alone valsartan had no effects on NOS activity and NO level. 4.The action of Angll that decreased NOS activity and NO leve could be reversed by valsartan. The result suggested that valsartan could improve the function of human coronary artery endothelial cells and prevent atherosclerosis.