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OBJECTIVE:To study the effect and mechanism of physcion 8-O-β-glucopyranoside(PG)on the apoptosis of skin melanoma A375 cells. METHODS:After A375 cells were treated by PG with 0,10,20,50 μg/mL for 24,48,72 h,CCK-8 method was adopted to determine the survival rate of cells. After A375 cells were treated by PG with 0(control),20,50 μg/mL for 48 h,flow cytometry was used to detect the apoptosis rate of cells with membrane protein Ⅴ/propidium iodide (PI) double staining. Immunoblotting was used to detect the protein expressions of Caspase-3 and polyadenyl adenine diphosphate ribose poly-merase (PARP) and protein expressions of cytochrome C inside and outside mitochondria. After A375 cells were treated by PG with 0 (control),5,10 μmol/L for 48 h,enzyme substrate method was used to determine the activities of Caspase-8 and Cas-pase-9. RESULTS:PG can effectively decrease the survival rate of A375 cells. Compared with control,apoptosis rate of cells was obviously increased after treated by PG with 20,50 μg/mL(P<0.01);protein expressions of Caspase-3,PARP in cells and cyto-chrome C in cell matrix were obviously enhanced(P<0.05 or P<0.01);and protein expression of cytochrome C in mitochondria was obviously weakened(P<0.05 or P<0.01). Caspase-9 activity in cells was obviously enhanced after treated by PG with 5,10μmol/L(P<0.05 or P<0.01);and Caspase-8 activity had no obvious changes. CONCLUSIONS:PG can inhibit activity of A375 cells and promote its apoptosis,and its pro-apoptotic effects is achieved by destructing mitochondrial membrane potential and pro-moting cytochrome C outflow.
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OBJECTIVE:To study the effect and mechanism of physcion 8-O-β-glucopyranoside(PG)on the apoptosis of skin melanoma A375 cells. METHODS:After A375 cells were treated by PG with 0,10,20,50 μg/mL for 24,48,72 h,CCK-8 method was adopted to determine the survival rate of cells. After A375 cells were treated by PG with 0(control),20,50 μg/mL for 48 h,flow cytometry was used to detect the apoptosis rate of cells with membrane protein Ⅴ/propidium iodide (PI) double staining. Immunoblotting was used to detect the protein expressions of Caspase-3 and polyadenyl adenine diphosphate ribose poly-merase (PARP) and protein expressions of cytochrome C inside and outside mitochondria. After A375 cells were treated by PG with 0 (control),5,10 μmol/L for 48 h,enzyme substrate method was used to determine the activities of Caspase-8 and Cas-pase-9. RESULTS:PG can effectively decrease the survival rate of A375 cells. Compared with control,apoptosis rate of cells was obviously increased after treated by PG with 20,50 μg/mL(P<0.01);protein expressions of Caspase-3,PARP in cells and cyto-chrome C in cell matrix were obviously enhanced(P<0.05 or P<0.01);and protein expression of cytochrome C in mitochondria was obviously weakened(P<0.05 or P<0.01). Caspase-9 activity in cells was obviously enhanced after treated by PG with 5,10μmol/L(P<0.05 or P<0.01);and Caspase-8 activity had no obvious changes. CONCLUSIONS:PG can inhibit activity of A375 cells and promote its apoptosis,and its pro-apoptotic effects is achieved by destructing mitochondrial membrane potential and pro-moting cytochrome C outflow.
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OBJECTIVE:To observe the correlation of vitamin D with the incidence of cardiovascular disease and risk factors in elderly patients with type 2 diabetes mellitus. METHODS:94 patients,aged 60 years old above,with type 2 diabetes mellitus were collected retrospectively. Those patients were divided into diabetes group(41 cases)and diabetes complicated with cardiovas-cular disease group(53 cases)according to the condition of disease. 25(OH)D levels and lab indicators of 2 groups were detect-ed. The relationship of cardiovascular lesion with risk factor was analyzed by Multivariate Logistic regression;the correlation be-tween 25(OH)D and risk factors was analyzed by pearson analysis. RESULTS:There was statistical significance in the levels of 25 (OH)D,TC,TG,UA and Ca between diabetes group and diabetes complicated with cardiovascular disease group,with statistical significance(P<0.05). pearson analysis showed that the level of 25(OH)D was negatively related to age,FIB,TC and UA,and positively related to D-dimer and HDL-C,especially correlated with FIB and TC significantly,with statistical significance (P<0.01). Multivariate Logistic regression analysis showed that the deficiency of 25(OH)D was pathogenic factors of type 2 diabetes mellitus. CONCLUSIONS:There is a correlation between the level of 25(OH)D and cardiovascular risk factors as age,FIB,TC, UA,D-dimer,HDL-C. Vitamin D deficiency is the independent risk factors for cardiovascular disease in elderly patients with type 2 diabetes mellitus.
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OBJECTIVE:To investigate the regulation effects of oleanolic acid on the mitochondrial nitric oxide synthase(mt-NOS) in human umbilical vein endothelial cells (HUVECs) with oxidative damage. METHODS:HUVECs in exponential phase were divided into normal group,model group and oleanolic acid low,medium and high dose groups(5,20 and 35 μmol/L). After drug acting for 24 h,all groups were given culture solution containing 100 μg/ml ox-LDL to reproduce oxidative damage except normal group. CCK-8 was used to detect cell viability. The mitochondria in cells were extrated,enzyme chemical method was used to detect mtNOS activity and mtNO content,fluorescence microplate method was used to detect fluorescence intensity of reactive oxygen species(ROS),and western blot was used to detect expression of cytochrome C(Cyto-C). RESULTS:Compared with nor-mal group,the cell viability in model group was decreased;mtNOS activity,mtNO content,ROS fluorescence intensity and Cy-to-C protein expression were increased,with significant differences (P<0.05). Compared with model group,the cell viability in oleanolic acid low,medium and high dose groups was increased;mtNOS activity,mtNO content,ROS fluorescence intensity and Cyto-C protein expression were decreased,with significant differences(P<0.05),and they had positive correlation with concentra-tions. CONCLUSIONS:Oleanolic acid can decrease the mtNOS activity of HUVECs,reduce the production of mtNO and Cyto-C, by a mechanism that may be related to the decrease of ROS expression.