title: Increased iNOS activity in vascular smooth muscle cells from diabetic rats: potential role of Ca(2+)/calmodulin-dependent protein kinase II delta 2 (CaMKIIδ(2)).
abstract: OBJECTIVE
  Inducible nitric oxide synthase (iNOS) expression may be increased by cytokine plasma levels contributing to vascular damage in diabetes. Besides transcriptional regulation, Ca(2+)/CaMKII may play a role in post-translationally controlled iNOS activity. We accordingly investigated the involvement of the Ca(2+)/CaMKIIδ(2) signaling pathway in regulating lipopolysaccharide (LPS)-induced iNOS activity in cultured aortic vascular smooth muscle cells (VSMCs) from diabetic rats.
  
  
  RESULTS
  VSMCs obtained from 10 diabetic rats (DR) and 10 control rats (CR) were stimulated with 20 μg/ml LPS. After 24 h, iNOS protein levels were 1.37 fold increased in DR- vs CR-VSMCs (p < 0.05; Western Blot), while iNOS activity (conversion l-((3)H)-arginine into l-((3)H)-citrulline) and intracellular nitrotyrosine levels (immunofluorescence) were about 2.7 fold greater in DR- than in CR-VSMCs. Interestingly, LPS increased intracellular Ca(2+) levels (Fluorescence video imaging) more markedly in DR- than in CR-VSMCs. This was associated with CaMKII activation by phosphorylation, a decreased amount of co-immunoprecipitating iNOS/CaMKIIδ(2) (Western Blot) and increased iNOS activity. The CaMKII inhibitor KN-93 abolished all the LPS-effects.
  
  
  CONCLUSIONS
  These results indicate that the Ca(2+)/CaMKIIδ(2) signaling pathway may be an important regulator of iNOS activity in diabetes, and hence contribute to the potential development of innovative therapeutic strategies for vascular complications in diabetes.
authors.affiliation: Department of Biomedical Sciences, G. d'Annunzio University, Chieti-Pescara, Italy; Aging Research Center, Ce.S.I., G. d'Annunzio University Foundation, Chieti-Pescara, Italy.
authors.forename: Natalia
authors.initials: N
authors.lastname: Di Pietro
authors.forename: Pamela
authors.initials: P
authors.lastname: Di Tomo
authors.forename: Sara
authors.initials: S
authors.lastname: Di Silvestre
authors.forename: Annalisa
authors.initials: A
authors.lastname: Giardinelli
authors.forename: Caterina
authors.initials: C
authors.lastname: Pipino
authors.forename: Caterina
authors.initials: C
authors.lastname: Morabito
authors.forename: Gloria
authors.initials: G
authors.lastname: Formoso
authors.forename: Maria Addolorata
authors.initials: MA
authors.lastname: Mariggiò
authors.forename: Assunta
authors.initials: A
authors.lastname: Pandolfi
mesh_terms: D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000502:physiology*; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology*; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000201:enzymology*; D052247:Nitric Oxide Synthase Type II / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D052247:Nitric Oxide Synthase Type II; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2