thioacetamide and Carcinoma--Hepatocellular

Liver fibrosis is a worldwide public health problem due to its life-threatening complications, including portal hypertension, liver failure, cirrhosis, and hepatocellular carcinoma (HCC). Liver fibrosis is the net result of a complex excessive accumulation of extracellular matrix (ECM). Activation of hepatic stellate cells (HSCs) are the cause of deposition of ECM and are commonly recognized as a key step in liver fibrosis. The aim of this study was to investigate the effect of foreskin-derived mesenchymal stem cells treated with boron compounds on liver fibrosis. Rats were injected intraperitoneally with thioacetamide (TAA) at a dose of 150 mg/kg except sham and control groups' rats. Thioacetamide (TAA), foreskin-derived mesenchymal stem cells (TAA + FSDMSC), FSDMSC treated with boric acid (TAA + FSDMSC + BA), FSDMSC treated with sodium pentaborate pentahydrate (TAA + FSDMSC + NaB), control and sham groups were studied. Boron compound treated foreskin-derived mesenchymal stem cells were injected into the tail vein, and evaluations were conducted after 4 weeks and liver tissues were obtained for structural, immunohistochemical, and western blot studies and blood samples were taken for biochemical analysis. FSDMSC (BA) alleviates TAA-induced rats liver fibrosis, and BA showed a positive effect on foreskin-derived mesenchymal stem cells viability. After using BA-treated mesenchymal stem cells, we observed that there was regression in the fibrotic areas at TAA-induced liver fibrosis. The result demonstrates that the contribution of TAA + FSDMSC and TAA + FSDMSC (NaB) at the level of structure is not effective in regression of fibrosis in TAA-generated liver fibrosis. We concluded that FSDMSC treated with BA may be a factor in the regression of fibrosis.

Topics: Animals; Carcinoma, Hepatocellular; Fibrosis; Foreskin; Liver; Liver Cirrhosis; Liver Neoplasms; Male; Mesenchymal Stem Cells; Rats; Thioacetamide

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death, so we should be concerned and look for effective/less-harmful treatments than chemotherapeutics already clinically in application. Aspirin works well ''in conjunction'' with other therapies for HCC since aspirin can boost the sensitivity of anti-cancer activity. Vitamin C also was shown to have antitumor effects. In this study, we examined the anti-HCC activities of synergistic combination (aspirin and vitamin C) vs. doxorubicin on HCC-bearing rats and hepatocellular carcinoma (HepG-2) cells.. Based on our results, aspirin plus vitamin C can be considered reliable, accessible, and efficient synergistic anti-HCC medication.

Topics: Animals; Ascorbic Acid; Aspirin; Carcinoma, Hepatocellular; Doxorubicin; Humans; Liver Neoplasms; Rats; Thioacetamide; Tumor Suppressor Protein p53; Vitamins

Sorafenib (Sora) represents one of the few effective drugs for the treatment of advanced hepatocellular carcinoma (HCC), while resistance and cardiotoxicity limit its therapeutic efficacy. This study investigated the effect of transient receptor potential melastatin 7 (TRPM7) inhibitor, carvacrol (CARV), on overcoming Sora resistance and cardiotoxicity in thioacetamide (TAA) induced HCC in rats.. TAA (200 mg/kg/twice weekly, intraperitoneal) was administered for 16 weeks to induce HCC. Rats were treated with Sora (10 mg/Kg/day; orally) and CARV (15 mg/kg/day; orally) alone or in combination, for six weeks after HCC induction. Liver and heart functions, antioxidant capacity, and histopathology were performed. Apoptosis, proliferation, angiogenesis, metastasis, and drug resistance were assessed by quantitative real time polymerase chain reaction, enzyme-linked immunosorbent assay, and immunohistochemistry.. CARV/Sora combination significantly improved survival rate, and liver functions, reduced Alpha-Fetoprotein level, and attenuated HCC progression compared with Sora group. CARV coadministration almost obviated Sora-induced changes in cardiac and hepatic tissues. The CARV/Sora combination suppressed drug resistance and stemness by downregulating ATP-binding cassette subfamily G member 2, NOTCH1, Spalt like transcription factor 4, and CD133. CARV boosted Sora antiproliferative and apoptotic activities by decreasing cyclin D1 and B-cell leukemia/lymphoma 2 and increasing BCL2-Associated X and caspase-3.. CARV/Sora is a promising combination for tumor suppression and overcoming Sora resistance and cardiotoxicity in HCC by modulating TRPM7. To our best knowledge, this study represents the first study to investigate the efficiency of CARV/ Sora on the HCC rat model. Moreover, no previous studies have reported the effect of inhibiting TRPM7 on HCC.

Topics: Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cardiotoxicity; Cell Line, Tumor; Cell Proliferation; Liver Neoplasms; Rats; Sorafenib; Thioacetamide; TRPM Cation Channels

Genistein is a recognized isoflavone present in soybeans with antioxidant, anti-inflammatory, antiangiogenic and antitumor activities. This study aimed to test ability of genistein in modulating versican/platelet derived growth factor (PDGF) axis in HCC.. HCC was experimentally induced in male Sprague-Dawley rats then treated with 25 or 75 mg/kg genistein. Antioxidant activities of genistein was assessed by measuring the gene expression of Nrf2 and the hepatic levels of malondialdehyde (MDA), superoxide dismutase (SOD) and reduced glutathione. Expression of versican, PDGF, protein kinase C (PKC) and ERK-1 protein was assessed by Western blotting and immunostaining.. HCC induced an elevation in oxidative stress, PDGF, versican, PKC and ERK protein expression levels. Genistein significantly reduced an HCC-induced increase in oxidative stress. Moreover, genistein dose-dependently reduced HCC-induced elevation of PDGF, versican, PKC and ERK protein expression levels. Moreover, genistein helped retain a normal hepatocyte structure and reduced fibrous tissue deposition, especially in high dose.. Genistein exerted antitumor and antioxidant effects and therefore suppress HCC development via inhibition of the PDGF/versican bidirectional axis, suppressing both ERK1 and PKC as downstream regulators. Therefore, genistein is a potential novel therapeutic candidate for improving the outcome of patients with HCC.

Topics: Animals; Carcinoma, Hepatocellular; Genistein; Liver Neoplasms; Male; Oxidative Stress; Platelet-Derived Growth Factor; Rats; Rats, Sprague-Dawley; Signal Transduction; Thioacetamide; Versicans

The current study investigates the anti-inflammatory and hepatoprotective effects of selenium (Se) formulated as nanoparticles (SeNPs) and in combination with quercetin (QCT) against thioacetamide (TAA)-induced hepatocellular carcinoma (HCC) in rats. ​​​​​​Seventy-two male Sprague-Dawley rats were divided into six groups (n = 12). Three control groups; normal, SeNPs; group received SeNPs only and HCC; group received TAA. In addition, three preventive groups; SeNPs + TAA, QCT + TAA, and QCT + SeNPs + TAA. Induction of HCC was detected histopathologically and by the raise of the serum level of alpha-fetoprotein (AFP). Oxidative stress was evaluated by the hepatic levels of reduced glutathione (GSH), glutathione peroxidase (GPx), and malondialdehyde (MDA) spectrophotometrically. The oncogenic pathway of p53/β-catenin/cyclin D1 was assessed by immunohistochemistry. The inflammatory markers; interleukin-33 (IL-33), IL-6, and IL-1β were assessed by enzyme-linked immune sorbent assay. SeNPs prevented the elevation of serum AFP and hepatic IL-33, IL-1β, and IL-6 in comparison to HCC or QCT + TAA groups. SeNPs + TAA exhibited a lower positive hepatic staining of p53, β-catenin, and cyclin D1 in comparison to HCC or QCT + TAA groups. Moreover, SeNPs improved the overall oxidative balance indicated by low hepatic MDA and enhanced GSH and GPx when compared to HCC or QCT + TAA groups. ​​SeNPs alone and in combination with QCT were found to suppress the progression of HCC in rats via the enhancement of the oxidative stress and then inflammatory status and the prevention of the deregulation of the oncogenic axis pathway of p53/β-catenin/cyclin D.

Topics: alpha-Fetoproteins; Animals; beta Catenin; Carcinoma, Hepatocellular; Cyclin D1; Inflammation; Interleukin-33; Interleukin-6; Liver; Liver Neoplasms; Male; Nanoparticles; Oxidative Stress; Quercetin; Rats; Rats, Sprague-Dawley; Selenium; Thioacetamide; Tumor Suppressor Protein p53

The first-line treatment for advanced hepatocellular carcinoma (HCC) is the multikinase inhibitor sorafenib (SOR). Sofafenib resistance is linked to protein kinase B/ mammalian target of rapamycin (AKT/mTOR) and nuclear factor kappa B (NF-κB) activation, apoptosis inhibition and oxidative stress. This study investigated selenium nanoparticles (SeNps) to overcome SOR resistance in thioacetamide (TAA) induced HCC in rats.. TAA (200 mg/kg/twice weekly, i.p.) was administered for 16 weeks to induce HCC.s. Rats were treated with oral SOR (10 mg/Kg daily), selenium, and SeNps (5 mg/kg three times/week) alone or in combination, for two weeks. Apoptosis, proliferation, angiogenesis, metastasis and drug resistance were assessed. Cleaved caspase 3 (C. CASP3), mTOR, and NF-κB were determined by western blotting. Expression of p53 gene and long-noncoding RNA-AF085935 was determined by qRT-PCR. Expression of B- Cell Leukemia/Lymphoma 2 (Bcl2), Bcl associated X protein (Bax)and glypican 3 (GPC3) was determined by enzyme-linked immunosorbent assay. Liver functions, antioxidant capacity, histopathology and CD34 immunohistochemistry were performed.. SOR/SeNps reversed TAA-induced HCC in rats, through reduction of oxidative stress, activation of p53, Bax and CASP3, and inhibition of Bcl2. SOR/SeNps ameliorated the HCC-induced effect on cell proliferation and drug resistance by targeting mTOR and NF-κB pathways. SOR/SeNps decreased CD34 immunostaining indicating a decrease in angiogenesis and metastasis. SOR/SeNps regulated HCC epigenetically through the lncRNA-AF085935/GPC3 axis.. SOR/SeNps are a promising combination for tumor suppression and overcoming sorafenib resistance in HCC by modulating apoptosis, AKT/mTOR and NF-κB pathways, as well as CD34 and lncRNA-AF085935/GPC3 axis.

Topics: Animals; Antineoplastic Agents; bcl-2-Associated X Protein; Carcinoma, Hepatocellular; Caspase 3; Cell Line, Tumor; Glypicans; Liver Neoplasms; Mammals; Nanoparticles; NF-kappa B; Proto-Oncogene Proteins c-akt; Rats; RNA, Long Noncoding; Selenium; Sorafenib; Thioacetamide; TOR Serine-Threonine Kinases

Hepatocellular carcinoma (HCC) arises in a cirrhotic, pro-angiogenic microenvironment. Inhibiting angiogenesis is a key mode of action of multikinase inhibitors and current non-cirrhotic models are unable to predict treatment response. We present a novel mouse cirrhotic model of xenotransplant that predicts the natural biology of HCC and allows personalized therapy.. Cirrhosis was induced in NOD Scid gamma mice with 4 months of thioacetamide administration. Patient derived xenografts (PDXs) were created by transplant of human HCC subcutaneously into non-cirrhotic mice and intra-hepatically into both cirrhotic and non-cirrhotic mice. The applicability of cirrhotic PDXs for drug testing was tested with 16 days of either sorafenib or lenvatinib. Treatment response was evaluated by MRI.. 8 out of 19 (42%) human HCC engrafted in the cirrhotic model compared with only 3 out of 19 (16%) that engrafted in the subcutaneous non-cirrhotic model. Tumor vasculature was preserved in the cirrhotic model but was diminished in the non-cirrhotic models. Metastasis developed in 3 cirrhotic PDX lines and was associated with early HCC recurrence in all 3 corresponding patients (100%), compared with only 5 out of 16 (31%) of the other PDX lines, P = .027. The cirrhotic model was able to predict response and non-response to lenvatinib and sorafenib respectively in the corresponding patients. Response to lenvatinib in the cirrhotic PDX was associated with reduction in CD34, VEGFR2 and CLEC4G immunofluorescence area and intensity (all P ≤ .03).. A clinically relevant cirrhotic PDX model preserves tumor angiogenesis and allows prediction of response to multikinase inhibitors for personalized therapy.

Topics: Adult; Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Hepatocellular; Cell Proliferation; Disease Models, Animal; Female; Humans; Liver Cirrhosis; Liver Neoplasms; Male; Mice; Mice, Inbred NOD; Mice, SCID; Middle Aged; Neovascularization, Pathologic; Phenylurea Compounds; Precision Medicine; Prognosis; Protein Kinase Inhibitors; Quinolines; Sorafenib; Thioacetamide; Tumor Cells, Cultured; Tumor Microenvironment; Xenograft Model Antitumor Assays

Hepatocellular carcinoma (HCC) is a highly invasive form of hepatic cancer. It is a highly intricate disease with multiple pathophysiological mechanisms underlying its pathogenesis.. The results of the current investigation shed light on the ability of saxagliptin (SAXA) (12.5 mg/kg) to defer HCC progression in an experimental model of thioacetamide (TAA)-induced hepatocarcinogenesis.. SAXA administration improved liver function biomarkers, with a concomitant histopathological recovery. Mechanistically, the observed hepatoprotective impact was associated with significant suppression of the hepatic content of Wnt3a, β-catenin, Notch1, Smo, and Gli2 and enhanced expression of GSK 3β. Nevertheless, the hepatic expression of PCNA, P53, and cyclin D1 was significantly enhanced, with a parallel increase in the tumor expression of caspase-3. Thus, it appears that SAXA significantly enhanced tumor apoptosis, with concomitant suppression of HCC proliferation.. SAXA deferred experimentally-induced HCC via suppressing Wnt/Hedgehog/Notch1 Signaling, with enhanced tumor apoptosis and suppressed proliferation.

Topics: Adamantane; Animals; Apoptosis; Carcinoma, Hepatocellular; Cell Proliferation; Dipeptides; Dipeptidyl-Peptidase IV Inhibitors; Hedgehog Proteins; Liver; Liver Neoplasms; Liver Neoplasms, Experimental; Male; Rats, Sprague-Dawley; Receptor, Notch1; Thioacetamide; Wnt Signaling Pathway

The results of the current study investigated the chemo-preventive effect of crocin against hepatocarcinogenesis in rats with particular focus on the evaluation of the modulatory impact of crocin on apoptotic and nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathways. Thioacetamide (TAA) (200 mg/kg, I.P.) was used for experimental induction of hepatocarcinogenesis in rats. Crocin administration significantly attenuated TAA-induced cancerous lesions with concomitant attenuation of impaired liver functions. This was associated with significant enhancement in hepatic Nrf2 and heme oxygenase-1 (HO-1) expression with parallel suppression in Keap-1 expression. Inline, crocin induced a significant improvement in hepatic oxidative status with enhanced antioxidant batteries. Crocin administration significantly suppressed the hepatic content of c-Jun N-terminal kinase (c-JNK) with significant upregulation in TNF-related apoptosis-inducing ligand (TRAIL) and caspase-8 protein expression as well as p53 gene expression; biomarkers of apoptosis. Moreover, hepatic expression of the apoptotic BAX significantly increased and the anti-apoptotic Bcl-2 significantly decreased in the liver specimen; biomarkers of intrinsic apoptosis. In conclusion; crocin attenuates experimentally induced hepato-carcinogenesis via modulation of oxidative/apoptotic signaling. Namely, crocin induced hepatic expression of Nrf2 with downstream modulation of endogenous HO-1 and Keap-1 signaling with modulation of various key players of apoptosis including; c-JNK, p53, TRAIL, caspase-8, BAX, and Bcl-2.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinogenesis; Carcinoma, Hepatocellular; Carotenoids; Heme Oxygenase (Decyclizing); JNK Mitogen-Activated Protein Kinases; Kelch-Like ECH-Associated Protein 1; Liver; Liver Neoplasms; Male; NF-E2-Related Factor 2; Protective Agents; Proto-Oncogene Proteins c-bcl-2; Rats, Sprague-Dawley; Signal Transduction; Thioacetamide

Given the enormous impact of HCC on the patients' quality of life and healthcare economics, the current study was conducted to investigate the potential ability of adiponectin to reverse established HCC and to investigate the underlying mechanisms which control the chemotherapeutic and hepatoprotective effects. HCC was induced in Male Sprague Dawely rats by I.P. injection of thioacetamide(200 mg/kg) 3 times/week for 14 weeks.HCC development was confirmed by histopathological examination and assessment of serum levels of α-fetoprotein (AFP). Adiponectin was administered (5 μg/kg, I.P.) starting from week 13 of the experiment and for further 4 weeks. Adiponectinadministration revealed a significant antitumor activity with significant improvement in liver functions and oxidative status. Nevertheless, pathological features as cirrhosis, dysplastic changes, and tumoral nodules were significantly attenuated with significant enhancement in hepatic caspase-3 immunostaining. Mechanistically, adiponectin administration was associated with significant restoration of p53 activity; which increased by 133%, with a reduction in HCC-induced expression of-JNK which decreased by 53%as well as a significant enhancement of hepatic TRAIL and caspase-8 activities which increased by 27% and 20% respectively. In conclusion; Adiponectin can be proposed as a promising therapy for HCC. Adiponectin's tumoricidal activity can be partially mediated by blocking HCC-induced reduction in p53 expression as well as reactivation of TRAIL signaling and induction of apoptotic pathway providing more protection for the body against the tumor.

Topics: Adiponectin; Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Caspase 8; Liver; Liver Neoplasms; Male; Rats, Sprague-Dawley; Signal Transduction; Thioacetamide; TNF-Related Apoptosis-Inducing Ligand; Tumor Suppressor Protein p53

Improvement of gut microbiota may help in preventing the progression of cirrhosis. We supposed that Lactobacillus Plantarum (L. Plantarum) protects the cirrhotic liver through suppression of TLR4/ CXCL9/ PREX-2.. Rats were divided into two groups. Group I, lasts for six weeks and Group II lasts for 12 weeks. Each group was subdivided into: naïve, Lactobacillus Plantarum (L. Plantarum), thioacetamide (TAA) and TAA + L. Plantarum. Liver function tests, α fetoprotein (AFP) levels, CXCL9, PREX-2 and TLR4 expression were assessed. Histological studies were performed.. TAA induced significant deterioration in liver functions and increased AFP. There was periportal cirrhosis, vacuolated hepatocytes, decrease hepatocyte parrafin-1 (hep par-1) expression, increase proliferating cell nuclear antigen (PCNA) positive nuclei and cytokeratin AE1/AE3. The PCR results showed significant increase in TLR4, CXCL9 and PREX-2 expression. Early administration of L. Plantarum significantly decreased the expression of TLR4, CXCL9 and PREX-2 together with improvement in liver function and prevented the pathological changes.. The cirrhotic complications induced by TAA are through activation of TLR4/ CXCL9/ PREX-2 pathway and could be prevented by the early administration of L. Plantarum.

Topics: Animals; Carcinoma, Hepatocellular; Chemokine CXCL9; Lactobacillus plantarum; Liver; Liver Cirrhosis, Experimental; Liver Neoplasms; Male; Microbiota; Probiotics; Rats; Rats, Wistar; Signal Transduction; Thioacetamide; Toll-Like Receptor 4

Impaired apoptosis and systemic toxicity of chemotherapeutic drugs make cancer treatment suboptimal. Thus, there is urgency for drug repurposing which facilitates discovery of safe and effective combination therapy. This study aimed to evaluate chloroquine's (CQ) ability to trigger TRAIL/TRAILR2 apoptotic pathway in thioacetamide (TAA)-induced hepatocellular carcinoma (HCC) either alone or in combination with doxorubicin (DOX). Moreover, its ability to attenuate DOX-induced cardiotoxicity was investigated. TAA was injected in male Sprague Dawely rats (200 mg/kg; ip; 2 times/week) for 16 weeks. After the 16th week, rats were further divided into different groups (n = 10) and treated for 7 weeks. CQ group (received CQ 25 mg/kg/day; orally), DOX group (received DOX 1 mg/kg; ip; 2 times/week) and CQ/DOX group. Liver function biomarkers, AFP, hepatic levels of MDA and GSH, serum CK-MB and LDH enzymes activity were measured. Quantitative, Real-Time PCR was used to measure TRAIL, TRAILR2, caspase-8, caspase-9, caspase-3, BCL-2 and TGF-β1 genes expression levels. Necroinflammation and fibrosis were scored by histopathological examination. CQ improved liver functions, reduced AFP level and attenuated HCC progression. CQ induced apoptosis via upregulation of TRAIL/TRAILR2, caspase-8, caspase-3 and caspase-8 genes and downregulation of BCL-2 gene. Moreover, CQ/DOX showed marked decrease in hepatic MDA level, serum CK-MB, LDH enzymes activity, as well as marked increase in hepatic GSH level. In conclusion, this work assess the in vivo efficacy of CQ/DOX combination therapy in this HCC model that not only has enhanced anti-tumor activity but it also protects against DOX-induced cardiotoxicity. Nevertheless, more studies should be performed to illustrate the molecular mechanism of CQ's cardioprotective effect.

Topics: Animals; Carcinoma, Hepatocellular; Caspases; Chloroquine; Doxorubicin; Gene Expression Regulation; Liver Neoplasms; Male; Proto-Oncogene Proteins c-bcl-2; Rats; Receptors, TNF-Related Apoptosis-Inducing Ligand; RNA, Messenger; Thioacetamide; TNF-Related Apoptosis-Inducing Ligand; Up-Regulation

The study was conducted for evaluation of the antitumor activity of SSTN. Sixty male Sprague-Dawley rats were randomized into four equal groups: Control, SSTN. SSTN

Topics: Angiogenesis Inhibitors; Animals; Carcinoma, Hepatocellular; Cell Proliferation; Down-Regulation; Integrin alphaVbeta3; Liver Function Tests; Liver Neoplasms, Experimental; Male; Neovascularization, Pathologic; Oxidative Stress; Rats; Rats, Sprague-Dawley; Signal Transduction; Syndecan-1; Thioacetamide

The urgent unmet need for hepatocellular carcinoma (HCC) therapies is addressed here by characterising a novel mouse model of HCC in the context of ongoing liver damage and overnutrition. Male C57Bl/6J mice were treated with diethylnitrosamine (DEN) and thioacetamide (TAA), and some were provided with an atherogenic high fat diet (HFD). Inflammation, steatosis, fibrosis, 87 genes, liver lesions and intratumoural leukocyte subsets were quantified up to 24 weeks of age. Adding HFD to DEN/TAA increased fibrosis, steatosis and inflammation, and the incidence of both HCC and non-HCC dysplastic lesions. All lesions contained α-SMA positive fibroblasts. Macrophage marker F4/80 was not significantly different between treatment groups, but the macrophage-associated genes Arg-1 and Cd47 were differentially expressed. Fibrosis, cancer and cell death associated genes were upregulated in DEN/TAA/HFD livers. Fewer Kupffer cells and plasmacytoid dendritic cells were in tumours compared to control liver. In conclusion, combining a hepatotoxin with an atherogenic diet produced more intrahepatic tumours, dysplastic lesions and fibrosis compared to hepatotoxin alone. This new HCC model provides a relatively rapid means of examining primary HCC and potential therapies in the context of multiple hepatotoxins including those derived from overnutrition.

Topics: Alkylating Agents; Animals; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; Diet, High-Fat; Diethylnitrosamine; Gene Expression Regulation, Neoplastic; Liver Neoplasms, Experimental; Male; Mice; Mice, Inbred C57BL; Thioacetamide

Angiogenesis plays important roles in progression of hepatocellular carcinoma. The antiangiogenic mechanisms of vitexicarpine are not fully defined. Therefore, we conducted the following study to evaluate the antiangiogenic mechanism and antitumor activity of vitexicarpine in vivo model of hepatocellular carcinoma through modulation of vascular endothelial growth factor signaling pathway. Hepatocellular carcinoma was induced in Sprague Dawley rats by thioacetamide. Hepatocellular carcinoma was assessed by measuring serum alpha-fetoprotein and investigating liver sections stained with hematoxylin/eosin. Hepatocellular carcinoma rats were injected with vitexicarpine (150 mg/kg) for 2 weeks. Hepatic vascular endothelial growth factor was measured by enzyme-linked immunosorbent assay. Protein and expression of hepatic phospho-Ser473-AKT (p-AKT) and phospho-Tyr419-Src (p-Src) were determined. The apoptotic pathway was evaluated by assessment of protein expression of caspase-3. Vitexicarpine increased rats' survival time and decreased serum alpha-fetoprotein as well as it ameliorated fibrosis and massive hepatic tissue breakdown. It attenuated hepatocellular carcinoma-induced protein and gene expression of vascular endothelial growth factor, p-AKT, p-Src, and caspase-3. In conclusion, this study suggests that vitexicarpine possesses both antiangiogenic and antitumor activities through inhibition of vascular endothelial growth factor, p-AKT/AKT, and p-Src with subsequent inhibition of apoptotic pathway.

Topics: alpha-Fetoproteins; Angiogenesis Inhibitors; Animals; Apigenin; Apoptosis; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Gene Expression Regulation, Neoplastic; Humans; Liver Neoplasms; Neoplasm Proteins; Neovascularization, Pathologic; Rats; Signal Transduction; Thioacetamide; Vascular Endothelial Growth Factor A; Xenograft Model Antitumor Assays

Wnt3a and Wnt5a are ligands orchestrating the canonical and non-canonical pathways, respectively, with involvement in hepatocellular carcinoma (HCC). Hesperidin (HP) is a natural product found in citrus fruits and reputed for its antitumor activity. The present study aims to investigate the potential hepatoprotective effect of HP against thioacetamide (TAA)-induced HCC focusing on its potential role on Wnt3a and Wnt5a signaling pathways.. Forty rats were equally divided into groups; normal control, HP control (receiving HP, 150mg/kg/day), HCC (receiving TAA, 200mg/kg twice weekly for 14weeks) and HP-HCC (receiving HP and TAA). Gene expressions of Wnt3a, Wnt5a, β-catenin and Cyclin D1 were assessed by qPCR, while their protein levels, along with active caspase-3 level, were quantified by ELISA and immunohistochemistry. Liver functions, oxidative stress parameters and myeloperoxidase activity were measured. MTT assay of hepG2 cells treated with recombinant Wnt3a (10ng/ml) in presence or absence of HP (100μM) was performed.. HCC group exhibited a significant increase in Wnt3a, β-catenin, Cyclin D1 and Wnt5a gene expressions, as well as, their protein levels. HP significantly prevented TAA-activated Wnt3a/β-catenin and Wnt5a pathways. Moreover, HP exerted hepatoprotective effect by significantly improving the oxidative imbalance, inflammation and liver function parameters, serum ALT, AST activities, and albumin level.. Our study is the first to report the possible role of Wnt3a/β-catenin and Wnt5a pathways in TAA-induced early HCC model in rats. HP has a prophylactic effect against hepatocarcinogenesis via preventing the induction of both canonical and non-canonical Wnt pathways.

Topics: Animals; Carcinoma, Hepatocellular; Gene Expression Regulation; Hesperidin; Liver Neoplasms; Male; Oxidative Stress; Rats; Rats, Sprague-Dawley; Thioacetamide; Wnt Signaling Pathway; Wnt-5a Protein; Wnt3A Protein

To elucidate the role of STAT3 in hepatocarcinogenesis and biliary ductular proliferation following chronic liver injury.. We investigated thioacetamide (TAA)-induced liver injury, compensatory hepatocyte proliferation, and hepatocellular carcinoma (HCC) development in hepatic STAT3-deficient mice. In addition, we evaluated TAA-induced biliary ductular proliferation and analyzed the activation of sex determining region Y-box9 (SOX9) and Yes-associated protein (YAP), which regulate the transdifferentiation of hepatocytes to cholangiocytes.. Both compensatory hepatocyte proliferation and HCC formation were significantly decreased in hepatic STAT3-deficient mice as compared with control mice. STAT3 deficiency resulted in augmentation of hepatic necrosis and fibrosis. On the other hand, biliary ductular proliferation increased in hepatic STAT3-deficient livers as compared with control livers. SOX9 and YAP were upregulated in hepatic STAT3-deficient hepatocytes.. STAT3 may regulate hepatocyte proliferation as well as transdifferentiation into cholangiocytes and serve as a therapeutic target for HCC inhibition and biliary regeneration.

Topics: Adaptor Proteins, Signal Transducing; Animals; Biliary Tract; Carcinogenesis; Carcinoma, Hepatocellular; Cell Cycle Proteins; Cell Proliferation; Cell Transdifferentiation; Chemical and Drug Induced Liver Injury; Hepatocytes; Liver; Liver Neoplasms; Liver Neoplasms, Experimental; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Phosphoproteins; Phosphorylation; Regeneration; SOX9 Transcription Factor; STAT3 Transcription Factor; Thioacetamide; Up-Regulation; YAP-Signaling Proteins

The present study identified genes showing promoter region hypermethylation by CpG island microarrays in the liver of rats treated with hepatocarcinogen thioacetamide (TAA) for 28days. Among 47 hypermethylated genes, Hist1h2aa, Tmem70, Ube2e2, and Slk were confirmed to show hypermethylation by methylation-specific quantitative polymerase chain reaction (PCR) and pyrosequencing analyses as well as downregulation of transcript levels by real-time reverse transcription-PCR analysis in the livers of rats treated with TAA. All gene products of the 4 selected genes showed decreased immunoreactivity forming negative liver cell foci in a subpopulation of glutathione S-transferase placental form (GST-P)

Topics: Adenoma, Liver Cell; Animals; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; Down-Regulation; Epigenesis, Genetic; Gene Expression Regulation, Neoplastic; Liver; Liver Neoplasms; Membrane Proteins; Rats; Thioacetamide; Ubiquitin-Protein Ligases

Most hepatocellular carcinomas (HCCs) develop in the context of chronic liver inflammation. Oxidative stress is thought to play a major role in the pathogenesis of HCC development. In this study, we examined whether cold-inducible RNA-binding protein (Cirp) controls reactive oxygen species (ROS) accumulation and development of HCC by using murine models of hepatocarcinogenesis and human liver samples. Cirp expression, ROS accumulation, and CD133 expression were increased in the liver of tumor-harboring mice. Cirp deficiency reduced production of interleukin-1β and interleukin-6 in Kupffer cells, ROS accumulation, and CD133 expression, leading to attenuated hepatocarcinogenesis. Thioacetamide treatment enhanced hepatic expression of CD133 and phosphorylated signal transducer and activator of transcription 3 (STAT3), which was prevented by treatment with the antioxidant butylated hydroxyanisole. Intriguingly, the risk of human HCC recurrence is positively correlated with Cirp expression in liver. Cirp appears to play a critical carcinogenic function and its expression might be a useful biomarker for HCC risk prediction.

Topics: AC133 Antigen; Animals; Antigens, CD; Antioxidants; Butylated Hydroxyanisole; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Glycoproteins; Humans; Interleukin-1beta; Interleukin-6; Kupffer Cells; Liver; Liver Neoplasms; Mice; Mice, Inbred C57BL; Mice, Knockout; Neoplasm Recurrence, Local; Neoplastic Stem Cells; Oxidative Stress; Peptides; Phosphorylation; Reactive Oxygen Species; RNA-Binding Proteins; SOXB1 Transcription Factors; STAT3 Transcription Factor; Thioacetamide; Tissue Fixation

Cirrhosis is characterized by an excessive accumulation of extracellular matrix components including hyaluronic acid (HA) and is widely considered a preneoplastic condition for hepatocellular carcinoma (HCC). 4-Methylumbelliferone (4MU) is an inhibitor of HA synthesis and has anticancer activity in an orthotopic HCC model with underlying fibrosis. Our aim was to explore the effects of HA inhibition by 4MU orally administered on tumor microenvironment. Hepa129 tumor cells were inoculated orthotopically in C3H/HeJ male mice with fibrosis induced by thioacetamide. Mice were orally treated with 4MU. The effects of 4MU on angiogenesis were evaluated by immunostaining of CD31 and quantification of proangiogenic factors (vascular endothelial growth factor, VEGF, interleukin-6, IL-6 and C-X-C motif chemokine 12, CXCL12). IL-6 was also quantified in Hepa129 cells in vitro after treatment with 4MU. Migration of endothelial cells and tube formation were also analyzed. As a result, 4MU treatment decreases tumor growth and increased animal survival. Systemic levels of VEGF were significantly inhibited in 4MU-treated mice. Expression of CD31 was reduced after 4MU therapy in liver parenchyma in comparison with control group. In addition, mRNA expression and protein levels of IL-6 and VEGF were inhibited both in tumor tissue and in nontumoral liver parenchyma. Interestingly, IL-6 production was dramatically reduced in Kupffer cells isolated from 4MU-treated mice, and in Hepa129 cells in vitro. Besides, 4MU was able to inhibit endothelial cell migration and tube formation. In conclusion, 4MU has antitumor activity in vivo and its mechanisms of action involve an inhibition of angiogenesis and IL-6 production. 4MU is an orally available molecule with potential for HCC treatment.

Topics: Administration, Oral; Animals; Antineoplastic Agents; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Movement; Chemokine CXCL12; Endothelial Cells; Gene Expression Regulation, Neoplastic; Hymecromone; Interleukin-6; Kupffer Cells; Liver Cirrhosis; Liver Neoplasms; Male; Mice; Mice, Inbred C3H; Neovascularization, Pathologic; Platelet Endothelial Cell Adhesion Molecule-1; Signal Transduction; Survival Analysis; Thioacetamide; Vascular Endothelial Growth Factor A

Hepatocellular carcinoma represents one of the most-rapidly spreading cancers in the world. In the majority of cases, an inflammation-driven fibrosis or cirrhosis precedes the development of the tumor. During malignant transformation, the tumor microenvironment undergoes qualitative and quantitative changes that modulate the behavior of the malignant cells. A key constituent for the hepatic microenvironment is the small leucine-rich proteoglycan decorin, known to interfere with cellular events of tumorigenesis mainly by blocking various receptor tyrosine kinases (RTK) such as EGFR, Met, IGF-IR, PDGFR and VEGFR2. In this study, we characterized cell signaling events evoked by decorin deficiency in two experimental models of hepatocarcinogenesis using thioacetamide or diethyl nitrosamine as carcinogens. Genetic ablation of decorin led to enhanced tumor occurrence as compared to wild-type animals. These findings correlated with decreased levels of the cyclin-dependent kinase inhibitor p21(WAF1/CIP1) and a concurrent elevation in retinoblastoma protein phosphorylation via cyclin dependent kinase 4. Decreased steady state p21(Waf1/Cip1) levels correlated with enhanced expression of transcription factor AP4, a known transcriptional repressor of p21(Waf1/Cip1), and enhanced c-Myc protein levels. In addition, translocation of β-catenin was a typical event in diethyl nitrosamine-evoked tumors. In parallel, decreased phosphorylation of both c-Myc and β-catenin was observed in Dcn(-/-) livers likely due to the hindered GSK3β-mediated targeting of these proteins to proteasomal degradation. We discovered that in a genetic background lacking decorin, four RTKs were constitutively activated (phosphorylated), including three known targets of decorin such as PDGFRα, EGFR, IGF-IR, and a novel RTK MSPR/RON. Our findings provide powerful genetic evidence for a crucial in vivo role of decorin during hepatocarcinogenesis as lack of decorin in the liver and hepatic stroma facilitates experimental carcinogenesis by providing an environment devoid of this potent pan-RTK inhibitor. Thus, our results support future utilization of decorin as an antitumor agent in liver cancer.

Topics: Animals; Blotting, Western; Carcinogenesis; Carcinoma, Hepatocellular; Cyclin-Dependent Kinase Inhibitor p21; Decorin; Diethylnitrosamine; ErbB Receptors; Gene Expression Regulation, Neoplastic; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Liver Neoplasms; MAP Kinase Signaling System; Mice; Models, Biological; Real-Time Polymerase Chain Reaction; Receptor Protein-Tyrosine Kinases; Receptor, Platelet-Derived Growth Factor alpha; Receptors, Somatomedin; Statistics, Nonparametric; Thioacetamide

Epigallocatechin-gallate (EGCG) claims a plethora of health benefits including protection against neoplastic diseases. Meanwhile, heparan-sulfate proteoglycans (HSPGs) have defensive role against tumour cell invasion. Therefore, the chemopreventive and hepatoprotective effects of EGCG were studied in hepatocellular carcinoma (HCC) in vivo and in vitro and compared with strong water soluble antioxidant, sodium ascorbate.. HCC was induced in SD rats by thioacetamide (200 mg/Kg). Some rats were treated with EGCG (20 mg/Kg) or sodium ascorbate (100 mg/Kg). Liver impairment was assessed by measuring serum α-fetoprotein and investigating liver sections stained with H/E. Hepatic HSPGs, syndecan-1 and matrix metalloproteinase-9 (MMP-9) were measured by ELISA. Gene expression of fibroblast growth factor (FGF)-2 was measured. Cell death was assessed by caspase-3 activity. In addition, all markers were measured in human hepatocellular carcinoma cell line (HepG2).. EGCG increased the animal survival and decreased both α-fetoprotein and HepG2 viability. In addition, EGCG ameliorated fibrosis and massive hepatic tissue breakdown. EGCG restored HSPGs and reduced expression of MMP-9, syndecan-1 and FGF-2 in-vivo and in-vitro. Sodium ascorbate showed significantly lower results than EGCG.. Besides antioxidant activity, other mechanisms are involved in the chemopreventive and hepatoprotective effects of EGCG including restoration of HSPGs receptors and inhibition of vascular invasion.

Topics: alpha-Fetoproteins; Animals; Antineoplastic Agents, Phytogenic; Antioxidants; Ascorbic Acid; Camellia sinensis; Carcinoma, Hepatocellular; Catechin; Fibroblast Growth Factor 2; Gene Expression; Hep G2 Cells; Heparan Sulfate Proteoglycans; Humans; Liver; Liver Neoplasms; Male; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Phytotherapy; Plant Extracts; Rats, Sprague-Dawley; Syndecan-1; Thioacetamide

We previously reported a toxicogenomics-based prediction model for hepatocarcinogens in which the expression patterns of signature genes following repeated doses of either genotoxic or non genotoxic compounds were similar. Based on the results of our prediction model, we hypothesized that repeated doses of non-genotoxic carcinogens might have initiating potential. Here, we conducted a two stage hepatocarcinogenesis study in rats exposed to the initiating agent nitrosodiethylamine (DEN), and hepatotoxic compounds thioacetamide (TAA), methapyrilene (MP) and acetaminophen (APAP) for 1-2 weeks followed by the liver tumor promoter phenobarbital (PB). The duration of initial treatment was determined based on positive results from our prediction model. Combined treatment of 3 or 30 mg/kg of genotoxic DEN and PB induced marked increases in altered hepatocellular foci and a DEN dose-dependent increase in the number and area of glutathione S-transferase-placental form (GST-P)-positive foci. A low number of altered hepatocellular foci were also observed in rats treated with TAA at a dose of 45 mg/kg.MP at a dose of 100 mg/kg induced a very low number of foci, but APAP did not. Hierarchical clustering analysis using gene expression data revealed that 2-week treatment with TAA at a dose of 30 mg/kg and MP at 45 mg/kg induced specific expression of DNA damage-related genes, similar to 1-week treatment with DEN at a dose of 30 mg/kg. These results suggest that TAA and MP induce DNA damage, which partially supports our hypothesis. Although this study does not indicate whether tumor growth in response to these compounds can be assessed in this model, our results suggest that cumulative treatment with non genotoxic TAA might have initiating potential in the liver.

Topics: Acetaminophen; Animals; Carcinoma, Hepatocellular; Diethylnitrosamine; Disease Models, Animal; DNA Damage; Dose-Response Relationship, Drug; Glutathione Transferase; Liver Neoplasms; Male; Methapyrilene; Mutagenicity Tests; Oxidative Stress; Phenobarbital; Rats, Sprague-Dawley; Thioacetamide

Recombinant vesicular stomatitis virus (VSV) shows promise for the treatment of hepatocellular carcinoma (HCC), but its safety and efficacy when administered in a setting of hepatic fibrosis, which occurs in the majority of clinical cases, is unknown. We hypothesized that VSV could provide a novel benefit to the underlying fibrosis, due to its ability to replicate and cause cell death specifically in activated hepatic stellate cells. In addition to the ability of VSV to produce a significant oncolytic response in HCC-bearing rats in the background of thioacetamide-induced hepatic fibrosis without signs of hepatotoxicity, we observed a significant downgrading of fibrosis stage, a decrease in collagen content in the liver, and modulation of gene expression in favor of fibrotic regression. Together, this work suggests that VSV is not only safe and effective for the treatment of HCC with underlying fibrosis, but it could potentially be developed for clinical application as a novel antifibrotic agent.

Topics: Animals; Carcinoma, Hepatocellular; Cell Line, Tumor; Hep G2 Cells; Humans; Infusions, Intra-Arterial; Liver Cirrhosis; Liver Cirrhosis, Experimental; Liver Neoplasms; Male; Oncolytic Virotherapy; Oncolytic Viruses; Rats; Rats, Inbred BUF; Thioacetamide; Vesicular stomatitis Indiana virus

Liver cirrhosis is characterized by an excessive accumulation of extracellular matrix components, including hyaluronan (HA). In addition, cirrhosis is considered a pre-neoplastic disease for hepatocellular carcinoma (HCC). Altered HA biosynthesis is associated with cancer progression but its role in HCC is unknown. 4-Methylumbelliferone (4-MU), an orally available agent, is an HA synthesis inhibitor with anticancer properties. In this work, we used an orthotopic Hepa129 HCC model established in fibrotic livers induced by thioacetamide. We evaluated 4-MU effects on HCC cells and hepatic stellate cells (HSCs) in vitro by proliferation, apoptosis and cytotoxicity assays; tumor growth and fibrogenesis were also analyzed in vivo. Our results showed that treatment of HCC cells with 4-MU significantly reduced tumor cell proliferation and induced apoptosis, while primary cultured hepatocytes remained unaffected. 4-MU therapy reduced hepatic and systemic levels of HA. Tumors systemically treated with 4-MU showed the extensive areas of necrosis, inflammatory infiltrate and 2-3-fold reduced number of tumor satellites. No signs of toxicity were observed after 4-MU therapy. Animals treated with 4-MU developed a reduced fibrosis degree compared with controls (F1-2 vs F2-3, respectively). Importantly, 4-MU induced the apoptosis of HSCs in vitro and decreased the amount of activated HSCs in vivo. In conclusion, our results suggest a role for 4-MU as an anticancer agent for HCC associated with advanced fibrosis.

Topics: Animals; Antineoplastic Agents; Apoptosis; Carcinoma, Hepatocellular; Cell Line, Tumor; Cell Proliferation; Drug Screening Assays, Antitumor; Fibroblasts; Glucuronosyltransferase; Humans; Hyaluronan Receptors; Hyaluronan Synthases; Hyaluronic Acid; Hymecromone; Liver; Liver Cirrhosis, Experimental; Liver Neoplasms, Experimental; Male; Mice; Mice, Inbred C3H; Neoplasm Transplantation; Thioacetamide; Tumor Burden

Liver cirrhosis and hepatocellular carcinomas are two major causes of morbidity and mortality worldwide, and can synergistically interact to expedite the tumor progression. How fibrosis promotes the hepatoma growth remains completely unexplained. Using an in situ murine hepatoma model together with fibrosis induction by thioacetamide (TAA), the hepatoma growth and the immune factors in the fibrotic liver were analyzed. We found that TAA-fibrosis induction enhanced hepatoma cell growth in the liver and increased the mortality of hepatoma-bearing mice. The tumor-infiltrating CD4(+) or CD8(+) T cells are downregulated by fibrosis induction. The Foxp3(+) regulatory T cells (Treg) cells were induced. We conclude that fibrosis induction causes further immunosuppression, in which Treg cells exert a downregulation effect on the antitumor immunity.

Topics: Animals; Carcinogens; Carcinoma, Hepatocellular; CD4-Positive T-Lymphocytes; Cell Line, Tumor; Cell Proliferation; Forkhead Transcription Factors; Immune Tolerance; Liver Cirrhosis; Liver Neoplasms, Experimental; Lymphocyte Count; Male; Mice; Mice, Inbred BALB C; Neoplasms; T-Lymphocytes, Regulatory; Thioacetamide; Tumor Escape

Hepatocellular carcinoma (HCC) is one of the common cancers worldwide, caused by Hepatitis C virus (HCV) and hepatotoxins. Here we report the development of HCC in wild type as well as HCV core protein (HCP)-transgenic zebrafish upon treatment with a hepatotoxin, thioacetamide (TAA). Two-fold accelerated HCC development could be achieved in the TAA-treated transgenic fish, that is, the progression of the disease in TAA-treated wild type zebrafish developed HCC in 12 weeks whereas that of HCP-transgenic zebrafish shortened the HCC progression to 6 weeks. Histopathological observation showed the specific pathological features of HCC. The HCC progression was confirmed through RT-PCR that revealed an up and down regulation of different marker genes at various stages of HCC progression such as, steatohepatitis, fibrosis and HCC. Moreover, HCV core protein expressed in the HCP-transgenic zebrafish and TAA synergistically accelerate the HCC development. It must be mentioned that, this is the first report revealing HCV core protein along with TAA to induce HCC in zebrafish, particularly, in a short period of time comparing to mice model. As zebrafish has already been considered as a good human disease model and in this context, this HCC-zebrafish model may serve as a powerful preclinical platform to study the molecular events in hepatocarcinogenesis, therapeutic strategies and for evaluating chemoprevention strategies in HCC.

Topics: Animals; Animals, Genetically Modified; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; DNA Primers; Fatty Liver; Hepacivirus; Liver; Liver Cirrhosis; Liver Diseases; Liver Neoplasms; Microscopy, Confocal; Reverse Transcriptase Polymerase Chain Reaction; Thioacetamide; Viral Core Proteins; Zebrafish

Mice primed by feeding griseofulvin or diethyl 1,4-dihydro 1,4,6-trimethyl 3,5-pyridine decarboxylate for 5 months followed by drug withdrawal for 1 month (drug-primed mice) were given thioacetamide intraperitoneally, and the livers were subsequently studied at intervals up to 7 days. The hepatocellular proliferative response was measured by immunostaining for proliferative cell nuclear antigen. Necrosis was followed by measuring ALT. Mallory bodies were identified by immunoperoxidase stains for ubiquitin and cytokeratin. Preneoplastic foci were localized using immunofluorescence stain for glutathione S-transferase (GST mu) and histochemical stain for gamma glutamyl transpeptidase (GGT). The results showed that the preneoplastic foci selectively proliferated and expanded and formed nodules as indicated by quantitation of nuclei stained positive for proliferating cell nuclear antigen after thioacetamide treatment. Data support the hypothesis that the preneoplastic foci consisted of clones of hepatocytes which preferentially express GST mu, GGT and Mallory bodies. These preneoplastic cells selectively proliferate in response to the promoter effects of necrosis-induced liver cell regeneration ("chemical partial hepatectomy").

Topics: Animals; Carcinoma, Hepatocellular; gamma-Glutamyltransferase; Glutathione Transferase; Griseofulvin; Hepatocytes; Liver; Liver Neoplasms; Liver Regeneration; Mice; Precancerous Conditions; Pyridines; Thioacetamide

To examine the property of hepatocellular carcinoma (HCC), rat HCC cells were implanted into normal and cirrhotic rat livers. Implanted HCC grew much more progressively in cirrhotic livers than in normal livers. Kupffer cells were decreased profoundly in cirrhotic livers, resulting in markedly impaired phagocytic activity. Furthermore, production of Kupffer cell-related cytokines, such as interleukin-1 beta and tumor necrosis factor-alpha, was decreased profoundly in cirrhotic livers. Our results indicate that liver cirrhosis is a prominent promoting factor in HCC progression, and that markedly depressed Kupffer cell activity may play a role in augmented HCC progression in cirrhotic livers.

Topics: Animals; Carcinoma, Hepatocellular; Disease Models, Animal; Disease Progression; Interleukin-1; Kupffer Cells; Liver; Liver Cirrhosis; Liver Neoplasms; Male; Phagocytosis; Precancerous Conditions; Rats; Rats, Sprague-Dawley; Thioacetamide; Tumor Necrosis Factor-alpha

Heparin-binding epidermal growth factor-like growth factor is an hepatotrophic factor expressed in non-parenchymal liver cells but not in hepatocytes in regenerating rat liver after partial hepatectomy. Human hepatocellular carcinoma cells also produce this growth factor. In this study, the expression of the growth factor in the hepatocytes of fibrotic liver during hepatocarcinogenesis was investigated.. Hepatic fibrosis was induced in rats by oral administration of 0.05% thioacetamide. Hepatocytes were isolated by in situ perfusion methods. Growth factor gene and protein expression were investigated by northern hybridization and immunohistochemistry, respectively. Expression of glutathione s-transferase P, which is expressed when hepatocytes undergo neoplastic transformation, was also investigated.. Some hepatocytes in fibrotic liver, but not in normal liver, stained positively by immunohistochemistry for heparin-binding epidermal growth factor-like growth factor. The growth factor and glutathione s-transferase P gene transcript were present in hepatocytes isolated from fibrotic liver, but not in those isolated from normal liver. Immunohistochemical localization of both proteins in fibrotic liver revealed similar patterns.. In essence, hepatocytes in fibrotic rat liver produce heparin-binding epidermal growth factor-like growth factor. Expression of this growth factor may occur as hepatocytes are transformed to a neoplastic phenotype.

Topics: Animals; Carcinoma, Hepatocellular; Electrophoresis, Agar Gel; Epidermal Growth Factor; Glutathione Transferase; Heparin-binding EGF-like Growth Factor; Immunohistochemistry; Intercellular Signaling Peptides and Proteins; Liver; Liver Cirrhosis, Experimental; Liver Neoplasms; Male; Rats; Rats, Sprague-Dawley; Thioacetamide

Rats were treated with low doses of the hepatocarcinogens dimethylnitrosamine or thioacetamide, and livers were examined 48 h later. These treatments are known to produce altered RNA compartmentation, wherein a class of repetitive RNA sequences normally restricted to the nucleus appears in the cytoplasm. Reverse transcription-PCR amplifications demonstrated that the sequences showing altered compartmentation consisted largely of a subfamily of the rodent B2 sequence family, the counterpart of human Alu sequences involved in retrotransposition. Northern blot analyses showed that these B2 sequences were found in cytoplasmic RNA as 170- to 360-nucleotide "sense" transcripts, and competition hybridization experiments established that B2 sequences represented most (if not all) of the sequences showing altered compartmentation. The major increase in B2 transcriptions in cytoplasmic RNA was not associated with any change in B2 transcription by RNA polymerase III. In situ hybridizations showed that the altered compartmentation of B2 sequences occurred in well-delineated foci within the rat liver; these foci consisted of a central region containing a prominent infiltrate of macrophages admixed with small hepatocytes and a peripheral region of histologically normal hepatocytes that showed evidence of oxidative damage. Altered compartmentation of B2 sequences may represent an important focal initiatory change in a subset of hepatocytes, whereas subsequent retrotranspositional events (associated with Alu-like sequences) could predispose initiated cell foci to alterations in promotion/progression phases.

Topics: Animals; Base Sequence; Blotting, Northern; Carcinogens; Carcinoma, Hepatocellular; Cell Compartmentation; Dimethylnitrosamine; In Situ Hybridization; Liver Neoplasms; Male; Molecular Sequence Data; Nucleic Acid Conformation; Phosphorus Radioisotopes; Rats; Rats, Sprague-Dawley; Repetitive Sequences, Nucleic Acid; RNA, Antisense; RNA, Messenger; Thioacetamide; Time Factors

2-(Aminomethyl)-4-aminobutyric acid (isoornithine), 3-methylisoornithine, and 2,3-dimethylisoornithine were not decarboxylated by liver ornithine decarboxylase (ODC, EC 4.1.1.17) of thioacetamide-treated rats but were good competitive inhibitors of the enzyme (Ki ranged from 0.72 to 1.79 mM). When assayed in vivo in the treated rats, the above mentioned isoornithines were also found to inhibit liver ODC when administered 1 h before sacrifice. When the methylputrescines formally derived from the decarboxylation of several isoornithines were assayed on rat liver ODC, it was found that only 2,3-dimethylputrescine decreased the enzymatic activity. When assayed in vivo, it was found to decrease ODC activity by 60%, when the latter was measured 1 h after administration. The effect was reverted 4 h after administration of the drug. Isoornithines were not taken up by H-35 hepatoma cells; hence they did not affect their ODC activity. 2,3-Dimethylputrescine however, was transported into the cells and significantly decreased its ODC activity.

Topics: Animals; Binding, Competitive; Carcinoma, Hepatocellular; Enzyme Inhibitors; Liver; Liver Neoplasms; Ornithine; Ornithine Decarboxylase Inhibitors; Putrescine; Rats; Structure-Activity Relationship; Thioacetamide; Tumor Cells, Cultured

Out of 56 thioacetamide (TAA)-fed rats hepatocellular carcinoma were found in four instances after 300, 360, 450 and 495 days in the present experiment. Of the four tumours, two showed lung metastasis. The size of TAA-treated hepatocyte nucleoli increased greatly after several weeks of feeding. Two enzymes, succinate dehydrogenase and glucose-6-phosphatase were found in both cytoplasma and nucleoli of hepatocytes of which nucleolar localizations are quite new. In hepatocellular carcinoma, malignant cells lost glucose-6-phosphatase activity completely in cytoplasm and nucleoli though succinate dehydrogenase activity was present in these organelles.

Topics: Acetamides; Animals; Carcinoma, Hepatocellular; Glucose-6-Phosphatase; Liver Neoplasms; Male; Rats; Succinate Dehydrogenase; Thioacetamide

Topics: Animals; Carcinoma, Hepatocellular; Cell Nucleolus; Glucose-6-Phosphatase; Liver Neoplasms; Male; Rats; Succinate Dehydrogenase; Thioacetamide

As part of a continuing comparison of nuclear proteins of tumors and other tissues, 32P-labeled nuclear proteins were extracted successively with 0.15 and 0.35 m NaC1 from the nuclei of normal, regenerating, and thioacetamide-treated rat liver as well as Novikoff hepatoma 3 hr after injection of 32Pi into rats. Separation of proteins of these fractions with aqueous phenol was carried out before two-dimensional electrophoresis on polyacrylamide gels. By autoradiography many common spots were found, but four 32P-labeled protein spots, CU', C13p, C21p, and CMp, were found in the Novikoff hepatoma and not in the various liver samples studied. Two spots, B6 and B10, were found in the liver patterns and not in the tumor. Sot B33 was very dense in regenerating liver but was only a faint spot in thioacetamide-treated liver. The greater density of Spots CU', C13p, C21p, and CMp in the tumor patterns is consistent with the increased density reported earlier for spots of the C-region of a variety of tumors.

Topics: Animals; Autoradiography; Carcinoma, Hepatocellular; Cell Nucleus; Chromosomal Proteins, Non-Histone; Electrophoresis, Polyacrylamide Gel; Liver; Liver Neoplasms; Liver Regeneration; Male; Neoplasms, Experimental; Phosphoproteins; Rats; Thioacetamide

The activity of rRNA methylases was stimulated by high-energy precursors of RNA (ribonucleoside triphosphates) and inhibited by degradation products of RNA (ribonucleotides and oligoribonucleotides). The response of methylases from rat Novikoff ascites tumor and liver to these metabolites was strikingly different. The highly active tumor enzymes responded preferentially to inhibition by catabolic metabolites, whereas the less active liver enzymes responded exclusively to stimulation by anabolic metabolites. When the activity of rRNA methylases was assayed in response to increasing concentration of S-adenosylmethionine, the tumor enzymes responded with a hyperbolic substrate dependence curve and the liver enzymes with a sigmoidal curve. In the presence of an inhibitory dinucleotide, ApA, the tumor enzymes responded with a sigmoidal curve; in the presence of a stimulator, adenosine 5'-triphosphate, the liver enzymes responded with a hyperbolic substrate concentration curve. When normal rats were subject to a series of treatments by thioacetamide, a hepatocarcinogen, the liver nucleolar rRNA methylases became responsive to inhibition by ApA and relatively unresponsive to stimulation by adenosine 5'-triphosphate. When tumor-bearing rats were treated with polyinosinate:polycytidylate, an antitumor agent, the tumor nucleolar rRNA methylases became unresponsive to inhibition by ApA and more responsive to stimulation by adenosine 5'-triphosphate. A correlation was noted between increased methylation efficiency in vivo and increased stability of nucleolar RNA during incubation in vitro, or vice versa. These results are interpreted to indicate that rRNAmethylases are regulated by cellular metabolites during the nucleolar biosynthesis of ribosomes and that rRNA methylases may provide a favorable site for selective action by cancer chemotherapeutic agents.

Topics: Adenosine Triphosphate; Animals; Carcinoma, Hepatocellular; Cell Nucleolus; Hydrogen-Ion Concentration; Kinetics; Liver; Liver Neoplasms; Neoplasms, Experimental; Oligoribonucleotides; Orotic Acid; Rats; Ribonucleotides; Ribosomes; S-Adenosylmethionine; Thioacetamide; tRNA Methyltransferases

Six-week-old male Swiss mice were given 0.03% thioacetamide (TAA) in the diet 24, 72, and 168 hours after partial hepatectomy. TAA-treated mice from all three groups were killed when they were 4, 9, and 13 months old. Intact and partially hepatectomized animals on normal diets served as controls. None of the controls evidenced neoplasms at any age. All three experimental groups developed liver tumors earlier than did intact mice treated with the TAA diet. Progressive metabolic studies on the livers or tumor tissues of treated mice showed that the levels of glucose-6-phosphatase, fructose-1,6-diphosphatase, and glycogen decreased significantly in the 4-month-old treated group when there was no significant alteration in liver histology. These parameters were lowest in the tumor tissues of treated mice.

Topics: Acetamides; Age Factors; Animals; Carcinoma, Hepatocellular; Diet; Fructose-Bisphosphatase; Hepatectomy; Liver; Liver Glycogen; Liver Neoplasms; Male; Mice; Neoplasms, Experimental; Precancerous Conditions; Thioacetamide

Studies were made on the effects of alpha-amanitin, cycloheximide, and thioacetamide on synthesis and content of low molecular weight nuclear RNA. Cycloheximide, an inhibitor of protein synthesis and the synthesis of 45S pre-rRNA and 5S RNA, also inhibited synthesis of nuclear U1 and U3 RNAs. alpha-Amanitin, an inhibited the synthesis of U1 and U2 low molecular weight nuclear RNA. Thioacetamide, which induces nucleolar hypertrophy and increased nucleolar RNA polymerase activity, markedly increased synthesis of 5.8S RNA and U3 RNA. These results show that syntheses of individual low molecular weight nuclear (LMWN) RNAs are controlled by different regulatory mechanisms. In particular, there appears to be a specific relationship between U3 RNA and functional states of the nucleolus.

Topics: Acetamides; Amanitins; Animals; Carcinoma, Hepatocellular; Cell Nucleus; Cycloheximide; Dose-Response Relationship, Drug; Liver; Liver Neoplasms; Male; Molecular Weight; Neoplasms, Experimental; Phosphates; Rats; RNA; RNA, Neoplasm; Thioacetamide

Topics: Animals; Carcinoma 256, Walker; Carcinoma, Hepatocellular; Chromatin; Electrophoresis, Polyacrylamide Gel; Kidney; Liver; Liver Neoplasms; Liver Regeneration; Neoplasm Proteins; Neoplasms, Experimental; Nucleoproteins; Rats; Thioacetamide

In the tumorous phase of TAA-induced cirrhosis of the liver comparative studies on the mitotic index, on the frequency of binucleated liver cells and on nuclear volume showed approximately similar values in the small pseudolobuli and in several hepatomas. Obviously "minimum deviation hepatomas" had developed. Decrease of ploidy characteristic for hepatomas was observed. In thioacetamide hepatomas the phenomenon of quadrupling of the cell nuclei due to dysfunctional nuclear edema occurred whereas the normal nuclei of liver cells can only double in volume in all phases of thioacetamide-intoxication. Particular karyometric aspects of the nuclear-nucleolar metabolism of hepatoma cells following thioacetamide intoxication are discussed.

Topics: Animals; Carcinoma, Hepatocellular; Cell Nucleus; Karyometry; Liver Cirrhosis; Liver Neoplasms; Rats; Thioacetamide

Topics: Acetamides; Animals; Carcinoma 256, Walker; Carcinoma, Hepatocellular; Cell Nucleus; Citrates; Electrophoresis, Polyacrylamide Gel; Liver; Liver Neoplasms; Liver Regeneration; Male; Neoplasm Proteins; Neoplasms, Experimental; Proteins; Rats; Sulfuric Acids; Thioacetamide

Topics: Acetamides; Adenosine Triphosphate; Animals; Carcinogens; Carcinoma, Hepatocellular; Cell Nucleus; Centrifugation, Density Gradient; Cytoplasm; DNA, Neoplasm; Liver; Liver Neoplasms; Male; Proteins; Rats; RNA, Neoplasm; Temperature; Thioacetamide; Time Factors; Tritium

Topics: Acetamides; Aflatoxins; Animals; Biological Transport; Carcinogens; Carcinoma, Hepatocellular; Cell Nucleus; Cytoplasm; Dimethyl Sulfoxide; DNA, Neoplasm; Fluorenes; Liver; Liver Neoplasms; Male; Methane; Neoplasms, Experimental; Nitrosamines; Nucleic Acid Hybridization; p-Dimethylaminoazobenzene; Rats; RNA; RNA, Neoplasm; Structure-Activity Relationship; Thioacetamide; Transcription, Genetic

Topics: Carcinoma, Hepatocellular; Cell Nucleus; Diploidy; DNA; Hepatectomy; Humans; Karyotyping; Liver; Liver Neoplasms; Microscopy, Electron; Thioacetamide

Topics: Acetamides; Animals; Carcinoma, Hepatocellular; Liver; Liver Glycogen; Liver Neoplasms; Male; Mice; Mice, Inbred C3H; Thioacetamide

Topics: Animals; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; Endoplasmic Reticulum; Glycogen; Hyperplasia; Liver; Liver Cirrhosis; Liver Neoplasms; Male; Microscopy, Electron; Rats; Ribosomes; Thioacetamide; Time Factors; Water

Topics: Acetamides; Animals; Carcinoma 256, Walker; Carcinoma, Hepatocellular; Cell Nucleolus; Electrophoresis, Polyacrylamide Gel; Liver; Liver Neoplasms; Liver Regeneration; Neoplasm Proteins; Proteins; Rats; Thioacetamide

Topics: Animals; Biological Transport; Carbon Radioisotopes; Carcinoma, Hepatocellular; Cell Nucleus; Centrifugation, Density Gradient; Chromatin; Dose-Response Relationship, Drug; Fluorenes; Liver; Liver Neoplasms; Male; Microscopy, Electron; Orotic Acid; p-Dimethylaminoazobenzene; Phenotype; Rats; RNA; Thioacetamide; Time Factors

Topics: Acetamides; Animals; Carcinoma, Hepatocellular; Cell Nucleus; Chemical and Drug Induced Liver Injury; Hepatectomy; Liver; Liver Cirrhosis; Liver Neoplasms; Liver Regeneration; Mitosis; Neoplasms, Experimental; Precancerous Conditions; Rats; Thioacetamide