The CXC chemokine IL-8, which promotes adhesion, activation, and transmigration of polymorphonuclear neutrophils (PMN), has been associated with production of tissue injury in reperfused myocardium. Hypoxia-inducible factor-1 (HIF-1) is a heterodimeric peptide that is a key regulator of genes such as heme oxygenase (HO)-1 expressed under hypoxic conditions. We hypothesized that HO-1 plays an important role in regulating proinflammatory mediator production under conditions of ischemia-reperfusion. HIF-1 was activated in the human microvascular endothelial cell line (HMEC-1) with the prolyl hydroxylase inhibitor dimethyloxalylglycine (DMOG). DMOG significantly attenuated cytokine-induced IL-8 promoter activity and protein secretion and cytokine-induced PMN migration across human microvascular endothelial cell line HMEC-1 monolayers. In vivo studies in a rabbit model of myocardial ischemia-reperfusion showed that rabbits pretreated with a 20 mg/kg DMOG infusion (n ϭ 6) 24 h before study exhibited a 21.58 Ϯ 1.76% infarct size compared with 35.25 Ϯ 2.06% in saline-treated ischemia-reperfusion animals (n ϭ 6, change in reduction ϭ 39%; P Ͻ 0.001). In DMOG-pretreated (20 mg/kg) animals, plasma IL-8 levels at 3 h after onset of reperfusion were 405 Ϯ 40 pg/ml vs. 790 Ϯ 40 pg/ml in saline-treated ischemiareperfusion animals (P Ͻ 0.001). DMOG pretreatment reduced myocardial myeloperoxidase activity, expressed as number of PMN per gram of myocardium, to 1.43 Ϯ 0.59 vs. 4.86 Ϯ 1.1 (P ϭ 0.012) in saline-treated ischemia-reperfused hearts. Both in vitro and in vivo DMOG-attenuated IL-8 production was associated with robust HO-1 expression. Thus our data show that HIF-1 activation induces substantial HO-1 expression that is associated with attenuated proinflammatory chemokine production by microvascular endothelium in vitro and in vivo.hypoxia-inducible factor 1; cardiac ischemia-reperfusion; interleukin-8 SIGNIFICANT CLINICAL AND EXPERIMENTAL research has focused on protection of ischemic myocardium. Successful protection strategies are diverse and have included sublethal ischemia and certain pharmacological approaches (23,27,35). The biological process now recognized as "preconditioning" enhances endogenous cellular mechanisms within the myocardium and results in protection against postischemic injury. Identifying new pharmacological agents that promote long-lasting protection against ischemia-reperfusion injury is an important goal. In this study, we examine a novel pharmacological strategy for cardiac preconditioning using an inhibitor of prolyl hydroxylase, a key enzyme controlling stabilization, and activation of hypoxia-inducible factor-1 (HIF-1). HIF-1 is a heterodimer consisting of HIF-1␣ and HIF-1␤ subunits (38, 39), and its activity is determined by stable expression of the ␣-subunit (11, 12). HIF-1␣ subunit activity is negatively regulated in normoxic cells by hydroxylation of proline residues that signal ubiquitination and degradation through the proteasome pathways (9, 10). Both prolyl hydroxylase inhibition and ...

A lcohol-use disorders (AUDs) encompass a spectrum of disorders, including excessive use, abuse, dependence, and addiction. Abuse occurs when a patient experiences adverse socioeconomic or health consequences related to the use of the substance. 1 Dependence is present either when the patient experiences withdrawal symptoms on discontinuation of the substance or when larger amounts are necessary to achieve the desired effect. 1 Addiction is indicated by the patient experiencing a compulsive craving for a substance. 1 At any one time, a patient may have more than one of these disorders simultaneously present. The National Institutes of Health 2 have put forth guidelines that quantify excessive and unhealthy use of alcohol; a level of use above this threshold increases a patient's risk for health problems. 3 Men aged Յ 65 years who consume more than four standard drinks in a typical day or . 14 standard drinks in a typical week have excessive and unhealthy alcohol consumption. These volumes are halved for women of all ages and men aged .

Microvascular endothelial cells provide a critical regulatory interface between blood constituents and tissue. Hypoxia inducible factor-1 (HIF-1) is a key transcription factor required for expression of hypoxia-dependent genes. We employed a model of hypoxia and reoxygenation (H/R) using the dermal microvascular endothelial cell line HMEC-1 to examine the effects of altered oxygen concentrations on microvascular HIF-1 expression and nitric oxide (NO) formation. Hypoxia increased inducible NO synthase (iNOS) mRNA in a time-dependent manner in HMEC-1. However, endothelial NO synthase mRNA progressively declined during hypoxia. H/R promoted significant increases in cellular nitrite levels that were significantly abrogated by the specific iNOS inhibitor N6-(1-iminoethyl)-L-lysine, di hy drochloride. Exogenous NO promoted stabilization of the alpha subunit of HIF-1 and produced functional DNA binding. Exposure of HMEC-1 to H/R resulted in previously unrecognized biphasic HIF-1alpha stabilization during reoxygenation. When the iNOS gene was silenced through the use of iNOS-specific small interfering RNA, HIF-1alpha stabilization and HIF-1 activation were dramatically diminished, suggesting that inducible NOS-derived NO is a key factor sustaining HIF-1 activation during both hypoxia and reoxygenation.

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