Induction of Glutathione Transferase in Insulin-Like Growth Factor Type I Receptor-Overexpressed Hepatoma Cells

Abstract

Insulin-like growth factor type I receptor (IGF-IR) is frequently overexpressed in human hepatocellular carcinoma cells (HCC), and this overexpression has been correlated with increased tumor growth. The protective response of HCC to reactive oxygen species (ROS) produced by chemotherapeutic agents is mediated with the induction of phase II detoxifying genes including glutathione transferase (GST). To understand the roles of IGF-IR overexpression in HCC in terms of its detoxifying effect on ROS and conferred resistance to chemotherapy, we analyzed whether IGF-IR overexpressions affect IGF-1-inducible GST expression. GST␣ was induced by exposure to IGF-1 in IGF-IR cells but not in cells expressing normal levels of IGF-IR. Furthermore, IGF-IR-overexpressed HCCs (IR-HCC) are more resistant to doxorubicin than control HCC cells, which was associated with the increased GST induction by IGF-1. Molecular analyses using GSTA2 promoter supported the involvement of xenobiotic response element (XRE) in GST␣ induction. IGF-1 caused the nuclear translocation of CCAAT/ enhancer-binding protein ␤ (C/EBP␤), which might be responsible for XRE activation. In addition, IGF-1 increased the activities of phosphatidylinositol 3-kinase (PI3-kinase) and extracellular signal-regulated kinase in IR-HCCs. Moreover, the inhibition of PI3-kinase completely abolished the nuclear translocation of C/EBP␤ and the up-regulation of GST␣ protein in IR-HCC treated with IGF-1. However, specific inhibitors against extracellular signal-regulated kinase, c-Jun N-terminal kinase, or p38 kinase did not alter IGF-1-inducible GST␣ expression. These results provide evidence that one of the pathological consequences of IGF-IR overexpression in HCCs is the potentiation of GST␣ inducibility by IGF-1. Moreover, this potentiation of GST may be associated with decreased susceptibility to chemotherapeutic agents such as doxorubicin.

Enhancement of vascular endothelial growth factor–mediated angiogenesis in tamoxifen-resistant breast cancer cells: role of Pin1 overexpression

“…The fractionated proteins were then transferred electrophoretically onto nitrocellulose paper and immunoblotted with specific antibodies. Nuclear extracts were prepared as previously described (15).…”

Novel Role of Pin1 Induction in Type II Collagen-Mediated Rheumatoid Arthritis

“…HTB-94 and human chondrocytes stably expressing Pin1 were established using murine stem cell virus (MSCV)-GFP retrovirus system (21). Briefly, Pin1 cDNA was subcloned into MSCV-GFP retroviral vector, and Phoenix cells (a packaging cell line) were transfected with MSCV-GFP (control) or MSCV-Pin1-GFP (Pin1 overexpression) plasmid.…”

“…Pellets containing crude nuclei were resuspended in 100 l of extraction buffer (20 mM HEPES (pH 7.9), 400 mM NaCl, 1 mM EDTA, 1 mM DTT, and 1 mM PMSF), incubated for 30 min on ice, and centrifuged at 15,800 ϫ g for 10 min; the supernatants containing the nuclear extracts were collected and stored at Ϫ80°C until required. SDS-PAGE and immunoblot analyses were performed, as described previously (21). Cell lysates were fractionated by 10% gel electrophoresis, and electrophoretically transferred to nitrocellulose membranes.…”

Identification of a region in the human IRS2 promoter essential for stress induced transcription depending on SP1, NFI binding and ERK activation in HepG2 cells

“…The IRS2 branch of insulin/IGF1 signaling controls both hepatic insulin response as well as pancreatic b-cell growth and function. Moreover, IGF1Rs are frequently overexpressed in human hepatocellular cancer (HCC), and this overexpression has been correlated with increased tumor growth (Lee et al 2007). Inhibition of the IGF1R tyrosine kinase induces growth inhibition, apoptosis, and cell cycle arrest in human HCC cell lines.…”