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

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.