RESUMO
Objective:To explore the effect of differentiation of cardiac stem cells(CSC)mediated by pilose antler polypeptides on the expressions of terminal myocardial differentiation genes atrial natriuretic polypeptide (ANP)and myosin light chain 2v(MLC-2v),and to clarify the mechanism of repairing the damaged myocardium.Methods:The healthy male Wistar rats born 2 d were selected to extract the CSC.The surface marker c-Kit and the purity of CSC were identified by immunofluorescence and flow cytometry.The culture dish was used as the experimental unit and the extracted cells were divided into the following four groups:blank control group(the same amount of buffer was added for induction),5-azacytidine group(induced with 3 μmol·L-15-azacytidine),pilose antler polypeptides group(induced with 800 mg·L-1pilose antler polypeptides)and combined group(induced with 800 mg·L-1pilose antler polypeptides and 3 μmol·L-15-azacytidine);the cells were incubated for 48 h in the condition of 37℃ and 5% CO2.The expression levels of ANP and MLC-2v in supernatant of the cells in various groups were detected by ELISA method.The expression levels of ANP and MLC-2v mRNA in the cells in various groups were detected by RT-PCR method.Results:CSC were prepared with the purity>95%.The results of ELISA showed that the expression levels of ANP and MLC-2v in 5-azacytidine,pilose antler polypeptides and combined groups were significantly increased compared with blank control group(P<0.05).The expression levels of ANP and MLC-2v in combined group were increased compared 5-azacytidine and pilose antler polypeptides groups,but there were no significant differences(P>0.05).The result of RT-PCR showed that the expression levels of ANP and MLC-2v mRNA in 5-azacytidine,pilose antler polypeptides and combined groups were significantly increased compared with blank control group(P<0.05).Conclusion:Pilose antler polypeptides can induce the differentiation of CSC into cardiac cells by promoting the expressions of ANP and MLC-2v,and they can reduce the cytotoxicity induced by 5-azacytidine in some degrees.
RESUMO
Objective To evaluate effects of Lycium barbarum polysaccharide (LBP) on expression of vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1α (HIF-1α) in ultraviolet B (UVB)-radiated HaCaT cells.Methods Conventionally cultured HaCaT cells were divided into control group and LBP groups,which were firstly treated with DMEM,12.5,25.0,50.0 and 100 μg/ml LBP solution respectively for 4 hours,and then were irradiated by UVB at different intensity of 0,20,40,60 mJ/cm2 separately.After 24-hour continuing culture,CCK-8 assay was performed to determine the cell survival rate,and an enzymatic-biochemical method to estimate the activity of superoxide dismutase (SOD).RT-PCR and Western blot analysis were conducted to measure the mRNA and protein expression of HIF-1α and VEGF respectively.Results Compared with the control group at the same UVB radiation dose,the 12.5-,25.0-and 100.0-μ,g/ml LBP groups showed different extents of increase in survival rates of UVB-radiated cells (P ＜ 0.05),and the 50.0-μg/ml LBP group showed the highest cell survival rate (P ＜ 0.01).Among all the LBP groups,SOD activity was highest in the 50.0-μg/ml LBP group (P ＜ 0.01).Along with the increase of UVB radiation dose,the mRNA and protein expression of HIF-1α and VEGF all gradually increased.Compared with the control group,the 50.0-μg/ml LBP group could effectively reduce the mRNA and protein expression of HIF-1α and VEGF in HaCaT cells (all P ＜ 0.05).Conclusion LBP may play a role in protecting cells from UVB radiation-mediated damage,likely by influencing the mRNA and protein expression of HIF-1α and VEGF in HaCaT cells.