The Effect of Procyanidin B2 on Alleviating Cellular Senescence Through GATA4/P53 Pathway in BNL.CL2 Cells
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Abstract:
The alleviation of BNL.CL2 hepatocyte senescence by procyanidin B2 was investigated. Palmitic acid (PA) was used to induce hepatocyte senescence. The MTT assay was used to identify the cell viability treated by PA and PCB2. The effect of PCB2 on cell senescence-specific β-galactosidase (SA-β-Gal) expression was detected by senescence-specific β-galactosidase assay kit. The effect of PCB2 on the expression of cell cycle-related genes CDK6 and p53 mRNA was examined by real-time quantitative PCR. The effect of PCB2 on P53 and GATA4 protein expression and nuclear size was detected by immunofluorescence staining. The results showed that the 100 μM PA inhibited cell proliferation (control: 1.289 vs PA: 0.655, p<0.001), 12.5 μg/mL PCB2 improved the cell proliferation (0.766, p<0.01) which was inhibited by PA. Compared with PA group, the number of positive cells expressing β-galactosidase in PCB2 group is significantly decreased. The nucleus size was increased after PA treatment (PA: 7995.1, p <0.05), while PCB2 reversed the effect (3848.8, p <0.05). Senescence associated gene p53 expression was raised (3.36, p <0.05), CDK6 gene expression decreased (0.49, p <0.05) in the PA group; PCB2 could effectively improve the senescence of pa induced liver cells (p53: 1.11; CDK6: 1.01, p <0.05). Immunofluorescence staining showed that p53 and GATA4 were co-localized in the nucleus of the PA-induced senescent cells. After the PCB2 treatment, expression of p53 and GATA4 proteins in nucleus decreased, and p53 and GATA4 proteins co-occurred in the cytoplasm. Therefore, we believe that PCB2 can alleviate hepatocyte senescence, which may be through the GATA4/p53 signaling pathway.
