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Revista Espanola de Enfermedades... Nov 2019We have read with great interest the study by Vera MC et al. on the intrasplenic transplantation of hepatocytes from donors receiving the carcinogenic drugs...
We have read with great interest the study by Vera MC et al. on the intrasplenic transplantation of hepatocytes from donors receiving the carcinogenic drugs diethylnitrosamine (DEN) and 2-actylaminofluorene (2-AAF). The most important conclusion of the study is that there were no signs of tumor progression in the recipient liver at seven and 60 days after hepatocellular transplantation and no differences were found in the animals with healthy hepatocyte transplants.
Topics: 2-Acetylaminofluorene; Animals; Diethylnitrosamine; Hepatocytes; Liver; Rats; Rats, Wistar
PubMed: 31663360
DOI: 10.17235/reed.2019.6270/2019 -
Stem Cell Research & Therapy Oct 2022Hepatic fibrosis is a common pathologic stage in chronic liver disease development, which might ultimately lead to liver cirrhosis. Accumulating evidence suggests that...
ADSCs-derived exosomes ameliorate hepatic fibrosis by suppressing stellate cell activation and remodeling hepatocellular glutamine synthetase-mediated glutamine and ammonia homeostasis.
BACKGROUND
Hepatic fibrosis is a common pathologic stage in chronic liver disease development, which might ultimately lead to liver cirrhosis. Accumulating evidence suggests that adipose-derived stromal cells (ADSCs)-based therapies show excellent therapeutic potential in liver injury disease owing to its superior properties, including tissue repair ability and immunomodulation effect. However, cell-based therapy still limits to several problems, such as engraftment efficiency and immunoreaction, which impede the ADSCs-based therapeutics development. So, ADSCs-derived extracellular vesicles (EVs), especially for exosomes (ADSC-EXO), emerge as a promise cell-free therapeutics to ameliorate liver fibrosis. The effect and underlying mechanisms of ADSC-EXO in liver fibrosis remains blurred.
METHODS
Hepatic fibrosis murine model was established by intraperitoneal sequential injecting the diethylnitrosamine (DEN) for two weeks and then carbon tetrachloride (CCl) for six weeks. Subsequently, hepatic fibrosis mice were administrated with ADSC-EXO (10 μg/g) or PBS through tail vein infusion for three times in two weeks. To evaluate the anti-fibrotic capacity of ADSC-EXO, we detected liver morphology by histopathological examination, ECM deposition by serology test and Sirius Red staining, profibrogenic markers by qRT-PCR assay. LX-2 cells treated with TGF-β (10 ng/ml) for 12 h were conducted for evaluating ADSC-EXO effect on activated hepatic stellate cells (HSCs). RNA-seq was performed for further analysis of the underlying regulatory mechanisms of ADSC-EXO in liver fibrosis.
RESULTS
In this study, we obtained isolated ADSCs, collected and separated ADSCs-derived exosomes. We found that ADSC-EXO treatment could efficiently ameliorate DEN/CCl-induced hepatic fibrosis by improving mice liver function and lessening hepatic ECM deposition. Moreover, ADSC-EXO intervention could reverse profibrogenic phenotypes both in vivo and in vitro, including HSCs activation depressed and profibrogenic markers inhibition. Additionally, RNA-seq analysis further determined that decreased glutamine synthetase (Glul) of perivenous hepatocytes in hepatic fibrosis mice could be dramatically up-regulated by ADSC-EXO treatment; meanwhile, glutamine and ammonia metabolism-associated key enzyme OAT was up-regulated and GLS2 was down-regulated by ADSC-EXO treatment in mice liver. In addition, glutamine synthetase inhibitor would erase ADSC-EXO therapeutic effect on hepatic fibrosis.
CONCLUSIONS
These findings demonstrated that ADSC-derived exosomes could efficiently alleviate hepatic fibrosis by suppressing HSCs activation and remodeling glutamine and ammonia metabolism mediated by hepatocellular glutamine synthetase, which might be a novel and promising anti-fibrotic therapeutics for hepatic fibrosis disease.
Topics: Ammonia; Animals; Carbon Tetrachloride; Carcinoma, Hepatocellular; Diethylnitrosamine; Exosomes; Fibrosis; Glutamate-Ammonia Ligase; Glutamine; Hepatic Stellate Cells; Homeostasis; Liver Cirrhosis; Liver Neoplasms; Mice; Stromal Cells; Transforming Growth Factor beta
PubMed: 36195966
DOI: 10.1186/s13287-022-03049-x -
Nature Communications Sep 2021Although psycho-social stress is a well-known factor that contributes to the development of cancer, it remains largely unclear whether and how environmental eustress...
Although psycho-social stress is a well-known factor that contributes to the development of cancer, it remains largely unclear whether and how environmental eustress influences malignant diseases and regulates cancer-related therapeutic responses. Using an established eustress model, we demonstrate that mice living in an enriched environment (EE) are protected from carcinogen-induced liver neoplasia and transplantable syngeneic liver tumors, owning to a CD8 T cell-dependent tumor control. We identify a peripheral Neuro-Endocrine-Immune pathway in eustress, including Sympathetic nervous system (SNS)/β-adrenergic receptors (β-ARs)/CCL2 that relieves tumor immunosuppression and overcomes PD-L1 resistance to immunotherapy. Notably, EE activates peripheral SNS and β-ARs signaling in tumor cells and tumor infiltrated myeloid cells, leading to suppression of CCL2 expression and activation of anti-tumor immunity. Either blockade of CCL2/CCR2 or β-AR signaling in EE mice lose the tumor protection capability. Our study reveales that environmental eustress via EE stimulates anti-tumor immunity, resulting in more efficient tumor control and a better outcome of immunotherapy.
Topics: Animals; B7-H1 Antigen; Carbon Tetrachloride; Chemokine CCL2; Diethylnitrosamine; Drug Resistance, Neoplasm; Hepatic Stellate Cells; Hepatocytes; Humans; Immune Checkpoint Inhibitors; Liver; Liver Neoplasms, Experimental; Male; Mice; Neuroimmunomodulation; Organoids; Receptors, Adrenergic, beta; Receptors, CCR2; Signal Transduction; Stress, Psychological; Sympathetic Nervous System; Tumor Escape; Tumor Microenvironment
PubMed: 34593796
DOI: 10.1038/s41467-021-25967-9 -
Current Protocols in Pharmacology Sep 2014Human hepatocellular carcinoma (HCC) develops most often as a complication of fibrosis or cirrhosis. While most human studies of HCC provide crucial insights into the...
Human hepatocellular carcinoma (HCC) develops most often as a complication of fibrosis or cirrhosis. While most human studies of HCC provide crucial insights into the molecular signatures of HCC, seldom do they address the etiology of HCC. Mouse models are essential tools for investigating the pathogenesis of HCC; however, the overwhelming majority of cancer models in rodents do not feature liver fibrosis. Detailed in this unit is a protocol for an experimental mouse model of HCC that arises in association with advanced liver fibrosis. The disease model is induced by a single injection of N-nitrosodiethylamine (DEN) followed by repeated administration of carbon tetrachloride (CCl4 ). A dramatic potentiation of liver tumor incidence is observed following administration of DEN and CCl4 , with 100% of mice developing liver tumors at 5 months of age. This model can be employed for studying the molecular mechanisms of fibrogenesis and HCC development, and in cancer hazard/chemotherapy testing of drug candidates.
Topics: Animals; Carbon Tetrachloride; Carcinogens; Carcinoma, Hepatocellular; Chemical and Drug Induced Liver Injury; Diethylnitrosamine; Disease Models, Animal; Liver; Liver Cirrhosis; Liver Neoplasms, Experimental; Male; Mice
PubMed: 25181010
DOI: 10.1002/0471141755.ph1430s66 -
Cellular and Molecular Gastroenterology... 2021Cancer-associated fibroblasts (CAFs) play a key role in the cancer process, but the research progress is hampered by the paucity of preclinical models that are essential...
BACKGROUND & AIMS
Cancer-associated fibroblasts (CAFs) play a key role in the cancer process, but the research progress is hampered by the paucity of preclinical models that are essential for mechanistic dissection of cancer cell-CAF interactions. Here, we aimed to establish 3-dimensional (3D) organotypic co-cultures of primary liver tumor-derived organoids with CAFs, and to understand their interactions and the response to treatment.
METHODS
Liver tumor organoids and CAFs were cultured from murine and human primary liver tumors. 3D co-culture models of tumor organoids with CAFs and Transwell culture systems were established in vitro. A xenograft model was used to investigate the cell-cell interactions in vivo. Gene expression analysis of CAF markers in our hepatocellular carcinoma cohort and an online liver cancer database indicated the clinical relevance of CAFs.
RESULTS
To functionally investigate the interactions of liver cancer cells with CAFs, we successfully established murine and human 3D co-culture models of liver tumor organoids with CAFs. CAFs promoted tumor organoid growth in co-culture with direct cell-cell contact and in a Transwell system via paracrine signaling. Vice versa, cancer cells secrete paracrine factors regulating CAF physiology. Co-transplantation of CAFs with liver tumor organoids of mouse or human origin promoted tumor growth in xenograft models. Moreover, tumor organoids conferred resistance to clinically used anticancer drugs including sorafenib, regorafenib, and 5-fluorouracil in the presence of CAFs, or the conditioned medium of CAFs.
CONCLUSIONS
We successfully established murine and human 3D co-culture models and have shown robust effects of CAFs in liver cancer nurturing and treatment resistance.
Topics: Animals; Antineoplastic Agents; Cancer-Associated Fibroblasts; Carcinoma, Hepatocellular; Coculture Techniques; Culture Media, Conditioned; Diethylnitrosamine; Drug Resistance, Neoplasm; Humans; Liver Neoplasms; Liver Neoplasms, Experimental; Mice; Organoids; Paracrine Communication; Primary Cell Culture; Tumor Cells, Cultured; Tumor Microenvironment; Xenograft Model Antitumor Assays
PubMed: 32932015
DOI: 10.1016/j.jcmgh.2020.09.003 -
Arhiv Za Higijenu Rada I Toksikologiju Mar 2021In 2018, some sartan medicinal products were reported to be contaminated with nitrosamine compounds, which are potent mutagenic carcinogens. Two nitrosamines received... (Review)
Review
In 2018, some sartan medicinal products were reported to be contaminated with nitrosamine compounds, which are potent mutagenic carcinogens. Two nitrosamines received particular attention: -nitrosodimethylamine (NDMA) and -nitrosodiethylamine (NDEA). These have since been confirmed in different types of medicinal products, including ranitidine and metformin. Consequently, the European Medicines Agency (EMA) started an investigation into the cause of contamination and an assessment of the risk to patients taking contaminated medicinal products. The main source of contamination were changes in production, which involves combinations of amines and nitrogen compounds and the use of specific catalysts and reagents. Withdrawals of medicinal products that took place in Croatia did not lead to a shortage of sartan- or metformin-containing medicines. Moreover, ranitidine had been preventively withdrawn all over the EU, including Croatia, creating shortages at the time, but was subsequently replaced with therapeutic alternatives.
Topics: Carcinogens; Diethylnitrosamine; Dimethylnitrosamine; Drug Contamination; Humans; Nitrosamines
PubMed: 33787187
DOI: 10.2478/aiht-2021-72-3491 -
Cellular and Molecular Gastroenterology... 2023Phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme of the de novo serine synthesis pathway (SSP), has been implicated in the carcinogenesis and metastasis...
BACKGROUND & AIMS
Phosphoglycerate dehydrogenase (PHGDH), the rate-limiting enzyme of the de novo serine synthesis pathway (SSP), has been implicated in the carcinogenesis and metastasis of hepatocellular carcinoma (HCC) because of its excessive expression and promotion of SSP. In previous experiments we found that SSP flux was diminished by knockdown of zinc finger E-box binding homeobox 1 (ZEB1), a stimulator of HCC metastasis, but the underlying mechanism remains largely unknown. Here, we aimed to determine how SSP flux is regulated by ZEB1 and the contribution of such regulation to carcinogenesis and progression of HCC.
METHODS
We used genetic mice with Zeb1 knockout in liver specifically to determine whether Zeb1 deficiency impacts HCC induced by the carcinogen diethylnitrosamine plus CCl. We explored the regulatory mechanism of ZEB1 in SSP flux using uniformly-labeled [C]-glucose tracing analyses, liquid chromatography-mass spectrometry, real-time quantitative polymerase chain reaction, luciferase report assay, and chromatin immunoprecipitation assay. We determined the contribution of the ZEB1-PHGDH regulatory axis to carcinogenesis and metastasis of HCC by cell counting assay, methyl thiazolyl tetrazolium (MTT) assay, scratch wound assay, Transwell assay, and soft agar assay in vitro, orthotopic xenograft, bioluminescence, and H&E assays in vivo. We investigated the clinical relevance of ZEB1 and PHGDH by analyzing publicly available data sets and 48 pairs of HCC clinical specimens.
RESULTS
We identified that ZEB1 activates PHGDH transcription by binding to a nonclassic binding site within its promoter region. Up-regulated PHGDH augments SSP flux to enable HCC cells to be more invasive, proliferative, and resistant to reactive oxygen species and sorafenib. Orthotopic xenograft and bioluminescence assays have shown that ZEB1 deficiency significantly impairs the tumorigenesis and metastasis of HCC, and such impairments can be rescued to a large extent by exogenous expression of PHGDH. These results were confirmed by the observation that conditional knockout of ZEB1 in mouse liver dramatically impedes carcinogenesis and progression of HCC induced by diethylnitrosamine/CCl, as well as PHGDH expression. In addition, analysis of The Cancer Genome Atlas database and clinical HCC samples showed that the ZEB1-PHGDH regulatory axis predicts poor prognosis of HCC.
CONCLUSIONS
ZEB1 plays a crucial role in stimulating carcinogenesis and progression of HCC by activating PHGDH transcription and subsequent SSP flux, deepening our knowledge of ZEB1 as a transcriptional factor in fostering the development of HCC via reprogramming the metabolic pathway.
Topics: Humans; Animals; Mice; Carcinoma, Hepatocellular; Liver Neoplasms; Phosphoglycerate Dehydrogenase; Diethylnitrosamine; Cell Line, Tumor; Carcinogenesis; Zinc Finger E-box-Binding Homeobox 1
PubMed: 37331567
DOI: 10.1016/j.jcmgh.2023.06.006 -
Advances in Experimental Medicine and... 2015Alcohol is a well-established risk factor for hepatocellular carcinoma, and the mechanisms by which alcohol liver cancer is complex. It has been suggested that ethanol... (Review)
Review
Alcohol is a well-established risk factor for hepatocellular carcinoma, and the mechanisms by which alcohol liver cancer is complex. It has been suggested that ethanol (EtOH) metabolism may enhance tumor progression by increasing hepatocyte proliferation. To test this hypothesis, ethanol (EtOH) feeding of male mice began 7 weeks post-injection of the chemical carcinogen diethylnitrosamine (DEN), and continued for 16 weeks, with a final EtOH concentration of 28% of total calories. As expected, EtOH increased the total number of cancerous foci and liver tumors identified in situ fixed livers from the EtOH+DEN group compared to corresponding pair-fed (PF)+DEN and chow+DEN control groups. In the EtOH+DEN group, tumor multiplicity corresponded to a 3- to 4-fold increase in proliferation and immunohistochemical staining of β-catenin in non-tumorigenic hepatocytes when compared to the PF+DEN and chow+DEN groups, p<0.05. Analysis of EtOH-treated livers from a previously published rat model of chronic liver disease revealed increases in hepatocyte proliferation accompanied by a hepatic depletion of retinol and retinoic acid stores (p<0.05), nuclear accumulation of β-catenin (p<0.05), increased cytosolic expression p-GSK3β (p<0.05), significant upregulation of soluble Wnts, Wnt2, and Wnt7a, and increased expression of several β-catenin targets involved in tumor promotion and progression, cyclin D1, c-myc, WISP1, and MMP7 (p<0.05). These data suggest that chronic EtOH consumption activates the Wnt/β-catenin signaling pathway, which increases hepatocyte proliferation thus promoting tumorigenesis following an initiating insult in the liver.
Topics: Alcohol Drinking; Animals; Diethylnitrosamine; Liver Neoplasms; Male; Mice; Mice, Inbred C57BL; Wnt Signaling Pathway; beta Catenin
PubMed: 25427908
DOI: 10.1007/978-3-319-09614-8_11 -
Gene Expression Aug 2018Activation of the Wnt/β-catenin signaling is reported in large subsets of hepatocellular carcinoma (HCC). Upregulation of Wnt genes is one contributing mechanism. In...
Activation of the Wnt/β-catenin signaling is reported in large subsets of hepatocellular carcinoma (HCC). Upregulation of Wnt genes is one contributing mechanism. In the current study, we sought to address the role of hepatocyte-derived Wnts in a model of hepatic injury, fibrosis, and carcinogenesis. We subjected hepatocyte-specific Wntless knockout mice (HP-KO), unable to secrete Wnts from hepatocytes, and littermate controls (HP-CON) to diethylnitrosamine and carbon tetrachloride (DEN/CCl4) and harvested at 3, 5, and 6 months for histological and molecular analysis. Analysis at 5 months displayed increased hepatic expression of several Wnts and upregulation of some, but not all, β-catenin targets, without mutations in Ctnnb1. At 5 months, HP-CON and HP-KO had comparable tumor burden and injury; however, HP-KO uniquely showed small CK19+ foci within tumors. At 6 months, both groups were moribund with comparable tumor burden and CK19 positivity. While HCC histology was indistinguishable between the groups, HP-KO exhibited increased active β-catenin and decreased c-Myc, Brd4, E-cadherin, and others. Hepatic injury, inflammation, and fibrosis were also indistinguishable at 3 months between both groups. Thus, lack of Wnt secretion from hepatocytes did not affect overall injury, fibrosis, or HCC burden, although there were protein expression differences in the tumors occurring in the two groups.
Topics: Animals; Cadherins; Carbon Tetrachloride; Carcinoma, Hepatocellular; Diethylnitrosamine; Hepatocytes; Liver Neoplasms; Male; Mice; Nuclear Proteins; Proto-Oncogene Proteins c-myc; Transcription Factors; Wnt Proteins; beta Catenin
PubMed: 29519268
DOI: 10.3727/105221618X15205148413587 -
Hepatology Communications Dec 2022Liver cancer, comprised primarily of hepatocellular carcinoma (HCC), is the third leading cause of cancer deaths worldwide and increasing in Western countries. We...
Liver cancer, comprised primarily of hepatocellular carcinoma (HCC), is the third leading cause of cancer deaths worldwide and increasing in Western countries. We previously identified the transcription factor zinc fingers and homeoboxes 2 (Zhx2) as a regulator of hepatic gene expression, and many Zhx2 target genes are dysregulated in HCC. Here, we investigate HCC in Zhx2-deficient mice using the diethylnitrosamine (DEN)-induced liver tumor model. Our study using whole-body Zhx2 knockout (Zhx2 ) mice revealed the complete absence of liver tumors 9 and 10 months after DEN exposure. Analysis soon after DEN treatment showed no differences in expression of the DEN bioactivating enzyme cytochrome P450 2E1 (CYP2E1) and DNA polymerase delta 2, or in the numbers of phosphorylated histone variant H2AX foci between Zhx2 and wild-type (Zhx2 ) mice. The absence of Zhx2, therefore, did not alter DEN bioactivation or DNA damage. Zhx2 livers showed fewer positive foci for Ki67 staining and reduced interleukin-6 and AKT serine/threonine kinase 2 expression compared with Zhx2 livers, suggesting that Zhx2 loss reduces liver cell proliferation and may account for reduced tumor formation. Tumors were reduced but not absent in DEN-treated liver-specific Zhx2 knockout mice, suggesting that Zhx2 acts in both hepatocytes and nonparenchymal cells to inhibit tumor formation. Analysis of data from the Cancer Genome Atlas and Clinical Proteomic Tumor Consortium indicated that ZHX2 messenger RNA and protein levels were significantly higher in patients with HCC and associated with clinical pathological parameters. Conclusion: In contrast to previous studies in human hepatoma cell lines and other HCC mouse models showing that Zhx2 acts as a tumor suppressor, our data indicate that Zhx2 acts as an oncogene in the DEN-induced HCC model and is consistent with the higher ZHX2 expression in patients with HCC.
Topics: Animals; Humans; Mice; Carcinoma, Hepatocellular; Diethylnitrosamine; Genes, Homeobox; Homeodomain Proteins; Liver Neoplasms; Mice, Inbred C57BL; Mice, Knockout; Proteomics; Transcription Factors; Zinc Fingers
PubMed: 36194180
DOI: 10.1002/hep4.2106