ABSTRACT
BACKGROUND: Oncogenic insulin-like growth factor-II (IGF-II) is overexpressed in hepatocellular carcinoma (HCC). The present study aimed to analyze the dynamic alteration of IGF-II CpG site methylation status and its molecular mechanism in HCC progression. METHODS: IGF-II alterations were observed in rat hepatocarcinogenesis models induced by 2-acetylaminofluorene. Liver IGF-II expression was compared by immunohistochemistry or tissue IGF-II specific concentration (nmol/mg protein). Status of human IGF-II promoter 3 (P3) or rat IGF-II P2 CpG site methylation was amplified by methylation-specific polymerase chain reaction (MSP). Serum IGF-II levels were quantitatively detected by an enzyme-linked immunosorbent assay. RESULTS: The levels of hepatic IGF-II expression were significantly elevated in the HCC group (P < 0.001). The unmethylation rate of IGF-II P3 CpG sites was 100% in the HCC-, 52.5% in the paracancerous-, and none (0%) in the distal noncancerous-tissues. Abnormal IGF-II expression was related to differentiation degree, tumor invasion, and positive HBV-DNA (all P < 0.001), with a negative correlation between P3 methylation degree and IGF-II expression. There was a positive correlation between liver IGF-II specific concentration and circulating IGF-II level (râ¯=â¯0.97, P < 0.001). Significantly negative correlation was found between IGF-II P2 CpG site methylation and circulating IGF-II (rs = -0.89, P < 0.001) or liver IGF-II level (rs = -0.84, P < 0.001). CONCLUSIONS: The increase of serum IGF-II and the alteration of oncogenic gene IGF-II methylation may be biomarkers for HCC diagnosis and DNA methylation may be the therapeutic target of HCC.
Subject(s)
Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Cell Transformation, Neoplastic/pathology , Insulin-Like Growth Factor II/genetics , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Adult , Analysis of Variance , Animals , Biopsy, Needle , Cell Transformation, Neoplastic/genetics , Cells, Cultured , Chi-Square Distribution , Cohort Studies , DNA Methylation , Disease Models, Animal , Gene Expression Regulation, Neoplastic , Hepatocytes/pathology , Humans , Immunohistochemistry , Liver Cirrhosis/pathology , Liver Cirrhosis/physiopathology , Middle Aged , Random Allocation , Rats , Rats, Sprague-DawleyABSTRACT
OBJECTIVE: To investigate the dynamic expression of hypoxia inducible factor-1alpha (HIF-1alpha) and its clinical values in hepatocellular carcinoma (HCC). METHODS: The dynamic changes of liver pathology, HIF-1alpha transcription and expression were observed through the hepatoma model. The self-control specimens from 35 human HCC patients were collected and the expression, cellular distribution, and clinicopathological features of HIF-1alpha and its gene was analyzed by immunohistochemistry, western blotting and nested- PCR, respectively. RESULTS: Both levels of hepatic HIF-1alpha and HIF-1alpha mRNA expression increased during the HCC development course. The incidence of HIF-1alpha and the ratio of HIF-1alpha to beta-actin was 0% and 0.16+/-0.02 in the control rats, 77.8% and 0.29+/-0.04 in the denatured rats, 88.9% and 0.52+/-0.03 in the precancerous rats, and 100% and 0.84+/-0.02 in the cancerous rats respectively, with significant difference between the control group and any of the experimental groups (P = 0.000). The positive HIF-1alpha was brown and granule-like and mainly presented in cytoplasm and few in nucleus. The incidence of HIF-1alpha was 80% (28/35) in HCC and 100% (35/35) in its surrounding tissues. The clinical pathological features indicated HIF-1alpha expression associated with tumor size and differentiation degree the of HCC. No correlation was found between HIF-1alpha and tumor numbers or positive-HBsAg. CONCLUSIONS: HIF-1alpha expression is associated with occurrence and development of HCC, and is perhaps a target molecule for HCC therapy.
Subject(s)
Carcinoma, Hepatocellular/metabolism , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Liver Neoplasms/metabolism , Adult , Aged , Animals , Carcinoma, Hepatocellular/pathology , Female , Humans , Liver/metabolism , Liver/pathology , Liver Neoplasms/pathology , Male , Middle Aged , RNA, Messenger/genetics , Rats , Rats, Sprague-DawleyABSTRACT
BACKGROUND: Hepatocellular carcinoma (HCC) is characterized by multicause, obvious multistage and multifocal processes of tumor progression. The development of HCC is related intimately to overexpression and signal transduction of many cellular factors. This study was undertaken to investigate the dynamic expression and alteration of heat shock protein (HSP) gp96 along with its gene during HCC development. METHODS: A rat model of hepatoma induced with 2-fluorenylacetamide (2-FAA, 0.05%) was established in male Sprague-Dawley rats. Total RNA and pathological changes were observed during hepatocarcinogenesis. Total RNAs were transcribed into cDNA by reverse transcription and the gene fragment of gp96 was amplified by nested RT-PCR. The gp96 expression in rat liver tissues was semi-quantitatively analyzed by immunohistochemistry. RESULTS: Histological examination suggested that hepatocytes in rats fed with 2-FAA showed vacuole-like denaturation at the early stages, then dysplastic nodules appeared at the middle stage, and finally progressed to tubercles of cancerous nests. A tendency of increasing liver gp96 protein level was found from normal liver to precancerous to cancerous tissues during hepatoma development (P<0.01), and was in accordance with the changes in gp96 mRNA (P<0.05). CONCLUSION: HSP gp96 is involved in HCC development and its overexpression may be a useful marker for early diagnosis.
