ABSTRACT
To explore the underlying mechanism(s) of pulmonary arterial hypertension in endotoxic shock, the roles of N-acetylcysteine (NAC), nitric oxide (NO) and carbon monoxide (CO) were investigated. Pulmonary arterial rings (3-mm width) were prepared from 24 rabbits. Lipopolysaccharide (LPS), after 7-hour incubation, decreased the endothelium-dependent relaxation response of the arterial ring (pre-contracted with phenylephrine) to acetylcholine (1 mumol/L), but did not affect the endothelium-independent relaxation response to sodium nitroprusside. The LPS effects were reduced by a concomitant incubation with the free radical scavenger (NAC), NO donor (L-arginine), and CO donor (hemin), respectively. On the other hand, the LPS effects were enhanced by applying heme oxygenase-1 (HO-1) inhibitor (zinc protoporphyrin) to block CO production. The response to acetylcholine changed from relaxation to contraction, however, the contractile response to phenylephrine increased significantly after pre-incubation with nitric oxide synthase (NOS) inhibitor (L-NAME) to block NO production, confirming the importance of CO and NO. These results show that LPS impairs endothelium-dependent relaxation of the pulmonary artery, which can be greatly reduced by the antioxidant, or by supplying with NO and CO. Thus, multiple factors are involved in this model of endotoxin-induced pulmonary hypertension.
Subject(s)
Animals , Male , Rabbits , Acetylcysteine , Metabolism , Carbon Monoxide , Metabolism , Hypertension, Pulmonary , Lipopolysaccharides , Toxicity , Nitric Oxide , Metabolism , Pulmonary Artery , Shock, SepticABSTRACT
This study, using cultured bovine pulmonary artery endothelial cells (BPAECs), was undertaken to investigate the roles of endogenous ONOO(-) in LPS-caused injury in endothelial cells. The fluorescent intensity of nitrotyrosine (NT), a specific marker of ONOO(-) generation, in BPAECs represented the content of endogenous ONOO(-) generation. The fluorescent intensity of NT and the number of NT positive cells were detected with flow cytometry (FCM), and the percentage of NT positive cells was calculated. The results are as follows. (1) LPS (1, 5 and 10 microg/ml) caused a marked increase in fluorescent intensity of NT in a dose-dependent manner, which was significantly increased compared to the vehicle group (P<0.01).The number and percentage of NT positive cells were markedly increased (both P<0.05 vs vehicle group). Aminoguanidine (AG), a selective inhibitor of inducible nitric oxide synthase (iNOS), inhibited LPS-induced increase in fluorescent intensity of NT in BPAECs. However, the number and percentage of NT positive cells had a tendency to reduce. (2) LPS brought about an enhancement in MDA content and the activity of LDH in cultured supernatant. AG reversed the enhancement in MDA content induced by LPS (P<0.01). In contrast, AG had a marginal effect on the activity of LDH. (3) LPS induced an increase in apoptotic rate in BPAECs in a dose-dependent manner. The number of apoptotic cells markedly increased as well. Some BPAECs stained with fluorescent probe ethidium bromide showed morphological features of apoptosis with chromatin condensation and nuclear fragmentation. AG reduced the apoptotic rate and the number of apoptotic cells, both of which were still higher than those of vehicle group (P<0.05). LPS led to inhibition of mitochondrial respiration and membrane potential in an accumulation manner. In conclusion, LPS caused injury to cultured BPAECs and increased the production of ONOO(-).The cytotoxicity of LPS may be mediated by the endogenous ONOO(-).
Subject(s)
Animals , Cattle , Cells, Cultured , Endothelial Cells , Cell Biology , Metabolism , Pathology , Lipopolysaccharides , Toxicity , Lung Injury , Peroxynitrous Acid , Physiology , Pulmonary Artery , Cell Biology , PathologyABSTRACT
<p><b>AIM</b>To investigate the alleviating effect of N-acetylcysteine (NAC) on lung injury induced by lipopolysaccharides (LPS) and its mechanism.</p><p><b>METHODS</b>The effects of NAC on changes of the pulmonary arterial reactivity and the ultrastructure of pulmonary arterial endothelium induced by LPS were observed with the isolated artery ring technique and scanning electron microscope (SEM). Malondialdehyde (MDA), nitric oxide (NO) contents and superoxide dismutase (SOD) activity of pulmonary artery tissues were detected.</p><p><b>RESULTS</b>The exposure of pulmonary artery to LPS (4 microg/ml, 7 h) led to reduction of endothelium-dependent relaxation response to acetylcholine (ACh), which was reversed by the concomitant exposure to NAC (0.5 mmol/L, 7 h), whereas NAC itself had no effect on the response. Significant structural injury were observed under SEM in LPS group and alleviated the changes in LPS + NAC group. The MDA, NO contents increased but SOD activity decreased in LPS group, which were reversed by the concomitant exposure to NAC.</p><p><b>CONCLUSION</b>NAC protects pulmonary artery endothelium and enhances endothelium-dependent relaxation response of pulmonary artery by antioxidation effect, which may be one of the mechanisms of its reversing pulmonary artery hypertension and following lung injury induced by LPS.</p>