Mechanisms of the chemopreventive effect of pequi oil on in vitro hepatocarcinogenesis

Author(s)

Simone Morais Palmeira

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina (FM/SBD)
• Defense date: 2022-10-20

Examining board members

Francisco Javier Hernandez Blazquez; Thiago Pinheiro Arrais Aloia; Durvanei Augusto Maria

• Advisor: Francisco Javier Hernandez Blazquez

Abstract

Pequi oil (Caryocar brasiliense, Camb) inhibits the development of hepatocarcinogenesis in mice, but the mechanisms of this effect are still unknown. It is important to understand how the oil acts in carcinogenesis chemoprevention, as this fruit, native to Brazil, has promising use in the prevention of liver cancer. Therefore, in this study, we sought to identify in vitro the mechanisms by which pequi oil inhibits hepatocarcinogenesis. To evaluate the possible oil mechanisms, an in vitro liver carcinogenesis model was established. The cell line AML12 and the carcinogen aflatoxin B1(AFB1) were used in the model. Cells were exposed to AFB1 (1M), pequi oil (5.62 g/mL) in combination with AFB1 (OP+AFB1) and vehicle control (0.01% DMSO) for 40 cycles, each cycle being 72h). Subcultures of cells treated with AFB1 and OP+AFB1 for 40 cycles were characterized according to the gain in phenotype or inhibition of the malignant phenotype and the following changes were evaluated: changes in growth by quantifying the proliferative index by immunostaining with BrdU and cell cycle distribution by immunoblotting; changes in cell death were evaluated by double-labeled apoptosis assay with annexin V and propidium iodide dyes analyzed by flow cytometry; and evaluation of oxidative stress was by measuring reactive oxygen species (ROS), the fluorescent dye DCFDA and immunostaining by fluorescence of antioxidant defense proteins were used. Furthermore, we evaluated the effect of pequi oil on oxidative stress and the antigenotoxic effect on HepG2 cells by quantification of reactive oxygen species (ROS) and a comet assay to determine DNA damage. As a result, we found that pequi oil reduces the generation of intracellular ROS in AML12 cells exposed to AFB1. It is possible that oil acts on endogenous antioxidant defenses that are responsible for the removal of ROS. Pequi oil also decreased ROS generation and reduced DNA strand breaks in HepG2 cells exposed to hydrogen peroxide (H2O2), strengthening its effect on reactive oxygen species control and indicating its antigenotoxic effect by inhibiting oxidative damage to the DNA of HepG2 cells exposed to hydrogen peroxide. Therefore, we conclude that C. brasiliense oil acts chemopreventively in the inhibition of oxidative stress and in the protection of genetic material against oxidative damage from hepatocytes by reducing ROS and DNA strand breaks, and it can potentially be applied in the prevention of liver cancer (AU)

• FAPESP's process: 16/01149-6 - Mechanisms of the chemopreventive activity of pequi oil (Caryocar brasiliense Camb) in hepatocarcinogenesis and its potential antineoplastic effect on HepG2 cell line
• Grantee: Simone Morais Palmeira
• Support Opportunities: Scholarships in Brazil - Doctorate