The Study Of Predicting Markers And Associated Molecular Mechanisms In Hepatocellular Carcinoma Bone Metastasis

Purpose:The aim of this study was to evaluate the correlation between CTGF and IL-11mRNA expression in HCC, and investigated its relationship to bone metastasis.Methods:The expression of connective tissue growth factor (CTGF) and interleukin-11(IL-11) in RNA extracted from127formalin-fixed, paraffin-embedded HCC specimens was examined by quantitative real-time polymerase chain reaction. Cox regression analysis of potential biomarkers for BM was performed between clinicopathological features and CTGF and IL-11expression.Results:Expression of CTGF was detected in all specimens, and the overall positive rate for IL-11expression was24.4%(31/127). Mean CTGF expression in the BM samples was3.63-times higher than in samples from non-metastatic patients. Multivariate Cox proportional hazards analyses revealed that significant independent prognostic factors for development of BM in HCC patients were tumor differentiation (p=0.043), vascular invasion (p=0.027), tumor-node-metastasis stage (p=0.005), intratumoral CTGF expression (p=0.010), and intratumoral IL-11expression (p=0.031).Conclusion:Expression levels of intratumoral CTGF or IL-11were independent prognostic factors for development of BM in HCC patients. Our findings demonstrated that intratumoral CTGF expression combined with IL-11may be useful in predicting the development of BM in HCC. Purpose:HCC specimens have been confirmed that CTGF and IL-11expression are closed related with development of BM in HCC patients. Here, we further study the expression of these two cytokines in HCC cell lines and analyse the molecular and cellular mechanisms underlying HCC with BM.Methods:CTGF and IL-11expression was measured in stepwise metastatic HCC cell lines. The effect of human recombinant CTGF and IL-11on migratory ability of the human HCC cell line was also analysed. CTGF and IL-11knock-down siRNA cell lines were also established to detect the underlying signal pathway.Results:The expression of CTGF and IL-11mRNA was detected, at various levels, in all five cell lines. Strong expression of CTGF and IL-11was observed in the highly metastatic HCC cell lines (MHCC97L, MHCC97H, and MHCCLM3) relative to that seen in weakly metastatic cell lines (HuH7and7721). Treatment with exogenous rhCTGF and rhIL-11enhances the migratory ability of human HCC LM3cells. CTGF could enhance cell migration by activating NF-κB, while rhIL-11increase the STAT3phosphorylation.Conclusion:CTGF and IL-11high expression indicated highly invasive and metastatic ability of HCC. CTGF and IL-11can stimulate different signaling and thus contribute to HCC metastasis. Purpose:Hypoxic microenvironments favor tumor growth and metastasis development, and affect many aspects of the biology of tumors and their responses to therapy. CTGF and IL-11are closely related with HCC metastasis. Here, we further study the correlation between hypoxia and expression of CTGF and IL-11.Methods:HCC97L cellular hypoxic model was established in vitro, conditioned medium (CM) was collected at different time course. The expressions of CTGF、HIF-1α、VEGF and OPG were examined by quantitative real-time polymerase chain reaction (PCR) and the release of these cytokines were also analysed by enzyme-linked immunosorbent assay (ELISA). rhCTGF、rhIL-11and CTGF siRNA were also established to detect the interaction between different molecules under hypoxia.Results:Hypoxic stimulation of HCC97L cells increased the level of CTGF mRNA by2.80-fold within1.5h, and hypoxia-inducible factor-1α mRNA levels in these cells could be increased by stimulation with recombinant CTGF protein. Furthermore, OPG and matrix metalloproteinase-2and-9levels were also induced under hypoxic conditions. Meanwhile, knockdown of CTGF inhibit production of VEGF and nuclear translocation of HIF-1αConclusion:Tumor hypoxia enhanced the expression of CTGF, which initiates the invasive angiogenesis cascade and enhances expression of many hypoxia-associated genes. Cellular release of OPG may play a role in tumor cell survival. The hypoxia-induced cascade in HCC cells may contribute to invasion and metastasis in vivo. Purpose:The interaction between parenchymal and stromal cells is important for tumor formation and development. Recently, it has been showed that activated hepatic stellate cells (HSCs) are the crucial factor responsible for liver fibrosis and involved in progrssion of hepatocellular carcinoma (HCC) by interaction with tumour cells. We hypothesized that disruption of the interaction of HCC cells with HSCs would lead to their sensitization to therapeutic agents such as Sorafenib or5-FU. Here, we investigate the interaction between HS cells (LX-2) and HCC cells (HuH7) in vitro.Methods:The expressions of CXCR4in different HCC cells were examined. The effects of HCC conditioned medium (HCC-CM) on activation and proliferation of HS cells (LX-2), the influences of HS cell CM (HS-CM) on proliferation and invasion of HCC cells (HuH7), and the effects of CXCR4inhibitor AMD3100on their interaction were assessed by using CM co-culture system. The effect of Lx-2on HCC apoptosis was also analysed using TUNEL detection method.Results:Tumour-CM could promote the proliferation and activation of LX-2cells, while HS-CM increased HCC cell proliferation, invasion and resistance to sorafenib. AMD3100induces disruption of the interaction of HCC cells with HSC and enhanced the ratio of apoptosis in the presence of HS-CM, and significantly inhibited the HCC cell migration in response to HS-CM.Conclusion:This study indicates that HCC cells can promote the activation of HSCs and stromal HSC promotes HCC progression. By using agents such as AMD3100that disrupt interaction with the microenvironment may enhance the efficacy of cytotoxic agents in cancer therapy and lead to more effective therapeutic approaches.