The effect and underlying mechanism of Timosaponin B-II on RGC-5 necroptosis induced by hydrogen peroxide

Abstract

BackgroundNecroptosis is an important mode of cell death, which is due to oxidant stress accumulation. Our previous study indicated that oxidant stresses could be reduced by Timosaponin B-II (TBII), a kind of Chinese herb RhizomaAnemarrhenae monomer extraction. We wonder the possible effect of Timosaponin B-II, whether it can protect cells from necroptosis via reducing the oxidant stress, in RGC-5 following hydrogen peroxide (H2O2) insult.MethodsRGC-5 cells were grown in DMEM, the model group was exposed in H2O2 with the concentration of 300 μM, and the experimental group was pre-treated with Timosaponin B-II at different concentrations (1 μM, 10 μM, 100 μM and 1000 μM) for 24 hrs. MTT assay was carried out to measure the cytotoxicity of H2O2, MDA concentration assay was executed to evaluate the degree of oxidative stress, TNF-α ELISA Assay was used to measure the concentration of TNF-α, finally, the degree of necrosis were analyzed using flow cytometry.ResultsWe first constructed the cell injury model of necroptosis in RGC-5 upon H2O2 exposure. Morphological observation and MTT assay were used to evaluate the degree of RGC-5 death. MDA assay were carried out to describe the degree of oxidant stress. Annexin V/PI staining was used to detect necroptotic cells pre-treated with or without Timosaponin B-II following H2O2 injury. TNF-α ELISA was carried out to detect the TNF-α accumulation in RGC-5. Upon using Timosaponin B-II with concentration of 100 μM, the percentage of cell viability was increased from 50% to 75%, and the necrosis of cells was reduced from 35% to 20% comparing with H2O2 injury group. Oxidant stress and TNF-α was reduced upon injury which decreased the ratio of RGC-5 necroptosis.ConclusionOur study found out that Timosaponin B-II might reduce necroptosis via inhibition of ROS and TNF-α accumulation in RGC-5 following H2O2 injury.

Pin1 Promotes Regulated Necrosis Induced by Glutamate in Rat Retinal Neurons via CAST/Calpain2 Pathway

“…Dysregulation of regulated necrosis contributes to the development of diseases, such as cancer, and neurodegeneration ( Galluzzi et al, 2014 ; Pasparakis and Vandenabeele, 2015 ). Although regulated necrosis includes several cell death types, such as, receptor interacting protein 3-dependent necroptosis that can be triggered by tumor necrosis factor-α ( Degterev et al, 2008 ; Jiang et al, 2014 ), erastin-induced ferroptosis that requires iron, mitochondrial permeability transition dependent regulated necrosis, etc. ( Gao et al, 2015 ; Kong et al, 2017 ).…”

Receptor interacting protein 3-induced RGC-5 cell necroptosis following oxygen glucose deprivation

“…Rosenbaum et al [ 6 ] found a number of necroptotic cells in the rat retinal ganglion cell layer at 6 h after aHIOP. Our previous studies also indicated that necroptosis occurred in RGC-5 cells at an early stage following elevated hydrostatic pressure in vitro [ 15 ] or 300 μM hydrogen peroxide (H 2 O 2 ) treatment [ 14 ]. Recently, in vitro models of OGD have been widely applied to simulate neuronal ischemia in vivo.…”

“…Son et al [ 13 ] showed that Nec-1 can significantly reduce ROS production in Theiler’s murine encephalomyelitis virus (TMEV)-infected macrophages. Our previous studies showed that Timosaponin B-II, an anti-oxidative monomer extracted from Rhizoma anemarrhenae , reduced retinal ganglion cell-5 (RGC-5) cell necroptosis by inhibiting ROS accumulation [ 14 ]. Together, these results suggested that ROS accumulation might directly lead to necrosis.…”

Protective effects of timosaponin BII on oxidative stress damage in PC12 cells based on metabolomics

“…It is extracted from Anemarrhena asphodeloides rhizome and shows a very good antioxidant effect (Feng, Chen, Lin, & Liu, ). Many studies have shown that timosaponin BII not only has an effect on scavenging oxygen free radicals, and that it can inhibit the secretion of inflammatory cytokines and shows an obvious anti‐inflammatory effect (Jiang et al, ; Kim et al, ). In addition, T. J. Li, Qiu, Yang, Rui, and Chen () found that timosaponin BII can aid repair of impairment of learning and memory caused by cerebral ischemia through anti‐oxidation.…”

“…BII not only has an effect on scavenging oxygen free radicals, and that it can inhibit the secretion of inflammatory cytokines and shows an obvious anti-inflammatory effect (Jiang et al, 2014;Kim et al, 2009). In addition, T. J.…”