Effect Of Hh Signaling Pathway In Microenvironment Formation Of Mesenchymal Ovarian Cancer

Background and aims:Ovarian cancer is the most malignant tumor in the female reproductive system,and its incidence is third in the female reproductive system cancer,but the mortality rate is the highest in gynecological cancer.The current first-line treatment of ovarian cancer is surgery combined with chemotherapy,75%of patients will have a short-term recurrence and drug resistance.Studies have shown that ovarian cancer is a malignant tumor with heterogeneity in origin,pathological disease generation,clinical stage and prognosis,and can be divided into four subtypes(differentiated,proliferative,immunoreactive and mesenchymal)at the molecular level.Therefore,it is necessary to carry out research on various subtypes of ovarian cancer to find new therapeutic clues.Compared with other subtypes,mesenchymal ovarian cancer has a unique tumor microenvironment,with high expression of factors associated with promoting mesenchymal production(such as FAP that contribute to fibroblast proliferation,etc).And cancer-associated fibroblasts(CAFs)in the microenvironment can promote tumor proliferation,angiogenesis,metastasis,and drug resistance.But there is still a lack of in-depth study on the interrelationship between tumor cells and CAFs in the environment and their effect on tumor phenotype.This study will address the mutual feedback between cancer cells and fibroblasts and evaluate their potential application in cancer therapy.Methods:1.Analysis of the microenvironmental composition of mesenchymal ovarian cancer based on TCGA ovarian cancer data;screening for its molecular characteristics to distinguish other subtypes;2.Model of GLi1 gene overexpression in ovarian cancer cells(OVCAR3?A2780)was established,the changes of GLi1 and downstream target genes were analyzed by fluorescence real-time quantitative and Western Blot detection,and the effect of Hh signal on ovarian cancer cell phenotype was evaluated by six days proliferation rate and clone formation;3.Establish the culture system of supernatant culture CAFs of ovarian cancer cells(OVCAR3?A2780),and analyze the changes of iconic genes by fluorescence real-time quantitative detection and Western Blot detection.To evaluate the effect of serum on CAFs phenotype of ovarian cancer by EDU?six-day proliferation rate and clonal formation;4.Establish the culture system of ovarian cancer cells(OVCAR3?A2780)cultured in CAFs supernatant,and analyze the changes of GLi1 and downstream target genes in ovarian cancer cells by fluorescence real-time quantitative detection and Western Blot detection.The effects of supernatant on the phenotype of ovarian cancer cells were assessed by EDU>CAFs six-day proliferation rate and clonal formation:Results:1.Based on the transcriptome data of 308 cases of ovarian cancer,we found that the immune cells decreased significantly and the CAFs cells increased significantly in the microenvironment of mesenchymal ovarian cancer.2.qRT-PCR results showed that the mRNA levels of GLil and downstream CXCL12 were significantly up-regulated after overexpression of genes in ovarian cancer cells.Western blot results also showed a significant increase in protein expression CXCL12 after GLi1 overexpression.3.After Co-culture,the phenotypic experiments of cancer-associated fibroblasts and ovarian cancer cells showed an obvious trend of promoting proliferation.Conclusion:1.In the microenvironment of interstitial ovarian cancer,the number of immune cells decreased significantly,while the number of CAFs cells increased significantly.2.The activation of Hh signal pathway can promote the proliferation of ovarian cancer cells in microenvironment.3.The exudates of tumor-associated fibroblasts can induce the proliferation of ovarian cancer cells.4.Ovarian cancer cells can promote the proliferation of tumor-related fibroblasts.