| Background:Gastric cancer(GC)is one of the most common malignant tumors in China,its incidence in our country ranks second and mortality ranks third in all kinds of tumors.Due to its low early diagnosis rate,low degree of differentiation and high metastasis rate,the 5-year survival rate of GC has not improved much in recent years compared with other tumors such as lung cancer and breast cancer.Therefore,GC is still the main tumor that endangers human health.HEF1(Human Enhancer of Filamentation 1)is a member of the CAS(Crk-associated substrate)family of scaffolding proteins.HEF1 is similar to CAS in physiological mechanisms,suggesting that HEF1 may play roles in differentiation.At present,reports on the molecular mechanisms of HEF1 in GC differentiation are limited.Thus,this study mainly explores HEF1 expression in GC tissues or cells,prognostic value of HEF1 and the molecular mechanism of its regulating GC cell differentiation.Methods:In this study,we collected 108 GC patients’ information and tissue specimens,according to the immunohistochemical staining,the univariate and multivariate analysis was performed by Cox proportional hazards model and the survival curves were drawn by Kaplan-Meier methods to investigate the relationship between HEF1 expression and clinical pathological parameters and prognosis of GC patients.To analyze the relationship between HEF1 and GC cell differentiation,Real-time PCR(qRT-PCR) was used to detect the expression changes of GC differentiation markers(CD133 and E-cadherin).HEF1 expression in GC cells was analyzed by Western blot.QRT-PCR was used to screen the related Wnt ligands in Wnt signaling pathway.Enzyme linked immunosorbent assay(Elisa)was used to analyze the Wnt ligand expression to explore the relationship between HEF1 and the Wnt ligand.To determine the relationship between HEF1 and nuclear translocation of β-catenin levels,Western blot was used to analyze the expression changes of β-catenin.To explore the relationship between HEF1 and Wnt signaling pathway,immunofluorescence assay was used to detect the expression changes of major factors Wnt5a,β-catenin,c-myc and Cyclin D1 in Wnt signaling pathway.QRT-PCR was used to detect the expression changes of GC differentiation markers(CD133 and E-cadherin)to explore the relationship between Wnt signaling pathway and GC cell differentiation.Results:Compared with normal adjacent gastric tissues,HEF1 expression was higher in GC tissues.HEF1 expression was weakly in well differentiated GC tissues,while was strongly in poor differentiated GC tissues.Among all clinical pathological parameters,tumor differentiation was significantly correlated with HEF1 expression(P<0.001).However,no significant differences were observed between HEF1 expression and other clinical pathological parameters.GC patients with HEF1 high expression had a higher risk of death(HR:2.431,95%CI:1.273-4.640,P=0.007;HR:2.052,95%CI:1.026-4.103,P=0.042).GC patients with HEF1 high expression showed significantly poorer disease-free and overall survival than GC patients with HEF1 low expression(median disease-free survival time:37.6and 63.7 months,Log Rank χ~2=8.89,P<0.01;median overall survival time:43.8 and 65.3 months,Log Rank χ~2=14.57,P<0.01).HEF1 expression was higher in poor differentiated GC cells than in well differentiated GC cells(P<0.05),HEF1 expression related to GC cell differentiation and HEF1 regulated GC cell differentiation.Upregulation of HEF1 increased Wnt5a expression and the nuclear translocation of β-catenin(P<0.05),thereby resulting in the expression changes of downstream genes in the Wnt signaling pathway(c-myc and Cyclin D1).Besides,Wnt signaling pathway regulated GC cell differentiation(P<0.05).Conclusions:Therefore,our findings suggested that HEF1 expression was higher in poor differentiated GC tissues and cells.HEF1 expression related to prognosis of GC patients,and HEF1 may be an independent prognostic factor for GC patients.Notably,HEF1 may regulate GC cell differentiation through the Wnt5a/β-catenin signaling pathway. |
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