Effects of Carboxymethylcellulose and Polysorbate 80 food additive on Nonalcoholic Fatty Liver Disease-associated hepatocarcinogenesis

• Grant number: 22/16633-1
• Support Opportunities: Regular Research Grants
• Start date: April 01, 2023
• End date: March 31, 2026
• Field of knowledge: Health Sciences - Medicine - Pathological Anatomy and Clinical Pathology
• Principal Investigator: Luís Fernando Barbisan
• Grantee: Luís Fernando Barbisan
• Host Institution: Instituto de Biociências (IBB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil
• Associated researchers: Bruno Cogliati ; Daniel Rodrigues Cardoso ; Maria Angel Garcia Chaves

Abstract

Hepatocellular carcinoma (HCC) features as the 6th/3rd most incident and deadliest, respectively, among all kinds of cancers worldwide, and it has been linked to chronic liver diseases like non-alcoholic fatty liver disease (NAFLD). NAFLD affects ~25% of the global population and contributes to increasing the HCC-related incidence by 26%. Adherence to a western diet (WD) - mostly composed of saturated fatty acids, sugars, and food additives - has been targeted as a driver of NAFLD, also increasing the risk of developing HCC by 80%. Ultra-processed food (UPF) is characterized by its hypercaloric and food additive-enriched profile, of which carboxymethylcellulose (CMC) and polysorbate 80 (P80) show potential effects on the liver-gut-adipose axis and prospecting as potential drivers of HCC. We sought to assess the effects of CMC and/or P80 on both in vivo and in vitro models of HCC-associated NAFLD. Male C57BL/6 mice will receive intraperitoneal injections of diethylnitrosamine [DEN, 25mg/Kg of body weight (b.w.), 1x/week, for 4 weeks] or vehicle, starting on the 14th post-natal day. At the 6th week of life, mice also received a WD (hypercaloric chow: 30% of saturated fat/20% of sucrose/0.2% of cholesterol; and a high-sugar solution: 55/45% of d-frcutose/d-glucose, for drinking) or a basal diet, for further 24 weeks. Simultaneously, mice received intragastric injections of CMC (370.0 or 740.0 mg/Kg of b.w., 5x/week, for 24 weeks) and/or P80 (100.0 or 200.0 mg/Kg of b.w., 5x/week, for 24 weeks) and were euthanized at the end of 24th week of the protocol. Neoplastic and hepatic (histopathological, lipid/collagen content, Ki67, cleaved caspase-3, and ±-smooth muscle actin immunoreactions, metabolomic, and mRNAseq), adipose tissue (histopathological and CD68 immunoreaction), small intestine (zonula occludens-1 and occludin immunoreaction, toluidine blue, and mucin density), feces (microbiome profiling), and serum (glucose tolerance test, alanine aminotransferases, total cholesterol, and triglycerides levels) samples will be further assessed. In a co-culture spheroid model (C3A e LX-2 cells) exposed to a fatty acid-supplemented medium, cell viability, lipid/collagen content, will be assessed. Additionally, based on our findings of mRNA sequencing, a western blotting strategy will be performed in spheroids for further validation of the in vivo findings. Data will be analyzed by one-way ANOVA or Kruskal-Wallis and Tukey pos hoc test and will be considered statistically different when p<0.05. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

ROMUALDO, GUILHERME RIBEIRO; HEIDOR, RENATO; BACIL, GABRIEL PRATA; MORENO, FERNANDO SALVADOR; BARBISAN, LUIS FERNANDO. Past, present, and future of chemically induced hepatocarcinogenesis rodent models: Perspectives concerning classic and new cancer hallmarks. Life Sciences, v. 330, p. 16-pg., 2023-10-01. (23/08751-7, 22/16633-1, 22/13402-9, 22/06082-8)

VALENTE, LETICIA CARDOSO; BACIL, GABRIEL PRATA; RIECHELMANN-CASARIN, LUANA; BARBOSA, GIULLIA CAVICHIOLLI; BARBISAN, LUIS FERNANDO; ROMUALDO, GUILHERME RIBEIRO. Exploring in vitro modeling in hepatocarcinogenesis research: morphological and molecular features and similarities to the corresponding human disease. Life Sciences, v. 351, p. 13-pg., 2024-06-13. (22/13402-9, 23/05411-0, 23/08751-7, 22/16633-1, 23/17585-3)