The Protection And Mechanism Study Of Main Active Ingredients Of GuanXin Decoction On Human Umbilical Vein Endothelial Cells Stimulated By Tnf-α

Objective To observe the effect of main active ingredients of Guanxin Decoction on expression of NF-KB、eNOS、VCAM-1、ICAM-1and E-selectin of human umbilical vein endothelial cells(HUVECs) stimulated by TNF-α.Investigating, the protective effect and mechanism of Guanxin Decoction to provide an experimental basis for its clinical application.Methods To culture the HUVECs in vitro, and divide them into six groups randomly:the normal control group(Normal),the TNF-a positive control group(TNF-a),the Astragaloside IV group(AG),the Isorhamnetin group(IG),the Quercetin group(QG),the P-Sitosterol group(SH). HUVECs of the normal control group were conventional cultured twenty-four hours,HUVECs of the TNF-a positive control group were stimulated by the TNF-a of200ng/ml six hours after conventional cultured18hours first.HUVECs of the four active ingredients groups were cultured with culture medium containing Astragaloside IV (80ug/ml), Isorhamnetin (30ug/ml),Quercetin(20ug/ml),β-Sitosterol(20ug/ml) for eighteen hours,and then stimulated by the TNF-a of200ng/ml six hours.Finally, collecting the HUVECs of each group,the Western Blot was used to detect the protein quantitative of NF-Kb, eNOS, ICAM-1, VCAM-1and E-Selectin expressing from HUVECs.Results Compared with HUVECs of normal control group, Western Blot results showed that the TNF-α stimulation group,the expression of VCAM-1, ICAM-1, E-selectin and NF-κB were significantly increased,while the expression of eNOS decreased obviously, significant statistical significance(P<0.01).Compared with the TNF-α positive control group, the Astragaloside IV group,the Isorhamnetin group,the Quercetin group,the β-Sitosterol group can obviously inhibit the quantitative expression of VCAM-1, ICAM-1, E-Selectin and NF-κB (P<0.01), but also significantly up-regulate the expression of eNOS (P<0.01), especially the effects of the Astragaloside IV group was significant in the above, β-sitosterol group can inhibit the expression of VCAM-1,E-selectin,and up-regulate the expression of eNOS (P<0.01), it plays the weakest role,but it had no obvious effect on the inhibition of the expression of ICAM-1and NF-κB (P>0.05).Conclusion The Astragaloside IV, the Isorhamnetin and the Quercetin have a protective effect on inhibiting the expression of NF-κB, ICAM-1, VCAM-1and E-selectin, up-regulate the expression of eNOS from HUVECs, especially the effects of the Astragaloside IV were significant. β-Sitosterol can inhibit the expression of E-selectin,VCAM-1,up-regulate the expression of eNOS,but with weakly compared with other active ingredients,and it had no obvious effect on the inhibition of the expression of ICAM-1and NF-κB.The Mechanism of HUVEC stimulated by TNF-α can be mitigated by GuanXin Decoction. It may relate to the inhibition of NF-κB transcription activity, so as to reduce the expression of ICAM-1, VCAM-1and E-selectin, at the same time, enhance the activity of eNOS to make NO protecting the vascular endothelium.