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- Author or Editor: C. P. Coyne x
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SUMMARY
The antibiotic polymyxin B sulfate is a cationic polypeptide with a unique cyclical configuration and distinct cationic characteristics. In this investigation, polymyxin H was evaluated to determine its ability to prevent synthesis of lactic acid and tumor necrosis factor-α (tnf-α) by lipopolysaccharide-stimulated strain RAW 264.7 macrophage-like cell populations. In this context, gradient concentrations of polymyxin B were formulated in the presence of fixed concentrations of lipopolysaccharide fractions from Escherichia coli (B4:0111), E coli (J5), Klebsiella pneumoniae, Pseudomonas aeruginosa. Salmonella minnesota, and S typhimurium (Re). Quantitation of tnf-α was established by the application of a tissue culture-based biological assay system, using the WEHI 164 clone 13 indicator cell line. Investigations also included evaluation of the ability of gradient concentrations of lipopolysaccharide fractions from E coli (B4:0111), E coli Q5), K pneumoniae, P aeruginosa, S minnesota, and S typbimurium (Re) to form a complex with polymyxin B. This was established through application of high-performance thin-layer chromatography techniques. On the basis of the known molecular characteristics of lipopolysaccharide, its lipid A-core subfractions, and polymyxin B, these results imply that cytoprotective properties of polymyxin B are attributable to direct interaction and subsequent complex formation. More specifically, the mechanism by which polymyxin B exerts affinity for lipopolysaccharide fractions is proposed to occur through attractive ionic interactions established between the cationic diami-nobutyric acid residues of polymyxin B and the mono- or diphosphate group(s) of the lipid A-corc moiety. It is highly probable that this molecular phenomenon is accompanied by hydrophobic interactions established between the terminal methyloctanoyl or methylheptanoyl groups of polymyxin B and the saturated carbon chains of the lipid A-core subfraction of lipopolysaccharide fractions
Summary
A selected group of pharmaceutical compounds were evaluated for the ability to inhibit the biochemical activity of fibrinoligase (coagulation factor XIIIa*) in pooled equine plasma. Criteria for the pharmaceuticals selected were based on the mechanism of the transglutamination biochemical reaction mediated by coagulation factor XIIa*. These criteria were complemented by recognition of the molecular configuration and chemical composition of amino acid residue side chains involved in the process of covalent fibrin monomer polymerization (cross-linking, transglutamination) mediated by this enzyme. Each pharmaceutical was evaluated individually and in combination with other potential coagulation factor XIIIa* inhibitors in an effort to detect additive and synergistic phenomenon. In this context, pharmaceuticals with a carbonylamide (eg, cefuroxime, Girard's reagent-P, prolinamide) were applied in concert with compounds with a terminal amine (eg, d-arginine, l-lysine) functional group. In concept, this method theoretically served to competitively simulate glutamine and lysine amino acid residues within strands of fibrin monomer substrate involved in phase I (carbonylamide) and phase II (terminal amine) of the transglutamination reaction (covalent fibrin monomer cross-linking). Halogen-dinitro and ethylene compounds were also evaluated because of their reported ability to inactivate enzyme systems dependent on an intact sulfhydryl group located at their biochemically active site (eg, cystine amino acid residue). This group of pharmaceutical compounds failed to inhibit the biochemical activity mediated by coagulation factor XIIIa* in equine plasma.
SUMMARY
In this preliminary investigation, various hematologic variables potentially influential in determining the degree of blood viscosity were evaluated in 10 Thoroughbred horses subjected to competitive acute running exercise. Following completion of sprints over a distance of 1.25 miles, mean percent (± sd) increases in pcv (38.3 ± 12.9%), rbc (47.8 ± 15.3%), and rouleaux index (232.7 ± 176.8%) were recognized. Simultaneous increases in total plasma protein (28.3 ± 5.31%), serum albumin (26.7 ± 6.80%), α1-globulin (60.0 ± 49.0%), α2-globulin (25.5 ± 27.9%), β1-globulin (46.7 ± 21.1%), (32-globulin (35.0 ± 50.6%), γ1- and 2-globulins (38.7 ± 29.6%), and plasma fibrinogen (12.5 ± 10.4%) concentrations increased simultaneously. Horses also had consistent decreases in albumin:globulin ratio (- 10.0 ± 7.43%). Alterations in these hematologic values after acute running exercise in Thoroughbred horses accompanied increases in serum (69.3 ± 39.7%), plasma (39.7 ± 11.9%), and blood (134.7 ± 55.3%) viscosity.
Summary
Several pharmaceutical compounds were evaluated for their ability to selectively inhibit activated coagulation factor-XIII-like enzyme activity (eg, Xllla*) in pooled equine plasma. Presence of coagulation factor-XIIIa*-like enzyme activity in plasma was established by assay procedures involving incorporation of the fluorescent amine compound, monodansylcadaverine, into purified casein, which served as a protein substrate.
Pharmaceuticals inhibitory to coagulation factor-XIIIa*-like enzyme activity were recognized by plasma gel formation of high spectrophotometric transmittance (transparency), solubility of transparent fibrin gels in concentrated urea solution, in conjunction with simultaneous depletion of native fibrinogen fractions, and production of fibrin monomer. Compounds acting primarily as anticoagulants were recognized by lack of plasma gel formation, but retaining high spectrophotometric transmittance and no detectable depletion of native fibrinogen fractions. Compounds failing to inhibit either thrombin-mediated fibrinogen-fibrin transformation (ie, coagulation) or coagulation factor-XIIIa*-like enzyme activity were recognized by opaque plasma gels caused by fibrin polymerization, low spectrophotometric transmittance values, and coinciding with depletion of native fibrinogen fractions.
Pharmaceuticals capable of exerting selective inhibition of coagulation factor-XIIIa*-like enzyme activity were further classified as competitive inhibitors of phase 1 (carbamide) or phase 2 (terminal amine) of the transglutamination process.
Summary
Objectives of this investigation were to extract and isolate protein fractions inhibitory to the cytotoxic properties of tumor necrosis factor-α (tnf-α). In this context, mixed populations of wbc were harvested from equine blood and were stimulated with a combination of a synthetic chemotactic peptide and a calcium ionophore. Several methods were subsequently applied for the initial preparation of cell-free crude protein extracts, including fractional precipitation with gradient concentrations of ammonium sulfate and preparative-scale isoelectric focusing. In addition, protein fractions were harvested from extracts of concentrated equine urine. Protein extracts of urinary origin were further separated by gel-filtration column chromatography.
Identification of protein fractions possessing properties inhibitory to the cytotoxic characteristics of tnf-α was facilitated by a tissue culture-based technique for the biological assay of tnf-α-mediated cytotoxicity. Purified protein extracts possessed a marked ability to inhibit or neutralize the cytotoxic properties of tnf-α, on the basis of survival of murine fibrosarcoma cell populations, compared with appropriate negative and positive reference controls. Relative purity of inhibitors and estimation of approximate molecular weight were established by conventional reducing and nonreducing sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis. Equine inhibitory protein fractions from mixed wbc populations, purified in the manner described, had molecular weights of 70,000 to 80,000 and 28,000. An analogous protein fraction of 28 kDa also was isolated from equine concentrated urine. Estimated isoelectric point of tnf-α inhibitor protein fractions was between pH of 5.5 and 6.1. These physical characteristics of equine tnf-α inhibitor protein fractions were similar to those described for a membrane-associated tnf-α receptor protein shed from chemotaxin- and calcium-ionophor-stimulated human wbc populations.
Summary
Four hours prior to exercise on a high-speed treadmill, 4 dosages of furosemide (0.25, 0.50, 1.0, and 2.0 mg/kg of body weight) and a control treatment (10 ml of 0.9% NaCl) were administered iv to 6 horses. Carotid arterial pressure (cap), pulmonary arterial pressure (pap), and heart rate were not different in resting horses before and 4 hours after furosemide administration. Furosemide at dosage of 2 mg/kg reduced resting right atrial pressure (rap) 4 hours after furosemide injection. During exercise, increases in treadmill speed were associated with increases in rap, cap, pap, and heart rate. Furosemide (0.25 to 2 mg/kg), administered 4 hours before exercise, reduced rap and pap during exercise in dose-dependent manner, but did not influence heart rate. Mean cap was reduced by the 2-mg/kg furosemide dosage during exercise at 9 and 11 m/s, but not at 13 m/s. During recovery, only rap was decreased by furosemide administration. Plasma lactate concentration was not significantly influenced by furosemide administration. Furosemide did not influence pcv or hemoglobin concentration at rest prior to exercise, but did increase both variables in dose-dependent manner during exercise and recovery. However, the magnitude of the changes in pcv and hemoglobin concentration were small in comparison with changes in rap and pap, and indicate that furosemide has other properties in addition to its diuretic activities. Furosemide may mediate some of its cardiopulmonary effects by vasodilatory activities that directly lower pulmonary arterial pressure, but also increase venous capacitance, thereby reducing venous return to the atria and cardiac filling.
SUMMARY
Furosemide, which commonly is used as a prophylactic treatment for exercise-induced pulmonary hemorrhage in horses, may mediate hemodynamic changes during exercise by altering prostaglandin metabolism. To determine if furosemide's hemodynamic effects during exercise in horses could be reversed, cyclooxygenase inhibitors were administered with furosemide. Four treatments were administered 4 hours prior to treadmill exercise at 9 and 13 m/s. They included a control treatment (10 ml of 0.9% NaCl solution, iv), furosemide (1 mg/kg of body weight, iv) administered alone, and furosemide in combination with phenylbutazone (4 mg/kg, iv, q 12 h for 2 days) or with flunixin meglumine (1.1 mg/kg, iv, on the day of experiment). Five horses were randomly assigned to complete all treatments. Physiologic variables at rest prior to exercise were not influenced by treatments. Furosemide, administered alone, reduced mean right atrial pressure and mean pulmonary artery pressure during exercise. The combinations of furosemide and flunixin meglumine or furosemide and phenylbutazone, at both levels of exercise intensity, returned mean right atrial pressure and mean pulmonary artery pressure to the value of the control treatment. During rest and exercise, plasma lactate concentration, pcv, heart rate, mean carotid artery pressure, oxygen consumption, carbon dioxide elimination, and cardiac output were not altered by any of the treatments. At 5 minutes after exercise, the administration of furosemide, alone or with phenylbutazone, reduced mean right atrial pressure. Other measured variables were not significantly influenced by treatments during recovery from exercise. These results suggested that cyclooxygenase inhibition partially reverses the decrease in mean right atrial pressure or pulmonary artery pressure induced by furosemide during exercise. Furosemide may mediate some of its physiologic activities in exercising horses through the cyclooxygenase pathway.
Summary
The ability of polysulfated glycosaminoglycans (psgag) to inhibit the complement cascade was evaluated. The role of complement in inflammation and infection has been well documented. Inhibition of the complement cascade by psgag could explain why intra-articularly administered psgag diminish diarthrodial joint inflammation and potentiate septic arthritis in horses.
Hemolytic complement testing was performed to evaluate the effect of psgag on the equine classical and alternate pathways of complement, using rabbit erythrocytes as the target cells. Concentration of psgag between 0.2 mg/ml and 0.6 mg/ml significantly (P< 0.05) inhibited equine complement in dose-related fashion. Further increase in complement inhibition was not observed at psgag concentration >0.6 mg/ml. Difference was not apparent in the extent of inhibition of complement from each of the 4 horses tested. Polysulfated glycosaminoglycans appeared to inhibit the classical and alternate complement pathways equally, indicating possible effect on complement components common to both pathways. Heat inactivation of complement function completely inhibited (P<0.01) the hemolytic activity of the serum from all horses.
Abstract
OBJECTIVE
To investigate the effects and duration of orally administered prednisolone on renal function evaluated by glomerular filtration rate (GFR) determination and creatinine (Cr) and symmetric dimethylarginine (SDMA) concentrations as well as on urinalysis, electrolytes, and hydric status in healthy dogs.
ANIMALS
14 healthy Beagles.
PROCEDURES
In this prospective double-masked placebo-controlled study, dogs were randomized after baseline evaluation to receive a 7-day course of either prednisolone (1.5 to 2.0 mg/kg, PO, q 12 h) or a placebo. A repeated-measure design was performed, each dog participating in 4 successive sampling sessions. Clinical data, systolic blood pressure, CBC, and biochemical analyses including serum SDMA concentration, GFR determination, urine output quantification, and complete urinalysis were performed for all dogs the day before (D0) and at the end of steroid administration (D7) as well as 2 weeks (D21) and 4 weeks (D35) after the end of treatment.
RESULTS
At D7, when compared with baseline, GFR increased significantly in treated dogs, whereas creatinine and SDMA concentrations decreased significantly. GFR and Cr but not SDMA modifications persisted significantly at D21. None of the variables differed significantly from baseline at D35. The OR of presenting an albumin band on urine electrophoresis was 2.4 times as high in treated versus control dogs (OR, 36; 95% CI, 1.8 to 719.4; P = 0.02).
CLINICAL RELEVANCE
A short-term course of immune-suppressive prednisolone treatment in healthy dogs leads to a sustained but reversible renal hyperfiltration state. Modification in electrolytic variables can affect the clinical interpretation of blood work in such patients.
