MHC class II expression in the bovine mammary gland.

The distribution of major histocompatibility complex (MHC) class II positive cells within the connective tissue and the epithelium of the involuted bovine mammary gland has been determined. The effect of intramammary administration of the antigens ovalbumin and formalin killed Streptococcus uberis on the distribution pattern has also been investigated. Infusion of formalin killed S. uberis increased cellular expression of class II antigens when compared with quarters either infused with ovalbumin, not infused at all, or from which minor pathogens had been isolated. The increased expression occurred particularly in the area of the gland cistern-secretory tissue junction.     

Disposition of gentamicin in the genital tract of cows

The distribution of gentamicin (G) in plasma and uterine lumen was studied following intramuscular (i.m.) and intrauterine (i.u.) treatment. A Foley catheter was inserted into one uterine horn and retained in place by inflation of the cuff. This provided a closed system for collection of uterine lumen samples and analysis of the concentration of gentamicin for 6 h following treatment. Four normal cycling and healthy cows in dioestrus were given i.m. injections of 4 mg gentamicin/kg BW and another two were given i.m. injections of 2 mg gentamicin/kg BW gentamicin. The uteri were infused with 50 ml saline containing phenolsulphonphthalein (PSP) indicator. Blood and infused solution (IS) samples were periodically collected during the 6-h period following i.m. administration. Six hours after injection, approximately 183.7 micrograms gentamicin and 39.4 micrograms gentamicin were accumulated in the uterine lumen of cows receiving 4 mg gentamicin/kg BW and 2 mg gentamicin/kg BW, respectively. The amount of gentamicin reaching the blood stream after i.m. administration of 4 mg gentamicin/kg BW was 2.89 times that reached after administration of 2 mg gentamicin/kg BW based on the area under the curve of plots of plasma concentration of gentamicin versus time. Four normal-cycling and healthy cows in dioestrus were given i.u. infusions of gentamicin (225-275 mg) diluted in 50 ml saline containing PSP indicator using a Foley catheter in a closed system. Samples from the IS and blood were collected at various intervals for 6 h after infusion. Following i.u. infusion of gentamicin, an average of 29.4% of the dose was absorbed into the bloodstream. The majority of the dose of gentamicin (70.6%) remained in the uterine lumen throughout the 6-h period.     

MHC class II expression by follicular keratinocytes in canine demodicosis--an immunohistochemical study

MHC class II proteins present fragments of extra cellular antigen to stimulate CD4(+) T lymphocytes. Aim of this study was the detection of MHC class II antigens on different cutaneous cells in canine demodicosis. Histopathological and immunohistochemical examination of skin biopsies from 44 dogs with demodicosis is reported. The control group consisted of skin biopsies taken from 10 necropsied dogs without obvious skin lesions. The immunohistological assessment of the MHC class II expression revealed MHC class II proteins on different cell types of infiltrating inflammatory cells, i.e. APCs (antigen-presenting cells), macrophages, T lymphocytes and B lymphocytes. The plasma cells, however, only showed expression in 32 (73%) of 44 cases. Generally it was noticeable that most plasma cells but never all of them expressed MHC class II. Neutrophils, mast cells and eosinophils were MHC class II negative. Furthermore, in 39 biopsies (89%) from dogs with demodicosis MHC class II positive follicular keratinocytes were found. The control group did not show MHC class II expression on epithelial cells. Concerning the endothelial cells, a total of 25 biopsies (57%) showed MHC class II expression in which different vascular plexuses were affected by staining. This examination shows that MHC class II expression in the skin of dogs suffering form demodicosis is elevated. Especially the MHC class II expression by follicular keratinocytes seems to be conspicuous. We hypothesize that this is in association with the development and the maintenance of follicular inflammation.     

Distribution of chloramphenicol in the genital tract of postpartum cows

Chloramphenicol was administered by constant IV infusion to 7 healthy postpartum cows at rates predicted to approach a steady-state plasma concentration of 5 micrograms/ml. After 8 hours of constant IV infusion, uterine tissues were removed surgically and were assayed for chloramphenicol concentrations. Mean plasma-to-tissue ratios of chloramphenicol concentrations were 3.05, 3.63 (6 cows only), and 3.22 for caruncles, endometrium, and uterine wall, respectively. Plasma-to-tissue ratios of the 3 tissues were not significantly different (P greater than 0.10). Intrauterine (IU) injections of chloramphenicol (20 mg/kg of body weight) were administered to 3 healthy post-partum cows. The mean value of the fraction of the drug absorbed from the uteri of these cows was 0.40. Mean concentrations of chloramphenicol were 43.8 micrograms/g in caruncles, 34.6 micrograms/g in endometrium, 2.8 micrograms/g in uterine wall, and 2.9 micrograms/ml in plasma 8 hours after IU injections. Chloramphenicol has now been banned for use in food-producing animals in the United States because of its potential for causing toxicosis in human beings. It is illegal to use chloramphenicol in food-producing animals in the United States and in some other countries as well. This includes use by the IU route of administration because chloramphenicol and most drugs are absorbed from the uterus into the bloodstream and are distributed to milk and tissues.     

奶牛生殖道内益生乳酸菌的筛选及分子鉴定

为了筛选出用于制作微生态制剂的候选菌株,选取15头产后30⁶0d的健康荷斯坦奶牛,使用MRS琼脂培养基厌氧技术从生殖道分泌物中分离得到47株革兰阳性菌株。随后对其抑菌效果和产过氧化氢能力进行测试,从中挑选出抑菌效果和产过氧化氢能力相对较强的菌株4株,通过生化试验和16SrDNA序列分析方法对其进行鉴定。鉴定结果为:1株为魏斯氏菌,1株为大芬戈尔德菌,1株为乳酸明串珠菌,1株为营养泥杆菌。最后测试混合菌株的抑菌效果得出以下结论:大芬戈尔德菌、乳酸明串珠菌、营养泥杆菌混合之后抑菌效果在这几组混合液中抑菌效果是最好的。     

A study of dendritic cell and MHC class II expression in dogs with immunomodulatory-responsive lymphocytic-plasmacytic pododermatitis

The term immunomodulatory-responsive lymphocytic-plasmacytic pododermatitis (ImR-LPP) has previously been proposed to denote a sub-population of dogs with idiopathic pododermatitis. The objective of this study was to investigate dendritic cell (DC) and MHC class II antigen expression in lesional skin of dogs with ImR-LPP (n = 47). Median epidermal CD1c⁺ cell counts were 37.8 and 12.5 mm⁻¹ in ImR-LPP dogs and healthy controls (n = 27), respectively (P < 0.01), while the corresponding dermal cell counts were 180.9 and 45.0 mm⁻², respectively (P < 0.01).Intra-epidermal clusters of DCs were observed in 18/47 dogs with ImR-LPP. Median epidermal MHC class II⁺ cell counts were 32.5 and 10.5 mm⁻¹ in ImR-LPP dogs and healthy controls, respectively (P < 0.01), while the corresponding dermal cell counts were 216.9 and 46.9 mm⁻², respectively (P < 0.01). Dermal MHC class II⁺ staining was primarily associated with DCs (47/47 dogs), mononuclear inflammatory cells (45/47), fibroblast-like cells (19/47) and vascular endothelium (14/47). The DC hyperplasia and increased MHC class II expression in lesional ImR-LPP skin are consistent with enhanced antigen presentation, and suggest that both parameters may contribute to the pathogenesis of ImR-LPP through the priming and activation of CD4⁺ T cells. Equally, it is possible that the enhanced DC numbers observed in this study may contribute to the immunoregulation of steady-state pathology in lesional ImR-LPP skin through additional expanded, although as yet unresolved, mechanisms.     

Molecular characterization of MHC class II region in guinea fowl

1. The MHC class II gene was amplified, cloned and sequenced in guinea fowl. 2. The NumeMHC II sequence of 754 nucleotides included complete exon 1 (91 nt), exon 2 (270 nt), exon 3 (282 nt) and exon 4 (110 nt). 3. The size of β(1) and β(2), domains were 89 and 93 amino acids, respectively in guinea fowl. 4. High amino acid variability (38·2%) was observed within guinea fowl in β(1) domain, while in β(2) domain, amino acid variability (6·3%) was low. 5. Among poultry species, the percent amino acid identity between guinea fowl and chicken, quail, pheasant and duck was 38·8, 42·2, 44·4 and 58·8 in β(1) domain; and 13·8, 17·0, 13·8 and 27·6 in β(2) domain, respectively. 6. Sequence alignment with mammalian and avian MHC showed that many of the conserved features of MHC class II glycoprotein was conserved in guinea fowl. 7. Within-species genetic distances (Poisson correction) based on cumulative amino acid variability in β(1) domain and β(2) domains was 0·141 in guinea fowl. 8. Guinea fowl showed low and similar genetic distances with all the poultry species (0·255-0·268) except duck (0·456). 9. Guinea fowl made separate branch within the major cluster having chicken, quail and pheasant, showing equal distance from these poultry species, whereas duck MHC II clustered separately.     

Detection of porcine MHC class II antigens in different immunoassays

A monoclonal antibody (Mab), Mab 4-24-11, to human class II major histocompatibility antigens has been tested in commonly used immunoassays for detection of porcine class II major histocompatibility antigens (SLA-D). In a radioimmunoprecipitation assay, Mab 4-24-11 reacted with proteins from mitogen-stimulated porcine mononuclear cells (MNC). The molecular weights of the precipitated proteins were approximately 34 and 28 Kd. In indirect immunofluorescence, approximately 25% of porcine MNC in suspension reacted with Mab 4-24-11. This percentage diminished to 14% when Ig bearing MNC were removed, while it increased to 36% when adherent MNC were enriched. Thus, it was concluded that Mab 4-24-11 cross-reacts with SLA-D. In a immunohistochemical study, Mab 4-24-11 reacted with cells in acetone fixed cryostat sections from the gastric mucosa and endometrium. These properties of Mab 4-24-11 make it useful as a tool for further studies on the porcine immune system.     

MHC class II+ cells in the gills of Atlantic salmon (Salmo salar L.) affected by amoebic gill disease

Amoebic gill disease (AGD) is characterised by the association of Neoparamoeba sp. with hyperplastic gill tissue of affected fishes, however, the identity and role of host cells associated with AGD lesions are not known. Here, we investigated cells with an immunological role that were associated with AGD lesions by locating cellular MHC class II β chain. A tank housing Atlantic salmon (Salmo salar) was inoculated with Neoparamoeba sp., and MHC class II β chain expression in the gills was qualitatively assessed by immunohistochemistry. In AGD-naïve control fish, MHC class II⁺ cells were detected basolateral to the interlamellar epithelium as well as upon the interlamellar and secondary epithelium. In the gills of AGD affected fish MHC class II⁺ cells were observed in both affected and unaffected tissue. Within AGD lesions, numerous MHC class II⁺ cells were present and these cells exhibited variable levels of expression suggesting that like mammals, MHC class II expression is highly regulated. The presence of MHC class II⁺ cells within gill lesions is indicative of immune cell trafficking and these cells could contribute in an antigen presentation capacity to the development of an antibody response in fish chronically affected by AGD.     

Distribution and ultrastructure of mast cells in the equine respiratory tract

Mast cells in the equine respiratory mucosa were studied at both light--and transmission electron--microscope levels. Mast cells were identified at all levels of the tract, with the greatest cell density in the nasopharynx. The majority (57 to 94 per cent) of this cell population were located within the connective tissue of the lamina propria. Up to 20 per cent of these cells were associated with the mucosal glandular tissue, whilst small numbers were present within the surface epithelium and in association with nodular lymphoid tissue. In the peripheral lung tissue 20 per cent of the mast cell population was associated with the small airways, 25 per cent with the pleura, and 32 per cent with blood vessels. The fine and ultrastructural features of these mast cells resemble those described in other species, and their granules consist of two types which resemble human mast cells.     

Structure and expression of class I MHC genes in the miniature swine

The genome of the miniature swine, unlike other species, contains a relatively small class I MHC gene family, consisting of only seven members. This provides an excellent system in which to identify and characterize the regulatory mechanisms which operate to both coordinately and differentially regulate the expression of a multi-gene family. The structure of class I SLA genes, like other class I genes, consists of eight exons encoding a leader sequence, three extracytoplasmic domains, a transmembrane domain and intracytoplasmic domains. Despite the common structure, two sub-families of class I genes can be distinguished within the SLA family. One, containing the closely related PD1 and PD14 genes, encodes the classical transplantation antigens. Another contains the highly divergent PD6; the functions of the products of this subfamily, if any, are not known. The class I SLA genes share some common regulatory mechanisms, as evidenced by the fact that all three genes analyzed are transcribed in mouse L cells. Furthermore, interferon treatment of transfected mouse L cells enhances expression of all three genes. Both PD1 and PD6 are transcribed in vivo, where the highest levels of expression are observed in lymphoid tissues. Superimposed on the common patterns of class I gene expression are distinct ones, as evidenced by the findings that PD1 is preferentially expressed in B cells, whereas PD6 is preferentially expressed in T cells. These differences may reflect the extensive divergence of the 5' flanking sequences of these genes. Future studies will be aimed at elucidating the precise molecular interactions and mechanisms which give rise to the observed differential expression.     

Requirement for MHC class II positive accessory cells in an antigen specific bovine T cell response

Purified populations of bovine antigen presenting cells (APCs) and T cells have been isolated from peripheral blood and characterised using various monoclonal antibodies (mAbs) for cell surface markers. Bovine APCs were found in an adherent cell fraction and were non-specific esterase positive, phagocytic and expressed bovine major histocompatibility complex (MHC) class II determinants, all of which are typical macrophage characteristics. T cells were rigorously depleted of accessory cell function before being used in an antigen presenting cell assay. The generation of T helper cells in response to the soluble antigen, ovalbumin, was entirely dependent upon a critical number of APCs. Further the proliferative response was inhibited by several mAbs to bovine MHC class II molecules. Thus the interaction between bovine APCs and helper/inducer T lymphocytes (TH/I) appears to be similar to that in other species.     

Down-regulation of MHC class II receptors of DH82 cells, following infection with Ehrlichia canis

In order to evaluate whether infection with E. canis alters the expression of major histocompatibility complex (MHC) class I and/or MHC class II receptors, and by doing so alters the immune response to the organism, flow cytometry was performed on DH82 cells infected with Ehrlichia canis (90% infection) and on uninfected DH82 cells of the same passage, using anti-canine MHC class I and II antibodies. MHC class II expression was evident in 47.6 and 46.2% (mean 46.9%) of uninfected DH82 cells using two different anti-MHC class II antibodies, while no MHC class II expression was evident in DH82 cells infected with E. canis. The present results indicate that infection of DH82 macrophages with E. canis down-regulates their MHC class II receptors. These results suggest a possible mechanism by which E. canis evades the immune system.     

Histopathology of genital besnoitiosis of cows in Israel

Histological examination of genital organs of 16 cows affected with Besnoitia besnoiti showed that cysts, with or without surrounding granulomatous reaction, were present in 6 of these animals. The role of these organisms in the pathogenesis of bovine abortions and infertility seems to be minor, if any.     

Genotyping of feline MHC (FLA) class II DRB by PCR-RFLP method using group-specific primers

For genotyping of feline major histocompatibility complex (FLA) class II DRB, the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) method using group-specific primers was tried. Sixty-six DRB genes were classified into 8 groups according to differences in the first 5' amino acid sequences. The group-specific primers were designed as forward ones, which were specific for 5' base sequences of genes in each group. Three to 7 appropriate restricted enzymes were selected by computer analysis for RFLP typing of the genes divided into each group. In 6 out of 9 cats, the results of DRB typed by direct sequence method agreed with results of the PCR-RFLP method using group-specific primers. In the other 3 cats, the number of genes amplified by group-specific primers was I or 2 more than those detected by direct sequence method. The direct sequence method in 9 cats identified 5 new FLA-DRB genes. The PCR-RFLP method using group-specific primers could divide 66 genes into 37 genes and 10 subgroups from the RFLP pattern. One to 6 genes in each cat, and a total of 203 genes and subgroups were detected in 68 domestic cats. The genes detected might be biased to the subgroup G1-1a (28.8%), DRB*0501 (10.3%), G1-2a (9.4%) and G6b (7.4%). The PCR-RFLP method using group-specific primers may be useful in typing FLA class II DRB.