Aloperine protects human retinal pigment epithelial cells against hydrogen peroxide-induced oxidative stress and apoptosis through activation of Nrf2/HO-1 pathway

Abstract

Age-related macular degeneration (AMD) is a complex multifactorial disease associated with the dysfunction of retinal pigment epithelium (RPE). Aloperine is a quinolizidine alkaloid that has been proven to possess broad pharmacological activities. However, the effects of aloperine on AMD remain unclear. In the present study, we used hydrogen peroxide (H₂O₂) to induce oxidative injury in human RPE cells (ARPE-19 cells). ARPE-19 cells were pretreated with different concentrations of aloperine for 2 h, followed by H₂O₂ exposure. Cell cytotoxicity was determined using lactate dehydrogenase (LDH) release assay. Cell viability was measured using Cell Counting Kit-8 (CCK-8) assay. The reactive oxygen species (ROS) generation, malondialdehyde (MDA) level, superoxide dismutase (SOD) activity and glutathione peroxidase (GSH-PX) activity were detected to reflect oxidative status. Western blot was performed to detect the expressions of bcl-2, bax, nuclear factor-erythroid 2-related factor 2 (Nrf2), and heme oxygenase-1 (HO-1). The activity of caspase-3 was also assessed to indicate cell apoptosis. In addition, ARPE-19 cells were transfected with siNrf2 to knock down Nrf2. Our results showed that pretreatment with aloperine elevated the reduced cell viability of H₂O₂-induced ARPE-19 cells in a dose-dependent manner. Aloperine greatly decreased the production of ROS and MDA, and increased the activities of SOD and GSH-PX in H₂O₂-stimulated ARPE-19 cells. H₂O₂-caused a decrease in bcl-2 expression and increases in bax expression and caspase-3 activity were mitigated by aloperine. Moreover, aloperine treatment enhanced the expression levels of Nrf2 in nuclear fraction and the HO-1 expression in lysates. Knockdown of Nrf2 reversed the protective effects of aloperine on H₂O₂-induced ARPE-19 cells. In conclusion, these findings demonstrated that aloperine protected ARPE-19 cells from H₂O₂-induced oxidative stress and apoptosis in part via activating the Nrf2/HO-1 signaling pathway. The findings suggested a therapeutic potential of aloperine for the treatment of ADM.

Keywords: Age-related macular degeneration (AMD); Nrf2/HO-1 signaling pathway; aloperine; oxidative stress; retinal pigment epithelium (RPE).