Effect And Mechanism Of Ganoderma Lucidum Polysaccharides On Cultured Human Umbilical Vein Endothelial Cells Senescence Induced By AngiotensinⅡ

BackgroudA major risk factors of atherosclerotic cardioascular disease is aging. The morbidity of hypertension, coronary disease and shock rise signif ieantly. Although AS is known as chronic inflammatory disease, more and more evidence suggested that cell seneseence is one of the main causes of organism aging and diseases including atherosclerosis and hypertension. Therefore, studies on cell senescence and anti-senescence effects of GLP is significant to prevention and cure of AS.Object iveMany studies have now confirmed that endothelial cell senescence plays an important role in the vaseular aging proeess. This experiment establish the endothelial cell senescence model by angiotensin-IIindueing human umbilieal vein endothelial cells(HUVECs) aging, and explore the influence of GLP on endothelial cell senescence、caveolin-1、eNOS expression and NO seeretion indueed by AngⅡto HUVECs, compared with the control drug Irbesartan(Irb), to provide experimental and theoretical basis of GLP for clinical therapy of AS.MethodsWe culture HUVECs in vitro. Experiment was divided into:(1) control group:no intervention of factors;(2) AngⅡgroup:10-6mol/L AnⅡ;(3) Irb group:10-6mol/L Irb+10-6mol/L AngⅡ (4) GLP group:100mg/L GLP+10-6mol/L AngⅡ. cells and the culture medium were collected48hours after the intervention. Senescence β-galstaining、Human telomerase activity by PCR-ELISA method and cell cyele analysis were used to identify cell aging status. The caveolin-1protein and eNOS protein expression of endothelial cell were analyzed by Western blotting. And the cell culture medium were used to analyze the nitrie oxide(NO) content by using NO detection kit.Results1.Flow cytometry results of cell cycle analysis:compared to the control group(G0/G1phase and S phase), G0/G1phase of AngⅡgroup increased and S phase deereased, suggesting that Individual to join AngⅡon endothelial cells can induced G0/G1phase arrest andS phase decreased; Irb or GLP group compared with Ang II group (G0/G1phase and S phase) showed signifcant difference(P<0.05), Irb.. GLP inhibited AngⅡ on the endothelial cell cycle, making the reduetion of G0/G1phase and the inerease of S phase; but Irb group and GLP group(P>0.05)showed not statistically significant inhibiting Ang Ⅱ on the endothelial cell cycle.2.Cell senescence β-galaetosidase staining analysis showed that:compared to the control group, Ang Ⅱ group P<0.05, suggesting that Individual to Ang Ⅱon endothelial cells can enhance cell β-galaetosidase activity; Irb or GLP group compared with Angllgroup showed signifcant difference (P<0.05), Irb、 GLP inhibited Ang Ⅱ role in the enhancement of cell senescence β-galaetosidase activity; but Irb group and GLP group(P>0.05) showed not statistically significant inhibiting Ang Ⅱ role in the enhancement of cell senescence0-galaetosidase activity.3. PCR-ELISA results of telomerase activity analysis:compared to the control group, AngⅡgroup P<0.05, suggesting that Individual to Angllon endothelial cells can lower telomerase activity; Irb、GLP group compared with AngⅡgroup showed signifcant difference(P<0.05), Irb or GLP inhibited Angllrole in the lowerment of cell telomerase activity; but Irb group and GLP group(P>0.05)showed not statistically significant inhibiting AngⅡrole in the lowerment of cell telomerase activity.4. The results of Western Blot:AngⅡ inhibited markly eNOS protein expression decreased in HUVECs and promoted significantly caveolin-I protein expression (P<0.05). On the other hand, GLP enhanced the expression of eNOS protein dramatically and suppressed the expression of caveolin-1protein significantly in HUVECs (P<0.05). the effect of Irb was similar to that of GLP (P>0.05).5.The results of NO test showed that:compared to the control group, AngⅡ group P<0.05, suggesting that Individual to Ang Ⅱ on endothelial cells can inhibit the secretion of NO;Irb、GLP group compared with AngⅡgroup showed signifcant difference(P<0.05), suggesting that Irb or GLP inhibited Ang Ⅱ role in redueing cell NO generation on endothelial cells; but Irb group and GLP group(P>0.05) showed not statistically significant redueing AngⅡrole in in redueing cell NO generation on endothelial cells.Conculsion1. Ang Ⅱ can induce human umbilieal vein endothelial cells (HUVECs) senescence.2. GLP can inhibit endothelial cell senescence induced by AngⅡ.3. Caveolin-1might play a key role in attenuating eNOS activity. The role o f GLP in endothelial cell senescence indueed by AngⅡmay be related to the down-regulated expression of caveolin-1protein in endothelial cells, and prompte the expression of eNOS protein, causing the amount of NO to elevate.