A study was performed to determine whether equine peritoneal macrophages produce interleukin 6 (IL-6) in vitro in response to endotoxin. Peritoneal fluid was collected from 14 clinically normal adult horses and was used as the source of peritoneal macrophages. Macrophages from each horse were isolated and cultured separately in vitro in the absence or presence of various concentrations (0.5, 5, or 500 ng/ml) of endotoxin (lipopolysaccharide from Escherichia coli 055:B5). Culture medium supernatants were collected after 3, 6, 12, and 24 hours' incubation and were frozen at - 70 C until assayed for IL-6 activity. Supernatant IL-6 activity was determined by use of a modified colorimetric assay and the murine hybridoma cell line B13.29 clone B.9, which is dependent on IL-6 for survival. Results indicated that equine peritoneal macrophages produce IL-6 in vitro and that supernatant medium IL-6 activity was significantly (P < 0.05) increased by exposure to endotoxin. Significant (P < 0.05) time and treatment effects on macrophage IL-6 production were apparent. The IL-6 activity peaked at 6 or 12 hours' incubation, then remained high through 24 hours' incubation, regardless of endotoxin exposure. Medium IL-6 activity during 3 and 6 hours' incubation was significantly (P < 0.05) greater in macrophages exposed to 5 or 500 ng of endotoxin/ml than in those exposed to 0.5 ng of endotoxin/ml; however peak IL-6 activity was similar among all endotoxin concentrations. Endotoxin concentration did not have an effect on medium IL-6 activity from macrophages exposed to endotoxin for 12 or 24 hours.

A series of experiments was performed in vitro and in vivo to determine the influence of FK-565, a heptanoyl tripeptide, on lymphocyte and macrophage function in swine. Compared with values for control cultures, mitogen-stimulated lymphocyte blastogenesis and interleukin-2 production were unaffected in cells preincubated with 0.1, 1.0, and 10.0 microgram of FK-565/ml. Natural killer cell activity was increased by preincubation with 1.0 microgram of FK-565/ml; however, this increase was not statistically significant. In vitro treatment of porcine alveolar macrophages with FK-565 did not enhance cytolytic activity or bactericidal activity. In in vivo experiments, FK-565 given orally to pigs at concentrations of 6 or 60 microgram-kg-l.-d-1 for 5 days did not affect lymphocyte blastogenesis, interleukin-2 production, or alveolar macrophage bactericidal activity. A trend toward increased natural killer cell activity was evident in pigs treated with FK-565. In contrast, pigs treated with 6 microgram-kg-1.-d-1 had significantly (P less than 0.01) decreased alveolar macrophage cytolytic activity. These data indicate that at the dosages tested, FK-565 is not a suitable immunomodulator for enhancement of nonspecific immunity in swine.

Opsonized Rhodococcus equi activated the respiratory burst of resident alveolar macrophages (AM) from adult horses in a logarithmic-linear, mass-related manner. The effect of R equi was not significantly different from that of equal masses of opsonized zymosan A. Therefore, R equi does not appear to attenuate the respiratory burst of equine AM. The stimulatory effect of R equi was not reflected by increased production of superoxide anion (O2-), but increased activity of the hexose monophosphate shunt was observed. These results suggest a similarity between the respiratory burst of Am from horses and that of AM from rabbits. We concluded that resident AM from adult horses do not produce O2- concurrently with an increase in activity of the hexose monophosphate shunt when stimulated with either opsonized zymosan A or opsonized R equi. This suggests that O2- is not an important component of the antibacterial defenses of equine AM. Whether equine AM are incapable of producing O2- or require different stimuli to produce it was not determined.

Electron microscopy revealed several unique features in canine bone marrow, compared with that of other species. The marrow was fatty and extensively trabeculated and was enclosed by a complete layer of endosteal bone-lining cells. Branched reticular cells were closely associated with each other and, occasionally covered part of the sinus wall as an adventitial layer. The extent of adventitial coverage varied markedly and was less extensive, compared with that of other species. On average, only 23% of the sinus wall was covered by adventitial layer, in contrast to 65% reported in laboratory animals. Unilaminar sinuses, with no adventitial coverage, accounted for greater than 38% of all sinuses. Quantitative analysis indicated that 60% of the latter sinuses contained apertures, as opposed to 35% of sinuses with adventitial coverage (P less than 0.05). Moreover, the number of apertures in unilaminar sinuses was significantly (P less than 0.009) greater than that in multilaminar sinuses. Apertures were observed every 59 micromoles in unilaminar sinuses, in contrast to every 109 micromoles in multilaminar sinuses. Approximately 75% of the apertures were occupied by cells in transit, and only 25% were free of cells. Macrophages were distributed throughout the marrow and were closely associated with all blood cell lines. Occasionally, cells that entered the lumen were not fully mature. Erythroblasts were seen migrating across the wall and within the lumen of sinuses. The less-extensive adventitial coverage in canine bone marrow might indicate that the rate of cell delivery from the marrow into the circulation was relatively high in this species. The prevalence of unilaminar sinuses, along with the larger number of apertures, suggested that these sinuses were more accessible to the migrating cells and that the cellular traffic across them was intense.

Animals recovered from viral diseases represent an important model to study the host cellular and humoral immune responses to the etiologic agents. This is particularly important for African swine fever virus (ASFV) infections in which antibodies have little or no virus-neutralizing effect. Pigs surviving experimental infection with the naturally occurring low-virulent, nonhemadsorbing ASFV/NH/P68 (NHV) isolate did, however, exhibit virus-specific T-cell activities, as measured by a variety of assays. A strong virus-induced, antigen-specific blastogenic response was observed only with blood mononuclear cells (BMC) from ASF-recovered swine, whereas cells from recovered and naive swine responded similarly to the mitogens concanavalin A and phytohemagglutinin. The ASFV-induced blastogenesis was dependent on virus dose and on the presence of adherent cells. Blood mononuclear cells cultured with antigenically related hemadsorbing ASFV isolates of different virulence characteristics, the highly virulent L60 isolate and moderately virulent DRII isolate, exhibited a similar magnitude of blastogenesis to cells infected with the low-virulent NHV isolate. Virus-infected cells proved to be an efficient inducer of interleukin-2 (IL-2) activity to cells from recovered swine, but not from naive swine, whereas T-cell-specific lectins induced production of similar amounts of IL-2 activity from cells of naive and recovered swine. Correlated with the appearance of virus-induced IL-2 activity in the culture supernatant was the induction of promiscuous killing in cells exposed to prolonged (7 days) virus stimulation. This lymphokine-activated killing could be induced experimentally early in the virus stimulatory process (3 days) by the addition of exogenous lymphokines to the cultures. It was concluded that swine inoculated with low-virulent ASFV isolates are a useful model for identifying and characterizing ASFV immune mechanisms in vitro. Furthermore, this ASFV model implicates lymphokines as inducers of nonspecific cell-mediated immunity; in fact, lymphokine-activated killer type responses may contribute to recovery from this viral infection. More important, ASFV-specific blastogenic and cytotoxic T-cells are prime candidates for the cells inducing and/or conferring protective immunity against challenge ASFV infection.

Ponies with recurrent airway obstruction (principal ponies) and their controls were given aerosolized Micropolyspora faeni antigen via endotracheal tube during a period when the principal ponies were in disease remission. In both groups of ponies, we performed bronchoalveolar lavage (BAL) and measured pulmonary function at base line, and 5 hours after aerosol administration of 30 ml of 0.9% NaCl solution or 30 ml of 1% w/v particulate M faeni antigen in 0.9% NaCl solution. In both groups of ponies, aerosolized M faeni antigen increased WBC count, neutrophil numbers, and albumin concentration in BAL fluid, but macrophage numbers decreased. In the principal ponies, BAL mast cell numbers were decreased 5 hours after administration of M faeni antigen. The M faeni antigen had no effect on the mechanical properties of the lungs or on gas exchange in the control ponies, but did increase respiratory frequency minute ventilation and pulmonary resistance, and decreased arterial oxygen tension in the principal ponies. Changes in pulmonary function were apparent only in the principal ponies, which suggests that neutrophils, per se, do not cause pulmonary dysfunction and that M faeni may be one of the etiologic agents involved in chronic obstructive pulmonary disease.

Ischemia/reperfusion(I/R) injury of the rat testis causes germ cell death and infiltration of inflammatory cells. To investigate the mechanism of germ cell death in torsion of the rat testis, apoptosis and macrophage activation were studied using the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling(TUNEL) method and immunohistochemistry in the testes of Sprague-Dawley rats subjected to 1.5 h of ischemia, followed by 0, 1, 3, 6, 12, 24, 48 and 96 h of reperfusion.

OBJECTIVE To evaluate acute changes of the liver by use of shear wave elastography (SWE) and CT perfusion after radiofrequency ablation (RFA). ANIMALS 7 healthy Beagles. PROCEDURES RFA was performed on the liver (day 0). Stiffness of the ablation lesion, transitional zone, and normal parenchyma were evaluated by use of SWE, and blood flow, blood volume, and arterial liver perfusion of those regions were evaluated by use of CT perfusion on days 0 and 4. All RFA lesions were histologically examined on day 4. RESULTS Examination of the SWE color-coded map distinctly revealed stiffness of the liver tissue, which increased from the normal parenchyma to the transitional zone and then to the ablation zone. For CT perfusion, blood flow, blood volume, and arterial liver perfusion decreased from the transitional zone to the normal parenchyma and then to the ablation zone. Tissue stiffness and CT perfusion variables did not differ significantly between days 0 and 4. Histologic examination revealed central diffuse necrosis and peripheral hyperemia with infiltration of lymphoid cells and macrophages. CONCLUSIONS AND CLINICAL RELEVANCE Coagulation necrosis induced a loss of blood perfusion and caused tissue hardening (stiffness) in the ablation zone. Hyperemic and inflammatory changes of the transitional zone resulted in increased blood perfusion. Acute changes in stiffness and perfusion of liver tissue after RFA could be determined by use of SWE and CT perfusion. These results can be used to predict the clinical efficacy of RFA and to support further studies, including those involving hepatic neoplasia.

The objective of this study was to investigate the normal cytological evaluation of bronchoalveolar lavage (BAL) fluid in healthy adult ferrets (N = 12). These ferrets underwent bronchoscopy and BAL using sterile saline [1.5 mL/kg body weight (BW)]. Percentage of fluid recovered, total leukocyte count, differential leukocyte count, and cell count of the epithelial lining fluid (ELF) were determined. The mean percentage of lavage volume recovered from the right lung and left lung were 67.8 ± 14.9% and 69.7 ± 20.0%, respectively. Gender (P = 0.12) and weight (P = 0.17) did not significantly affect the mean percentage of recovered volume. The mean percentage of recovered volume (P = 0.47) and the mean leukocyte count (P = 0.17) from the right and left lung were not significantly different. Macrophages were the main leukocyte component of the lavages, followed by neutrophils, lymphocytes, and eosinophils. The mean proportion of ELF in BAL fluid was 9.3 ± 3.7% v/v. Bronchoscopy is clinically useful for collecting good quality BAL samples for cytological analysis in ferrets. The leucocyte differential was established, which may help veterinarians to make better clinical decisions when treating respiratory disease. Further studies are required with a larger group in order to establish the healthy reference intervals for BAL values in ferrets.

OBJECTIVE To investigate the anti-inflammatory and immunomodulatory properties of tulathromycin in vitro and in experimental models of Actinobacillus pleuropneumoniae–induced pleuropneumonia and zymosan-induced pulmonary inflammation in pigs. ANIMALS Blood samples from six 8- to 30-week-old healthy male pigs for the in vitro experiment and sixty-five 3-week-old specific pathogen–free pigs. PROCEDURES Neutrophils and monocyte-derived macrophages were isolated from blood samples. Isolated cells were exposed to tulathromycin (0.02 to 2.0 mg/mL) for various durations and assessed for markers of apoptosis and efferocytosis. For in vivo experiments, pigs were inoculated intratracheally with A pleuropneumoniae, zymosan, or PBS solution (control group) with or without tulathromycin pretreatment (2.5 mg/kg, IM). Bronchoalveolar lavage fluid was collected 3 and 24 hours after inoculation and analyzed for proinflammatory mediators, leukocyte apoptosis, and efferocytosis. RESULTS In vitro, tulathromycin induced time- and concentration-dependent apoptosis in neutrophils, which enhanced their subsequent clearance by macrophages. In the lungs of both A leuropneumoniae– and zymosan-challenged pigs, tulathromycin promoted leukocyte apoptosis and efferocytosis and inhibited proinflammatory leukotriene B4 production, with a concurrent reduction in leukocyte necrosis relative to that of control pigs. Tulathromycin also attenuated the degree of lung damage and lesion progression in A pleuropneumoniae–inoculated pigs. CONCLUSIONS AND CLINICAL RELEVANCE Tulathromycin had immunomodulatory effects in leukocytes in vitro and anti-inflammatory effects in pigs in experimental models of A pleuropneumoniae infection and nonmicrobial-induced pulmonary inflammation. These data suggested that in addition to its antimicrobial properties, tulathromycin may dampen severe proinflammatory responses and drive resolution of inflammation in pigs with microbial pulmonary infections.