| BackgroundGastric cancer is the most common tumors of digestive system in world,and its incidence rate ranks fourth among all cancers,the mortality rate ranks second in the world.Although there are a variety of new chemotherapeutic drugs and molecular targeted drugs in clinical,but the efficacy of advanced gastric cancer is still poor,the rate of 5 year survival has been around 15%-25%.Therefore,it is important to find the early biomarkers and therapeutic targets for gastric cancer.Estrogen related receptors(ERRs)are characterized by the conserved domain of nuclear receptors,which is the first orphan nuclear receptors are discovered through hybridization technique,and includes ERRα,ERRβ and ERRγ.ERRs were completely different from the traditional ligand dependent pathway,which did not combine with estrogen,and did not respond to estrogen.The biological characteristics of ERRs show that without the ligands,ERRs also execute estrogen signaling pathways,and have the same target genes in many organs(breast and skeletal tissues)as estrogen receptors.In recent years,many literatures have reported that the expression level of ERRα and ERRγ could be used as early molecular markers in the diagnosis of breast cancer.In addition,there is increasing evidence that ERRs participates in the regulation of cell energy metabolism,proliferation and apoptosis,which is closely related with the occurrence and development of tumor.ERRβ as a member of the ERRs family,in addition to play an important role in the above physiological activities,but also expression in many tumors,for instance,breast cancer,ovarian cancer,prostate cancer,etc.On the one hand,ERRβ may play a promoter gene function in the development of tumor.In addition,some scholars have reported that the expression of ERRβ in prostate cancer decreased.Therefore,we hypothesized that ERRβ may vary with the type of cancers;the expression of ERRβ is also different.At present,there is no report about the expression and function of ERRβ in gastric cancer.AimsTo detect the expression of ERRβ in gastric cancer,and to investigate the relationship between expression level and clinical parameters and prognosis of patients with gastric cancer;To explore the effects of ERRβ on proliferation of gastric cancer cells and its molecular mechanism,which is important for prevention and treatment of gastric cancer to find the target.Methods1.We collected 109 cases of gastric cancer tissue and 42 cases of adjacent tissue samples,and made the paraffin tissue chip.Then,the expression level of ERRβ in gastric cancer was detected by immunohistochemical method.In addition,10 cases of fresh and fresh gastric cancer were collected,and the mRNA and protein of ERRβ were detected by PCR and WB methods.2.According to the immunohistochemical scoring results,the relationship between the expression levels of ERRβ and clinical parameters in patients with gastric cancer was analyzed.After survival analysis,follow-up data and multivariate regression model were performed to analyze the relationship between the expression of ERRβ and survival time.3.Detection of mRNA and protein levels of ERRβ in different gastric cancer cell lines and gastric mucosal epithelial cell lines.The high expression of ERRβ in gastric cancer cell line was screened out,and the cell model of down regulated ERRβ expression was established by virus transfection.4.MTT method was used to detect the effect of down regulating the expression of ERRβ on the proliferation of XN0422 gastric cancer cell line in vitro.5.Tumorigenicity in nude mice was used to detect the effect of down regulating the expression of ERRβ on the ability of XN0422 gastric cancer cell line in vivo.6.The WB method was used to detect down regulating the expression of ERRβ on self renewal effect in gastric cancer cells.Results1.ERRβ expression in gastric cancer was higher than that in the adjacent issuesImmunohistochemical results showed ERRβ staining was mainly detected in the nucleus,and ERRβ expression increased in gastric cancer(P<0.05).And with the decrease of the degree of differentiation of gastric cancer,the expression intensity of ERRβ was increased.Consistent with the immunohistochemistry result,the mRNA and protein level of ERRβ in the fresh gastric cancer was much higher than that in the control group.2.ERRβ expression is closely related to the depth of invasion,lymph node and differentiation of gastric cancerAccording to the results of ERRβ,the relationship between the expression level of ERRβ and the clinicopathological parameters of gastric cancer was analyzed.The results showed that ERRβ was closely related to the depth of invasion(P<0.05),lymph node metastasis(P<0.05)and differentiation(P<0.05),but not with age,sex,distant metastasis and tumor size(P>0.05).3.ERRβ expression is the independent factors for prognosis in gastric cancer.Survival analysis showed that the survival time of ERRβ positive patients was significantly lower than that of ERRβ negative patients(P<0.05),indicating that the expression of ERRβ is closely related to the prognosis of gastric cancer patients.Univariate and multivariate analysis revealed that ERRβ expression was one of the independent factors influencing the prognosis of gastric cancer.4.ERRβ expression is increased in poorly differentiated gastric cancer cell linesWe detect the RNA and protein expression levels of ERRβ in different gastric cancer cell lines and gastric mucosal epithelial cells(GES-1,AGS)were detected by PCR and WB respectively.The results show that ERRβ in gastric epithelial cells(GES-1,AGS)was low,and ERRβ high differentiation gastric cancer cell lines expression was next,while in the low differentiation of gastric cancer cell lines SGC7901,MGC803,MKN45,KATO-III,XN0422 was high expression level.5.Downregulation of ERRβ reduces the proliferation of gastric cancer cell linesThe MTT assay was used to detect the proliferation of XN0422 gastric cancer cells after changing the expression of ERRβ.Compared with the control group,the proliferation ability of XN0422 gastric cancer cells was significantly decreased after inhibiting the expression of ERRβ.6.Downregulation of ERRβ inhibits the growth of gastric cancer cells in vivoThe results showed that after the down-regulation of ERR expression,the tumor volume and weight of the infected group were significantly decreased than that in the control group,which indicated that the expression of ERRβ could inhibit the tumor growth in vivo.7.Down regulation of ERRβ can inhibit the expression of tumor self-renewal related indicatorsThe results showed that after down regulation of ERRβ expression,the levels of Nanog and Bmi1 were decreased in the interference group compared with the control group.Conclusions1.ERRβ expression was significantly increased in gastric cancer,and correlated with clinicopathological characteristics and prognosis in gastric cancer patients.2.Downregulation of ERRβ inhibits the proliferation of gastric cancer cells,which may be associated with tumor self-renewal. |
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