Diabetic retinopathy is driven by oxidative stress-mitochondrial damage. Activation of ROS producing cytosolic NADPH oxidase 2 (Nox2) in diabetes precedes retinal mitochondrial damage, initiating a vicious cycle of free radicals. Elevated ROS levels peroxidize membrane lipids increasing damaging lipid peroxides (LPOs). While glutathione peroxidase 4 (GPx4) neutralizes LPOs, an imbalance in its generation-neutralization leads to ferroptosis, which is characterized by increased LPOs, free iron and decreased GPx4 activity. Mitochondria are rich in polyunsaturated fatty acids and iron and have mitochondrial isoform of GPx4. Our aim was to investigate mitochondrial ferroptosis in diabetic retinopathy, focusing on Nox2 mediated ROS production. Using human retinal endothelial cells, incubated in 5 mM or 20 mM D-glucose for 12-96 h, with or without Nox2 inhibitors (100 μM apocynin, 5 μM EHop-016 or 5 μM Gp91 ds-tat), or ferroptosis inhibitors (1 μM ferrostatin-1, 50 μM deferoxamine) or activator (0.1 μM RSL3), cytosolic and mitochondrial ROS, LPOs, iron, GPx4 activity, mitochondrial integrity (membrane permeability, oxygen consumption rate, mtDNA copy numbers) and cell death were quantified. High glucose significantly increased ROS, LPOs and iron levels and inhibited GPx4 activity in cytosol, and while Nox2 and ferroptosis inhibitors prevented glucose-induced increase in ferroptosis markers, mitochondrial damage and cell death, RSL3, further worsened them. Furthermore, high glucose also increased ferroptosis markers in the mitochondria, which followed their increase in the cytosol, suggesting a role of cytosolic ROS in mitochondrial ferroptosis. Thus, targeting Nox2-ferroptosis should help break down the self-perpetuating vicious cycle of free radicals, initiated by the damaged mitochondria, and could provide novel therapeutics to prevent/retard the development of diabetic retinopathy.

2024-10-14·Vascular Biology

Whole-body insulin resistance leads to accelerated atherosclerosis: role for Nox2 NADPH oxidase

Article

作者: Maqbool, Azhar ; Sukumar, Piruthivi ; Bridge, Katherine ; Kearney, Mark T ; Yuldasheva, Nadira Yusupovna ; Cubbon, Richard M ; Wheatcroft, Stephen B ; Imrie, Helen ; Muminov, Shovkat Kadirovich ; Shawer, Heba ; Makava, Natallia ; Skromna, Anna ; Griffin, Kathryn ; Viswambharan, Hema

Insulin resistance underpins the progression of type 2 diabetes mellitus and leads to a collection of risk factors for the development of atherosclerosis. Whether or not insulin resistance at a whole-body level per se leads to accelerated atherosclerosis is unclear. To answer this question, we generated atherosclerosis-prone mice with whole-body insulin resistance secondary to haploinsufficiency of the insulin receptor (IR+/−) deficient in ApoE−/− (IR+/−/ApoE−/−). IR+/−/ApoE−/− and ApoE−/− littermates had similar weight, lipids, and glucose tolerance at baseline. After 12 weeks of Western high-cholesterol diet, IR+/−/ApoE−/− had significantly more atherosclerosis in the thoracoabdominal aorta and at the level of the aortic sinus than ApoE−/− littermates. Excess Nox2 NADPH oxidase (Nox2) derived superoxide has been suggested to underpin diabetes-related atherosclerosis. In IR+/−/ApoE−/− we examined the effect of inhibiting Nox2 using genetic or pharmacological approaches on the development of atherosclerosis. To genetically delete Nox2, we generated IR+/−/ApoE−/−/Nox2−/y and to inhibit Nox2 pharmacologically, we treated IR+/−/ApoE−/− with the peptide Nox2 inhibitor gp91dstat. IR+/−/ApoE−/−/Nox2−/y had significant disruption of the aortic wall with increased thoracoabdominal atherosclerosis when compared to IR+/−/ApoE−/−/Nox2+/y littermates. Inhibition of Nox2 using gp91dstat reduced atherosclerosis in the thoracoabdominal aorta of IR+/−/ApoE−/−. Whole-body insulin resistance accelerates the development of atherosclerosis. Genetic inhibition of Nox2 leads to disruption of the aortic wall in IR+/−/ApoE−/− mice with accelerated atherosclerosis, whereas pharmacological Nox2 inhibition reduces atherosclerosis in IR+/−/ApoE−/− without disruption of the arterial wall.

2024-02-01·Hypertension

Prenatal Nicotine Exposure Raises Male Blood Pressure via FTO-Mediated NOX2/ROS Signaling

Article

作者: Jiang, Siyi ; Liu, Bailin ; Li, Yong ; Zhang, Lubo ; Xiao, Daliao ; Xu, Zhice ; Xia, Liang ; Shao, Xuesi M ; Yu, Wansu

BACKGROUND:Cigarette smoking/nicotine exposure in pregnancy shows an increased risk of hypertension in offspring, but the mechanisms are unclear. This study tested the hypothesis that m6A RNA hypomethylation epigenetically regulates vascular NOX (NADPH oxidase) and reactive oxygen species production, contributing to the fetal programming of a hypertensive phenotype in nicotine-exposed offspring.METHODS:Pregnant rats were exposed to episodic chronic intermittent nicotine aerosol (CINA) or saline aerosol control from gestational day 4 to day 21, and experiments were performed in 6-month–old adult offspring.RESULTS:Antenatal CINA exposure augmented Ang II (angiotensin II)–stimulated blood pressure response in male, but not female offspring. Moreover, CINA increased vascular NOX2 expression and superoxide production exclusively in male offspring. Inhibition of NOX2 with gp91ds-tat, both ex vivo and in vivo, mitigated the CINA-induced elevation in superoxide production and blood pressure response. Notably, CINA enhanced the expression of vascular m6A demethylase FTO (fat mass and obesity-associated protein), while reducing the total vascular m6A abundance and specific m6A methylation of the NOX2 gene. Additionally, ex vivo inhibition of FTO with FB23-2 attenuated CINA-induced increases in vascular NOX2 expression. In vitro experiments using human umbilical vein endothelial cells demonstrated that nicotine dose-dependently upregulated FTO and NOX2 protein abundance, which were reversed by treatment with the FTO inhibitor FB23-2 or FTO knockdown using siRNAs.CONCLUSIONS:This study uncovers a new mechanism: m6A demethylase FTO-mediated epigenetic upregulation of vascular NOX2 signaling in CINA-induced hypertensive phenotype. This insight could lead to a therapeutic target for preventing and treating developmental hypertension programming.

100 项与 gp91ds-Tat 相关的药物交易

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