Mechanisms Of NLRC5 In The Cell Proliferation, Migration And Invasion Of Hepatocellular Carcinoma By Targeting The Wnt/β-catenin Signaling Pathway

Hepatocellular carcinoma(HCC) is the fifth most common malignancy in the world, and its incidence and mortality increased rapidly in recent decades. The relationship between liver cancer and inflammation has been the focus of controversy for a long time. The body has formed a state of chronic inflammation in the long-term stimulation the infectious(hepatitis B virus or hepatitis C virus). It is favorable to promote the occurrence, development and metastasis of hepatocellular carcinoma under this background. Innate immunity was known as the first barrier against infection. It can cause chronic inflammation, and even cause cancer if its response was sustained for too long. Mutations of several NLRs, including NOD1, NOD2, NLRP1, NLRP3, NLRP7 and NLRP12, have been implicated in tumorigenesis. NLRC5(NLR family, CARD domain containing 5) is a member of the NLR protein family. NLRC5 has been reported to regulate immune responses and be associated with chronic inflammatory diseases. However, the biological function of NLRC5 in HCC has not yet been well demonstrated. Therefore, our studies are the first to observe the changes of NLRC5 expression in hepatocellular carcinoma and further explore its effect on the role and the molecular mechanism of hepatocellular carcinoma cell proliferation, migration and invasion.Methods1. Human liver tissues were detected by immunohistochemistry. Normal liver cell line L02 and hepatocellular carcinoma cell line Hep G2, SMMC-7721 and BEL-7402will be detected to research the function of NLRC5.2. Transfecting with p EGFP-C2-NLRC5 plasmid or NLRC5-si RNA was applied to verify the change of NLRC5 expression.3. The role of NLRC5 in hepatocellular carcinoma cell proliferation, migration and invasion capacities were evaluated by using MTT, flow cytometry, wound healing, transwell assay. Western blot analysis and q PCR assay were performed to assess NLRC5 interacting with the activation of Wnt/β-catenin signaling pathway.4. The effect of i CRT3 on migration and proliferation of hepatocellular carcinoma cells were observed. There was also investigated the changes of Wnt/β-catenin signaling pathway related proteins.Results1. We demonstrate that NLRC5 was highly expressed in HCC.2. Overexpression of NLRC5 promoted the proliferation, migration and invasion of HCC cells in vitro and increased the activation of C-myc, cyclin D1 and MMP-7 in HCC cells.3. Knockdown of NLRC5 significantly inhibited cell proliferation, reduced cell migration and invasion, and arrested the cell cycle at G0/G1 phase. Down-regualtion of NLRC5 by si RNA was related to a decrease of the expressions of C-myc, cyclin D1 and MMP-7 at proteins levels in HCC cells.4. Up-regulation of NLRC5 not only positively correlates with the increase of β-catenin but also coordinates the activation of downstream Wnt/β-catenin signaling pathway.ConclusionThe study indicated that NLRC5 may play a crucial role in the proliferation, migration and invasion of HCC cells through Wnt/β-catenin signaling pathway.