Impact of grape seed proanthocyanidin B2 pretreatment on mitochondrial oxidative stress, endoplasmic reticulum stress, and apoptosis in renal tubular epithelial cells during in-vitro hypoxia-reoxygenation.

Abstract

Purpose: To investigate the impact of grape seed proanthocyanidin B2 (GSPB2) pretreatment on hypoxia-reoxygenation model of HK-2 cells in vitro.

Methods: The experiment was divided into five groups: control group (control), GSPB2 group (GSPB2), hypoxia-reoxygenation group (HR), GSPB2 + HR group (GSPB2+HR), and GSPB2 + brusatol (BRU) + HR group (GSPB2 + BRU + HR). Flow cytometry was used to detect apoptosis of cells. Transmission electron microscopy was employed to observe ultrastructural changes of cells. Mitochondrial membrane potential (MMP) was measured. Cellular immunofluorescence was used to assess intracellular Ca²+ concentration. Immunofluorescence staining and Western blotting were conducted to detect expression levels of Nrf2, HO-1, glucose-regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), and cleaved-caspase3.

Results: Compared to HR group, GSPB2 + HR group showed significantly increased cell viability, and reduced mitochondrial damage in the cytoplasm. MMP in GSPB2 + HR group was significantly restored, and intracellular Ca²+ concentration was significantly decreased. The expression of Nrf2 and HO-1 proteins was significantly upregulated, while the expression of GRP78, CHOP, and cleaved-caspase3 proteins was markedly downregulated.

Conclusion: GSPB2 pretreatment can alleviate oxidative stress damage, mitochondrial injury, and endoplasmic reticulum stress induced by hypoxia-reoxygenation in HK-2 cells in vitro. This effect may be related to the ability of GSPB2 pretreatment to activate the endogenous antioxidant system, particularly through the activation of the Nrf2/HO-1 signaling pathway.