| Cervical cancer is the second most common cancer in women worldwide. High-risk human papillomavirus (HPVs) have been well established as the major cause of cervical cancer. The E6and E7viral proteins inactivate the key tumor suppressors, p53and pRB respectively. Cell cycle disorder results in abnormal proliferation and malignant transformation of cells. Previous study of us has indicated that progranulin (PGRN) is highly expressed in cervical cancer tissue and cell lines, suggested PGRN may contribute to development and progression of cervical cancer.Hepatocellular carcinoma (HCC) is the third most common malignant tumor in the world, and the morbidity of HCC is the second place in China. According to the world health organization, about1million people die of liver cancer each year in the world. HCC is easily recurrence and metastasis after surgery, therefore the treatment of liver cancer is still difficulty in clinical research. High expression of PGRN was found in over70%of HCCs studied. PGRN is related to liver cancer cells proliferation, invasion and tumorigenicity, which provides a potential that PGRN may be a therapeutic target of HCC.PGRN, a secreted growth factor, is widely distributed in various tissues. It is involved in embryonic development, wound repair, inflammation, and tumorigenesis. Earlier study indicate that,PGRN is highly expressed in aggressive cancer cell lines and clinical specimens such as breast, ovarian, prostate, HCC and glioblastoma. The elevated expression of PGRN promotes the activation of MAPK/Erk, PI3K/Akt/mTOR, and FAK signaling pathways, by which subsequently lead to enhanced protein synthesis, acceleration of cell proliferation and invasion, and inhibition of cell apoptosis. Akt/mTOR signaling pathway is related with various physiological processes including cell growth, proliferation and differentiation. Amount of studies have demonstrated that alteration of the Akt/mTOR signaling pathway are frequent in human cancers including gastric, small cell lung, cervical and liver cancer. Therefore, Akt/mTOR signaling pathway has been extensively studied as novel targets for cancer therapeutics. At present, the molecular mechanism of PGRN in the tumorigenesis of cervical cancer and liver cancer remains poorly understood. In this study, we investigated PGRN regulated Akt/mTOR signaling pathway in cervical ande liver cancer cell lines, and the contribution of Akt/mTOR signaling pathway in PGRN stimulated cell proliferation.1. Role of Akt/mTOR signaling pathway in PGRN promoted proliferation of cervical cancer cells.Objective To investigate the activation of Akt/mTOR and Erk signaling in cervical cancer cells stimulated by PGRN, and it’s role in the proliferation of cervical cells. Methods Activation of Akt, Erk, mTOR, and p70S6K were assessed in recombinant human PGRN (rhPGRN) stimulated SiHa and HeLa cells by western blot assay. CCK-8assay, colony-formation assays in soft agar or in medium with different concentration of FBS were used to examine the proliferation, anchorage-independent growth, and survival of cervical cells treated with rhPGRN in the presence or absence of inhibitors of PI3K/Akt, MEK/Erk or mTOR. Results The levels of phosphorylated Akt, Erk, mTOR, and p70S6K cells were elevated in SiHa and HeLa treated with rhPGRN. LY294002, U0126or rapamycin pretreatment inhibited PGRN enhanced proliferation of H8and HeLa cells, and anchorage-independent growth, survival in low concentration FBS of H8cells. Conclusions PGRN stimulates the activation of Akt/mTOR and Erk signaling pathway in cervical cancer cells, and PGRN enhanced proliferation, transformation and survival of cervical cells depend on the activation of Akt/mTOR and Erk signaling pathway.2. Role of Akt/mTOR signaling pathway in PGRN promoted cell proliferation and tumor formation of liver cancer cells. Objective To explore the role of activation of Akt/mTOR signaling pathway in PGRN stimulated proliferation of liver cancer cells. Methods Activation of Akt, Erk, mTOR, and p70S6K were assessed in recombinant human PGRN (rhPGRN) stimulated HepG2cells by western blot assay. CCK-8assay, colony-formation assay in medium with different concentration of FBS, and Tumor growth assay were used to examine the proliferation and survival of cervical cells treated with rhPGRN in the presence or absence of inhibitors of PI3K/Akt, MEK/Erk or mTOR. Results rhPGRN treatment promoted phosphorylation of Akt, Erk, mTOR and p70S6K. Akt/mTOR inhibitors reduced PGRN enhanced proliferation of HepG2cells. rhPGRN administration increased tumor growth of HepG2in nude mice, which was slowed by mTOR inhibitor rapamycin in vivo. Conclusions PGRN stimulates the activation of Akt/mTOR and Erk signaling pathway in liver cancer cells, and Akt/mTOR signaling pathway mediates PGRN enhanced cell proliferation in vitro, and tumor growth in vivo.In summary, we observed (1) PGRN stimulates the phosphorylation of Akt, Erk and mTOR in cervical cancer SiHa and HeLa cells, and subsequently promotes the activation of p70S6K, a downstream protein of mTORC1;(2) The regulatory role of PGRN in cell proliferation, anchorage-independent growth and survival depend, at least partially, on activation of Akt/mTOR signaling pathway;(3) rhPGRN treatment activates Akt/mTOR and Erk signaling pathway in liver cancer HepG2cells;(4) mTOR inhibitor, rapamycin, can inhibit PGRN stimulated cell proliferation and tumor growth in nude mice of HepG2cells.The determination of phosphorylate level of endocellular signaling molecules response to rhPGRN stimulation indicate that PGRN can increase the phosphorylation of mTOR, and activate the Akt/mTOR signaling pathway in cervical and liver cancer cells. Studies using inhibitors of signaling pathway also proves that Akt/mTOR signaling pathway play a crucial role in PGRN-regulated cell behaviors, which provides a theoretical and experimental basis to the molecular mechanism of PGRN in development and progression of cancer and therapeutic strategies of cancer targeting PGRN. |
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