Journal of Functional Foods (Apr 2022)
Puerarin promotes apoptosis and senescence of bladder cancer cells
Abstract
Objective: This study aims to explore the mechanism of puerarin regulating the proliferation, apoptosis and senescence of T24 and EJ cells via miR-139-5p/CCNB1 and PI3K/AKT pathways. Methods: Initially, Cell Counting Kit (CCK)-8, flow cytometry, β-galactosidase detection kit, and RT-qPCR are used to assess the effect of puerarin on the proliferation apoptosis and senescence of bladder cancer cells by regulating miR-139-5p, respectively. Then, potential complementary binding sites between miR-139-5p and the 3′ untranslated region (3′ UTR) of the CCNB1 gene are obtained by the bioinformatics software, starBase database. Further, a dual-luciferase reporter system is used to assess the targeted relationship of miR-139-5p to CCNB1. CCK-8, flow cytometry, and β-galactosidase detection kit were used to detect the influence of CCNB1 on the proliferation, apoptosis, and senescence of bladder cancer cells. RT-qPCR and Western blotting are used to determine the regulation of CCNB1 and PI3K/AKT pathways by miR-139-5p. Finally, the subcutaneous tumor formation in nude mice and immunohistochemical experiments are performed to assess the effect of puerarin on the proliferation of bladder cancer cells in vivo. Results: Puerarin induced proliferation inhibition, apoptosis, and senescence of bladder cancer cells in vitro. Puerarin down-regulated the expression of CCNB1, p-PI3K, and p-AKT proteins by overexpressing miR-139-5p, thereby exerting a tumor suppressor effect in bladder cancer. Compared with the control group, the volume and weight of subcutaneous tumors of nude mice in the puerarin administration group are reduced. Moreover, the ki-67 positive staining rate and the expression level of CCNB1, p-PI3K, and p-AKT protein are reduced. Conclusion: The experimental results showed that the anti-tumor effect of puerarin against bladder cancer might be achieved by inhibiting CCNB1 and PI3K/AKT pathway via up-regulating miR-139-5p.
