13 research outputs found
Hepatocyte specific GP73/GOLM1 knockout mice and human hepatocellular carcinoma cell lines provide insights into the potential roles of GP73 HCC serum biomarker
No full textGolgi Protein -- 73kDa (GP73/GOLM1) is a resident cis-Golgi Type II membrane protein that is nearly undetectable in normal hepatocytes, but its expression increases in hepatocytes during hepatocellular carcinoma (HCC) and its serum levels increase in patients with chronic hepatitis C infection and HCC, yet its biological role in the liver and in the pathogenesis of HCC remains unclear. In order to investigate possible biological roles for GP73, hepatocyte-specific GOLM1 null mice (C57BL/6) were created using the Cre-loxP system, with floxed GOLM1 gene exon 3 and Cre recombinase driven by the albumin promoter. Mouse genotypes were confirmed by PCR analysis. GOLM1Fl/Fl/Cre(+) animals showed no obvious biological phenotype compared to their Cre(-) littermates. They exhibited normal growth, behaviors, and mated successfully, suggesting that hepatic GP73 is not vital for normal physiological function. Consistent with clinical observations in humans, GP73 expression increased in a chemically induced model of mouse liver fibrosis and carcinogenesis. Examination of GP73 expression in 14 human HCC cell lines revealed that GP73 is consistently expressed in mesenchymal HCC cells (modeling metastatic) but not in most epithelial HCC cell lines. Cell invasion assays indicated that mesenchymal HCC with high GP73 expression were more invasive suggesting a potential role for this protein in metastasis. Silencing in invasive mesenchymal HCC cells suggested that GP73 is necessary for enhanced cell proliferation and over-expression in a non-invasive epithelial HCC cell line suggested that GP73 is sufficient to induce epithelial to mesenchymal transition (EMT) and metastasis. Collectively the results suggest GP73 will serve as a potential therapeutic target for treatment of advanced HCC and that the hepatocyte-specific GP73 knockout mice will serve as a valuable tool to investigate the role of GP73 in HCC development
Abstract 5248: Pioglitazone prevents hepatocellular carcinoma development in a rat model of cirrhosis
No full textAbstract
Introduction: Advanced hepatocellular carcinoma (HCC) is a leading cause of mortality worldwide with limited treatment options. There is a readily identifiable cohort of cirrhosis patients at risk and they are ideal candidates for chemoprevention. Anti-hyperglycemic agents have garnered interest for their potential anti-fibrotic as well as chemo-preventive effects. Pioglitazone, a selective PPAR-γ agonist, has been shown to reduce non-alcoholic steatohepatitis (NASH), but its role as an anti-fibrotic and chemopreventive agent has yet to be elucidated. The hypothesis of this study is that Pioglitazone reduces cirrhosis and subsequent HCC development in rats with diethylnitrosamine (DEN)-induced cirrhosis.
Methods: Male Wistar received DEN 50mg/kg by intraperitoneal injection. DEN injury reliably recapitulates histological and molecular features of human HCC development with induction of hepatic fibrosis at 8 weeks, cirrhosis at 12 weeks, and HCC by 18 weeks. DEN-injured rats were randomized to receive oral gavage of pioglitazone at 3mg/kg/day (n=9) or vehicle control (n=9). Initiation of pioglitazone coincided with the development of liver fibrosis at 8 weeks. All animals were sacrificed at 18 weeks.
Results: As expected, repeated injections of DEN in rats resulted in progressive fibrosis, cirrhosis, followed by HCC formation. Treatment with pioglitazone resulted in a 56% reduction of surface nodules relative to treatment with vehicle (7.4±4.9 vs. 17±7; p<0.005). Liver sections were stained by picrosirius red to assess fibrosis and pioglitazone significantly reduced collagen deposition in DEN-injured rats (collagen proportional area = 3.2±1.8% vs. 9.2±2%; p<0.035). This histologic observation was further confirmed by gene expression analysis with reductions in collagen-I, α-smooth muscle actin, and transforming growth factor beta in rats treated with pioglitazone. Finally, weekly injection of DEN also caused a significant decrease in overall body weight in comparison to untreated rats (398.1±60 vs. 598±46 grams; p<0.015), and pioglitazone treatment resulted in a trend for better protection of body weight relative to vehicle (398.1±60 vs. 427.5±56.3 grams).
Conclusion: Overall our data supports the hypothesis that the anti-diabetic agent pioglitazone may be repurposed as a drug to reduce fibrosis and prevent HCC. This could be beneficiary in patient management given the low cost as well as minimal side effects.
Citation Format: Shen Li, Sarani Ghoshal, Gunisha Arora, Derek J. Erstad, Michael Lanuti, Kenneth K. Tanabe, Bryan C. Fuchs. Pioglitazone prevents hepatocellular carcinoma development in a rat model of cirrhosis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5248. doi:10.1158/1538-7445.AM2017-5248</jats:p
The farnesoid x receptor agonist edp-305 reduces interstitial renal fibrosis in a mouse model of unilateral ureteral obstruction.
No full textFarnesoid X receptor (FXR) is a nuclear receptor that has emerged as a key regulator in the maintenance ofhepatic steatosis, inflammation, and fibrosis. However, the role of FXR in renal fibrosis remains to be established.Here, we investigate the effects of the FXR agonist EDP-305 in a mouse model of tubulointerstitial fibrosis viaunilateral ureteral obstruction (UUO). Male C57Bl/6 mice received a UUO on their left kidney. On postoperative d 4,mice received daily treatment by oral gavage with either vehicle control (0.5% methylcellulose) or 10 or 30 mg/kgEDP-305. All animals were euthanized on postoperative d 12. EDP-305 dose-dependently decreased macrophageinfiltration as measured by the F4/80 staining area and proinflammatory cytokine gene expression. EDP-305 alsodose-dependently reduced interstitial fibrosis as assessed by morphometric quantification of the collagen pro-portional area and kidney hydroxyproline levels. Finally, yes-associated protein (YAP) activation, a major driver offibrosis, increased after UUO injury and was diminished by EDP-305 treatment. Consistently, EDP-305 decreasedTGF-b1–induced YAP nuclear localization in human kidney 2 cells by increasing inhibitory YAP phosphorylation.YAP inhibition may be a novel antifibrotic mechanism of FXR agonism, and EDP-305 could be used to treat renalfibrosis.—Li, S., Ghoshal, S., Sojoodi, M., Arora, G., Masia, R., Erstad, D. J., Ferriera, D. S., Li, Y., Wang, G., Lanuti,M., Caravan, P., Or, Y. S., Jiang, L.-J., Tanabe, K. K., Fuchs, B. C. The farnesoid X receptor agonist EDP-305 reducesinterstitial renal fibrosis in a mouse model of unilateral ureteral obstruction
Abstract 1323: Liver-specific knockout mice and liver-derived cell lines provide insights into potential roles for the GP73/GOLM1 HCC serum biomarker - association with sustained cell proliferation
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Abstract 4004: The H2 receptor antagonist nizatidine inhibits carcinogenesis in two rodent models of hepatocellular carcinoma
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SAMHD1 knockout hiPSC model enables high lentiviral transduction efficiency in myeloid cell types
No full textRecent advances in functional genomics tools have ushered in a new era of genetic editing to identify molecular pathways relevant to developmental and disease biology. However, limited model systems are available that adequately mimic cell states and phenotypes associated with human disease pathways. Here, we quantitatively analyzed the founder population bottleneck effect and demonstrated how the population changes from human induced pluripotent stem cells (hiPSCs) to hematopoietic stem cells and to the final induced macrophage population. We then engineered a key gene encoding an enzyme in the myeloid cell antiviral pathway-SAMHD1-knockout (KO) hiPSCs and characterized the hiPSC line with RNA-Seq and induced macrophages from two distinct protocols with functional analysis. We then generated SAMHD1 KO CRISPR-dCAS9 KRAB hiPSCs through lentiviral transduction aiming to increase the efficiency of lentiviral mediated gene transfer. We demonstrated increased lentiviral transduction efficiency in induced macrophages, as well as microglia induced with two distinct protocols. This model allows for efficient gene knockdown, as well as large-scale functional genomics screens in mature hiPSC-derived macrophages or microglia with applications in innate immunity and chronic inflammatory disease biology. These experiments highlight the broad applicability of this platform for disease-relevant target identification and may improve our ability to run large-scale screens in hiPSC-derived myeloid model systems
Simultaneous capture of single cell RNA-seq, ATAC-seq, and CRISPR perturbation enables multiomic screens to identify gene regulatory relationships
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Hepatocellular carcinoma chemoprevention by targeting the angiotensin-converting enzyme and EGFR transactivation
No full textHepatocellular carcinoma (HCC) is a leading cause of death among cirrhotic patients, for which chemopreventive strategies are lacking. Recently, we developed a simple human cell-based system modeling a clinical prognostic liver signature (PLS) predicting liver disease progression and HCC risk. In a previous study, we applied our cell-based system for drug discovery and identified captopril, an approved angiotensin converting enzyme (ACE) inhibitor, as a candidate compound for HCC chemoprevention. Here, we explored ACE as a therapeutic target for HCC chemoprevention. Captopril reduced liver fibrosis and effectively prevented liver disease progression toward HCC development in a diethylnitrosamine (DEN) rat cirrhosis model and a diet-based rat model for nonalcoholic steatohepatitis–induced (NASH-induced) hepatocarcinogenesis. RNA-Seq analysis of cirrhotic rat liver tissues uncovered that captopril suppressed the expression of pathways mediating fibrogenesis, inflammation, and carcinogenesis, including epidermal growth factor receptor (EGFR) signaling. Mechanistic data in liver disease models uncovered a cross-activation of the EGFR pathway by angiotensin. Corroborating the clinical translatability of the approach, captopril significantly reversed the HCC high-risk status of the PLS in liver tissues of patients with advanced fibrosis. Captopril effectively prevents fibrotic liver disease progression toward HCC development in preclinical models and is a generic and safe candidate drug for HCC chemoprevention
