transforming-growth-factor-beta and Hepatoblastoma

The β-catenin mutation is frequently observed in hepatoblastoma (HB), but the underlying mechanism by which Wnt/β-catenin signaling induces HB tumor formation is unknown. Here we show that expression of growth regulation by estrogen in breast cancer 1 (GREB1) depends on Wnt/β-catenin signaling in HB patients. GREB1 is localized to the nucleus where it binds Smad2/3 in a competitive manner with p300 and inhibits TGFβ signaling, thereby promoting HepG2 HB cell proliferation. Forced expression of β-catenin, YAP, and c-Met induces HB-like mouse liver tumor (BYM mice), with an increase in GREB1 expression and HB markers. Depletion of GREB1 strongly suppresses marker gene expression and HB-like liver tumorigenesis, and instead enhances TGFβ signaling in BYM mice. Furthermore, antisense oligonucleotides for GREB1 suppress the formation of HepG2 cell-induced tumors and HB-like tumors in vivo. We propose that GREB1 is a target molecule of Wnt/β-catenin signaling and required for HB progression.

Topics: Adolescent; Animals; Antineoplastic Agents; beta Catenin; Carcinogenesis; Cell Line, Tumor; Cell Proliferation; Child; Child, Preschool; Gene Expression Regulation, Neoplastic; Hepatoblastoma; Humans; Infant; Infant, Newborn; Liver Neoplasms; Male; Mice; Mice, Nude; Molecular Targeted Therapy; Neoplasm Proteins; Neoplasm Transplantation; Oligonucleotides, Antisense; Transforming Growth Factor beta; Wnt Signaling Pathway

This study analyzed gene expression patterns and global genomic alterations in hepatocellular carcinomas (HCC), hepatoblastomas (HPBL), tissue adjacent to HCC and normal liver tissue derived from normal livers and hepatic resections. We found that HCC and adjacent non-neoplastic cirrhotic tissue have considerable overlap in gene expression patterns compared to normal liver. Several genes including Glypican 3, spondin-2, PEG10, EDIL3 and Osteopontin are over-expressed in HCC vs. adjacent tissue whereas Ficolin 3 is the most consistently under-expressed gene. HCC can be subdivided into three clusters based on gene expression patterns. HCC and HPBL have clearly different patterns of gene expression, with genes IGF2, Fibronectin, DLK1, TGFb1, MALAT1 and MIG6 being over-expressed in HPBL versus HCC. In addition, specific areas of the genome appear unstable in HCC, with the same regions undergoing either deletion or increased gene dosage in all HCC. In conclusion, a set of specific genes and areas of genomic instability are found across the board in liver neoplasia.

Topics: Adaptor Proteins, Signal Transducing; Apoptosis Regulatory Proteins; Calcium-Binding Proteins; Carcinoma, Hepatocellular; Carrier Proteins; Cell Adhesion Molecules; Cluster Analysis; DNA-Binding Proteins; Fibronectins; Fibrosis; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genome; Glycoproteins; Glypicans; Heparan Sulfate Proteoglycans; Hepatoblastoma; Humans; Insulin-Like Growth Factor II; Intercellular Signaling Peptides and Proteins; Lectins; Liver Neoplasms; Membrane Proteins; Osteopontin; Proteins; Repressor Proteins; RNA-Binding Proteins; Sialoglycoproteins; Transforming Growth Factor beta; Transforming Growth Factor beta1

We previously showed a positive correlation between nephromegaly and plasma hepatocyte growth factor (HGF)/transforming growth factor beta1 (TGF-beta1) ratio in children with biliary atresia. The purpose of this study is to examine the possible reversibility of nephromegaly in patients with biliary atresia.. We evaluated kidney volume in 13 patients with biliary atresia before and after liver transplantation, 6 patients with hepatoblastoma, and 26 healthy children. Plasma HGF and TGF-beta1 levels were determined for all children.. We noted significant nephromegaly in children with biliary atresia before liver transplantation compared with healthy children and children after liver transplantation (P < 0.001 and P = 0.006 for intercepts, P = 0.064 and P = 0.753 for slopes by analysis of covariance, respectively). The highest plasma HGF levels and HGF/TGF-beta1 ratios and the lowest TGF-beta1 concentrations were found in children with biliary atresia before liver transplantation (P < 0.001). No statistically significant nephromegaly was observed in children with biliary atresia after liver transplantation or those with hepatoblastoma despite the presence of a mildly increased plasma HGF level and HGF/TGF-beta1 ratio. Plasma HGF/TGF-beta1 ratio correlated positively with degree of nephromegaly in all patients (r = 0.717; P < 0.001).. Our data suggest that liver transplantation reverses the nephromegaly present in children with biliary atresia and that plasma HGF/TGF-beta1 ratio may be associated with the development of nephromegaly in patients with biliary atresia.

Topics: Biliary Atresia; Biomarkers; Child; Child, Preschool; Female; Hepatoblastoma; Hepatocyte Growth Factor; Humans; Infant; Kidney; Liver Neoplasms; Liver Transplantation; Male; Organ Size; Postoperative Period; Transforming Growth Factor beta; Transforming Growth Factor beta1; Ultrasonography

Apolipoprotein M (apoM) is a novel apolipoprotein presented mostly in high-density lipoprotein (HDL) in human plasma, and is exclusively expressed in liver and in kidney. The pathophysiological function of apoM has not yet been elucidated. Apolipoprotein B (apoB), the characteristic apolipoprotein of low-density lipoprotein (LDL), is like apoM, a very hydrophobic protein, and thereafter they both must co-circulate with lipoprotein particles in plasma. The cytokine, transforming growth factor-beta (TGF-beta), has been shown to decreased apoB secretion in HepG2 cells, and we hypothesized that TGF-beta may have the same effects on apoM expression in HepG2 cells. In the present study, we used real-time RT-PCR to analyze apoM and apoB mRNA levels during administration of TGF-beta, as well as TGF-alpha, epidermal growth factor (EGF) and hepatic growth factor (HGF). TGF-beta significantly inhibited both apoM and apoB mRNA expression in HepG2 cells. The inhibitory effects of TGF-beta were dose-dependent, i.e. 1 ng/ml of TGF-beta decreased apoM mRNA levels by 30%, and 10 or 100 ng/ml of TGF-beta decreased apoM mRNA levels more than 65%. The effect of TGF-beta on apoB mRNA expression was slightly weaker than that of apoM, with a maximum effect at 10 or 100 ng/ml TGF-beta where apoB mRNA levels decreased about 55%. The inhibitory effects of TGF-beta on apoM and apoB mRNA levels also increased with increasing incubation time, where the maximum effect was obtained at 24 h. Moreover TGF-alpha, EGF and HGF all decreased both apoM and apoB mRNA levels, but to a less extent than TGF-beta. Further, all four cytokines had more pronounced effects on apoM mRNA expression than apoB mRNA expression. The present study suggested that apoM, like apoB, may be involved in the hepatic lipoprotein assembly in vivo.

Topics: Apolipoproteins; Apolipoproteins B; Apolipoproteins M; Down-Regulation; Epidermal Growth Factor; Gene Expression Regulation; Hepatoblastoma; Hepatocyte Growth Factor; Humans; Lipocalins; Liver Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor alpha; Transforming Growth Factor beta; Tumor Cells, Cultured

BAMBI is a transmembrane glycoprotein related to the transforming growth factor-beta (TGF-beta)-family type I receptors and functions as a negative regulator of TGF-beta signaling during development. BAMBI is induced by BMP signaling through the evolutionary conserved BMP-responsive elements in its promoter. Furthermore, we have recently shown that Wnt/beta-catenin signaling activates transcription of BAMBI and that BAMBI expression is aberrantly elevated in most colorectal carcinomas. Here, we show that BAMBI is also directly induced by TGF-beta signaling, through the three tandem repeats of 13 bp sequences containing the SMAD-binding elements, which are distinct from the BMP-responsive element. Our findings suggest that BAMBI transcription is regulated by TGF-beta signaling through direct binding of SMAD3 and SMAD4 to the BAMBI promoter.

Topics: Cell Line, Tumor; DNA-Binding Proteins; Genes, Regulator; Hepatoblastoma; Humans; Liver Neoplasms; Membrane Proteins; Nerve Growth Factors; Promoter Regions, Genetic; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad Proteins; Smad3 Protein; Smad4 Protein; Trans-Activators; Transcriptional Activation; Transforming Growth Factor beta; Xenopus Proteins

Hepatitis C virus (HCV) is a leading cause of chronic liver disease (CLD) worldwide. The chronicity is a result of viral persistence and the ability of the virus to escape from the immune mechanisms of the host. Transforming growth factor (TGF)-beta is a cytokine thought to be responsible for viral persistence and liver fibrogenesis.. The present study examined the levels of TGF-beta messenger (m)RNA by reverse transcription polymerase chain reaction (RT-PCR) in 35 liver biopsies and HCV-transfected HepG2 cells.. Transforming growth factor-beta mRNA was detected in nine liver biopsies from patients with chronic HCV infection, but was not detected in patients with non-HCV-related CLD or controls. On quantitation by semiquantitative PCR, TGF-beta mRNA levels ranged from 10-4.75 to 10-12.8 amol (10-7.46 +/- 3.771) in liver biopsies of HCV-related CLD. No significant difference in TGF-beta receptor levels was observed by RT-PCR in HCV- or non-HCV-related CLD by immunohistochemistry. To correlate these findings with in vitro experiments, levels of TGF-beta mRNA and its receptors were determined by RT-PCR in HepG2 cells transfected with HCV and hepatitis B virus (HBV) constructs, using mock-transfected cells as control. The TGF-beta protein levels were quantitated in these cell supernatants by enzyme immunoassay. The TGF-beta mRNA and protein levels were two logs and approximately 30 times higher in HCV-transfected HepG2 cells than in HBV- and mock-transfected cells, respectively. The TGF-beta receptors in HepG2 cells were also downregulated in HCV-transfected cells as compared with mock-transfected cells.. These observations suggest upregulation of TGF-beta in HCV infection and a probable role for TGF-beta in the pathogenesis of HCV-related CLD.

Topics: Adult; Aged; Chronic Disease; Female; Hepacivirus; Hepatitis C, Chronic; Hepatoblastoma; Humans; In Vitro Techniques; Liver Neoplasms; Male; Middle Aged; Receptors, Transforming Growth Factor beta; Reverse Transcriptase Polymerase Chain Reaction; Risk Factors; RNA, Messenger; Transforming Growth Factor beta; Tumor Cells, Cultured

To study the inhibitory effect of Kangxian Recipe (KXR) on TGF-beta 1 induced hepatocyte apoptosis.. The in vitro model of hepatocyte apoptosis was established by cell biologic methods, utilizing the characteristics of TGF-beta 1 to observe the inhibitory effect of KXR on hepatocyte apoptosis.. TGF-beta 1 induced apoptosis of hepatocyte in a dose-dependent manner. The apoptosis rate of 2.2.15 cells was 63% when 500 ng/L TGF-beta 1 was used, while for HepG2 cell, it was merely 44%. After treatment of 20 ng/L KXR, the apoptosis rate of the two kinds of cell lines lowered to 33% and 24% respectively. The inhibition rate of both groups was about 50%.. KXR had strong inhibitory effect on hepatocyte apoptosis induced by TGF-beta 1.

Topics: Apoptosis; Drug Combinations; Drugs, Chinese Herbal; Hepatitis B; Hepatoblastoma; Hepatocytes; Humans; Liver Neoplasms; Transforming Growth Factor beta; Tumor Cells, Cultured

Furin is recognized as being one of the main convertases of the cellular constitutive secretion pathway but the mechanisms regulating its expression are still unknown. We have previously demonstrated that TGFbeta1 up-regulates its own converting enzyme, furin, creating a novel activation/regulation cycle of potential importance in a variety of physiological and pathophysiological conditions. The fur (fes upstream region) gene is regulated via three alternative promoters; P1, P1A, and P1B. To gain insight into the molecular mechanism(s) underlying this up-regulation, we performed transient cell transfections with P1, P1A, and P1B promoter luciferase constructs. Transfection experiments in HepG2 cells revealed that fur P1 promoter is the strongest and the most sensitive to TGFbeta1 stimulation (5 ng/ml) (3.2-fold vs. 2.4-fold for P1A and 2.1-fold for P1B). Cotransfection with either a dominant negative mutant form of Smad2 [Smad2(3SA)] or a known Smad inhibitor [Smad7] inhibit constitutive and TGFbeta1-induced luciferase activity indicating the participation of endogenous Smads. Increased levels of TGFbeta1-induced transcriptional activation of the P1 promoter by overexpression of Smad2 and/or Smad4 is greatly reduced in the presence of Smad2(3SA) and completely inhibited by Smad7, suggesting the participation of endogenous Smad2/Smad4 complexes. Furthermore, the fork-head activin signal transducer (FAST-1), known to interact with Smad2/Smad4 complexes, is a potent stimulator of TGFbeta1-induced transactivation of the fur P1 promoter. Five prime-deletion analysis of this promoter identified the proximal region (between positions -8734 and -7925), as the nucleotide stretch that carries most of the transcriptional activation of fur P1 promoter by Smad2. Overall, the present data demonstrate that Smad2 and Smad4 possibly in complex with FAST-1 or other DNA binding partners participate in the constitutive and inducible transactivation of the fur P1 promoter. This represents the first detailed study of the transcriptional regulation of the fur gene.

Topics: Blotting, Northern; DNA-Binding Proteins; Furin; Gene Expression Regulation, Enzymologic; Hepatoblastoma; Humans; Liver Neoplasms; Luciferases; Promoter Regions, Genetic; RNA, Messenger; Smad2 Protein; Smad4 Protein; Smad7 Protein; Subtilisins; Trans-Activators; Transcription, Genetic; Transfection; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Cells, Cultured

We examined the effects of various peroxisome proliferators (PPs) such as the hypolipidaemic agents clofibric acid (CLO), bezafibrate (BEZA), ciprofibrate (CIPRO) and nafenopin (NAFE) and the plasticizer di-(2-ethylhexyl)phthalate (DEHP) on peroxisomal enzyme activities, apoptosis and DNA synthesis in rat FaO and human HepG2 hepatoma cell lines. Both growing and confluent cultures were treated with PPs (250 microM) for 48 or 72 h. In accordance with our previous observations in PP-treated primary hepatocyte cultures of rat and human origin, the various PPs increased peroxisomal enzyme activities in rat FaO cells but not in human HepG2 cells. PPs strongly induced apoptosis in FaO cells. They did not affect TGFbeta-induced apoptosis, with the exception of DEHP and NAFE, respectively blocking and increasing induced apoptosis in confluent cultures. Moreover, PPs produced a minor, but significant, decrease in DNA synthesis in FaO cells. PPs also decreased DNA synthesis in growing HepG2 cells, and CLO, CIPRO and NAFE induced apoptosis in confluent HepG2 cultures. This is in opposition with the effects of PPs on primary hepatocyte cultures, i.e. inhibition of both spontaneous and TGFbeta-induced apoptosis and increases in DNA synthesis in rat hepatocytes, and unchanged mitosis-apoptosis balance in human hepatocytes.

Topics: Acyl-CoA Oxidase; Animals; Apoptosis; Carcinoma, Hepatocellular; Carnitine O-Acetyltransferase; Cell Division; Cell Nucleus; DNA; DNA Replication; Hepatoblastoma; Liver; Liver Neoplasms; Oxidoreductases; Peroxisome Proliferators; Peroxisomes; Rats; Transforming Growth Factor alpha; Transforming Growth Factor beta; Tumor Cells, Cultured

Hepatoblastoma is a rare pediatric liver tumor. While much progress has been made in the treatment of the disease, very little is known about the moleculer events underlying the pathogenesis of this disease. We sought to investigate a series of hepatoblastomas for alterations in gene expression patterns with emphasis on important cell regulatory genes, including chromatin modifying enzymes, cyclin dependent kinase inhibitors, growth factors, oncogenes and cell cycle regulators. Total RNA was extracted from a series of sporadic hepatoblastomas with matched normal liver, some unmatched tumors and fetal livers, and gene expression was measured for various genes using RNase Protection Analysis (RPA). The results of this analysis show that the expression of many important regulatory genes are distinctly altered in these tumors, and a subset of tumors can be distinguished on the basis of these gene expression differences and histopathological features. Because the molecular events underlying the pathogenesis of this rare tumor are so poorly understood, this study represents a first step in determining some of the possible mechanisms involved which may provide future avenues of research.

Topics: Case-Control Studies; Cell Cycle Proteins; Child, Preschool; Cyclin-Dependent Kinases; Enzyme Inhibitors; Female; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Genes, p53; Genes, Retinoblastoma; Hepatoblastoma; Histone Deacetylases; Humans; Infant; Liver; Liver Neoplasms; Male; Proto-Oncogenes; Reference Values; RNA Probes; Transcription Factors; Transforming Growth Factor beta