Mechanism of Bushen Huayu Shengxin Decoction in Delaying Senescence of Bone Marrow Mesenchymal Stem Cells by Improving Ischemic-hypoxic Microenvironment / 中国实验方剂学杂志

ABSTRACT

Objective:To explore the mechanism of Bushen Huayu Shengxin decoction in delaying senescence of bone mesenchymal stem cells(BMSCs) by improving cellular microenvironment and regulating p16/pRb and p53/p21 signaling pathways. Method:The cells were cultured in serum-free 1640 medium and hypoxic cell workstation for 24 hours to establish the cell model of ischemic-hypoxic microenvironment in vitro, then randomized into control group (with complex medium), model group (with complete medium), and treatment group (with serum medium-containing Bushen Huayu Shengxin decoction), and all were cultured in hypoxic cell workstation for 24 hours. The normal group was added with control culture for complete medium, The cell cycle of BMSCs was detected by flow cytometry, the expressions of p16INK4a, p53, p21 and Survivn, cysteine aspartic acid protease-3 (Caspase-3), polyadenosine diphosphate ribose polymerase (PARP) mRNA were analyzed by Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR), and the levels of β-catenin protein and glycogen synthase kinase-3β（GSK-3β） protein were detected by Western blot. Result:Compared with the normal group, the proportion of S phase cells increased, while that at the G0/G1 phase decreased significantly in the model group (P<0.05). Compared with the model group, the proportion of S phase decreased, whereas that at the G0/G1 phase gradually increased in the treatment group (P<0.05). Compared with the normal group, mRNA expressions of p16INK4a, p53, p21 and Survivn, Caspase-3, PARP in the model group increased significantly (P<0.05). Compared with the model group, mRNA expressions of p16INK4a, p53, p21 and Survivn, Caspase-3, PARP in the treatment group decreased significantly (P<0.05). Compared with the normal group, protein expressions of β-catenin and GSK-3β in the model group increased significantly (P<0.05). Compared with the model group, protein expressions of β-catenin and GSK-3β in the treatment group decreased significantly (P<0.05). Conclusion:Bushen Huayu Shengxin decoction could delay the senescence of BMSCs by improving ischemic-hypoxic microenvironment and regulating p16/pRb and p53/p21 signaling pathways.