Functional Regulation of an Oxidative Stress Mediator, Rac1, in Diabetic Retinopathy
Early activation of cytosolic NADPH oxidase-2 (Nox2) in diabetes increases retinal ROS production, damaging their mitochondria. The assembly of Nox2 holoenzyme requires activation of a small molecular weight G protein Rac1. Rac1 activation is regulated by guanine exchange factors and guanine nucleotide-dissociation inhibitors, and post-translational modifications assist in its association with exchange factors and dissociation inhibitors. The goal of this study is to investigate the mechanisms of Rac1 activation in the development of diabetic retinopathy.
The levels of the dissociation inhibitor, prenylating enzyme (farnesyltransferase, FNTA), and exchange factor Vav2 were quantified in human retinal endothelial cells, incubated in normal or high glucose for 96 h. The roles of prenylation and Vav2 in Rac1-Nox2-ROS mitochondrial damage were confirmed in FNTA-siRNA–transfected cells and using the Vav2 inhibitor EHop, respectively. Retinal histopathology and functional changes associated with diabetic retinopathy were analyzed in diabetic mice receiving EHop for 6 months. Key parameters of Rac1 activation were confirmed in the retinal microvasculature from human donors with diabetic retinopathy.
In HRECs, glucose increased FNTA and Vav2 and decreased the dissociation inhibitor. FNTA-siRNA and EHop inhibited glucose-induced activation of Rac1–Nox2–ROS signaling. In diabetic mice, EHop ameliorated the development of retinopathy and functional/structural abnormalities and attenuated Rac1–Nox2–mitochondrial damage. Similar alterations in Rac1 regulators were observed in retinal microvasculature from human donors with diabetic retinopathy. In diabetes, Rac1 prenylation and its interactions with Vav2 contribute to Nox2–ROS–mitochondrial damage, and the pharmacological inhibitors to attenuate Rac1 interactions with its regulators could have the potential to halt/inhibit the development of diabetic retinopathy.
KeywordsDiabetic retinopathy NADPH oxidase Oxidative stress Post-translational modifications Ras-related C3 botulinum toxin substrate 1, Rac1
GM: researched data, literature search and manuscript editing; AJD: researched data, literature search and manuscript editing; AK: literature search, manuscript writing/editing; RAK: experimental plan, literature search, manuscript writing/editing. RAK is the guarantor of this work and, as such, had full access to all the data in this manuscript.
This study was supported in parts by grants from the National Institutes of Health (RAK: EY014370 and EY017313; RAK, AK: EY022230), Thomas Foundation (RAK), Department of Veterans Affairs (1BX000469, AK), and an unrestricted grant to the Ophthalmology Department from Research to Prevent Blindness. AK is the recipient of a Senior Research Career Scientist Award from the Department of Veterans Affairs (13S-RCS-006).
Compliance with Ethical Standards
The treatment of animals conformed to the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research and was approved by the Wayne State University’s Institutional Animal Care and Use Committee.
Conflict of Interest
The authors declare that they have no conflict of interest.
- 11.Bai Y, Xiang X, Liang C, Shi L (2015) Regulating Rac in the nervous system: molecular function and disease implication of Rac GEFs and GAPs. Biomed Res Int 2015:632450Google Scholar
- 15.Abdrabou A, Wang Z (2018) Post-translational modification and subcellular distribution of Rac1: an update. Cells 7Google Scholar
- 28.Bokoch GM, Bohl BP, Chuang TH (1994) Guanine nucleotide exchange regulates membrane translocation of Rac/Rho GTP-binding proteins. J Biol Chem 269:31674–31679Google Scholar
- 31.Kern TS, Miller CM, Tang J, Du Y, Ball SL, Berti-Matera L (2010) Comparison of three strains of diabetic rats with respect to the rate at which retinopathy and tactile allodynia develop. Mol Vis 16:1629–1639Google Scholar
- 34.Kowluru RA (2005) Diabetic retinopathy, oxidative stress and antioxidants. Curr Top Nutraceut Res 3:209–218Google Scholar
- 40.Chen GP, Zhang XQ, Wu T, Li L, Han J, Du CQ (2015) Alteration of mevalonate pathway in proliferated vascular smooth muscle from diabetic mice: possible role in high-glucose-induced atherogenic process. J Diabetes Res 2015:379287Google Scholar