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  • Author or Editor: Jana L. Cargile x
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SUMMARY

Tumor necrosis factor-α (tnf) is an important mediator of endotoxin-induced pathologic changes. To help define the role of tnf in equids with endotoxemia, the effects of pretreatment with a murine monoclonal antibody (mab) against equine tnf were evaluated in Miniature Horses given endotoxin. Five horses were given tnf mab at a dosage of 1.86 mg/kg of body weight, iv, and 5 were given control mab. Five minutes later, lipopolysaccharide (lps; Escherichia coli O55:B5), 0.25 µg/kg, was given to all horses by bolus iv infusion. Clinical signs of disease were monitored at intervals up to 24 hours after lps infusion, and blood was taken for determination of hematologic and clinical responses of horses given wbc count, pcv, plasma total protein concentration, plasma tnf activity, and serum mab concentration. Reduction of plasma tnf activity in anti-tnf-treated horses was highly significant (P < 0.001), compared with that in control horses. Horses given tnf mab had significantly improved clinical abnormality score (P < 0.010), lower heart rate (P < 0.001), and higher wbc count (P < 0.001), compared with horses given control mab. Rectal temperature, respiratory rate, pcv, and plasma total protein concentration were not significantly different between groups. Serum mab concentration peaked at 68 µg/ml 30 minutes after the end of antibody infusion in both groups. Neutralization of lps-induced tnf activity reduced the hematologic and clinical responses of horses given lps iv.

Free access
in American Journal of Veterinary Research

SUMMARY

A monoclonal antibody (mab) against equine tumor necrosis factor-α (Eq tnf) was used to investigate the role of tnf in cytokine, eicosanoid, and metabolic responses of Miniature Horses given endotoxin. Plasma concentrations of interleukin 6 (il-6), lactate, thromboxane A2 metabolite, and prostacyclin metabolite (6-keto-pgf ) were measured in 10 Miniature Horses given 0.25 µg of lipopolysaccharide (lps; Escherichia coli O55:B5)/kg of body weight. Five horses were given Eq tnf mab and 5 were given isotype-matched mab as control. All horses were given 1.86 mg of antibody/kg by iv infusion, 5 minutes before lps was given iv. Blood samples were taken 20 minutes before and at multiple intervals for 24 hours after lps was given. Interleukin 6 bioactivity in plasma was measured, using il-6-dependent cell line (B9). Eicosanoid activities were assessed by enzyme immunoassay, and plasma lactate concentration was determined enzymatically. Data were analyzed by anova and Tuke's honest significant difference test for significant (P < 0.05) effect of treatment. Horses given Eq tn mab had significantly (P < 0.050) lower peak mean ± sem il-6 (59 ± 29 U/ml), lactate (16 ± 2.00 mg/dl), and 6-keto-pgf (254 ± 79 pg/ml) values then did horses given control mab (880 ± 375 U/ml for il-6; 26 ± 0.04 mg/dl for lactate; and 985 ± 290 pg/ml for 6-keto-pgf ). There was no effect of anti-tnf treatment on lps-induced thromboxane A2 metabolite production. Tumor necrosis factor mediated il-6, lactate, and prostacyclin responses, without affecting thromboxane production in horses given lps.

Free access
in American Journal of Veterinary Research

SUMMARY

Six horses received intra-articular injections of a mixture of 1 μg of endotoxin/5 mg of equine tumor necrosis factor (eqTNF) monoclonal antibody in 1 antebrachiocarpal joint and an equal volume (2 ml) of 1 μg of endotoxin/5 mg of control antibody in the opposite joint. Synovial fluid sample collection (1 ml) was accomplished by use of an indwelling, intra-articular catheter at postinjection hours (pih) 0, 1, 1.5, 2, 5, and 8, and by arthrocentesis at pih 24. Joint fluid samples were analyzed for nucleated cell count, protein concentration, and tnf, interleukin 6 (il-6), il-1, and il-1-inhibitory activities. To monitor local inflammation, each carpus was graded semiquantitatively for swelling prior to each sample collection.

Tumor necrosis factor, il-1, or il-1-inhibitory activity was not detected in any synovial fluid sample collected before endotoxin/antibody was administered. However, low il-6 activity (< 100 U/ml) was found in 2 of 12 preinjection samples. In joints injected with endotoxin/control antibody mixture, maximal mean ± sem activities for tnf (1,019 ± 310 U/ml), il-1 (173 ± 102 U/ml), and il-6 (10.8 ± 3.1 × 104 U/ml) were observed at pih 2, 5, and 8, respectively. Tumor necrosis factor and il-1 activities returned to baseline values by pih 8 and 24, respectively; however, il-6 activity remained high. Interleukin 1-inhibitory activity (27.4 ± 2.25 IU/ml) was detected in all pih-24 samples from control joints, but was not detected at any other time in control joints (limit of detection, 20 IU/ml).

Tumor necrosis factor activity was not detected in any synovial fluid sample from joints treated with endotoxin/eqTNF antibody. In contrast, endotoxin-induced mean synovial il-1 and il-6 activities were not reduced significantly by eqTNF antibody. Mean il-1-inhibitory activity (pih 24) was higher in eqTNF antibody-treated joints (41.0 ± 7.7 IU/ml) than in control joints, but the difference was not significant. Mean wbc count and protein concentration in control and treated joints were maximal at pih 8. The curves for mean values of wbc count and total protein concentration were not significantly different in treated versus control joints. Swelling in each treated joint was either less than or the same as that in the opposite control joint at every time in the initial 8 pih. There was significant (P = 0.043) difference between treated and control joints at pih 5 and 8. These results describe a profile of synovial fluid tnf, il-1, il-6 bioactivities, and il-1-inhibitory activity during the initial 24 hours of synovitis induced by intra-articular administration of endotoxin in horses. Our eqTNF monoclonal antibody was effective in neutralizing tnf activity in synovial fluid when administered intra-articularly with endotoxin in horses. The induction of il-1, il-1-inhibitory activity, il-6, wbc, and total protein concentration responses are largely independent of tnf activity in synovial fluid of horses receiving endotoxin intra-articularly.

Free access
in American Journal of Veterinary Research