Effects Of Panax Notogiseng Saponins And Monomer Combination On The Endothelial Function And Monocyte-Endotheliocyte Adhension

As a common pathological foundation for most cardiovascular diseases, atherosclerosis is serious threaten for people's health and life. It has been indicated that injury of VEC is the precondition and the predominant mechanism in the initial stage of atherosclerosis. The oxidized low-density lipoprotein (ox-LDL), which is an independent risk factor in AS, can damage the vessel intima, upregulate the expression of endothelial adhesive molecules, induce the circulating monocytes migrating and adhering to intima, promote the macrophage to phagocytize lipid to form foam cells. The pathological processs described as above play an important role in the development of AS. Therefore, it is an key choice to prevent and treat the AS by protecting VEC and inhibiting the adhesion between EC and monocyte.Panax notoginseng saponins (PNS) are the principal ingredient extracted from the traditional Chinese medicine P.notoginseng. Epidemiological and clinical studies have shown that PNS have extensive and significant effects on the cardiovascular system. It has been observed in our laboratory previously that PNS could inhibit the development of atherosclerosis in rabbits effectively. There are more than 20 monomers extracted and identificated from PNS. Which of them play the critical role on the atherosclerosis? In other words, what is the material basis of the effects of PNS? That is key to understand the pharmacyological principle of PNS and to develop new drug for the treatment of atherosclerosis and other cardiovascular diseases.Considered the importance of EC in atherogensis, experiments were designed to investigate the effects of PNS and its momomers on the HUVEC injured by ox-LDL and the increase of adhesion of monocyte-endothelial cell induced by ox-LDL. At the same time, the regularity of compatibility of traditional Chinese medicine, the substace foundation and molecule mechanism of PNS on atherosclerosis were also studied.METHODS:The human umbilical vein endothelail cells (HUVEC) were isolated from umbilical vein by digestion with collagenase and identified by morphological methods and presence of von willebrand factor. The experiment was carried out with the ox-LDL to induce cell injury. Different concentration of PNS was added to the media, the cell viability, the injury of cells, and the content of ET and NO were determined with MTT assay, LDH assay, radio immunoassay and nitrate reductase assay, respectively. The monocyte-endothelial cell adhesion was measured by dye-bond protein method . The orthogonal experiment was designed to select the optimal monomer combination. Expression of ICAM-1 in endothelial cells were analyzed by western blotting and RT-PCR.RESULTS:1. After treated with ox-LDL for 24 hours, the viability of HUVEC decreased significantly compared with normal group (P<0.01). PNS alleviated the morphological damage and increased cell survival rate markedly.2. The LDH activity in ox-LDL group was more higher than that in normal group; PNS decreased LDH activity significantly compared with ox-LDL group.3. At the dosage of 300μg/ml, PNS increased the NO level and decreased ET level significantly by comparison of ox-LDL group, while PNS (30,100μg/ml) had no obvious effects (P>0.05).4. ox-LDL enhanced the adhesion of the monocyte to endothelial cell significantly compared with normal group(P<0.01), the treatment of PNS resulted in significant decline of adhesion between monocyte and endothelial cell(P<0.05 or0.01).5. In orthogonal experiment, Rg1, Rb1and R1 are main factors, the interaction between they was significant (P<0.05), the optimized combination was Rg1(10-5mol/L),Rb1(10-4mol/L), R1(10-5mol/L).6. It was showed that ox-LDL (100μg/ml) increased the expression of ICAM-1 mRNA and protein significantly compare with normal group (P<0.01). PNS and the optimal monomer combination could reduce ICAM-1 expression remarkably (P<0.05 or 0.01). CONCLUSION:1. ox-LDL (100μg/ml) could injury the HUVEC significantly. PNS could protect the VEC from damage and regulate the secretion of vascular active substance such as NO and ET. It indicated that PNS could interfere with the initial stage of the AS.2. PNS could inhibit the adhesion between monocyte and endothelium cell through down-regulating the expression of ICAM.3. The optimal monomer combination which could protect VEC and inhibit the monocyte-endothelial cell adhesion was Rg1(10-5mol/L), Rb1(10-4mol/L),R1(10-5mol/L).They may be the substace foundation for the antiatherogenic effect of PNS.