Hedgehog signaling pathway mediates invasion and metastasis of hepatocellular carcinoma via ERK pathway

Wei Wei,Wei He,Jing-Tao Lu,Wen-Di Zhao

doi:10.1038/aps.2012.24

Abstract

To investigate the role of Hedgehog (Hh) signaling pathway in the invasion and metastasis of human hepatocellular carcinoma (HCC). Eighty six HCC tissues samples and HCC cell line Bel-7402 were examined. The protein expression of sonic hedgehog (Shh), nuclear glioma-associated oncogene-1 (Gli1), MMP-9 and p-ERK1/2 in HCC was analyzed using immunohistochemistry and Western blot analysis. Boyden chamber assay and wound-healing assay were used to quantify the invasion and metastasis of Bel-7402 cells. In 86 HCC tissue samples, the positive ratio of Shh and nucleus Gli1 was 67.44% (58/86) and 60.47% (52/86), respectively; the expression of nucleus Gli1 was correlated with the tumor pathological grade (P=0.034), and with the ability of the tumor to invade and metastasize (P=0.001); the expression of nucleus Gli1 was also correlated with p-ERK1/2 (P=0.031) and with MMP-9 (P=0.034). Neither Shh, nor nucleus Gli1 was observed in normal liver tissue. KAAD-cyclopamine (KAAD-cyc), a specific inhibitor of the Hh pathway, at the concentrations of 1 and 4 μmol/L inhibited the invasion and migration of Bel-7402 cells and decreased the expression of Gli1 in nucleus and MMP-9, p-ERK1/2 proteins in Bel-7402 cells. On the other hand, Shh, a ligand of the Hh pathway, at the concentration of 0.5 μg/mL produced opposite effects. The MAPK pathway inhibitors U0126 and PD98059 at the concentrations of 5 and 10 μmol/L inhibited invasion and metastasis of Bel-7402 cells induced by Shh, and decreased the expression of p-ERK1/2 and MMP-9. However, U0126 and PD98059 had no effect on the expression of Gli1. Hh signaling pathway mediates invasion and metastasis of human HCC by up-regulating the protein expression of MMP-9 via ERK pathway.

Highlights

Hepatocellular carcinoma (HCC) is the common primary cancer with a multifaceted molecular pathogenesis
We investigated the mechanisms of the Hh signaling pathway in invasion and metastasis of HCC, focused on the correlation between the Hh signaling pathway and the MAPK signaling pathway
Overexpression of sonic hedgehog (Shh), glioma-associated oncogene-1 (Gli1), p-ERK1/2, and Matrix metalloproteinases (MMPs)-9 in HCC liver tissues with invasion and metastasis compared with nonmetastasis HCC liver tissue We detected Shh, Gli1 MMP-9, and p-ERK1/2 expressions in 86 cases of HCC liver tissues with or without invasion and metastasis by IHC staining

Summary

Introduction

Hepatocellular carcinoma (HCC) is the common primary cancer with a multifaceted molecular pathogenesis. Many signaling pathways are thought to be involved in the development and invasion of HCC, including the MAPK pathway[6, 7], Hh signaling pathway is a highly conserved system, which plays a crucial role in tissue patterning, cell differentiation and proliferation[14]. Hedgehog, including sonic hedgehog (Shh), Indian hedgehog (Ihh), and desert hedgehog (Dhh), appear to bind to a transmembrane receptor protein, known as Patched (Ptc), which in the absence of Shh exerts an inhibitory effect on the seven transmembrane receptor smoothened (Smo). Binding of Shh to Ptc alleviates the inhibitory effect of Ptc on Smo. Once activated, Smo induces a complex series of intracellular reactions that targets the glioma-associated oncogenes (Gli) transcription factor families, the zincfinger transcription effectors[15, 16].

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