The Research Of The Regulation Of The Extract Of Luoheng To Membrane Attack Complex C5b-9

This paper aimed to determine the sublytic dose of complementmembrane attack complex C5b-9on human umbilical vein endothelial cells(HUVEC), and to build the endothelial cell injury model. On this model, itis to research the regulation of the main ingredients in Luoheng, includingCynanchum, benzoin and different ratios of compound to inflammatorymediators lactate dehydrogenase (LDH), tumor necrosis factor (TNF-α)and interleukin-8(IL-8), then to explore the mechanism of their treatmentof atherosclerosis.HUVEC was cultured in vitro, and C5b-9was established on itssurface. Then the assembly results were observed by laser confocalscanning microscope (LCSM). And then by increasing the concentration ofcomplement, C5b-9was established on HUVEC again. With the qualitativeobservation by LCSM and quantitation detection of the concentration ofLactate Dehydrogenase in cell supernatant. Through the above the sublyticdose of C5b-9could be determined. Then the sublysis dose of C5b-9wasassembled on human umbilical vein endothelial cell surface in vitro by purecomplement proteins, and endothelial injury model was established.Groups were built; cells were cultured in different concentration of drugs.Then the concentration of LDH, TNF-α and IL-8in cell culture supernatantwas measured by UV and Elisa kits.The LCSM results showed that C5b-9had been successfullyassembled in HUVEC surface. When the concentration of C5b6came to1.6mg L-1, LDH’s concentration surged, and in this dose, the cellmembrane ruptured. After drug treatment, it could be measured that as to CynanchumGroup, LDH and IL-8activity were reduced in the high-dose group(119.854.12,33.652.48) significantly vs model control group(200.493.31,40.980.60)(P<0.01), TNF-α levels were reduced inhigh-dose (94.753.19) and medium-dose groups (92.811.78) vs modelcontrol group (108.002.30)(P<0.01). About Benzoin, the activity ofLDH(115.232.48) and IL-8(30.000.42) activity were reduced in extracthigh-dose group significantly vs model control group(200.493.31,40.980.60)(P <0.01), TNF-α(97.001.74) levels decreased inmedium-dose groups vs model control group (P <0.01). In combination(Cynanchum︰Benzoin)1︰1group, LDH and IL-8activity was reducedsignificantly vs model control group(200.493.31,40.980.60)(P <0.01),in the high-dose (113.842.62) and medium-dose (119.802.87) group theLDH levels were closed to blank control group (117.851.65), there wasno statistical difference, In extract high-dose group the IL-8(33.101.67)level was closed to the blank control group, and there was no statisticaldifference, TNF-α (98.692.01) level was similar to blank control group(98.692.01) in low-dose groups.In this paper, complement membrane attack complex C5b-9has beenestablished on human umbilical vein endothelial cells (HUVEC)successfully and its sublytic dose has been determined. The endothelial cellinjury model has been built, too. And through the research, it could befound that Cynanchum and Benzoin in Luoheng may repair endothelialinjury cells for therapy atherosclerosis by reducing the activity ofinflammatory mediators LDH, TNF-α and IL-8. When the mixture of thetwo drugs could produce geometric synergy, it may steadily reduce theactivity of various inflammatory mediators.