PurposeChronic ethanol (EtOH) ingestion increases the incidence of the acute respiratory distress syndrome and causes oxidative stress and cellular dysfunction in the lung. The mechanisms of EtOH-induced oxidative stress in the lung remain to be defined. We sought to determine if chronic EtOH ingestion alters the expression of lung NADPH oxidase, a major enzymatic source of superoxide generation, in an in vivo rat model of chronic EtOH ingestion.MethodsMale Sprague-Dawley rats were fed liquid diets containing EtOH (36% of calories) or an isocaloric diet substituting maltin-dextrin for EtOH (control) with or without the angiotensin-converting enzyme (ACE) inhibitor lisinopril for 6 weeks. Prior to sacrifice, blood pressure was monitored for 15-20 minutes. After sacrifice, lung frozen sections were stained with dihydroethidium (DHE) to detect superoxide production. Expression of specific components of the renin-angiotensin system (RAS) and specific NADPH oxidase components were then examined in lung homogenates.ResultsChronic EtOH ingestion in the rat had no significant effect on blood pressure but increased superoxide formation in lung parenchyma measured as DHE fluorescence, an effect inhibited by lisinopril. Chronic EtOH ingestion failed to increase lung ACE expression, but increased angiotensinogen, angiotensin II type 1 (AT1) and type 2 (AT2) receptor expression. Chronic EtOH ingestion also increased expression of the NADPH oxidase subunit, gp91phox, an effect inhibited by lisinopril.ConclusionsThese results indicate that chronic EtOH ingestion alters superoxide production and specific NADPH oxidase subunit expression in the lung by a RAS-dependent pathway. These findings provide new insights into mechanisms by which EtOH causes oxidative stress in the lung.

PurposeChronic ethanol (EtOH) ingestion increases the incidence and severity of the acute respiratory distress syndrome (ARDS), a pulmonary disorder characterized by disruption of the alveolar-capillary barrier. Nitric oxide (NO) has been implicated in regulation of endothelial barrier function. Because previous reports have demonstrated that exposure to EtOH for periods up to 6 hours increases endothelial NO production, we sought to determine if more chronic EtOH exposure could directly modulate endothelial nitric oxide synthase (eNOS) activity and endothelial NO production, thereby contributing to altered lung responses during chronic EtOH ingestion.MethodsTo test this hypothesis, porcine pulmonary artery endothelial cells (PAEC) were treated with 0.1% (w/v) EtOH for 72 hours in sealed chambers to prevent evaporation.ResultsEtOH caused dose-dependent increases in NO production and increased eNOS expression, effects that were attenuated by wortmannin, a specific PI-3 kinase inhibitor. EtOH also increased heat shock protein 90 (hsp90)-eNOS interaction, and geldanamycin, an hsp90 inhibitor, attenuated ETOH-stimulated eNOS-hsp90 interaction as well as NO production. In contrast to acute EtOH stimulation, chronic EtOH treatment did not stimulate Akt activation or phosphorylation of eNOS ser1177.ConclusionsThese results indicate that chronic EtOH exposure increases endothelial NO production via PI-3 kinase-mediated increases in eNOS expression and enhanced binding of hsp90 to eNOS. This study highlights potential differences in the mechanisms regulating endothelial NO production during acute and chronic EtOH exposure.

PurposeThe antidiabetic thiazolidinedione medications activate the peroxisome proliferator-activated receptor gamma (PPARγ) and exert vascular protective effects through incompletely defined mechanisms. We recently reported that the PPARγ ligands 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) and ciglitizone increased cultured endothelial cell (EC) nitric oxide (NO) release without increasing the expression of endothelial nitric oxide synthase (eNOS).MethodsTo further characterize the molecular mechanisms underlying PPARγ ligand-stimulated increases in EC NO production, human umbilical vein endothelial cells (HUVEC) were treated with 10 μM rosiglitazone for 24 hours followed by analysis of NO production, eNOS and heat shock protein 90 (hsp90) activation.ResultsRosiglitazone increased EC NO release without altering eNOS or hsp90 expression. However, rosiglitazone increased hsp90-eNOS protein-protein interactions. The hsp90 antagonist geldanamycin attenuated rosiglitazone-induced increases in NO production as well as eNOS-hsp90 association. Rosiglitazone also increased the phosphorylation of eNOS at ser1177.ConclusionsThese findings demonstrate that the PPARγ ligand rosiglitazone stimulates EC NO release by mechanisms involving eNOS-protein interactions and alterations in eNOS phosphorylation. These findings provide further insight into mechanisms of thiazolidinedione-induced vascular protection.

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measurements. Measures of ejection dynamics were then derived from the time series data, including stroke volume, cardiac output, ejection time, acceleration time, mean acceleration and net developed force. Summary of Results: Compared to gestational age matched normal fetuses, the CHD fetuses demonstrated a decrease in the net force developed (1.255 Ϯ 0.694, p < .03), prolonged acceleration times (0.243 Ϯ 0.075, p < .002) in the pulmonary artery (0.281 Ϯ 0.075, p < .004) in the aorta), and normal ejection times (0.183 vs. 0.185). Conclusions: Normal ejection times suggest that afterload is not a critical determinate in the observed decreases in cardiac output, stroke volume, and ejection fraction in the fetuses with CHD. The decrease in the net developed force and prolonged acceleration times suggests decreased myocardial function as the major cause of the altered in utero hemodynamics in this population.