[Interleukin-2 for the treatment of cows with malignant catarrhal fever]. / Behandlung von Rindern mit Bösartigem Katarrhalfieber mit Interleukin-2.

The goal of this study was to investigate whether administration of interleukin-2 (IL-2) would improve the outcome of cows with malignant catarrhal fever (MCF). The study population consisted of ten healthy control cows and 22 cows with MCF. Nineteen cows with MCF and all of the controls were treated with either 2'500 U IL-2 or 25'000 U IL-2, administered intravenously. Three cows with MCF were not treated with IL-2 (MCF controls). All of the cows with MCF received danofloxacin, flunixin meglumine and intravenous fluid therapy. Blood samples for haematological and biochemical evaluation were collected once daily for six days in all cows. Of the 19 cows treated with IL-2, 13 were eutha nized because of deterioration. All cows with MCF that did not receive IL-2 died. The clinical condition of six cows treated with 2'500 U IL-2 gradually improved. Sur viving cows had significantly higher total leukocyte counts than cows that died or were euthanized. The main reason for leukopenia in non-surviving vs. surviv ing cows was persistent lymphopenia. Use of the lower IL-2 dose was associated with clinical recovery in some cows and this treatment might therefore be considered in valuable cows, provided that the lymphocyte count is within the reference interval.

Surgical stress influences cytokine content in autologous conditioned serum.

REASONS FOR PERFORMING STUDY: No recommendations have been made regarding the relative timing of blood collection for autologous conditioned serum (ACS) preparation and surgical procedures. OBJECTIVES: 1) To identify effects of surgical stress on cytokine levels in ACS, 2) identify haematological markers for prediction of cytokine production in ACS and 3) investigate the necessity for specialised ACS containers when preparing a cytokine-rich serum. STUDY DESIGN: Experimental in vitro study. METHODS: Blood was drawn from 15 stallions admitted for elective castration preoperatively and 22-24 h post operatively and incubated in ACS containers and plastic vacutainer tubes containing Z Serum Clot Activator. Concentrations of interleukin (IL)-1 receptor agonist (IL-1Ra), IL-10, IL-1ß, tumour necrosis factor (TNF)-α, insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-ß were determined in all serum samples and compared between preparation methods and sampling time by ANOVA. Changes in cytokine levels induced by incubation, defined as delta cytokine, were calculated by subtracting the baseline levels from the levels in incubated samples. Based on post operative serum amyloid A (SAA), horses were grouped into 'mild', moderate' and 'marked' surgical stress; delta cytokine levels in post operative samples were compared between these groups by ANOVA. RESULTS: Delta IGF-1 was significantly lower in post operative samples compared with preoperative. Horses in the 'marked' surgical stress group had significantly lower delta IL-1Ra and delta TGF-ß than the 'moderate' group and significantly lower delta IGF-1 than the 'mild' group. No association between cytokine levels and haematology variables were identified. Cytokine levels were comparable between serum prepared in blood tubes and in specialised ACS containers. CONCLUSIONS: Surgical stress influences the cytokine content in ACS. Useful predictors of cytokine production in ACS were not identified. Specialised ACS containers may not be necessary for preparation of a cytokine-rich serum.

Intestinal strictures, fibrous adhesions and high local interleukin-10 levels in goats infected naturally with Mycobacterium avium subsp. paratuberculosis.

This study describes pathological findings and their association with the production of interferon (IFN)-γ and interleukin (IL)-10 in goats infected naturally with Mycobacterium avium subsp. paratuberculosis (MAP). Twenty-seven goats were subjected to pathological examination. More than half of the animals had severe, diffuse, transmural granulomatous enteritis, often with abundant acid-fast bacilli (AFB), which was most evident in the proximal jejunum. Jejunal strictures and fibrous, peritoneal adhesions were findings that are not often reported in animals with paratuberculosis. Immunohistochemical labelling of IL-10 was seen within diffuse, granulomatous lesions and this may have prevented optimal local IFN-γ production and exacerbated the disease. However, since IFN-γ production was detected in cells from blood, jejunum and jejunal lymph nodes of goats with severe lesions by enzyme-linked immunosorbent assay, intracellular labelling and in-situ hybridization, the up-regulation of IL-10 might have been a consequence rather than a cause of the severe disease. The IL-10 labelling was co-localized with major histocompatibility complex (MHC) class II(+) cells, but rarely with CD4(+) cells. Comparable numbers of CD4(+) and CD8(+) T cells were recruited to both severe, diffuse lesions and small to moderate granulomatous lesions, while few T cells expressing the γδ form of the T-cell receptor were associated with both types of lesions.

Characterization of macrophages and occurrence of T cells in intestinal lesions of subclinical paratuberculosis in goats.

The granulomatous lesions of subclinical paratuberculosis of goats were examined with emphasis on phenotypic characteristics of macrophages and the presence of different subpopulations of T cells. The macrophages in the granulomatous lesions were morphologically homogeneous in histological sections but showed varying expression of the macrophage marker CD68 (a glycoprotein found mainly in late endosomal and lysosomal membranes) and varying acid phosphatase activity. The lesional macrophages showed decreased expression of complement receptor 3 and major histocompatibility complex proteins, which are markers associated with phagocytosis and antigen-presentation, respectively. The granulomas showed low proliferation activity as measured by the proliferation-associated protein Ki-67, indicating that most cells were recruited to the lesions. Few apoptotic cells were demonstrated by the TUNEL technique, suggesting a low cell turnover in the lesions. CD4(+) T cells constituted the main T-cell population among the CD68(+) macrophages in the granulomatous lesions, and few CD8(+) T cells and gamma delta T cells were observed within the lesions, suggesting the limited ability of these cells to influence the granulomatous lesions in caprine subclinical paratuberculosis. Both WC1(+) and WC1(-) gamma delta T cells were present in the small intestinal wall, but the latter were the more numerous. No difference in the numbers of these cells was observed between the subclinically infected animals and control animals.

Lesions in subclinical paratuberculosis of goats are associated with persistent gut-associated lymphoid tissue.

The organized gut-associated lymphoid tissue (the Peyer's patches [PPs] of domestic ruminants) is an important site of lesions caused by Mycobacterium avium subsp. paratuberculosis. To investigate the association between PP morphology and the lesions of paratuberculosis in goats, two experiments were performed. Five healthy kids aged 4-5 weeks were examined and the morphology of organized lymphoid tissue in the small intestine was described. Morphological similarities were observed between the ileocaecal-valve PP (ICVPP) and the jejunal PPs (JPPs), with pear-shaped follicles, large submucosal interfollicular T-cell areas, and many intraepithelial leucocytes in the follicle-associated epithelium. The ileal PP (IPP) consisted of elongated follicles, small T-cell areas and few intraepithelial leucocytes. The association between these three locations of PPs and lesions of paratuberculosis was then studied in seven goats inoculated with M. a. paratuberculosis at 5-8 weeks of age and killed 2 years later, while in the subclinical phase of infection. Gross lesions were recorded in five animals and microscopic lesions were observed in the intestine and mesenteric lymph nodes of six animals. The lesions in the small intestine were mainly located in the PPs of the mid-jejunum (JPPs) and ICVPP. Lesions were not present in the intestinal segments that had contained IPP, which had undergone involution during the first 12-18 months of life. These observations indicate that the persistent organized lymphoid tissue in the JPPs and ICVPP, but not the involuted IPP, sustains the development of granulomatous inflammation due to paratuberculosis during the subclinical phase of infection.

Localisation of CD25+ cells and MHCII+ cells in lymph nodes draining Mycobacterium avium subsp. paratuberculosis vaccination granuloma and the presence of a systemic immune response.

Vaccination of goat kids against paratuberculosis protects against lesions and clinical disease. The systemic cellular response was studied in goat kids 3-9 weeks after vaccination. Peripheral blood cells showed increased interferon-gamma production and expression of interleukin-2 receptor (CD25) after stimulation with Mycobacterium avium subsp. paratuberculosis antigens. The lymph node draining the vaccination granuloma was studied three weeks after vaccination in a parallel group of goat kids. In deep cortex, MHCII+ cells were observed surrounded by CD4+ T-cells, while follicular hypertrophy and hyperplasia were prominent in the subcapsular region and along connective tissue trabecula. Comparison of the local and systemic immune responses revealed an inverse relationship between CD25+ T-cells in the lymph node deep cortex and cells in peripheral blood that up-regulate CD25 upon in vitro stimulation, suggesting that activated and regulatory T-cells in the local lymph node influence the level of circulating antigen-specific T-cells following vaccination against paratuberculosis in goats.

Detection of caprine arthritis--encephalitis virus RNA in macrophages by in situ hybridization using fluorescein-labelled single-stranded RNA probes.

The use of in situ hybridization (ISH) for the detection of caprine arthritis-encephalitis virus (CAEV) RNA with fluorescein-11-UTP-labelled single-stranded RNA probes is described. Three different probes were made by PCR amplification of proviral CAEV DNA (strain 75-G63). The PCR products were cloned into the plasmid pAM-18, and labelled single-stranded RNA probes were synthesized by the use of RNA polymerase. The LTR probe was able to detect viral RNA in CAEV-infected, cultured caprine macrophages, while probes based on the genes for the matrix and transmembrane proteins failed to do so. A few macrophages were positive for CAEV RNA 24 h post infection (p.i.) while most cells were positive 96 h p.i. The use of fluorescein-labelled RNA probes made this method feasible for kinetic in vitro studies of CAEV.