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Vascular inflammation contributes to cardiovascular diseases such as aortic aneurysm and dissection. However, the precise inflammatory pathways involved have not been clearly defined. We have shown here that subcutaneous infusion of Ang II, a vasopressor known to promote vascular inflammation, into older C57BL/6J mice induced aortic production of the proinflammatory cytokine IL-6 and the monocyte chemoattractant MCP-1. Production of these factors occurred predominantly in the tunica adventitia, along with macrophage recruitment, adventitial expansion, and development of thoracic and suprarenal aortic dissections. In contrast, a reduced incidence of dissections was observed after Ang II infusion into mice lacking either IL-6 or the MCP-1 receptor CCR2. Further analysis revealed that Ang II induced [CCR2.sup.+][CD14.sup.hi][CD11b.sup.hi]F4/80- macrophage accumulation selectively in aortic dissections and not in aortas from [Il6.sup.[-/-]] mice. Adoptive transfer of [Ccr2.sup.[+/+]] monocytes into [Ccr2.sup.[-/-]] mice resulted in selective monocyte uptake into the ascending and suprarenal aorta in regions of enhanced ROS stress, with restoration of IL-6 secretion and increased incidence of dissection. In vitro, coculture of monocytes and aortic adventitial fibroblasts produced MCP-1- and IL-6-enriched conditioned medium that promoted differentiation of monocytes into macrophages, induced CD14 and CD11b upregulation, and induced MCP-1 and MMP-9 expression. These results suggest that leukocyte-fibroblast interactions in the aortic adventitia potentiate IL-6 production, inducing local monocyte recruitment and activation, thereby promoting MCP-1 secretion, vascular inflammation, ECM remodeling, and aortic destabilization., Introduction Aneurysms and aortic dissections are major diseases affecting the aorta, accounting for almost 16,000 deaths in the United States annually. Aortic aneurysms, including both abdominal and thoracic aneurysms, are [...]

Endothelial nitric oxide synthase (eNOS) activation with subsequent inducible NOS (iNOS), cytosolic phospholipase [A.sub.2] ([cPLA.sub.2]), and cyclooxygenase-2 (COX2) activation is essential to statin inhibition of myocardial infarct size (IS). In the rat, the peroxisome proliferator-activated receptor-[gamma] agonist pioglitazone (Pio) limits IS, upregulates and activates [cPLA.sub.2] and COX2, and increases myocardial 6-keto-PG[F.sub.1[alpha]] levels without activating eNOS and iNOS. We asked whether Pio also limits IS in [eNOS.sup.-/-] and [iNOS.sup.-/-] mice. Male C57BL/6 wild-type (WT), [eNOS.sup.-/-], and [iNOS.sup.-/-] mice received 10 mg x [kg.sup.-1] x [day.sup.-1] Pio (Pio+) or water alone (Pio-) for 3 days. Mice underwent 30 min coronary artery occlusion and 4 h reperfusion, or hearts were harvested and subjected to ELISA and immunoblotting. As a result, Pio reduced IS in the WT (15.4 [+ or -] 1.4% vs. 39.0 [+ or -] 1.1%; P < 0.001), as well as in the [eNOS.sup.-/-] (32.0 [+ or -] 1.6% vs. 44.2 [+ or -] 1.9%; P < 0.001) and [iNOS.sup.-/-] (18.0 [+ or -] 1.2% vs. 45.5 [+ or -] 2.3%; P < 0.001) mice. The protective effect of Pio in [eNOS.sup.-/-] mice was smaller than in the WT (P < 0.001) and [iNOS.sup.-/-] (P < 0.001) mice. Pio increased myocardial Ser633 and Ser1177 phosphorylated eNOS levels in the WT and [iNOS.sup.-/-] mice. iNOS was undetectable in all six groups. Pio increased [cPLA.sub.2], COX2, and PG[I.sub.2] synthase levels in the WT, as well as in the [eNOS.sup.-/-] and [iNOS.sup.-/-], mice. Pio increased the myocardial 6-ketoPG[F.sub.1[alpha]] levels and [cPLA.sub.2] and COX2 activity in the WT, [eNOS.sup.-/-], and [iNOS.sup.-/-] mice. In conclusion, the myocardial protective effect of Pio is iNOS independent and may be only partially dependent on eNOS. Because eNOS activity decreases with age, diabetes, and advanced atherosclerosis, this effect may be relevant in a clinical setting and should be further characterized. endothelial nitric oxide synthase; inducible nitric oxide synthase; peroxisome proliferator-activated receptor-[gamma]

Endothelial nitric oxide synthase (eNOS) activation with subsequent inducible NOS (iNOS), cytosolic phospholipase A2 (cPLA2), and cyclooxygenase-2 (COX2) activation is essential to statin inhibition of myocardial infarct size (IS). In the rat, the peroxisome proliferator-activated receptory agonist pioglitazone (Pio) limits IS, upregulates and acti- vates cPLA2 and COX2, and increases myocardial 6-keto-PGF1~. levels without activating eNOS and iNOS. We asked whether Pio also limits IS in eNOS~' and iNOS' mice. Male C57BL16 wild-type (WT), eNOS", and iNOS' mice received 10 mgkg~ day~ Pio (Pio+) or water alone (Pio-) for 3 days. Mice underwent 30 mm coronary artery occlusion and 4 h reperfusion, or hearts were harvested and subjected to ELISA and immunoblotting. As a result, Pio reduced IS in the WT (15.4 ± 1.4% vs. 39.0 ± 1.1%; P < 0.001), as well as in the eNOS~ (32.0 ± 1.6% vs. 44.2 ± 1.9%; P < 0.001) and iNOS' (18.0 ± 1.2% vs. 45.5 ± 2.3%; P < 0.001) mice. The protective effect of Pio in eNOS~ mice was smaller than in the WT (P <0.001) and iNOS' (P < 0.001) mice. Pio increased myocardial Ser633 and Sen 177 phosphonylated eNOS levels in the WI and iNOS~ mice. iNOS was undetectable in all six groups. Pio increased cPLA2, COX2, and PGI2 synthase levels in the WT, as well as in the eNOS' and iNOS', mice. Pio increased the myocardial 6-keto- PGF1~. levels and cPLA2 and COX2 activity in the WT, eNOS~', and iNOS" mice. In conclusion, the myocardial protective effect of Pio is iNOS independent and may be only partially dependent on eNOS. Because eNOS activity decreases with age, diabetes, and advanced atherosclerosis, this effect may be relevant in a clinical setting and should be further characterized. [ABSTRACT FROM AUTHOR]

Evidence of diabetes-induced nuclear factor-kappaB (NF-kappaB) activation has been provided with DNA binding assays or nuclear localization with immunohistochemistry, but few studies have explored mechanisms involved. We examined effects of diabetes on proteins comprising NF-kappaB canonical and noncanonical activation pathways in the renal cortex of diabetic mice. Plasma concentrations of NF-kappaB-regulated cytokines were increased after 1 month of hyperglycemia, but most returned to control levels or lower by 3 months, when the same cytokines were increased significantly in renal cortex. Cytosolic content of NF-kappaB canonical pathway proteins did not differ between experimental groups after 3 months of diabetes, while NF-kappaB noncanonical pathway proteins were affected, including increased phosphorylation of inhibitor of kappaB kinase-alpha and several fold increases in NF-kappaB-inducing kinase and RelB, which were predominantly located in tubular epithelial cells. Nuclear content of all NF-kappaB pathway proteins was decreased by diabetes, with the largest change in RelB and p50 (approximately twofold decrease). Despite this decrease, measurable increases in protein binding to DNA in diabetic versus control nuclear extracts were observed with electrophoretic mobility shift assay. These results provide evidence for chronic NF-kappaB activation in the renal cortex of db/db mice and suggest a novel, diabetes-linked mechanism involving both canonical and noncanonical NF-kappaB pathway proteins. [ABSTRACT FROM AUTHOR]

Angiotensin-II (Ang-II), the major effector peptide of the renin angiotensin system, potently accelerates progression of atherosclemsis and vascular remodeling, and induces activation of NF-κB, a nuclear transcription factor regulating expression of numerous proinflammatory and proangiogenic cytokines, chemokines, growth factors, and adhesion molecules. To investigate Ang-II effects on diabetes-induced vascular inflammation, effects of chronic Ang-II infusion on NF-κB canonical and noncanonical proteins were studied in aorta of type 2, three month hyperglycemic db/db mice and congenic age-matched controls. Proteins were extracted from buffer (containing protease and phosphatase inhibitors)-perfused aortas obtained from four experimental groups, including untreated and Ang-II-infused controls, and untreated and Ang-II-infused db/db mice. Ang-II was infused subcutaneously via osmotic mini-pumps (0.5 mg/kg/day) for one month after two months of untreated diabetes. Protein levels of canonical (RelA/p65, IκBα, IκBβ, IKKβ, p50) and noncanonical (NIK, RelB, IKKα, p52, TRAF3) NF-κB proteins were quantified by Western blotting and normalized to tissue beta actin levels using infrared imaging. After 3 months of diabetes, most NF-κB canonical proteins were decreased in abundance, including IKKβ (40 % decrease), p50 (32 %), IκBα (26 %); RelA/p65 (20 %), and IκBβ (16%). Similar decreases were observed for noncaoonical NF-κB proteins, including IKKα (46 %), RelB (36%), and TRAF3 (32%), while p52 was unchanged and NIK was increased ∼60 %. In general, Ang-II infusion into diabetic mice produced further decreases in canonical NF-κB proteins as well as p52 and TRAF3 (both decreased ∼65 %), but increased RelB 30%. Ang-II increased the total amount of kinases, including IKKα (31%), IKKβ (61%), and NIK (45%). These results indicate that steady state levels of most NF-κB proteins are decreased in aorta of diabetic mice, and suggest that Ang-II has a divergent effect on canonical versus non-canonical NF-κB proteins. ADA-Funded Research [ABSTRACT FROM AUTHOR]