Standardization of diethylnitrosamine-induced hepatocellular carcinoma rat model with time based molecular assessment

• Published in: Experimental and molecular pathology 2021-12, Vol.123, p.104715-104715, Article 104715
• Main Authors: Ghufran, Hafiz, Azam, Maryam, Mehmood, Azra, Butt, Hira, Riazuddin, Sheikh
• Format: Article
• Language: English
• Subjects: Animals; Apoptosis - genetics; Carcinogenesis - genetics; Carcinoma, Hepatocellular - chemically induced; Carcinoma, Hepatocellular - genetics; Carcinoma, Hepatocellular - pathology; Cell Proliferation - genetics; Cytokines; Diethylnitrosamine; Diethylnitrosamine - toxicity; Disease Models, Animal; Fibrosis - chemically induced; Fibrosis - genetics; Fibrosis - pathology; Gene Expression Regulation, Neoplastic; Hepatocellular carcinoma; Humans; Inflammation - chemically induced; Inflammation - genetics; Inflammation - pathology; Liver - drug effects; Liver - pathology; Liver Cirrhosis - chemically induced; Liver Cirrhosis - genetics; Liver Cirrhosis - pathology; Liver Neoplasms - chemically induced; Liver Neoplasms - genetics; Liver Neoplasms - pathology; Molecular markers; Neoplasm Proteins - genetics; Neovascularization, Pathologic - chemically induced; Neovascularization, Pathologic - genetics; Neovascularization, Pathologic - pathology; Pathogenesis; Rat model; Rats
• ISSN: 0014-4800; 1096-0945
• DOI: 10.1016/j.yexmp.2021.104715

This study was intended (1) to develop a robust animal model for hepatocellular carcinoma (HCC) research, in which HCC tumors develop in a background of fibrosis or cirrhosis; and (2) to explore time-dependent regulatory changes in key molecular markers during disease advancement and HCC development. With the aim of establishing such HCC model, male Sprague-Dawley rats were injected with diethylnitrosamine (DEN) at a dose of 30 mg/kg twice a week for 10 weeks then once a week from 12th to 16th weeks. The rats were kept under observation until 18th week. At defined time intervals (2nd, 4th, 12th, and 18th week), serum biomarkers and microscopic components of tissue samples were used to investigate the chronic progression of liver disease, while gene and protein analysis was used to monitor expression patterns during HCC development. DEN-intoxicated rats manifested inflammation at week 4, fibrosis at week 12 and cirrhosis with early HCC tumors at week 18. Molecular analysis revealed that key markers of inflammation (Il-1β, Il-6, and Tnf-α), fibrosis (Tgf-β1, Col1α1, Col3α1, and Timp-1), and angiogenesis (Hif1-α and Vegf) were promptly (P ≤ 0.001) up-regulated at week 4, week 12 and week 18, respectively. Oxidative stress (iNos, Cyp2e1, and Sod1) and pro-apoptotic (Bax) markers showed significant upregulation from week 4 to week 12. However, Sod1 and Bax expressions dropped after week 12 and reached a minimum at 18th week. Strikingly, expressions of anti-apoptotic (Bcl-2) and cell proliferation (Pcna, Hgf, and Afp) markers were abruptly increased at week 18. Collectively, we describe an 18-week HCC model in DEN-intoxicated rats that exhibit chronic inflammation, oxidative imbalance, advance fibrosis/cirrhosis, halted apoptosis, and angiogenic sprouting, progressively. •DEN intoxication leads to HCC through a background of advanced fibrosis, or cirrhosis.•Metabolism of DEN in lobular hepatocytes causes imbalance in pro- and anti-oxidants.•Cancerous features develop due to imbalance between cell growth and death.•DEN intoxication dysregulates angiogenic pathways that lead to hypervascular HCC tumors.