Rat platelets adhere to human thrombin-treated rat lungs under flow conditions.

Platelet attachment to thrombin-treated endothelium was examined under flow conditions in the vascular bed of isolated perfused rat lungs. The infused thrombin bound to the pulmonary endothelium in a specific, competitive manner as shown by displacement of 125I-alpha-thrombin with an excess of cold diisopropylfluoro-phosphate-alpha-thrombin. alpha-Thrombin significantly increased the attachment of isolated 51Cr-labelled rat platelets to the perfused lungs. Scanning and high voltage electron microscopy showed single platelets in various stages of activation attached to pulmonary endothelium. The receptor binding site and catalytic activity of thrombin were essential to the enhancement of platelet attachment. Aspirin given to rats before the lung isolation had no effect on thrombin-induced platelet attachment. Thus, endothelial bound thrombin initiates platelet activation and enhances subsequent platelet attachment.

Vascular smooth muscle contains a depletable store of a vasodilator which is light-activated and restored by donors of nitric oxide.

Endothelium-denuded strips of rabbit thoracic aorta relax on exposure to light. This response is similar to endothelium-dependent relaxation as it is inhibited by hemoglobin and methylene blue, and is mediated by an increase in cyclic GMP. We now demonstrate that photorelaxation decreases on repeated exposure to light. The response can be restored by treating the depleted smooth muscle strips with acidified nitrite, but not nitrite alone, and with the nitric oxide donors, S-nitrosopenicillamine and glyceryl trinitrate, but not with hydralazine. These data indicate that photorelaxation is mediated in part by a "pool" of light-activated vasodilator(s) and suggest that this may act as a store of nitric oxide which could play a role in the regulation of vascular tone.

Platelet cGMP, but not cAMP, inhibits thrombin-induced platelet adhesion to pulmonary vascular endothelium.

We have compared the effects of intracellular pathways initiated by nitric oxide and prostacyclin on thrombin-induced platelet adhesion to endothelial cells. Platelet aggregate adhesion was enhanced when endothelial monolayers were pretreated with NG-monomethyl-L-arginine (L-NMMA), an inhibitor of nitric oxide production. In addition, decreased platelet aggregate adhesion was seen when platelets were pretreated with 8-bromoadenosine 3',5'-cyclic monophosphate (8-bromo-cAMP) or 8-bromoguanosine 3',5'-cyclic monophosphate (8-bromo-cGMP). Single platelet adhesion in isolated perfused lungs under flow conditions in the presence of shear was also assessed. Pretreatment of platelets with either Iloprost, in a dose sufficient to decrease platelet aggregation, or 8-bromo-cAMP did not affect platelet adhesion. However, pretreatment of platelets with 8-bromo-cGMP significantly reduced single platelet adhesion to endothelium. These studies illustrate that nitric oxide inhibits platelet adhesion to endothelium in the presence of shear. They further indicate that prostacyclin is also a regulator of this response but has effects more specifically related to the inhibition of platelet aggregation than platelet-endothelium interactions.

Non-steroidal anti-inflammatory drug-induced renal failure: a brief review of the role of cyclo-oxygenase isoforms.

Non-steroidal anti-inflammatory drugs are efficacious treatments for rheumatoid arthritis and osteoarthritis. However, an adverse effect of treatment with non-steroidal anti-inflammatory drugs is acute renal failure, particularly in a subset of patients that are in a state of effective volume depletion. The frequency of this side-effect in the general treated population is not known, but is probably less than 1% per year. Non-steroidal anti-inflammatory drugs act by inhibiting the synthesis of prostaglandins, which are important mediators of renal function. In the volume-depleted state prostaglandins may counter the vasoconstriction associated with the activation of the renin-angiotensin system. Cyclooxygenase is the rate-limiting enzyme involved in the synthesis of prostaglandins. Cyclooxygenase exists in two forms: a constitutive form (cyclooxygenase-1) and an inducible form (cyclooxygenase-2), which is associated with inflammation. Non-steroidal anti-inflammatory drugs are non-specific inhibitors of both forms of cyclooxygenase. New data are emerging regarding the role of cyclooxygenase-2 in the control of renal function. In normal rat and dog kidney, cyclooxygenase-2 is sparsely expressed in the macula densa, but expression is upregulated when animals are volume depleted. This review explores the possible role of cyclooxygenase-2 in the maintenance of normal renal function in volume depleted states.

Synthesis of nitric oxide in the bovine retina.

In the absence of light, high concentrations of cGMP open ion channels in the plasma membranes of rod outer segments. The source of stimulation of retinal guanylate cyclase is not known. Nitric oxide is a potent stimulator of guanylate cyclase in other cell systems. The present data demonstrate that nitric oxide synthase, an enzyme responsible for the production of nitric oxide, is present in retina, and specifically in the rod outer segments. This enzyme uses L-arginine as a substrate and is NADPH- and calcium- dependent. L-arginine-derived nitric oxide may be a source of activation of guanylate cyclase in the retina.

Leukocyte stimulation of intimal lesion formation is inhibited by treatment with diclofenac sodium and dexamethasone.

This investigation tested the effect of anti-inflammatory agents acting at different levels of the arachidonic acid cascade on leukocyte migration into the vessel wall. An animal model of vasculitis was used in which leukocytes stimulate migration of smooth muscle cells from the media into the intima resulting in formation of intimal lesions. In this model, an endotoxin-soaked thread is implanted in the adventitia along the ventral side of the rat femoral artery. At 2 days, subendothelial and medial leukocyte accumulation occurs exclusively in the ventral half of the vessel. At 14 days, intimal lesions composed primarily of smooth muscle cells are localized to the ventral half of the vessel. Treatment with neither the lipoxygenase inhibitor L-651,392 nor the dual cyclooxygenase and lipoxygenase inhibitor BW755C affected leukocyte migration into the vessel wall or subsequent lesion formation. Treatment with the cyclooxygenase inhibitor diclofenac sodium greatly reduced both leukocyte accumulation and lesion formation, whereas leukocyte migration and lesion formation were nearly totally inhibited by treatment with dexamethasone. Thus, it has been demonstrated that although leukocyte accumulation in the vessel wall stimulates intimal lesion formation, pharmacologic inhibition of leukocyte accumulation prevents lesion formation.

Thrombin induced platelet adhesion to endothelium is modified by endothelial derived relaxing factor (EDRF).

We investigated the hypothesis that thrombin-induced attachment of platelets to endothelial cells is modulated by EDRF. Thrombin significantly increased binding of radiolabelled platelets to cultured endothelium and to an intact pulmonary vasculature under flow conditions. These increases in binding were potentiated with hemoglobin (HB) and inhibited by superoxide dismutase (SOD) in both systems. We suggest that thrombin, in addition to enhancing platelet adhesion, elicits EDRF release from endothelial cells and that EDRF serves an antithrombotic function in the down regulation of platelet adhesion.

Interspecies differences in renal localization of cyclooxygenase isoforms: implications in nonsteroidal antiinflammatory drug-related nephrotoxicity.

Cyclooxygenase (COX) exists in 2 related but unique isoforms: one is constitutive (COX-1) and functions in normal cell physiology, and the other is inducible (COX-2) and is expressed in response to inflammatory stimuli. Nonsteroidal antiinflammatory drugs (NSAIDs) cause renal toxicity following inhibition of renal cyclooxygenases. Humans and animals exhibit differences in susceptibility to NSAID-related renal toxicity, which may be associated with differences in expression of 1 or both isoforms of COX in the kidney. In this study, we evaluated COX-1 and COX-2 expression in the kidneys of mixed-breed dogs, Sprague-Dawley rats, cynomolgus monkeys, and humans. In addition, the effect of volume depletion on renal COX expression was investigated in rats, dogs, and monkeys. COX expression was evaluated using 1 or more of the following procedures: reverse transcriptase polymerase chain reaction, in situ hybridization, and immunohistochemistry. We demonstrated that both COX isoforms are expressed in the kidneys of all species examined, with differences in the localization and level of basal expression. COX-1 is expressed at high levels in the collecting ducts and renal vasculature of all species and in a small number of papillary interstitial cells in rats, monkeys, and humans. Basal levels of COX-2 are present in the maculae densa, thick ascending limbs, and papillary interstitial cells in rats and dogs and in glomerular podocytes and small blood vessels in monkeys and humans. COX-2 expression is markedly increased in volume-depleted rats and dogs but not monkeys. These results indicate that significant interspecies differences exist in the presence and distribution of COX isoforms, which may help explain the difference in species susceptibility to NSAID-related renal toxicity.