| Objectives This experiment was designed to investigate the effects ofoxymatrine in human umbilical vein calcification of the smooth muscle cells andexplore the possible mechanisms on the basis of the human umbilical vein smoothmuscle cells （HUVSMCs） calcification model.Methods In the same time of the reproduction of vascular calcific model whichwas induced by β-Glycerophosphate, the cells were randomly divided into6groups:the control group（CON）,calcification group（CAL）,OMT group,theintervention groups:OMT L,OMT M,OMT H(OMT with10-6,10-4,10-2mol/L incalcified nutrient solution).Detection methods:①Identification of HUVSMCs:thecells were observed directly by binocular inverted phase contrastmicroscope;immunohistochemical stainned by anti-ɑ-smooth muscle actin（anti-ɑ-SMA）.②C alcium deposition inHUVSMCs were detected by Von Kossastaining.③Von Kossa staining of calcification percentage in positive cells werecalculated by micro-image analysis system.④The contents of alkaline phosphatase（ALP） in HUVSMCs were verified by pheny1diphosphate-2-sodium.⑤The totalcalcium contents in HUVSMCs were calculated by ultraviolet spectrophotometry.⑥A ctivity ofTGFβ1in the nutrient solution were determined by enzyme-linkedimmunosorbent assay, determining the change of the level of transforming growthfactor β1（TGFβ1） in HUVSMCs.⑦The expression of oseoprotegerin（OPG）, ligandreceptor activator of nuclear factor kappa B,（RANKL） in HUVSMCs were determined by Western-blot.⑧Osteocalcin （OC） contents in the nutrient solution weredetermined by radio immunoassay.Results①Reproduction of vascular calcific model: Von Kossa staining showedbrown calcium nudules in CAL group was significantly increased compared withCON group, the calcification cells percentage, calcium content and ALP of CALgroup were significantly increased than CON group（P<0.05）.②OMT inhibition ofHUVSMCs calcification: OMT can reduce the aggregation growth of calified smoothmuscle cell, Von Kossa staining showed brown calcium nudules in OMT interventiongroups was significantly reduced compared with CAL group,the calcification cellspercentage, calcium content and ALP of OMT intervention groups were significantlylower than CAL group（P<0.05）.③The mechanisms of OMT related with HUVSMCscalcification: the expression of TGFβ1and OC of OMT calcified group obviouslydecreased compared to CAL group, and the expression of OPG significantly increased,but the expression of RANKL significantly decreased.Conclusions①O MT could inhibitHUVSMCs calcification.②OMT couldinhibit HUVSMCs calcification with the appropriate concentration by weakeningTGFβ1,OC and the influencing the key signaling protein in OPG/RANK/RANKLsystem. |
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