The P2Y1 receptor-mediated leukocyte adhesion to endothelial cells is inhibited by melatonin.

Extracellular ATP (released by endothelial and immune cells) and its metabolite ADP are important pro-inflammatory mediators via the activation of purinergic P2 receptors (P2Y and P2X), which represent potential new targets for anti-inflammatory therapy. Endothelial P2Y1 receptor (P2Y1R) induces endothelial cell activation triggering leukocyte adhesion. A number of data have implicated melatonin as a modulator of immunity, inflammation, and endothelial cell function, but to date no studies have investigated whether melatonin modulates endothelial P2YR signaling. Here, we evaluated the putative effect of melatonin on P2Y1R-mediated leukocyte adhesion to endothelial cells and TNF-α production, using mesenteric endothelial cells and fresh peripheral blood mononuclear cells isolated from rats. Endothelial cells were treated with the P2Y1R agonist 2MeSATP, alone or in combination with melatonin, and then exposed to mononuclear cells. 2MeSATP increased leukocyte adhesion to endothelial cells and TNF-α production in vitro, and melatonin inhibited both effects without altering P2Y1R protein expression. In addition, assays with the Ca2+ chelator BAPTA-AM indicate that the effect of melatonin on 2MeSATP-stimulated leukocyte adhesion depends on intracellular Ca2+ modulation. P2Y1R is considered a potential target to control chronic inflammation. Therefore, our data unveiled a new endothelial cell modulator of purinergic P2Y1 receptor signaling.

Antioxidant effects of N-acetylserotonin: possible mechanisms and clinical implications.

This paper will review our recent data relevant to the antioxidant effects of N-acetylserotonin (NAS), the immediate precursor of melatonin, the pineal gland indole. Mechanisms of the antioxidant effects of NAS might involve interaction with melatonin type 3 receptors and nonreceptor mechanisms such as stimulation of glutathione peroxidase, an antioxidant enzyme; inhibition of lipid peroxidation; suppression of phospholipase A2 activation; attenuation of tumor necrosis factor-alpha production; prevention of pathological opening of the mitochondrial permeability transition pores; and inhibition of sepiapterin reductase, the key enzyme of biosynthesis of tetrahydrobiopterin, the essential cofactor of nitric oxide synthase. NAS actions on some of these enzymes might be receptor-mediated. Protective effects of NAS against oxidative damage are independent from the effect of melatonin and, depending on the model, are 5 to 20 times stronger than that of melatonin. Antioxidant effect of NAS might underpin its cognition-enhancing, antiaging, antidepressant, antihypertensive, and antitumor effects. NAS and its derivatives might be useful in protection against oxidative stress-related disorders (cell death, mutagenesis, aging) and diseases (sepsis, cancer, postischemic trauma, Alzheimer's disease, parkinsonism).

A novel inhibitor of N-ethylmaleimide-sensitive factor decreases leukocyte trafficking and peritonitis.

Endothelial exocytosis is an early stage in the process of leukocyte trafficking. N-ethylmaleimide-sensitive factor (NSF) plays a critical role in regulating exocytosis. We hypothesized that inhibitors of NSF decrease endothelial exocytosis and vascular inflammation. We designed a novel fusion polypeptide consisting of a human immunodeficiency virus transactivator of transcription (TAT) protein transduction domain joined to a NSF homohexamerization domain. We show that this TAT-NSF polypeptide inhibits the ATPase activity and the disassembly activity of NSF. Furthermore, the TAT-NSF polypeptide decreases endothelial cell exocytosis and reduces leukocyte adherence to endothelial cells in culture. Finally, the TAT-NSF polypeptide inhibits leukocyte rolling on murine venules in vivo and inhibits leukocyte trafficking into the peritoneal cavity in a murine model of experimental peritonitis. These data suggest that NSF is a critical regulator of leukocyte trafficking in vivo. Novel compounds that inhibit the exocytic machinery in endothelial cells may be useful anti-inflammatory drugs.

Mild hyperthermia down-regulates receptor-dependent neutrophil function.

Mild hypothermia impairs resistance to infection and, reportedly, impairs phagocytosis and oxidative killing of unopsonized bacteria. We evaluated various functions at 33 degrees-41 degrees C in neutrophils taken from volunteers. Adhesion on endothelial cells was determined using light microscopy. Adhesion molecule expression and receptors, phagocytosis, and release of reactive oxidants were assessed using flow cytometric assays. Adhesion protein CD11b expression on resting neutrophils was temperature-independent. However, up-regulation of CD11b with tumor necrosis factor (TNF)-alpha was increased by hypothermia and decreased with hyperthermia. Neutrophil adhesion to either resting or activated endothelial cells was not temperature-dependent. Bacterial uptake was inversely related to temperature, more so with Escherichia coli than Staphylococcus aureus. Temperature dependence of phagocytosis occurred only wi thopsonized bacteria. Hypothermia slightly increased N-formyl-L-methionyl-L-leucyl-phenylalanine receptors on neutrophils: hyperthermia decreased expression, especially with TNF-alpha. N-formyl-L-methionyl-L-leucyl-phenylalanine-induced H2O2 production was inversely related to temperature, especially in the presence of TNF-alpha. Conversely, phorbol-13-myristate-12-acetate, an activator of protein kinase C, induced an extreme and homogenous release of reactive oxidants that increased with temperature. In contrast to nonreceptor-dependent phagocytosis and oxidative killing, several crucial receptor-dependent neutrophil activities show temperature-dependent regulation, with hypothermia increasing function. The temperature dependence of neutrophil function is thus more complicated than previously appreciated.

Antioxidant activity of nitro derivative of aspirin against ischemia-reperfusion in hamster cheek pouch microcirculation.

Aspirin that has been chemically combined with a nitric oxide (NO) donor (NCX-4016) has been shown to inhibit cyclooxygenase and prostaglandin generation while maintaining the inhibitory effects of aspirin. The possible role of reactive oxygen species (ROS) in the action of NCX-4016 in ischemia-reperfusion (I/R) has not been studied. Furthermore, we were interested in comparing the effects of a conventional NO donor [2,2'-hydroxynitrosohydrazino-bis-etanamine (DETA/NO)] and NCX-4016 at the microvascular level in the hamster cheek pouch visualized by using an intravital fluorescent microscopy technique. Microvascular injury was assessed by measuring diameter change, the perfused capillary length (PCL), and leukocyte adhesion. Animals were treated with NCX-4016 (100 mg/kg or 30 mg.kg(-1).day(-1) for 5 days po) or DETA-NO (0.5 mg/kg). Mean arterial blood pressure increased slightly but significantly after NCX-4016 treatment. During 5- and 15-min reperfusion, lipid peroxides in the systemic blood increased by 72 and 89% vs. baseline, respectively, and were still higher than in basal conditions after 30-min reperfusion in the I/R group. Pretreatment with NCX-4016 maintained ROS at normal levels; increased arteriolar diameter, blood flow, and PCL; and decreased leukocyte adhesion (P < 0.05). DETA-NO decreased ROS during 30-min reperfusion; however, later there was a significant increase during reperfusion. DETA-NO decreased leukocyte adhesion (P < 0.05) but microvascular permeability increased after 30 min of reperfusion. In conclusion, NCX-4016 attenuates oxidative stress and prevents arteriolar constriction during I/R, whereas DETA-NO increases lipid peroxides in the systemic blood and permeability after reperfusion.

Probucol restores the defective leukocyte-endothelial interaction in experimental diabetes.

Defective leukocyte-endothelial interactions have been observed in experimental type 1 diabetes. One of the mechanisms involved in the late complications of diabetes mellitus is the formation of free radicals species. Antioxidant treatment has been demonstrated to have beneficial effects on the complications observed in this pathology. Using intravital microscopy to visualize venules of the internal spermatic fascia, we demonstrated that the defective leukocyte-endothelial interactions in alloxan-induced diabetic rats could be corrected by probucol treatment. The defects were quantitated by the number of leukocytes rolling along the venular endothelium, sticking to the venular wall after topical application of zymosan-activated plasma (10%-0.1 ml) or leukotriene B4 (1 microM/0.1 ml) and migrated after the application of a local irritant stimulus (carrageenan, 100 microg/0.1 ml). Leukocyte counts, erythrocyte velocity and venular shear rate, unaltered in diabetic rats, were not modified by this treatment. Reactive oxygen species formation by endothelial cells increased in diabetic preparations, and the reduced expression of adhesion molecules such as intercellular adhesion molecule-1 (ICAM-1) and P-selectin in cross-sections of the whole testis of the animals, were both restored by the antioxidant agent. Therefore, antioxidant treatment improves leukocyte-endothelial interaction in diabetic rats at least in part by restoring the expression of adhesion molecules in venules of diabetic rats.

Heme-oxygenase-1 mRNA expression affects hemorrhagic shock-induced leukocyte adherence.

BACKGROUND: Hemorrhagic shock-related leukocyte adherence to endothelial cells is a key step in microvascular injury-related organ damage. Heme-oxygenase-1 (HO-1) metabolizes heme, a potent cytotoxic agent, to carbon monoxide and biliverdin. We hypothesized that changing HO-1 expression would change leukocyte adherence after hemorrhagic shock. METHODS: Rats were administered hemin, zinc protoporphyrin, or vehicle 6 hours before surgery. HO-1 expression was determined by reverse-transcriptase polymerase chain reaction in various tissues. Shock was induced in urethane-anesthetized animals by decreasing mean arterial blood pressure to 40 mm Hg for 60 minutes, followed by standard resuscitation measures. Leukocyte adherence was measured by intravital microscopy in rat mesenteric venules. RESULTS: Hemin, hemorrhagic shock, and the combination resulted in significantly increased HO-1 expression, whereas zinc-protoporphyrin (ZNPP) resulted in significantly decreased leukocyte adherence. After hemorrhagic shock and hemin administration, leukocyte adherence was significantly decreased 60 minutes into resuscitation (7.92 +/- 2.29 vs. 4.84 +/- 0.71 cells/100 microm, p < 0.05) and significantly increased after ZNPP plus shock (14.08 +/- 3.95, p

Functional relevance of activated beta1 integrins in mercury-induced nephritis.

Cell adhesion through different adhesion molecules is a crucial event in the inflammatory response. Integrins can only bind and mediate cellular adhesion after their activation by different specific stimuli. The state of beta1 integrin activation can be assessed by a group of monoclonal antibodies (HUTS) that selectively recognize beta1 integrins in their active form. A similar activated epitope in the rat was defined using the anti-human monoclonal antibody HUTS-21, which recognizes an activation-dependent epitope on the beta1 chain. It was found that the divalent cations Mn(2+) and Hg(2+) were able to induce in vitro the activation of beta1 integrins on rat lymphocytes. The Hg(2+) cation induces an autoimmune disease in the Brown Norway rat characterized by synthesis and glomerular deposits of anti-glomerular basement membrane antibodies, proteinuria, and interstitial nephritis. Using the mercury model of nephritis, it was found that the expression of HUTS-21 epitope is induced in vivo in rat lymphocytes, and its appearance is correlated with the other parameters at the onset of the disease. In addition, the administration of HUTS-21 monoclonal antibody to HgCl(2)-treated rats offered evidence of its protective effects (1) against infiltration of renal interstitium by leukocytes, and (2) in the reduction of anti-glomerular basement membrane synthesis and glomerular deposition. Nevertheless, urinary protein values remained unaffected. These results demonstrate a key role of beta1-activated integrins in both leukocyte cell-cell interactions and leukocyte infiltration pathway mechanism, and also indicate that leukocyte migration may have less importance in the development of this disease than previously thought.

Influence of tolrestat on the defective leukocyte-endothelial interaction in experimental diabetes.

One of the most devastating secondary complications of diabetes is the blunted inflammatory response that becomes evident even in the very early stages of poorly controlled diabetes mellitus. While the etiology of this diminished response is not clearly understood, it has been linked to a decrease in the respiratory burst of neutrophils, as well as a decrease in microvessel response to inflammatory mediators and defective leukocyte-endothelial interactions. Using video microscopy to visualize vessels of the internal spermatic fascia, we have characterized leukocyte-endothelial interactions in alloxan-induced diabetic and in galactosemic rats by quantitating the number of leukocytes rolling along the venular endothelium and the number of leukocytes sticking to the vascular wall after topical application of zymosan-activated plasma or leukotriene B(4) (1 ng/ml), as well as after the application of a local irritant stimulus (carrageenan, 100 microg). We observed that while 33 days of alloxan-induced diabetes or 7 days of galactosemia had no effect on total or differential leukocyte counts and on the wall shear rate, both treatments significantly (P<0.001) reduced the number of leukocytes rolling along the venular endothelium by about 70% and the number of adhered leukocytes in postcapillary venules by 60%. These effects were not observed in diabetic and galactosemic animals treated with an aldose reductase inhibitor. The results suggest that impaired leukocyte-endothelial cell interactions are a consequence of an enhanced flux through the polyol pathway.

Nitric oxide synthase inhibitor augments post-ischemic leukocyte adhesion in the cerebral microcirculation in vivo.

The objective was to examine the effect of the nitric oxide synthase inhibitor, N omega-nitro-L-arginine methyl ester (L-NAME) on leukocyte adhesion in the cerebral microcirculation during reperfusion following partial forebrain ischemia in the rat. Intravital fluorescence video-microscopy through a closed cranial window was used to visualize leukocyte-endothelium interaction in small pial veins of 15-100 microns diameter. Forebrain ischemia was produced by the ligation of both common carotid arteries plus elevation of the intracranial pressure to 20 mmHg for 60 min. The number of leukocytes adhering to the endothelium for longer than 3 sec was determined during ischemia (5 min and 60 min) and during reperfusion (5 min and 60 min). Two experimental groups were treated with either L-NAME or its inactive enantiomer D-NAME (20 mg kg-1 i.v.) 30 min prior to reperfusion. In a third group, also treated with D-NAME, post-ischemic hyperemia was prevented by lowering the ICP without removing the occlusion of common carotid arteries (partial reperfusion). The velocity of flow adjacent to the endothelial surface of pial veins was measured by tracking the movement of fluorescently labeled red blood cells as flow markers before and after ischemia. During ischemia, the number of adhering leukocytes increased approximately two-fold at 5 min, and three-fold at 60 min. In the D-NAME-treated group with complete reperfusion, leukocyte adhesion returned to the baseline level by 60 min of reperfusion. However, in the L-NAME-treated group, leukocyte adhesion remained elevated at 60 min of reperfusion. Post-ischemic flow velocity was significantly decreased (-66%) from control after L-NAME treatment whereas it was increased (+53%) in the D-NAME-treated group. In the partial reperfusion group, leukocyte adhesion continued to increase after the first hour of ischemia and reached a level 2.7-fold over baseline at 60 min reperfusion. Flow velocity remained below control (-26%) at 60 min reperfusion. Leukocyte adhesion was absent in pial arteries and no plugging by leukocytes was observed in cortical capillaries. The results suggest that leukocyte adhesion in small pial veins increases during 1 h forebrain ischemia and continues to increase during reperfusion if the velocity of flow or shear rate is low. The increase in leukocyte adhesion is reversible if flow velocity is elevated during reperfusion. L-NAME prevents post-ischemic hyperemia and augments leukocyte adhesion principally via a decrease in velocity or shear rate.

Differential influence of tyrosine residues of the common receptor beta subunit on multiple signals induced by human GM-CSF.

BACKGROUND: GM-CSF induces the proliferation and differentiation of a broad range of hematopoietic cells. Stimulation with this cytokine results in the induction of rapid tyrosine phosphorylation of cellular proteins, including the common beta subunit of its receptor. However, the significance of beta subunit phosphorylation with regard to regulation of signaling molecules and the functions of GM-CSF is less well understood. OBJECTIVE: To investigate the role of cytoplasmic tyrosine residues, we performed mutational analyses of the beta subunit. METHODS: A series of beta subunit mutants with 1, some, or all of 8 cytoplasmic tyrosines converted to phenylalanines were constructed. These beta subunit mutants were expressed in murine BA/F3 cells together with the wild-type human alpha subunit, and their potentials to transmit signals were examined. RESULTS: A mutant beta subunit lacking all cytoplasmic tyrosines (Fall) activated JAK2, but not Shc, SHP-2, the Raf/ERK cascade, signal transducer and activator of transcription 5 (STAT5), or the c-fos promoter, nor did it effectively induce proliferation. Adding back each tyrosine to Fall revealed that (1) Tyr577 was necessary and sufficient for Shc phosphorylation; (2) Tyr577, Tyr612, and Tyr695 were involved in activation of SHP-2, the Raf/ERK cascade, and c-fos transcription; and (3) all tyrosines, but particularly Tyr612, Tyr695, Tyr750, and Tyr806, facilitated STAT5 activation. Impaired cell growth was also partly restored by any of the tyrosines. Interestingly, the Fall mutant was still capable of suppressing apoptosis and maintaining cell viability. CONCLUSION: These results provide evidence that beta subunit tyrosines possess distinct, yet overlapping, roles in the activation of multiple signaling pathways induced by GM-CSF. In addition, GM-CSF may activate both beta subunit tyrosine-dependent and -independent pathways, with the latter being sufficient to transmit survival signals.

The biologic role of B2 integrins in the host response to expanded polytetrafluoroethylene.

We hypothesized that the adherence of leukocytes to a vascular graft surface is mediated by B2 integrins. We studied integrin expression and monoclonal antibody inhibition of peripheral blood leukocyte (PBL) binding in vitro and CD11b expression in vivo. Human PBL were grown on ePTFE in culture and labeled with monoclonal antibodies to CD11a, -b, -c, and/or CD18. Percentage of monoclonal antibody binding and inhibition of leukocyte adherence was studied for up to 30 min using flow cytometry. ePTFE segments were implanted subcutaneously in SH-1 mice and PBL harvested 4 days later. PBL binding of monoclonal antibodies against CD11b was measured using flow cytometry. CD18 was constitutively expressed and CD11a decreased over time. CD11b increased from 41 to 62% and CD11c increased transiently (P < 0.003, P < 0.005). Inhibiton of PBL adherence was greatest by CD11b (34%) and CD11b + CD18 (57%) (P < 0.005, P < 0.0025). Implanted ePTFE caused a fourfold increase in PBL monoclonal antibody binding of CD11b (P < 0.001) compared to sham, Staphylococcus aureus alone, and combination of ePTFE and S. aureus. In conclusion, leukocyte integrins play a central role in the interaction between PBLs and ePTFE as measured by binding of monoclonal antibodies and inhibition of PBL adherence. This is also true in vivo since PBL increase CD11b expression four times when ePTFE is compared to sham. These observations indicate a potential role in vivo for B2 integrins in vascular prosthetic infections.

Peritoneal dialysis solution attenuates microvascular leukocyte adhesion induced by nitric oxide synthesis inhibition.

In the mesenteric microcirculation, inhibition of nitric oxide (NO) synthesis results in an inflammatory response through increased leukocyte adherence to the microvascular postcapillary venular endothelium. Recent studies have demonstrated that elevated concentrations of endogenous NO synthesis inhibitors are present in renal failure. How peritoneal dialysis solutions may affect leukocyte-endothelial interactions during inflammation induced by NO synthesis inhibition has been previously unknown. Using in vivo intravital microscopy of the rat mesenteric postcapillary venules, microvascular leukocyte adherence was quantitated during baseline conditions in which the mesentery was superfused with a buffer solution, followed by the superfusion of a NO synthesis inhibitor NG-nitro-L-ARGININE methyl ester (L-NAME) added to the buffer, followed by 4.25% Dianeal (4.25% D). When compared to baseline, L-NAME increased the mean number of adherent leukocytes by fivefold (2.2 +/- 0.9 vs 11.6 +/- 3.6 leukocytes/100 microns venule/10 min, p < 0.05), while 4.25% D quickly reversed the L-NAME-induced inflammatory response, returning the number of adherent leukocytes back to baseline values (11.6 +/- 3.6 vs 2.4 +/- 1.3 leukocytes/100 microns venule/ 10 min, p < 0.05). These results confirm that NO synthesis inhibition induces inflammation in mesenteric postcapillary venules. Superfusion of 4.25% D reverses leukocyte adhesion induced by NO synthesis inhibition. Thus, a standard peritoneal dialysis solution (4.25% D) reverses the leukocyte-adhesive effects of NO synthesis inhibition in the mesenteric microcirculation.

Biologic interventions in rheumatoid arthritis.

An increasing understanding of the immunopathogenesis of rheumatoid arthritis and advances in biotechnology have lead to the therapeutic application of immune-specific interventions. The complexity of this disorder has generated numerous investigations using a wide range of biologic interventions. This article presents the current and potential targets for such biopharmaceutical agents and discusses their utility in rheumatoid arthritis.

Inhibition of human monocyte adhesion to endothelial cells by the coumarin derivative, cloricromene.

1. The ability of the coumarin derivative cloricromene (8-monochloro-3-beta-diethylaminoethyl-4-methyl-7-ethoxy- carbonylmethoxycoumarin) to inhibit monocyte adhesion to human cultured umbilical vein endothelial cells (HUVEC) was investigated. 2. Cloricromene (10-200 microM) inhibited, in a concentration-dependent manner, the adhesion of both resting and activated monocytes to HUVEC. Significant inhibition was reached with drug concentrations ranging between 15 to 30 microM. 3. The inhibitory activity was, at least in large part, directed to monocytes since no inhibition was observed after selective preincubation of HUVEC with cloricromene and the drug maintained its effect also on monocyte adhesion to paraformaldehyde-treated HUVEC. 4. Inhibition was maximal after 1 min of exposure of monocytes to cloricromene and persisted even in the absence of the drug. 5. Both basal and chemoattractant-mediated monocyte adhesion was inhibited by cloricromene as it was by TS1/18, a monoclonal antibody (mAb) directed to beta 2 integrins; however, cytofluorimetric analysis showed that cloricromene was unable to modulate the expression of beta 2 integrins on the monocyte surface. 6. When monocyte adhesion was mediated by a large set of adhesive receptors, as obtained after treatment of HUVEC with either interleukin 1 beta (IL-1; 50 ng ml-1) or tumour necrosis factor-alpha (TNF; 100 u ml-1), the inhibitory effect of cloricromene was considerably reduced. 7. The results of this study show that cloricromene may regulate monocyte adhesion to HUVEC, an event relevant in vivo in the pathogenesis of inflammatory and atherosclerotic processes.

The involvement of Kupffer cells in carbon tetrachloride toxicity.

Carbon tetrachloride (CCl4) is a classical pericentral hepatotoxicant; however, precise details of its mechanism of action remain unknown. One possibility is that Kupffer cells participant in this mechanism since CCl4 elevates calcium, and the release of toxic eicosanoids and cytokines by Kupffer cells is calcium-dependent. Therefore, these studies were designed to evaluate the role of Kupffer cells in CCl4 toxicity in the rat in vivo. Kupffer cells were destroyed selectively with gadolinium chloride treatment (10 mg/kg GdCl3 iv) 1 day prior to administration of CCl4 (4 g/kg ig). Twenty-four hours after CCl4 treatment, rats were anesthetized, blood samples were drawn for aspartate aminotransferase (AST) determination, which is indicative of parenchymal cell damage, and trypan blue was infused into the liver to stain the nuclei of dead hepatocytes. AST levels were in the normal range and trypan blue staining was negligible in livers from vehicle- or GdCl3-treated rats. As expected, CCl4 treatment alone elevated AST levels to values over 4000 U/liter and caused massive cell death (60-90 trypan blue-positive cells/pericentral field). In dramatic contrast, the elevation in AST and cell death due to CCl4 were almost completely prevented by GdCl3 treatment. In attempts to understand this phenomenon, metabolic and detoxification pathways were assessed. CCl4 is metabolized via cytochrome P450 II.E.1; however, GdCl3 treatment did not alter this pathway as assessed from p-nitrocatechol formation from the selective substrate, p-nitrophenol. GdCl3 treatment also had no effect on hepatic glutathione levels. On the other hand, GdCl3 treatment significantly reduced infiltration of neutrophils resulting from exposure to CCl4. These data clearly support the hypothesis that Kupffer cells participate in the mechanism of toxicity of CCl4 in vivo, possibly by release of chemoattractants for neutrophils.

Therapeutic immunosuppression of T cells.

Current immunosuppressive therapy carries a range of unwanted side effects, and tends to penalize the whole immune system. It is desirable to develop therapies that are more selective for antigen-reactive cells. As T lymphocytes use a wide range of surface receptors to interact with antigen-bearing cells and with each other, much interest is devoted to trying to develop agents that selectively block the interaction of these receptors with their ligands, and others that could be used to reprogram the immune system so that it might become tolerant to the antigens rather than attack them.

Complete prevention of myocardial stunning, contracture, low-reflow, and edema after heart transplantation by blocking neutrophil adhesion molecules during reperfusion.

This study tested the hypothesis that preventing neutrophil adhesion during reperfusion, by blocking either the neutrophil membrane CD18 integrin complex or its endothelial and myocyte ligand, intercellular adhesion molecule-1 (ICAM-1), would reduce myocardial inflammation and edema and improve reflow and ventricular function after heart preservation and transplantation. After cardioplegia and insertion of a left ventricular balloon, rabbit hearts were heterotopically transplanted into recipient rabbits either immediately (immediate, n = 12) or after preservation in 4 degrees C saline (3 hours of ischemia, n = 33). Forty-five minutes before reperfusion, recipients of preserved hearts received intravenous infusions of either saline (vehicle, n = 13), anti-CD18 monoclonal antibody (Mab) R15.7 (2 mg/kg) (anti-CD18, n = 10), or anti-ICAM-1 Mab R1.1 (2 mg/kg) (anti-ICAM, n = 10). During 3 hours of reperfusion the slope of the peak-systolic pressure-volume relation and its volume-axis intercept, the exponential elastic coefficient of the end-diastolic pressure-volume relation, the unstressed ventricular volume, and the time constant of the exponential left ventricular pressure decay after dP/dtmin were serially measured. Myocardial blood flow was measured with microspheres from which coronary vascular resistance was calculated. After explanation, the degree of myocardial inflammation, estimated by tissue neutrophil sequestration (myeloperoxidase assay) and myocardial water content were determined. Within each group no significant differences in measurements made at 1, 2, and 3 hours of reperfusion were noted. Compared with the immediate transplantation group, the vehicle group demonstrated a significant increase in myeloperoxidase activity (3380 +/- 456 versus 1712 +/- 552 microU/gm, p < 0.05), coronary vascular resistance (115.5 +/- 13.4 versus 70.5 +/- 10.6 U/gm, p < 0.05), and myocardial water content (79.8% +/- 0.4% versus 75.6% +/- 1.3%, p < 0.05), a significant decrease in unstressed ventricular volume (a leftward shift in the end-diastolic pressure-volume relation) (-0.49 +/- 0.24 versus 0.28 +/- 0.21 ml, p < 0.05), and a marked prolongation in exponential left ventricular pressure delay after dP/dtmin (156.64 +/- 3.81 versus 37.25 +/- 3.34 msec, p < 0.01).(ABSTRACT TRUNCATED AT 400 WORDS)

Adhesion promotes transcellular leukotriene biosynthesis during neutrophil-glomerular endothelial cell interactions: inhibition by antibodies against CD18 and L-selectin.

Eicosanoid formation by transcellular routes can amplify the levels and types of lipid mediators within a local milieu. To evaluate the role of adhesion in this process, we assessed the influence of mAb against adhesion molecules on LTC4 generation by PMN-endothelial cell interaction. Transcellular LTC4 generation was initiated by addition of fMLP to coincubations of GM-CSF-primed PMN and TNF-activated endothelial cells cultured from kidney glomeruli. Both PMN-endothelial cell adhesion and transcellular LTC4 generation were inhibited by mAb against leukocyte L-selectin and CD18. These results indicate that cytokine-treated PMN and endothelial cells generate LTC4 via transcellular routes by receptor-triggered mechanisms. They suggest that adhesion promotes transcellular eicosanoid biosynthesis and that adhesion molecules may also be targets for blockade of transcellular biosynthesis of lipid mediators.