Progranulin Is Necessary For Epithelial Ovarian Cancer Progression And Regulates Epithelial-mesenchymal Transition

Background:Ovarian cancer (OC) is the most lethal gynecologic malignancy. Epithelial ovarian cancers (EOCs) accounted for 90% of all ovarian cancers. At present, the main treatment of epithelial ovarian cancer is cytoreductive surgery accompanied with platinum-based chemotherapy, but its five-year survival rate is only about 30%. The main reason is that majority of epithelial ovarian are not diagnosed until the disease are at advanced stage, when the multiple distant metastasis has occurred. It has been confirmed that distant metastasis of malignant tumor is closely related to the invasion and infiltration capacity of the tumor cell. Therefore, it is of great significance to clarify molecular alteration and the occurrence and development process of epithelial ovarian cancer, and to explore the mechanism of acquiring invasion and metastasis ability for epithelial ovarian cancer early diagnosis, treatment and prognosis.Progranulin (PGRN) is one of secretory growth factors discovered in the early 1990s, also called granulin/epithelin precursor(GEP), proepithelin(PEPI), PC cells-derived growth factor (PCDGF) or acrogranin, can be extracted in various tissue, different names were given according to the histologic type of extract. The gene of PGRN codes protein with 68.5 kDa in size, and protein molecular weight of high glycosylation of PGRN is 90 kDa. PGRN has a variety of biological functions and plays an important role in inflammatory reaction, embryonic development, wound repair, nerve nutrition, insulin resistance etc. In addition, the expression of PGRN increases in a variety of cancers, associated with the invasion, metastasis, drug resistance, stem cell characteristics of tumor cells in breast cancer, liver cancer, cervical cancer, adrenal carcinoma. The PI3K/AKT, ERK signaling pathways are involved in these biological processes. In addition, some studies confirm that PGRN can up-regulated GSK3P and cyclinDl, and subsequently participates in a series of biological processes. Recent studies have reported that the expression level of PGRN is closely related to the prognosis of the tumor, can be used as indicators to predict the prognosis of patients with tumor.Epithelial mesenchymal transition (EMT) can make epithelial cells obtain mesenchymal phenotype, give cell invasion and metastasis ability, play an important role in the process of the occurrence and development of tumor. At present, PGRN’s effect in epithelial mesenchymal transition has not yet been reported at home and abroad, the effect and its mechanism of action need further to be elaborated. Objective1. Researching on the different expression of PGRN between benign epithelial ovarian tumors and epithelial ovarian cancers;2. Researching on the PGRN’s effect to the ability of proliferation, metastasis, invasiveness in epithelial ovarian cancer cell;3. Researching on relationship between PGRN and epithelial mesenchymal transformation; testing molecular expression associated with EMT after the PGRN’s over expression in protein level;4. Researching on underlying molecular mechanisms of PGRN in the process of development in epithelial ovarian cancer.Methods1. The method of immunohistochemical was used to detecting the different expression of PGRN in benign epithelial ovarian tumor group (34 cases) and epithelial ovarian cancer tissue (42 cases), and to see if there are some correlation between PGRN and Snail in epithelial ovarian cancer tissue.2. Lentivirus transfection was used to build stable over-expression PGRN of epithelial ovarian cancer cell line (A2780, HO8910); CCK8 assay was used to detect the proliferation ability of cells; Transwell Chambers was used to detect the migration and invasiveness ability of cells;3. Western blotting was used to detect the changes of expression of epithelial-mesenchymal transition markers (E-cadherin, ZO-1, N-cadherin, Vimentin, Snail and Slug) on the protein level after PGRN was increased;4. Western blotting was used to detect the activity changes of Wnt/β-catenin, AKT/mTOR signaling pathway on the protein level after PGRN was increased; Results1. Immunohistochemical score results of benign epithelial ovarian tumor group and epithelial ovarian cancer group show that the immunohistochemical score of epithelial ovarian cancer group are obviously higher than the benign epithelial ovarian tumor group(3.2941±1.0317 vs.8.5500±0.7014) (p< 0.01). And there is a positive correlation property between PGRN and Snail in epithelial ovarian cancer group;2. The ability of cell proliferation, migration and invasion are significantly enhanced by over-expression of PGRN in epithelial ovarian cancer cells (A2780, HO8910)(p<0.05);3. The expression of epithelial phenotype marker (E-cadherin, ZO-1) are significantly decreased by over-expression of PGRN in epithelial ovarian cancer cells (A2780, HO8910), while the expression of mesenchymal phenotypic markers (N-cadherin, Vimentin, Snail and Slug) are significantly increased, which suggest that PGRN promoted epithelial ovarian cancer cells to transform from epithelial phenotype to mesenchymal phenotype.4. The expression of β-catenin is increased by over-expression of PGRN in epithelial ovarian cancer cells (A2780, HO8910); Meanwhile, the expression of p-GSK-3P (ser9) is significantly decreased, while GSK-3β expression had no obvious change; Those show that Wnt/β-catenin signaling pathway is activated by the increase of PGRN.5. The expression of p-mTOR、p-AKT is increased by over-expression of PGRN in epithelial ovarian cancer cells (A2780, HO8910); those suggest that AKT/mTOR signaling pathway is activated by the increase of PGRN.Conclusions1. The expression of PGRN increases in epithelial ovarian cancer tissue; and there is a positive correlation property between PGRN and Snail in epithelial ovarian cancer tissue.2. PGRN can enhance the ability of proliferation, metastasis, invasiveness in epithelial ovarian cancer cells;3. PGRN can induce epithelial-mesenchymal transition in epithelial ovarian cancer cells;4. PGRN may induce epithelial-mesenchymal transition through the activation of Wnt/β-catenin, AKT/mTOR signaling pathways in epithelial ovarian cancer cells.