title: Blockade of NADPH oxidase restores vasoreparative function in diabetic CD34+ cells.
abstract: OBJECTIVE
  The vasodegenerative phase of diabetic retinopathy is likely caused by endothelial dysfunction and reduced endothelial repair. Migration of endothelial progenitor cells (EPCs) into areas of vascular injury is critical to vascular repair. This key function, often defective in diabetes, is largely mediated by nitric oxide (NO), which is known to be inactivated by superoxide produced by NADPH oxidase. The authors tested the hypothesis that either increasing eNOS expression or inhibiting NADPH oxidase would restore the reparative function in diabetic EPCs.
  
  
  METHODS
  Peripheral blood was obtained from healthy (n = 27) and diabetic (n = 31) persons, and CD34(+) cells were isolated. Expression and activation of eNOS and NADPH oxidase and intracellular levels of NO, superoxide, and peroxynitrite were evaluated. cGMP production and migration to SDF-1α were also determined. Reparative function was evaluated in a mouse model of retinal ischemia-reperfusion injury.
  
  
  RESULTS
  Diabetic EPCs demonstrate reduced eNOS expression and decreased NO bioavailability and migration in response to SDF-1α. Increasing eNOS expression in diabetic cells by AVE3085 resulted in increased peroxynitrite levels and, therefore, did not enhance NO-mediated functions in vitro and in vivo. Expression of Nox2, NADPH oxidase activity, and superoxide levels were higher in diabetic than in nondiabetic EPCs. Pretreatment with apocynin or gp91ds-tat increased NO bioavailability without increasing eNOS activity in response to SDF-1α. Ex vivo NADPH oxidase inhibition in diabetic cells restored migratory function in vitro and enhanced their homing to ischemic retinal vasculature in vivo.
  
  
  CONCLUSIONS
  The NADPH oxidase system is a promising target for correcting vasoreparative dysfunction in diabetic EPCs.
authors.affiliation: Department of Pharmacology and Therapeutics, University of Florida, Gainesville, Florida 32610-0267, USA.
authors.forename: Yagna P R
authors.initials: YP
authors.lastname: Jarajapu
authors.forename: Sergio
authors.initials: S
authors.lastname: Caballero
authors.forename: Amrisha
authors.initials: A
authors.lastname: Verma
authors.forename: Takahiko
authors.initials: T
authors.lastname: Nakagawa
authors.forename: Margaret C
authors.initials: MC
authors.lastname: Lo
authors.forename: Qiuhong
authors.initials: Q
authors.lastname: Li
authors.forename: Maria B
authors.initials: MB
authors.lastname: Grant
mesh_terms: D000098:Acetophenones / Q000494:pharmacology; D000328:Adult; D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D052117:Benzodioxoles / Q000494:pharmacology; D054377:Chemokine CXCL12 / Q000378:metabolism; D006152:Cyclic GMP / Q000378:metabolism; D003930:Diabetic Retinopathy / Q000097:blood*; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D006023:Glycoproteins / Q000494:pharmacology; D006801:Humans; D007189:Indans / Q000494:pharmacology; D008297:Male; D051379:Mice; D008875:Middle Aged; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors*; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D030421:Peroxynitrous Acid / Q000378:metabolism; D015427:Reperfusion Injury / Q000201:enzymology / Q000503:physiopathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013481:Superoxides / Q000378:metabolism; D055815:Young Adult
chemical_list: C528910:2,2-difluorobenzo(1,3)dioxole-5-carboxylic acid indan-2-ylamide; D000098:Acetophenones; D018952:Antigens, CD34; D052117:Benzodioxoles; C515255:CXCL12 protein, human; D054377:Chemokine CXCL12; D004791:Enzyme Inhibitors; D006023:Glycoproteins; D007189:Indans; C473031:gp91ds-tat protein, chimeric; D013481:Superoxides; D030421:Peroxynitrous Acid; D009569:Nitric Oxide; C056165:acetovanillone; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III; D019255:NADPH Oxidases; D006152:Cyclic GMP