ANTIOXIDANT EFFECT OF SPINACH EXTRACT IN LIVER FIBROGENESIS ASSOCIATED TO ACTIVATION OF NRF2/HO-1 IN HYPERGLYCEMIC RATS

Abstract

Introduction and Objectives: In chronic hyperglycemia, increased oxidative stress plays an essential role in liver fibrogenesis. The spinach phytochemicals are shown to possess chemopreventive properties in this condition. Erythroid nuclear transcription factor 2 (Nrf2) is a transcriptional regulator expressing cytoprotective genes such as heme oxygenase-1 (ho-1). HO-1 is an enzyme present in the liver with antioxidant, anti-inflammatory, and anti-apoptotic capacities. However, the beneficial effect of spinach associated with endogenous activation of the Nrf2/HO-1 pathway is unknown. Objective: To evaluate the antioxidant effect of the spinach extract on injury hepatic associated with the Nrf2/HO-1 pathway in hyperglycemic rats. Materials and methods: The study was approved by the ethics committee from the ENCB-IPN (CONBIOETICA/09/CEI/002/20190327). We used hyperglycemic Wistar rats induced with 60 mg / kg of streptozotocin (HG; n = 7); intragastrical-treated HG with 400mg / kg of spinach methanolic extract (HG-EME; n = 7) and normoglycemic rats (NG; n = 7). Histological sections were obtained at 12 weeks and evaluated by Sirius red staining and immunohistochemistry (Nrf2, HO-1). The oxidative damage by lipid peroxidation was determined by the tissues' malonaldehyde formation (MDA) levels. The H-score system quantified the percentage of nuclear staining of Nrf2 and the intensities of Sirius red and HO-1. Statistical analysis: The data were analyzed by one-way ANOVA with the Tukey test using the GraphPad Prism program (Version 5.0). Kruskal-Wallis and Dunn's test established the MDA training levels. Statistical significance was considered with p <0.05. Results: Collagen fibers were formed mainly in the centrilobular zone. The percentage of collagen fibers staining in the HG group was 50 ± 10, compared to HG-EME (30 ± 10 p <0.05). In NG, the percentage of fibers was 20 ± 10. The nuclear Nrf2 (nNrf2) and cytoplasmic HO-1 staining were localized in the hepatocytes of region 3 Rappaport. The percentage of nNrf2 in HG was 30 ± 10, compared to HG-EME (70 ± 10 p <0.01). In NG, the staining percentage for nNrf2 was 15 ± 5. For HO-1 staining, the values in the HG-EME group were higher than in the HG group (p <0.01). Contrary, MDA levels in HG-EME decreased significantly compared to HG (p <0.05). Conclusions: Treatment with EME in hyperglycemic rats showed decreased liver damage generated by oxidative stress, associated with endogenous activation of the Nrf2/HO-1 pathway. The evaluation of liver biochemical tests can demonstrate a beneficial effect of EME. This work was supported by federal resources (HJM0713/19-I) and partially by INCICH.The authors declare that there is no conflict of interest.