Because hepatocyte-stimulating factor/interleukin 6 (IL-6) the principal inducer of acute-phase protein synthesis in the liver, quantification of its activity in blood provides an early and sensitive assessment of the acute-phase response. Circulating IL-6 activity was monitored in 4 adult horses for 72 hours after IV administration of endotoxin. In 4 experiments performed at weekly intervals and in randomized order, each horse was given endotoxin-1,000, 30, 1, and 0 ng/kg of body weight. Plasma IL-6 activity was quantified as the ability to promote growth of the IL-6-dependent B-cell hybridoma, B13.29 clone B9. Interleukin-6 activity (171 +/- 10.2 U/ml) was found in all pretreatment plasma samples and was significantly (P < 0.05) increased above baseline from 2 to 12 hours after 1,000 ng of endotoxin/kg was given and at 3 hours after 30 ng of endotoxin/kg was given. After 1,000- or 30-ng/kg dosage of endotoxin, peak plasma IL-6 activity (10,128 +/- 4,096 and 1,555 +/- 1,326 U/ml, respectively) was observed for 3 hours. The IL-6 response of endotoxin-treated horses began about 1 hour after tumor necrosis factor appeared in the circulation, and its course closely approximated the endotoxin-induced febrile reaction. Significant increase in plasma IL-6 activity was not detected in horses given 1 ng of endotoxin/kg or control buffer.

Systemic and pulmonary antibody responses of calves to Pasteurella haemolytica were evaluated by measuring immunoglobulin production in blood for 9 days and in pulmonary lavage fluid for 7 days after intrapulmonary inoculation. Clinical signs, pulmonary lesions, pulmonary and systemic inflammatory response, and amount of antigen in lavage fluid were used to evaluate the response of calves to challenge with P haemolytica. The pulmonary response consisted of production of IgG, IgE, and IgM antibodies to P haemolytica antigens and a 17- to 68-fold increase of cells in lavage fluid 8 hours after inoculation, with a gradual decrease toward normal. Antibodies of the IgM isotype to P haemolytica were demonstrated as early as 8 hours through 7 days after inoculation in 3 of 3 calves. Of the anti-P haemolytica isotypes, IgM was found in the highest concentration. In all of the inoculated calves, IgE was found 1 to 2 days after inoculation, and IgG was found in 2 of 3 inoculated calves from day 1 through 7 after inoculation. Detection of IgG correlated with smaller pulmonary lesions. Immunoglobulin A was not detected in lavage fluid. Serum was evaluated for IgG and IgM antibody response to P haemolytica. Specific IgM was detectable 5 days after inoculation, and IgG was detectable 7 days after inoculation. Pasteurella haemolytica antigens were not detected in serum or plasma. A transient increase in neutrophil count was found 8 hours after inoculation, with return to baseline values by 24 hours after inoculation. Antigen was detected in lavage fluid by use of monoclonal antibodies against selected P haemolytica capsular antigen, outer membrane antigens, and leukotoxin in all inoculated calves 8 hours after inoculation. The monoclonal antibody specific for P haemolytica capsule provided the best detection of antigen. The other monoclonal antibodies detected antigen, but were less consistent.

Neutrophil migration through bovine teat tissues into the teat cistern, after endotoxin infusion into the teat cistern, was determined in vivo by 2 experimental procedures, indium-111 labeling of blood neutrophils, and obtaining multiple biopsy specimens from the teat cistern tissues. In both experiments, the number of leukocytes in the teat cistern flushing samples was continuously measured. A lag phase of approximately 1 hour was required between endotoxin infusion into the teat cistern and the first observed neutrophil accumulation in the teat tissues. The rate of neutrophil accumulation in the teat tissues was highest between postinfusion (pI) hours 1 and 2, and the accumulation process ceased after PI hour 3. Neutrophils migrated toward the epithelium, and intraepithelial neutrophils were observed beginning approximately 2 hours after infusion, which coincided with the first influx of cells into the teat cistern. The cell influx into the teat cistern increased continuously up to PI hour 3, peaked between PI hours 3 and 5, and was close to preinfusion value at PI hour 22. Use of indium-111-labeled neutrophils in the study of the inflammatory process provides a reliable noninvasive method to quantify cell migration in vivo. Use of biopsies allows quantification of the number of cells in different tissue areas, but has the disadvantage of being invasive. These 2 procedures complement each other, and could be of use in future studies of the local inflammatory process.

Determination of antibodies to specific nuclear antigens, termed extractable nuclear antigen (ENA), was investigated in healthy dogs and in dogs with autoimmune, inflammatory, and neoplastic diseases. Using a counter-immunoelectrophoresis method, the dogs' sera were tested for antibodies against the nuclear antigens single-stranded DNA, Sm, Ro, La, ribonucleoprotein, Scl, and proliferating cell nuclear antigen. Antibodies to the Ro antigen were found in 1 dog with discoid lupus erythematosus, in 1 dog with pemphigus erythematosus, and in 1 dog with facial pyoderma and chronic superficial keratitis. In 15 dogs, antibodies were detected to ENA, but the precipitin lines were too weak to identify the specific ENA. These antibodies were found in some dogs with systemic lupus erythematosus, discoid lupus erythematosus, pemphigus erythematosus, dermatomyositis, vitiligo, lymphoma; in the dog with facial pyoderma and chronic superficial keratitis; and in 1 healthy dog. The highest percentage of dogs with antibodies to ENA in a large series (> 8) of this study was in dogs with systemic lupus erythematosus (4 of 13; 31%).

Systemic and pulmonary antibody responses of calves to Pasteurella haemolytica were evaluated by measuring immunoglobulin production in blood for 9 days and in pulmonary lavage fluid for 7 days after intrapulmonary inoculation. Clinical signs, pulmonary lesions, pulmonary and systemic inflammatory response, and amount of antigen in lavage fluid were used to evaluate the response of calves to challenge with P haemolytica. The pulmonary response consisted of production of IgG, IgE, and IgM antibodies to P haemolytica antigens and a 17- to 68-fold increase of cells in lavage fluid 8 hours after inoculation, with a gradual decrease toward normal. Antibodies of the IgM isotype to P haemolytica were demonstrated as early as 8 hours through 7 days after inoculation in 3 of 3 calves. Of the anti-P haemolytica isotypes, IgM was found in the highest concentration. In all of the inoculated calves, IgE was found 1 to 2 days after inoculation, and IgG was found in 2 of 3 inoculated calves from day 1 through 7 after inoculation. Detection of IgG correlated with smaller pulmonary lesions. Immunoglobulin A was not detected in lavage fluid. Serum was evaluated for IgG and IgM antibody response to P haemolytica. Specific IgM was detectable 5 days after inoculation, and IgG was detectable 7 days after inoculation. Pasteurella haemolytica antigens were not detected in serum or plasma. A transient increase in neutrophil count was found 8 hours after inoculation, with return to baseline values by 24 hours after inoculation. Antigen was detected in lavage fluid by use of monoclonal antibodies against selected P haemolytica capsular antigen, outer membrane antigens, and leukotoxin in all inoculated calves 8 hours after inoculation. The monoclonal antibody specific for P haemolytica capsule provided the best detection of antigen. The other monoclonal antibodies detected antigen, but were less consistent.

Postoperative abdominal fluid changes were compared in 2 groups of horses; those undergoing double small-colon resection and anastomosis (n = 10) and those undergoing exploratory celiotomy alone (n = 5). Peritoneal fluid was collected before surgery and on postoperative days 1, 3, 5, and 7. Total and differential nucleated cell counts, RBC numbers, and total protein and fibrinogen concentrations were evaluated. In both groups, all values were significantly higher than normal on the first postoperative day (after small-colon resection and anastomoses, WBC = 130,350 +/- 23,310 cells/microliter, RBC 7,389,000 +/- 6,234,000 cells/microliter, total protein = 3.63 +/- 0.16 g/dl; after exploratory celiotomy alone, WBC = 166,620 +/- 34,340 cells/microliter, RBC = 295,000 +/- 86,070 cells/microliter, total protein 4.38 +/- 0.54 g/dl). The number of total peritoneal nucleated cells and RBC significantly decreased after the first postoperative day, whereas total protein and fibrinogen concentrations, percent neutrophils, and percent mononuclear cells remained unchanged. None of the values had returned to normal by postoperative day 7 (after small-colon resection and anastomoses, WBC = 45,600 +/- 8,765 cells/microliter, RBC = 95,390 +/- 53,380 cells/microliter, total protein = 4.39 +/- 0.23 g/dl; after exploratory celiotomy alone, WBC = 43,340 +/- 7,746 cells/microliter, RBC = 12,860 +/- 11,790 cells/microliter, total protein = 3.92 +/- 2.20 g/dl.) The resection and anastomosis group had a significantly lower total protein concentration on the first postoperative day and a significantly higher mean total RBC count over the entire 7-day postoperative evaluation than did horses that underwent celiotomy alone. Other values in the 2 groups of horses did not differ significantly. As a result, there was insufficient evidence to conclude that resection and anastomosis of the small colon in healthy horses causes a different inflammatory response than does manipulation of the intestine alone.

Acyloxyacyl hydrolase (AOAH) is a lysosomal enzyme found in neutrophils and macrophages that acts to partially deacylate the lipid-A component of the endotoxin of gram-negative bacteria rendering it less toxic, yet maintaining much of its immunostimulatory potential. We have found that the activity of neutrophil AOAH per cell increased during localized inflammation. The purpose of this study was to determine the mechanism(s) responsible for these increases in neutrophil AOAH activity. Because changes in neutrophil maturity commonly are associated with inflammation, intravascular infusion of purified gram-negative bacterial lipopolysaccharide and SC injection of bovine recombinant granulocyte colony-stimulating factor was used to induce large numbers of circulating immature neutrophils. Immature neutrophils were found to have AOAH activity equal to that of mature cells; however, when neutrophils were stimulated in vitro with known activators, AOAH activity of activated cells was more than that of unstimulated cells. The increase in AOAH activity was inversely related to prestimulation activity. Increases in AOAH activity after neutrophil activation were not a result of de novo synthesis of the enzyme, because cycloheximide did not prevent activation-induced increases in activity.

Equine alpha1-acid glycoprotein (alpha-1AG) was isolated from equine serum by successive ammonium precipitation, anion- and cation-exchange chromatographies, and gel filtration. Purified equine alpha-1AG had a molecular weight of 46,000 +/- 1,000, and contained 31.4% carbohydrate. Gel isoelectric focusing revealed an isoelectric point range of 2.8 to 3.7. With immunoelectrophoresis, it was found that alpha-1AG migrated to the alpha-1-globulin region. Single radial immunodiffusion was used for quantitative measurement of alpha-1AG in equine serum. In clinically normal foals, serum alpha-1AG was undetectable (less than or equal to 20 micrograms/ml) in less than or equal to 7-day-old foals, but was detected by 14 days. The alpha-1AG concentration (mean +/- SD) increased to reach mean adult values of 99.23 +/- 26.90 micrograms/ml by 1 year of age. The alpha-1AG concentration in pregnant mares decreased at 2 to 3 months before parturition, then gradually increased until 1 day after parturition, when a brief decrease was observed. The concentration increased again at 2 weeks after foaling, then a decrease was observed, after which the alpha-1AG concentration increased again by 2 to 4 months after parturition. The concentration of serum alpha-1AG quickly rose to peak values 2 to 3 days after castration and jejunojejunostomy in adult horses, returning to baseline values by 14 to 28 clays after surgery. The alpha-1AG was concluded to be an acutephase reactive protein in horses.

Because hepatocyte-stimulating factor/interleukin 6 (IL-6) the principal inducer of acute-phase protein synthesis in the liver, quantification of its activity in blood provides an early and sensitive assessment of the acute-phase response. Circulating IL-6 activity was monitored in 4 adult horses for 72 hours after IV administration of endotoxin. In 4 experiments performed at weekly intervals and in randomized order, each horse was given endotoxin-1,000, 30, 1, and 0 ng/kg of body weight. Plasma IL-6 activity was quantified as the ability to promote growth of the IL-6-dependent B-cell hybridoma, B13.29 clone B9. Interleukin-6 activity (171 +/- 10.2 U/ml) was found in all pretreatment plasma samples and was significantly (P < 0.05) increased above baseline from 2 to 12 hours after 1,000 ng of endotoxin/kg was given and at 3 hours after 30 ng of endotoxin/kg was given. After 1,000- or 30-ng/kg dosage of endotoxin, peak plasma IL-6 activity (10,128 +/- 4,096 and 1,555 +/- 1,326 U/ml, respectively) was observed for 3 hours. The IL-6 response of endotoxin-treated horses began about 1 hour after tumor necrosis factor appeared in the circulation, and its course closely approximated the endotoxin-induced febrile reaction. Significant increase in plasma IL-6 activity was not detected in horses given 1 ng of endotoxin/kg or control buffer.