Effects of Wasabi (Wasabia japonica) ingestion on fibrosis-associated hepatocarcinogenesis in mice

• Grant number: 23/01362-5
• Support Opportunities: Scholarships in Brazil - Scientific Initiation
• Start date: May 01, 2023
• End date: February 29, 2024
• Field of knowledge: Health Sciences - Medicine - Pathological Anatomy and Clinical Pathology
• Principal Investigator: Guilherme Ribeiro Romualdo
• Grantee: Taís Carolina Paes Bronzato
• Host Institution: Faculdade de Medicina (FMB). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil

Abstract

Hepatocellular Carcinoma (HCC) associated with liver fibrosis/cirrhosis is the third type of cancer with the highest mortality worldwide, with an increasing incidence in eastern countries. Due to its poor prognosis, preventive strategies for this malignancy are encouraged, including modulation of the hepatocarcinogenesis process by nutraceuticals. The rhizome of wasabi (Wasabia japonica (Miq.) Matsum) - a vegetable of the Brassicaceae family - is widely used as an ingredient in Japanese culinary. The rhizome is rich in bioactive compounds, including allyl isothiocyanate (AITC), considered the most abundant compound. Although other less abundant wasabi isothiocyanates have already demonstrated inhibitory effects on HCC cells, the effects of this in natura food in in vivo models are unknown. Thus, we will evaluate whether exposure to wasabi or AITC attenuates the promotion step of fibrosis-associated hepatocarcinogenesis in animals. Thus, male C57BL6/J mice will be submitted to a chemically induced model, initiated by applications of diethylnitrosamine (DEN) genotoxic carcinogen, and promoted by multiple injections of the thioacetamide (TAA) fibrogenic agent. Concomitanty to TAA promotion (6 weeks), the animals will be exposed by intragastric administration (5×/week) to wasabi rhizome paste (50 or 100 mg/kg body weight, Minato Wasabi, LTDA, Brazil) or AITC (dose to be defined). The AITC dose will be proportional to the amount of this compound in the wasabi paste, which will be determined by gas chromatography coupled to mass spectrometry (GC/MS). At the end of the protocol, the animals will be euthanized, liver samples will be collected for histological analysis: (1) counting and morphometry of pre-neoplastic lesions (altered hepatocyte foci) in hematoxylin and eosin-stained slides; (2) morphometric analysis of collagen in Sirius red-stained slides; (3) immunohistochemistry for CD68 (macrophages), ±-SMA (hepatic stellate cells) and Ki-67 (hepatocyte proliferation) markers; biochemical analysis: (4) determination of malondialdehyde levels (oxidative stress) and antioxidant enzymes superoxide dismutase and catalase, and molecular analyses: (5) immunoblot for antioxidant Nrf2 and pro-inflammatory p65.The project will allow the comparison between the potential effects of the main bioactive compound, AITC, with those of the in natura food.