| Hepatocellular carcinoma(HCC)is one of the most prevalent cancer diseases in our country,whose occurrence and progress,including recurrence and metastasis,is an interactive process involving multi-genes,multi-steps and multi-factors.With the development of molecular biology,a growing number of critical signal pathways and their members have been clarified.Frizzled2(FZD2),a newly discovered tumor biomarker,was found to be overexpression in many kinds of malignancy.Studies showed FZD2 worsen the prognosis of patients with cancer by regulating a series of tumor biology,and played as a promising therapeutic target for oncotherapy.In the present study,we wanted to identify the oncogenic role of FZD2 in HCC,and its relationship with vasculogenic mimicry(VM)and cancer stem cells(CSCs),providing both a prognostic marker and therapeutic target for combating HCC metastasis.1.Expression of FZD2 in patients with HCC and its prognostic valueTo evaluate the prognostic value of FZD2 in HCC,we employed real-time quantitative PCR and immunohistochemical staining to detect the expression of FZD2 in 100 paired HCC tissues and their adjacent parts separately.We found that both the mRNA level and protein level of FZD2 expression was much greater in tumor tissues than in normal counterparts(P<0.05).When subjected to statistical analysis,expression of FZD2 was considered to be related to tumor size,histological differentiation,micro-embolus,TNM stage,BCLC stage,metastasis and early recurrence(P<0.05)of HCC.Survival analysis showed that overexpression of FZD2 predicted shorter relapse-free time and overall survival time of patients with HCC.Thus,we can expect that FZD2 may be an underlying prognostic marker for HCC.2.Effect of FZD2 on cell proliferation,migration,invasion and epithelial-mesenchymal transition(EMT)of HCCIn order to determined the oncogenic role of FZD2 in HCC,we employed cell models with FZD2 knockdown or overexpression.Cell proliferation,apoptosis,cell cycle,migration,invasion and EMT were examined by CCK-8 assay,plate colony formation assay,flow cytometry,transwell assay and Western blot successively.Afterwards subcutaneous xenotransplanted tumor models in nude mice were established by ejected with FZD2 stable knockdown HCC cell lines.Tumorigenic ability was observed and Ki-67 along with EMT related proteins staining was performed on xenotransplanted tumors.Finally,it turned out that FZD2 might promote cell proliferation,migration,invasion and EMT of HCC cell lines in vitro and in vivo.Also,silence of FZD2 induced HCC cell apoptosis in vitro.3.Effect of FZD2 on vasculogenic mimicry of HCCVM was regarded as a promoter of metastasis and recurrence of HCC,and its relationship with FZD2 was still vague.In present,we stained the HCC tissues xenotransplanted tumor tissues with CD34 and PAS and confirmed that the expression of VM was positively related with FZD2(P<0.05).In 3D culture,overexpression of FZD2 promoted the formation of tube-like structures of the HCC cell lines,while downregulation of FZD2 could inhibit their formation.Therefore,FZD2 was required for VM formation in HCC.4.Effect of FZD2 on cancer stem cells property of HCCCSCs property of HCC was another contributor to cancer metastasis and recurrence.Flow cytometry was utilized to screen the HCC cell lines and we found that knockdown of FZD2 decreased the CD44 positive stem cell subsets,which increased when FZD2 was upregulated.Meanwhile,CSCs transcriptional factors Nanog and SOX2 were found to be down-regulated in HCC cell lines and in mouse xenograft with low expression of FZD2,while overexpression of FZD2 would enhanced such stem cell phenotypes in HCC cell lines.Thus we convinced that FZD2 regulated CSCs property of HCC.Conclusion:FZD2 promotes metastasis and reccurrence of HCC by regulating EMT,VM and CSCs properity,and may be a promising predictor and therapeutic target for patients with HCC. |
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