Carbon monoxide (CO) is a product of the enzyme heme oxygenase (HO; EC 1.14.99.3). In vascular smooth muscle cells, exogenously administered CO increases cyclic guanosine 3',5'-monophosphate (cGMP), which is an important regulator ofvessel tone. We report here that smooth muscle cells produce CO via HO and that it regulates cGMP levels in these cells. Hypoxia, which has profound effects on vessel tone, significantly increased the transcriptional rate of the HO-1 gene resulting in corresponding increases of its mRNA and HO enzymatic activity. In addition, under the same conditions, rat aortic and pulmonary artery smooth muscle cells accumulated high levels of cGMP following a similar time course to that of HO-1 production. The increased accumulation of cGMP in smooth muscle cells required the enzymatic activity of HO, since it was abolished by a specific HO inhibitor, tin protoporphyrin. In contrast, NO-nitro-Larginine, a potent inhibitor ofnitric oxide (NO) synthesis, had no effect on cGMP produced by smooth muscle cells, indicating that NO is not responsible for the activation of guanylyl cyclase in this setting. Furthermore, conditioned medium from hypoxic smooth muscle cells stimulated cGMP production in recipient cells and this stimulation was completely inhibited by tin protoporphyrin or hemoglobin, an inhibitor of CO production and a scavenger of CO, respectively. This report shows that HO-1 is expressed by vascular smooth muscle cells and that its product, CO, may regulate vascular tone under physiologic and pathophysiologic (such as hypoxic) conditions.Regulation of blood vessel tone is critical to maintaining adequate tissue oxygenation and perfusion. This phenomenon involves a delicate balance between vasodilators and vasoconstrictors. Hypoxia, for example, has profound effects on blood vessel tone, principally through the release or inhibition of vasoactive mediators from endothelial cells (1, 2). One endothelium-derived mediator is nitric oxide (NO), a potent vasodilator that helps maintain normal vascular tone by stimulating guanylyl cyclase in smooth muscle cells (SMCs) and elevating cGMP levels. Endothelial NO was shown to be suppressed by a hypoxic state resulting in low cGMP levels (3). NO is normally produced by the body and serves as an important chemical messenger not only in the regulation ofvessel tone, but also in neuronal transmission. Like NO, carbon monoxide (CO) is an endogenously produced gas molecule that activates guanylyl cyclase (4). Although a role for CO has been suggested in neuronal signal transduction (5), it is not known whether CO plays a physiologic role in the vasculature.There are at least two endogenous sources of CO production, one of which is from the oxidation of organic molecules, but the predominant source is from the degradation of heme (6). Heme is metabolized to biliverdin and CO by hemeThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S...

Asthma is a prevalent disease of chronic inflammation in which endogenous counter-regulatory signaling pathways are dysregulated. Recent evidence suggests that innate lymphoid cells (ILCs), including natural killer (NK) cells and type 2 innate lymphoid cells (ILC2), can participate in the regulation of allergic airways responses, in particular airway mucosal inflammation. Here, we have identified both NK cells and ILC2 in human lung and peripheral blood in healthy and asthmatic subjects. NK cells were highly activated in severe asthma, linked to eosinophilia and interacted with autologous eosinophils to promote their apoptosis. ILC2 generated antigen-independent IL-13 in response to the mast cell product prostaglandin D2 (PGD2) alone and in a synergistic manner with the airway epithelial cytokines IL-25 and IL-33. Both NK cells and ILC2 expressed the pro-resolving ALX/FPR2 receptors. Lipoxin A4, a natural pro-resolving ligand for ALX/FPR2 receptors, significantly increased NK cell mediated eosinophil apoptosis and decreased IL-13 release by ILC2. Together, these findings indicate that ILCs are targets for lipoxin A4 to decrease airway inflammation and mediate the catabasis of eosinophilic inflammation. Because lipoxin A4 generation is decreased in severe asthma, these findings also implicate unrestrained ILC activation in asthma pathobiology.

Plasma homocystele levels are elevated in 20-30% of all patients with premature atherosclerosis. Al-though elevated homocystelne levels have been recognized as an Independent risk factor for myocardial Infarction and stroke, the m nism by which these elevated levels cause atherosclerosis is unknown. To understand the role ofhomocysteine In the pathogenesis of atherosclerosis, we ex the effect of homocysteine on the growth of both vascular smooth muscle cells and endothelial cells at concentrations similar to those observed In clinical sudis. As little as 0.1 mM homocysteine caused a 25% increase In DNA synthesis, and homocysteine at 1 mM increased DNA synthesis by 4.5-fold in rat aortic smooth muscle cells (RASMC). In contrast, homocysteine caused a dose-dependent decrease In DNA synthesis in human umbilical vein telll ces. Homocysteine increased mRNA levels of cyclin D1 and cyclin A in RASMC by 3-and 15-fold, respectively, inicating that homocysteine induced the mRNA of cycls important for the reentry of quiescent RASMC into the cell cycle. Furthermore, homocysteine promoted proliferation of quiescent RASMC, an effect markedly amplified by 2% serum. The growth-promoting effect of homocysteine on vascular smooth muscle cells, together with its inhibitory effect on endothelial cell growth, represents an important mechanism to explain homocysteine-induced atherosclerosis.

Nitric oxide (NO), which accounts for the biological properties of endothelium-derived relaxing factor, is generated by NO synthase (NOS). The vascular endothelium contains two types of NOS: one is constitutively expressed (cNOS), and the other is inducible. Endothelium-mediated vasorelaxation is impaired in atherosclerotic vessels. To determine whether tumor necrosis factor (TNF)-alpha, which is commonly found in atherosclerotic lesions, has an effect on NOS message, we measured cNOS mRNA levels in TNF-treated human umbilical vein endothelial cells (HUVECs) by RNA blot analysis with a cNOS cDNA probe. TNF-alpha markedly reduced cNOS mRNA levels in HUVECs in a dose- and time-dependent manner. In response to 3 ng/mL TNF-alpha, cNOS mRNA levels began to decrease at 4 hours and diminished to only 5% of control levels at 24 hours. As little as 0.1 ng/mL TNF-alpha reduced cNOS mRNA levels by 50%. This reduction in cNOS message in response to TNF-alpha depended on protein synthesis as it was blocked by cycloheximide. In nuclear runoff experiments, TNF-alpha did not change the rate of cNOS gene transcription. cNOS mRNA is very stable under basal conditions, with a half-life of 48 hours; however, treatment with TNF-alpha shortened this half-life to 3 hours. TNF-alpha thus appears to decrease cNOS mRNA levels by increasing the rate of mRNA degradation. TNF-induced reductions in cNOS mRNA levels may have an important effect on impaired endothelium-mediated vasorelaxation in atherosclerosis.