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Scientific Reports Jul 2018The 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...
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
PubMed: 29980757
DOI: 10.1038/s41598-018-28486-8 -
Mutation Research. Genetic Toxicology... Apr 2018Regulatory guidance documents stress the value of assessing multiple tissues and the most appropriate endpoints when evaluating chemicals for in vivo genotoxic...
Regulatory guidance documents stress the value of assessing multiple tissues and the most appropriate endpoints when evaluating chemicals for in vivo genotoxic potential. However, conducting several independent studies to consider multiple endpoints and/or tissue compartments is resource intensive. Furthermore, conventional approaches for scoring genotoxicity endpoints are slow, tedious, and less objective than what would be considered ideal. In an effort to address these issues with current practices, we attempted to i) employ flow cytometry-based methods to score liver micronuclei, blood micronuclei, and blood Pig-a gene mutation, and ii) integrate the endpoints into a common general toxicology study design-the rat 28-day repeat dose study. A proof-of-principle experiment was performed with 6-week old male Crl:CD(SD) rats exposed to diethylnitrosamine (DEN) for 28 consecutive days. One day later blood was collected for micronucleated reticulocyte (MN-RET) and Pig-a mutation assays, and liver tissue was obtained for micronucleated hepatocyte (MNHEP) scoring. MN-RET frequencies were not affected by DEN exposure, and mean Pig-a mutant cell frequencies were only slightly elevated. On the other hand, % MNHEP showed marked, dose-related increases (2.2, 7.2, and 9.1 mean fold-increase for 5, 10, 15 mg DEN/kg/day, respectively). Concurrent with MNHEP analyses, assessments of Ki-67-positive events and the proportion of 8n nuclei provided evidence for treatment-related changes to hepatocyte proliferation. Collectively, these results reinforce the importance of evaluating chemicals' genotoxic potential in liver in addition to hematopoietic cells, and suggest that several automated measurements can be successfully integrated into repeat-dose studies for higher efficiencies and better utilization of fewer animals.
Topics: Animals; Diethylnitrosamine; Dose-Response Relationship, Drug; Flow Cytometry; Hepatocytes; Male; Membrane Proteins; Micronucleus Tests; Mutagenicity Tests; Mutation; Rats, Sprague-Dawley; Reticulocytes
PubMed: 29555062
DOI: 10.1016/j.mrgentox.2018.02.005 -
Journal of Histotechnology Mar 2023The purpose of this study was to investigate the effect of combined therapy of diacerein and gold nanoparticles (AuNP) on diethylnitrosamine (DEN) induced hepatocellular...
Attenuation of diethylnitrosamine-induced hepatocellular carcinoma in a rat model by combination therapy of diacerein and gold nanoparticles: a histopathological and immunohistochemical study.
The purpose of this study was to investigate the effect of combined therapy of diacerein and gold nanoparticles (AuNP) on diethylnitrosamine (DEN) induced hepatocellular carcinoma (HCC) in a rat model. Normal healthy and DEN-induced (HCC) rats were divided into five groups. Group I healthy rats served as normal control, Group II untreated HCC rats, Group III HCC rats administered diacerein, Group IV HCC rats administered AuNP, and Group V HCC rats administered diacerein and AuNP. All treatments were given once daily for 4 weeks. Liver morphology and necroinflammation in all groups were evaluated using hematoxylin and eosin (H&E), Masson's trichrome for fibrosis, and immunohistochemistry assays for expression of TNF-α, IL-6, β-catenin, and caspase-3. Liver sections from Group II HCC rats showed loss of lobular architecture, thick fibrous tissue deposition, leukocyte infiltration, degenerated hepatocytes and HCC neoplastic nodules surrounded by extensive fibrosis. Group II had high expression of TNF-α, IL-6, and β-catenin, and low caspase-3 expression as compared to Group I. HCC rats treated with the combined therapy of diacerein and AuNP (Group V) showed markedly decreased HCC lesions, significant necroinflammation reduction (p ˂ 0.05) and 90% reduction in fibrosis as compared to Group II HCC + diacerein. This combined therapy also reduced (p ˂ 0.05) TNF-α, IL-6, β-catenin expression and increased caspase-3 expression. In conclusion, diacerein combined with AuNP synergistically attenuated the severity of HCC lesions by reducing necroinflammation and fibrosis, decreased TNF-α, IL-6, β-catenin expression, and increased caspase-3 expression for apoptosis.
Topics: Rats; Animals; Carcinoma, Hepatocellular; Caspase 3; Diethylnitrosamine; beta Catenin; Gold; Liver Neoplasms; Tumor Necrosis Factor-alpha; Interleukin-6; Metal Nanoparticles; Fibrosis
PubMed: 36214360
DOI: 10.1080/01478885.2022.2129935 -
Nature Communications Jan 2021Hepatocellular carcinoma (HCC) is the most predominant primary malignancy in the liver. Genotoxic and genetic models have revealed that HCC cells are derived from...
Hepatocellular carcinoma (HCC) is the most predominant primary malignancy in the liver. Genotoxic and genetic models have revealed that HCC cells are derived from hepatocytes, but where the critical region for tumor foci emergence is and how this transformation occurs are still unclear. Here, hyperpolyploidization of hepatocytes around the centrilobular (CL) region is demonstrated to be closely linked with the development of HCC cells after diethylnitrosamine treatment. We identify the CL region as a dominant lobule for accumulation of hyperpolyploid hepatocytes and preneoplastic tumor foci formation. We also demonstrate that upregulation of Aurkb plays a critical role in promoting hyperpolyploidization. Increase of AURKB phosphorylation is detected on the midbody during cytokinesis, causing abscission failure and hyperpolyploidization. Pharmacological inhibition of AURKB dramatically reduces nucleus size and tumor foci number surrounding the CL region in diethylnitrosamine-treated liver. Our work reveals an intimate molecular link between pathological hyperpolyploidy of CL hepatocytes and transformation into HCC cells.
Topics: Animals; Carcinoma, Hepatocellular; Cell Transformation, Neoplastic; Cells, Cultured; Diethylnitrosamine; Female; Hepatocytes; Humans; Liver; Liver Neoplasms; Male; Mice, Inbred C57BL; Mice, Inbred ICR; Mice, Inbred NOD; Mice, Knockout; Mice, SCID; Microscopy, Confocal; Polyploidy; Precancerous Conditions; Mice
PubMed: 33510150
DOI: 10.1038/s41467-020-20572-8 -
Journal of Toxicology and Environmental... 2018N-nitrosamines and their precursors found in cosmetics may be carcinogenic in humans. Thus the aim of this study was to carry out risk assessment for N-nitrosamines...
N-nitrosamines and their precursors found in cosmetics may be carcinogenic in humans. Thus the aim of this study was to carry out risk assessment for N-nitrosamines (N-nitrosodiethanolamine [NDELA], N-nitrosodiethylamine [NDEA]) and amines (triethanolamine [TEA], diethanolamine [DEA]) levels in cosmetics determined using validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) procedures. NDELA and NDEA concentrations were present at levels of "not detected" (N.D.) to 596.5 μg/kg and N.D. to 40.9 μg/kg, respectively. TEA and DEA concentrations ranged from N.D. to 860 μg/kg and N.D. to 26.22 μg/kg, respectively. The nitrite concentration (3-2250 mg/l), number of nitrosating agents to a maximum 5, and pH (3.93-10.09) were also assessed. The impact of N-nitrosamine formation on the levels of TEA, DEA, nitrite, and other nitrosating agents was also examined. N-nitrosamine concentrations correlated with the number of nitrosating agents and nitrite concentrations. Data demonstrated that higher nitrite concentrations and a greater number of nitrosating agents increased NDELA and NDEA yields. Further, the presence of TEA and DEA exerted a significant influence on N-nitrosamine formation. Risk assessments, including the margin of exposure (MOE) and lifetime cancer risk (LCR) for N-nitrosamines and margin of safety (MOS) for amines, were calculated using product type, use pattern, and concentrations. Exposure to maximum amounts of NDELA and NDEA resulted in MOE > 10,000 (based upon the benchmark dose lower confidence limit 10%) and LCR <1 × 10, respectively. In addition, TEA and DEA concentrations in cosmetic samples resulted in MOS values >100. Therefore, no apparent safety concerns were associated with cosmetic products containing NDELA, NDEA, TEA, and DEA in this study. However, since amines and nitrosating agents produce carcinogenic nitrosamines, their use in cosmetics needs to be minimized to levels as low as technically feasible.
Topics: Carcinogens; Chromatography, Liquid; Cluster Analysis; Cosmetics; Diethylnitrosamine; Ethanolamines; Multivariate Analysis; Nitrates; Nitrites; Risk Assessment; Tandem Mass Spectrometry
PubMed: 29694274
DOI: 10.1080/15287394.2018.1460782 -
International Journal of Medical... 2022Hepatocellular carcinoma (HCC) is a primary liver cancer commonly found in adults. Previously, we showed the anticancer effects of Thai herbal plant extract, Pierre...
Hepatocellular carcinoma (HCC) is a primary liver cancer commonly found in adults. Previously, we showed the anticancer effects of Thai herbal plant extract, Pierre (DM), in HCC-bearing rats. In the present study, we further examined the proposed mechanism of DM, including apoptosis and antioxidant activity. Moreover, we used RNA sequencing (RNA-seq) to analyze molecular pathways in the rat model in which HCC was induced by diethylnitrosamine (DEN) and thioacetamide (TAA). The HCC-bearing rats were then treated with 40 mg/kg of DM for 8 weeks, after which experimental and control rats were sacrificed and liver tissues were collected. The RNA-seq data of DEN/TAA-treated rats exhibited upregulation of 16 hallmark pathways, including epithelial mesenchymal transition, inflammatory responses, and angiogenesis (<0.01). DM extract expanded the Bax protein-positive pericentral zone in the tumor areas and decreased hepatic malondialdehyde levels, implying a decrease in lipid peroxidation in liver. However, DM treatment did not ameliorate the molecular pathways induced in DEN/TAA-treated livers. Our findings indicate that DM extract has antioxidant activity and exerts its pro-apoptotic effect on rat HCCs at the (post-)translational level.
Topics: Rats; Animals; Carcinoma, Hepatocellular; Thioacetamide; Diethylnitrosamine; Dioscorea; Antioxidants; Liver Neoplasms; Liver; Plant Extracts
PubMed: 36313224
DOI: 10.7150/ijms.72987 -
Human & Experimental Toxicology 2023Diethylnitrosamine (DEN), a hepatocarcinogen, is found in a variety of smoked and fried foods and was reported to be hepatotoxic in mice. Butylated hydroxytoluene (BHT)...
Diethylnitrosamine (DEN), a hepatocarcinogen, is found in a variety of smoked and fried foods and was reported to be hepatotoxic in mice. Butylated hydroxytoluene (BHT) is a potent antioxidant used in cosmetic formulations and as a food additive and preservative. As a result, BHT was studied as a potential inhibitor in the early stages of diethylnitrosamine (DEN)-induced HCC. Male Wistar albino rats ( = 24) were equally subdivided. Group 1 was the negative control; Group 2 and 3 administered BHT and DEN, respectively; Group 4 received BHT followed by DEN. Blood samples and rat livers were taken for biochemical and histological investigation. Hepatotoxicity was assessed by increased liver enzymes and HCC indicators, along with reduced antioxidant and pro-apoptotic factors. AFP, AFPL3, GPC3, GSH, SOD, MDA, CASP3 and BAX expression increased significantly after DEN treatment. DEN also reduced GPx, CAT, and CYP2E1 activity, and BCl-2 expression. Moreover, in the hepatic parenchyma, the DEN caused histological alterations. Pretreatment with BHT enhanced antioxidant status while preventing histopathological and most biochemical alterations. BHT pretreatment suppresses DEN-initiated HCC by decreasing oxidative stress, triggering intrinsic mitotic apoptosis, and preventing histopathological changes in liver tissue.
Topics: Rats; Male; Animals; Mice; Carcinoma, Hepatocellular; Antioxidants; Diethylnitrosamine; Butylated Hydroxytoluene; Liver Neoplasms; Rats, Wistar; Liver
PubMed: 36943693
DOI: 10.1177/09603271231165664 -
Scientific Reports Jul 2022Several fractions of Calotropis gigantea extracts have been proposed to have potential anticancer activity in many cancer models. The present study evaluated the...
Several fractions of Calotropis gigantea extracts have been proposed to have potential anticancer activity in many cancer models. The present study evaluated the anticancer activity of C. gigantea stem bark extracts in liver cancer HepG2 cells and diethylnitrosamine (DEN)-induced primary liver cancer in rats. The carcinogenesis model induced by DEN administration has been widely used to study pathophysiological features and responses in rats that are comparable to those seen in cancer patients. The dichloromethane (CGDCM), ethyl acetate, and water fractions obtained from partitioning crude ethanolic extract were quantitatively analyzed for several groups of secondary metabolites and calactin contents. A combination of C. gigantea stem bark extracts with doxorubicin (DOX) was assessed in this study to demonstrate the enhanced cytotoxic effect to cancer compared to the single administration. The combination of DOX and CGDCM, which had the most potential cytotoxic effect in HepG2 cells when compared to the other three fractions, significantly increased cytotoxicity through the apoptotic effect with increased caspase-3 expression. This combination treatment also reduced ATP levels, implying a correlation between ATP and apoptosis induction. In a rat model of DEN-induced liver cancer, treatment with DOX, C. gigantea at low (CGDCM-L) and high (CGDCM-H) doses, and DOX + CGDCM-H for 4 weeks decreased the progression of liver cancer by lowering the liver weight/body weight ratio and the occurrence of liver hyperplastic nodules, fibrosis, and proliferative cells. The therapeutic applications lowered TNF-α, IL-6, TGF-β, and α-SMA inflammatory cytokines in a similar way, implying that CGDCM had a curative effect against the inflammation-induced liver carcinogenesis produced by DEN exposure. Furthermore, CGDCM and DOX therapy decreased ATP and fatty acid synthesis in rat liver cancer, which was correlated with apoptosis inhibition. CGDCM reduced cleaved caspase-3 expression in liver cancer rats when used alone or in combination with DOX, implying that apoptosis-inducing hepatic carcinogenesis was suppressed. Our results also verified the low toxicity of CGDCM injection on the internal organs of rats. Thus, this research clearly demonstrated a promising, novel anticancer approach that could be applied in future clinical studies of CGDCM and combination therapy.
Topics: Adenosine Triphosphate; Animals; Calotropis; Carcinogenesis; Caspase 3; Diethylnitrosamine; Doxorubicin; Liver; Liver Neoplasms; Plant Bark; Plant Extracts; Rats
PubMed: 35840761
DOI: 10.1038/s41598-022-16321-0 -
Proceedings of the National Academy of... Mar 2021The CXC chemokine receptor type 4 (CXCR4) receptor and its ligand, CXCL12, are overexpressed in various cancers and mediate tumor progression and hypoxia-mediated...
The CXC chemokine receptor type 4 (CXCR4) receptor and its ligand, CXCL12, are overexpressed in various cancers and mediate tumor progression and hypoxia-mediated resistance to cancer therapy. While CXCR4 antagonists have potential anticancer effects when combined with conventional anticancer drugs, their poor potency against CXCL12/CXCR4 downstream signaling pathways and systemic toxicity had precluded clinical application. Herein, BPRCX807, known as a safe, selective, and potent CXCR4 antagonist, has been designed and experimentally realized. In in vitro and in vivo hepatocellular carcinoma mouse models it can significantly suppress primary tumor growth, prevent distant metastasis/cell migration, reduce angiogenesis, and normalize the immunosuppressive tumor microenvironment by reducing tumor-associated macrophages (TAMs) infiltration, reprogramming TAMs toward an immunostimulatory phenotype and promoting cytotoxic T cell infiltration into tumor. Although BPRCX807 treatment alone prolongs overall survival as effectively as both marketed sorafenib and anti-PD-1, it could synergize with either of them in combination therapy to further extend life expectancy and suppress distant metastasis more significantly.
Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Carcinoma, Hepatocellular; Cell Line, Tumor; Diethylnitrosamine; Drug Synergism; Humans; Liver Neoplasms; Liver Neoplasms, Experimental; Lymphocytes, Tumor-Infiltrating; Male; Mice; Molecular Docking Simulation; Rats; Receptors, CXCR4; Signal Transduction; Sorafenib; T-Lymphocytes, Cytotoxic; Tumor Microenvironment; Tumor-Associated Macrophages; Xenograft Model Antitumor Assays
PubMed: 33753481
DOI: 10.1073/pnas.2015433118 -
European Review For Medical and... Jan 2021To study the association of inflammatory factors and hepatocarcinoma stem cells of induced liver cancer rats.
OBJECTIVE
To study the association of inflammatory factors and hepatocarcinoma stem cells of induced liver cancer rats.
MATERIALS AND METHODS
30 SD male healthy rats were selected. 10 rats were given water as normal control group. 10 rats only were implemented laparotomy as sham operation group. The remaining 10 rats were the liver cancer model group and treated with diethylnitrosamine (DEN) to induce liver cancer. Real-time quantitative PCR was used to detect the related inflammatory factors in HCC tissues, including interleukin-6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), transforming growth factor-β1 (TGF-β), human interleukin-1α (IL-1α), human interleukin 1β (IL-1β) and levels of hepatocarcinoma stem cells indicators CD90, CD133, Alpha-fetoprotein (AFP). Correlation analysis was used to analyze the correlation between inflammatory factors and hepatocarcinoma stem cells markers CD90 and CD133.
RESULTS
The expression levels of IL-6, MCP-1 and TGF-β of HCC tissues in liver cancer model group were significantly higher than those in the control group and the sham operation group. The expression levels of CD90 and CD133 of tissues in the liver cancer model group were significantly higher than those in the control group and the sham operation group. The differences were statistically significant (p<0.001). By inhibiting related inflammatory factors, the growth, migration and invasion of liver cancer cells were significantly inhibited, and apoptosis was promoted. Correlation analysis results showed that the expression changes of IL-6, MCP-1 and TGF-β were significantly positively correlated with CD90 up-regulation (p<0.05), while the expression changes of IL-6, MCP-1 and TGF-β were significantly positively correlated with CD133 up-regulation (p<0.05).
CONCLUSIONS
The inflammatory factors IL-6, MCP-1 and TGF-β are closely related to hepatocarcinoma stem cells, which play an important role in promoting the occurrence and deterioration of liver cancer.
Topics: Administration, Oral; Animals; Apoptosis; Biomarkers, Tumor; Cell Proliferation; Chemokine CCL2; Diethylnitrosamine; Disease Models, Animal; Interleukin-6; Liver Neoplasms; Male; Neoplastic Stem Cells; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta1
PubMed: 33577025
DOI: 10.26355/eurrev_202101_24632