PGRN Contributes To The Migration And Invasion Of Cervical Cancer Cells By MTOR Signaling Pathway

Cervical cancer,as one of the most common malignancies in the female reproductive system,has become the second killer of women aged 15-44,second only to breast cancer.In 2012,more than 500,000 new cervical cancer cases,of which more than 85%occurred in developing countries,more than 260,000 cases of death.There is a lot of evidence that high-risk HPV infection is a key factor in cervical cancer,but only a few of HPV female infections develop cervical cancer,suggesting that other factors contribute to cervical cancer progression.The main features of malignant progression of cancer cells include growth signal self-sufficiency,growth inhibition signal tolerance,escape procedural death,infinite replication potential,persistent angiogenesis,tissue invasion and metastasis.Metastasis is the ultimate and most dangerous stage of tumor progression,with more than 90%of cancer patients dying from metastatic rather than primary tumors.Progress in cervical cancer is significant,including normal cervical,cervical intraepithelial neoplasia,carcinoma in situ,locally invasive and distant metastatic cancer.Early cervical cancer(?a-?a)patients generally better prognosis,but one-third of the death and recurrence,especially in distant organs or lymph node metastasis of cervical cancer(?b-? period)patients is difficult To achieve the purpose of radical.Pelvic lymph node metastasis is an important way to spread cervical cancer,lymph node metastasis with a direct impact on the prognosis of patients.To explore the mechanism of cervical cancer invasion and metastasis to enhance efficacy,prevent recurrence and improve the quality of life of patients is of great significance.Cancer cell metastases are divided into multiple steps,including cancer cells from the in situ tumor detachment,invading the surrounding tissue,penetrating into the blood vessels or lymphatic vessels,spreading with the bloodstream or lymphatic system,eventually infiltration and growth in the distal organs.Each step requires a unique molecular program involved,in which cancer cell adhesion and migration regulation and cytoskeleton properties play an important role.In addition,the initiation of tumor metastasis is associated with morphological changes in cancer cells,that is,epithelial-mesenchymal transition(EMT).Progranulin(PGRN)is a new multi-functional growth factor,which has the functions of promoting cell proliferation,survival,migration and anti-inflammation.It is involved in a variety of important physiological and disease processes.PGRN was originally identified as a growth factor for cancer cells,expressed in many tumors at high levels,associated with poor prognosis in tumors,and mediated the development and progression of breast cancer,ovarian cancer,prostate cancer,bladder cancer and liver cancer.PGRN is closely related to the invasion and metastasis of cancer cells,which can enhance the ability of fibroblasts to drive SW13 and MCF-7 cells through the matrix glue membrane to stimulate the production of vascular endothelial growth factor and angiogenin in breast cancer cells and promote cancer Cell matrix metalloproteinase production.etc.,and recent studies have found that PGRN can enhance the EMT program mediated ovarian cancer cell migration and invasionOur preliminary study reported for the first time that PGRN was highly expressed in cervical cancer as a tumor microenvironment factor and promoted malignant proliferation and transformation of cervical cancer cells.However,the role of PGRN in the migration and invasion of cervical cancer cells and the regulation of PGRN on the migration and invasion of cancer cells have not been reportedMethods:The cervical cancer cell lines(HeLa,SiHa)were treated with recombinant human PGRN(rhPGRN).The effects of PGRN on the motility,migration and invasion of cervical cancer cells were detected by scratch repair,transwell migration and invasion assays.The expression of PGRN and p-mTOR in normal cervical tissues and cervical cancer tissues was detected by immunoblotting and immunohistochemical staining.The role of mTOR signaling pathway in PGRN-stimulated cervical cancer cells migration and invasion was detected by using transwell assays with rapamycin pretreatment.The changes of cell structure related to the migration of cervical cancer cells were observed by using laser confocal microscopy after F-actin immunofluorescence staining.The activation of Rho small GTPase and phosphorylation of FAK,LIMK and cofilin were detected by western blot.The role of PGRN-stimulated FAK,Rho small GTPase signaling pathways in migration and invasion of cervical cancer cells were detected by using transwell assays with indicated inhibitors pretreatment.Results:rhPGRN treatment enhanced the motility,migration and invasion ability of cervical cancer cells.The effect of PGRN on the EMT of cervical cancer cells was weak,but PGRN promoted the cytoskeleton rearrangement,which is manifested as pressure fiber enhancement,plate pseudopodia and filopodia.PGRN activated mTOR signaling pathway through TNFR2 and the expression of PGRN was positively associated with p-mTOR levels in cervical cancer tissue.Inhibition of mTOR signaling by rapamycin reduced PGRN-stimulated migration and invasion of cervical cancer cells;PGRN activated cytoskeleton rearrangement-associated LIMK/cofilin signaling pathway,and enhanced FAK phosphorylation.PGRN-mediated migration and invasion of cervical cancer cells involved PGRN-activated FAK and Rho small GTPase.Conclusion:PGRN promotes the motility,migration and invasion of cervical cancer cells.It is found that PGRN mainly regulates the migration of cancer cells by promoting cytoskeletal rearrangement,and activation of FAK and mTOR-mediated Rho small GTPase signaling pathways contributes to PGRN-stimulated migration and invasion of cervical cancer cells.This study found that PGRN promotes the invasion and metastasis of cervical cancer and participates in the new mechanism of malignant progression of cervical cancer,which provides a new target for the control of cervical cancer invasion and metastasis.This study first proposed the regulation of PGRN on cytoskeleton rearrangement linking PGRN-mediated FAK and mTOR/LIMK/cofilin signaling pathway which provides a new perspective for PGRN signaling pathway and biological function research.