Martin A. Wasserman

Inflammatory mediators of experimental colitis in rats.

Colonic inflammation was induced in rats by intracolonic administration of 0.25 ml of 50% ethanol containing 30 mg of trinitrobenzene sulfonic acid (TNB). Control rats were treated with 0.25 ml of 50% ethanol or with 30 mg of TNB in 0.25 ml of saline. After 24 h, mucosal ulceration and hemorrhage were observed in TNB/ethanol-, 50% ethanol-, and to a lesser extent, in TNB/saline-treated rats. After 1 wk, mucosal damage was completely resolved in the 50% ethanol and TNB/saline-treated rats but the lesions in the TNB/ethanol-treated rats persisted and progressed to a chronic active inflammatory process after 3 wk. Myeloperoxidase activity was significantly elevated in mucosal scrapings from all treatment groups at all time intervals when macroscopic and microscopic mucosal injury was evident. Interleukin-1 was found to be the most sensitive indicator of mucosal inflammation, and its mucosal values correlated with myeloperoxidase activity. Leukotriene B4 was increased in control rats at 1 wk and in TNB/ethanol-treated rats at all time intervals. The maximal increase in leukotriene B4 was observed at 1 wk. Thromboxane B2 generation was reduced while platelet activating factor generation was not increased in TNB/ethanol-treated rats. These results indicate that in this TNB/ethanol model of gut inflammation, myeloperoxidase activity and interleukin-1 are reliable and sensitive indicators of colonic inflammation, and that thromboxane B2 is not involved in the acute lesions, whereas leukotriene B4 appears in the chronic active inflammatory response.

The effect of synthetic leukotrienes on tracheal microvascular permeability

The effect of synthetic leukotrienes (LT) C4, D4 and E4 on the permeability of the airway microvasculature to plasma albumin was quantitatively evaluated using an in situ guinea pig tracheal model. Vascular permeability was measured as extravascular albumin content by employing 125I-bovine serum albumin and, in order to correct for blood volume, 51Cr-erythrocytes were used. Intratracheal injection of synthetic LTC4, LTD4 and LTE4 (0.1-1000 ng) produced dose-dependent increases in tracheal extravascular albumin content. The leukotrienes were approximately 100-1000 fold more potent than histamine, although histamine did produce a greater maximal increase in extravascular albumin than the leukotrienes. Methacholine did not increase extravascular albumin content. The microvascular permeability effect of LTD4 was antagonized by FPL 55712 but not by mepyramine; conversely, the effect of histamine was antagonized by mepyramine and not by FPL 55712. Additionally, indomethacin did not alter the LTD4-induced increases in tracheal vascular permeability. These results suggest that the effect of LTD4 on tracheal microvascular permeability is directly mediated and is not the indirect result of cholinergic stimulation, histamine release or de novo synthesis of cyclooxygenase products.

Investigation of leukotriene involvement in the vasopermeability response associated with guinea pig tracheal anaphylaxis: Comparison with cutaneous anaphylaxis

A direct comparison of the role of leukotrienes in mediating the increase in microvascular permeability associated with guinea pig tracheal and cutaneous anaphylaxis was obtained by simultaneous administration of inflammatory stimuli to both trachea and ear. The SRS-A antagonist, FPL 55712, reduced the increase in tracheal extravascular albumin content evoked by LTC4, LTD4, and LTE4 but failed to significantly reduce the tracheal microvascular permeability response associated with local anaphylaxis. Moreover, the inhibitory effect of the histamine H1-receptor antagonist, mepyramine, was not augmented by the additional presence of FPL 55712. In contrast to tracheal anaphylaxis, a distinct leukotriene component was indicated in cutaneous anaphylaxis since the mepyramine-FPL 55712 combination produced a greater inhibition than mepyramine alone. These results suggest that the degree of leukotriene involvement in anaphylaxis may vary between tissues.

2-Nor-leukotriene analogs: Antagonists of the airway and vascular smooth muscle effects of leukotriene C4, D4 and E4

A structural analog of LTD4, 4R-hydroxy-5S-cysteinylglycyl-6Z-nonadecenoic acid (4R, 5S, 6Z-2-nor-LTD1) has been synthesized and pharmacologically characterized. It significantly antagonized the contractile action of LTD4, LTC4 and LTE4 in guinea pig airways. In addition, this compound antagonized the in vitro vasoconstrictive effects of LTD4 in the guinea pig pulmonary artery. The study of a series of structural analogs of 4R, 5S, 6Z-2-nor-LTD1 suggests that the spatial separation of the C-1 (eicosanoid) carboxyl relative to the hydroxyl is a critical determinant in LTD4 agonist/antagonist activity.

Enhancement of leukotriene D4-induced contraction of guinea-pig isolated trachea by platelet activating factor.

The effect of platelet activating factor (PAF), a potent lipid mediator of inflammation, was examined in the induction of airway hyperreactivity to known mediators of anaphylaxis. Concentration-dependent contractions of the isolated guinea-pig trachea to PAF (10(-7)-10(-5) M) were produced and an EC50 value was found to be 7.5 X 10(-7) M. Pretreatment for 30 min with a known PAF inhibitor, CV-3988 (10(-5) or 10(-4) M), produced significant inhibition of PAF contractions; however, at 10(-6) M, CV-3988 had no effect. In the presence of meclofenamic acid (10(-6) M), the concentration-response curve to PAF was shifted significantly upward and to the left. This potentiation could be reversed by pretreating the tissues with the peptidoleukotriene antagonists, FPL 55712 or SK&F 102922 (10(-5) M). Pretreatment with PAF concentrations having essentially no intrinsic activity (10(-8), 10(-7)) significantly enhanced the contraction of guinea-pig trachea to various concentrations of LTD4 and to certain concentrations of a thromboxane mimic (U-46619). Pretreatment with lyso-PAF failed to potentiate the LTD4 response, while pretreatment with CV-3988 reverse the potentiation by PAF of the lower concentrations of LTD4. However, PAF failed to enhance contractions (with or without the presence of meclofenamic acid) to acetylcholine, histamine, PGD2 or LTC4 (in the presence of serine borate). These results indicate a possible role for PAF as a mediator of airway hyperreactivity.

Flushing and haemodynamic responses to vasopressin peptides in the rhesus monkey

1 The mechanism of the flushing, hypotension and tachycardia associated with i.v. administration of desGlyd(CH2)5D‐Tyr(Et)VAVP (SK&F 101926; 25 μg kg−1) and the selective V2 antidiuretic agonist, desamino‐8‐D‐arginine vasopressin (dDAVP; 3 μg kg−1) was studied in ketamine‐anaesthetized rhesus monkeys. 2 The flushing associated with SK&F 101926 was reduced by pretreatment with a mast cell stabilizer and by repeated administration of peptide (within 2–4 weeks). A similar desensitization to dDAVP‐associated flushing was observed on repeated administration. 3 Treatment with dDAVP also resulted in reduced SK&F 101926‐associated flushing. 4 The hypotension associated with SK&F 101926 was not affected by pretreatment with a mast cell stabilizer. A similar degree of hypotension was observed with repeated administration of either SK&F 101926 or dDAVP. 5 The tachycardia associated with SK&F 101926 was reduced by pretreatment with a mast cell stabilizer or repeated administration of SK&F 101926. Repeated administration of dDAVP, however, resulted in an enhanced tachycardia. 6 Indomethacin (5 mg kg−1 i.v.) did not alter the flushing or the hypotension associated with the administration of either SK&F 101926 or dDAVP, but resulted in an enhanced tachycardia to SK&F 101926. 7 Administration of a selective V1 vasopressor antagonist did not result in flushing, hypotension or tachycardia. 8 It was concluded that the flushing response to vasopressin‐like peptides in rhesus monkeys may be due to an action on mast cells, whereas the haemodynamic responses are not, but probably involve direct vasodilator actions.

Synthesis and LTD4-antagonist activity of desamino-2-nor-leukotriene analogs

A series of desamino-2-nor-leukotriene analogs has been prepared by the reaction of various thiols with several methyl trans-4,5-epoxy-6Z-alkenoates, followed by deprotection. The products were assessed for their ability to antagonize the LTD4-induced contraction of the isolated guinea pig trachea. Several compounds displayed potent leukotriene antagonist activity, i.e., KB values in the sub-micromolar range, while only minimally affecting basal airway tone. The most potent analog, 4-hydroxy-5-(2-carboxyethylthio)-6Z-nonadecenoic acid, antagonized both LTD4- and LTE4-induced contractions of the trachea in an apparently competitive fashion. These agents possess increased potency relative to SK&F 101132, the first leukotriene analog identified as having LT-antagonist activity. Thus, these results demonstrate that deletion of the peptide amino group can produce leukotriene analogs which have minimal intrinsic contractile activity on the isolated guinea pig trachea, yet possess potent leukotriene-antagonistic effects.

Analysis of the antagonist profile of SK&F 88046 on guinea pig trachea

SK&F 88046 preferentially antagonized the contractions elicited by the functional thromboxane (Tx) A2 mimics and structural endoperoxide analogs, U-44069 and U-46619, on the guinea pig trachea. This concentration-dependent antagonism was described by pA2 values of 7.03 against U-46619 and 6.97 against U-44069; the slopes of both Schild plots were 0.9. Whereas SK&F 88046 did not antagonize the tracheal contractions elicited by leukotriene (LT) C4 or D4, carbachol or histamine, this agent did antagonize the contractions induced by carbocyclic thromboxane A2 (CTA2) and prostaglandin (PG) F2 alpha and D2. At 1 X 10(-5)M SK&F 88046, the antagonism was described by -log KB values of 5.9 for CTA2, 5.5 for PGF2 alpha, and 6.4 for PGD2. Thus, SK&F 88046 was 3 to 20-fold more potent an antagonist of U-44069 and U-46619, suggesting that SK&F 88046 may function primarily as a thromboxane/endoperoxide antagonist on the guinea pig trachea.

The effect of calcium antagonists on histamine and leukotriene-induced tracheal microvascular permeability in the guine pig

Abstract The effects of several calcium antagonists, i.e., nifedipine, verapamil adn 8-[N,N-diethylamino]-octyl 3,4,5-trimethoxybenzoate hydrochloride (TMB-8), were evaluated in situ on agonist-induced increases in permeability of the airway microvasculature in anesthetized guinea pigs. Vascular permeability was measured as tracheal extravascular albumin content by using 125 I-bovine serum albumin and the utilization of 51 Cr labelled-erythrocytes to correct for blood volume. Intratracheal injections of histamine (1, 10 and 100 μg) or leukotriene (LT) D 4 (1, 10 and 100 ng) produced dose-dependent increases in extravasated radiolabelled albumin in the trachea. Although histamine produced a greater maximal response than LTD 4 , the latter provocation was tent times more potent than the former. Nifedipine, a dihydropyridine calcium slow channel blocker, exhibited dose-dependent (30, 100 and 300 μg/kg) inhibitory activity against histamine-induced increases in extravascular albumin, while another calcium slow channel blocker, verapamil (100, 300 and 1000 μg/kg), exhibited much less activity. TMB-8, a purported intracellular calcium antagonist (1 and 10 mg/kg), was observed to have some inhibitory activity versus histamine. Similar doses of all three calcium antagonists failed to significantly inhibit increases in tracheal microvascular permeability evoked by LTD 4 . These results suggest that differences in mediator-induced microvascular permeability in the guinea pig trachea are evident depending upon the agonist selected and the pool of calcium utilized.

Differences in the ability of salbutamol to prevent and reverse LTC4-induced contractions of the guinea-pig isolated trachea: Influence of l-serine borate

In the presence of l-serine borate, salbutamol was much more effective in reversing rather than preventing LTC4-induced contractions of guinea-pig trachea. This suggests that different mechanisms are involved in initiating versus maintaining LTC4-induced contractions. In addition, the ability of salbutamol pretreatment to prevent LTC4-induced contractions was reduced substantially in the presence of l-serine borate, suggesting that the metabolites of LTC4 (LTD4 and LTE4) are more sensitive to inhibition by beta-adrenoceptor agonists than is LTC4 itself.