Inhibitory effect of genistein on cervical carcinoma cell through AMPK and mTOR signaling pathway

Abstract

Abstract: AIM: To observe the effect of genistein on growth and apoptosis of cervical carcinoma Caski cell lines , and to explore its potential mechanisms.METHODS: Cervical carcinoma cell lines Hela, SiHa, Caski, HeLa-S3, HeLa229 and C-33A were cultured and treated with genistein at different concentration and at different time points. The effect of genistein on cervical carcinoma cell lines proliferation was studied by means of Sulforhodamine B (SRB), the cell cycle and mitochondrial membrane potential were detected with flow eytometry;RT-PCR and western blot were used to detect the expressions of AMPK and mTOR signaling pathway related gene at mRNA and protein levels. Determination of TSC1/TSC2 association by immunoprecipitation assay.RESULTS: The rank order of genistein potency against cervical carcinoma is Caski>Hela> C-33A > SiHa>HeLa-S3>HeLa229. Genistein completely abolished cell-cycle progression released from double-thymidine-block synchronization and caused a subsequent apoptosis. The data were supported by down-regulation and reduced nuclear translocation of G₁-regulator proteins, including cyclin D1, cyclin E, Cdk4 and Cdk2. Further analysis showed that the mRNA expressions of the G₁-regulator proteins were not modified by Methyl jasmonate, indicating an inhibition of translational but not transcriptional levels. Genistein induced the assembly of tuberous sclerosis complex (TSC1/TSC2), leading to the blockade of cellular protein synthesis through inhibition of protein phosphorylation including mTOR (Ser2448), p70S6K (Thr421/Ser424 and Thr389) and 4E-BP1 (Thr37/Thr46 and Thr70). Furthermore, the AMPK activity was elevated by genistein. Compound C, a selective AMPK inhibitor, significantly reversed genistein-mediated effects suggesting the crucial role of AMPK. Besides, the loss of mitochondrial membrane potential and depletion of mitochondrial content indicated the mitochondrial stress caused by genistein.CONCLUSION: Genistein induces anticancer signaling cascades in a sequential manner. The exposure of cells to genistein induces mitochondrial stress and activation of AMPK that further induces the loss of mitochondrial membrane potential and activates TSC1/TSC2 association. Consequently, the mTOR mediated translational pathways are blocked, leading to G₁ arrest of the cell-cycle and subsequent cell death.

Key words: Genistein, Cervical carcinoma, AMPK, mTOR, G₁ arrest

CLC Number:

R965.2

LI Yong-zheng, CHEN Wei. Inhibitory effect of genistein on cervical carcinoma cell through AMPK and mTOR signaling pathway[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2014, 19(1): 15-22.

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