RESUMEN
Successful implantation is a highly coordinated process involving changes in cytokines, adhesion molecules, hormones, enzymes and growth factors. This study examines the expression of key cytokines and vascular surface molecules in the pregnant uterus of sheep around the time of implantation. Uterine tissues and uterine washings were collected from non-pregnant and pregnant sheep at 17-19 days post-coitus (dpc), 26-27 and 34-36 dpc. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of caruncular/placentomal tissues revealed that cytokines IL-2, IL-4 and IL-8, which were not detected in non-pregnant uterus, were induced more strongly at 26-27 dpc than at other stages of pregnancy tested. Cytokines LIF, IL-6, IL-10, TNF-alpha were also most highly expressed at 26-27 dpc, expression of them being lower at other time-points during early pregnancy and non-pregnancy. The cytokines IL-1beta, IFN-gamma and TGF-beta were expressed in all non-pregnant and pregnant tissues examined. Enzyme-linked immunosorbent assay (ELISA) performed on uterine washings clearly detected the presence of IL-1alpha protein at 26-27 and 34-36 dpc. Immunohistochemistry revealed that expression of vascular adhesion molecule VCAM-1 in endometrial endothelium was strongly induced at 26-27 dpc in the pregnant endometrium. Expression of CD5 on vascular endothelium was not induced in placentomal tissues until 26-27 dpc and was further increased by 34-36 dpc. These results demonstrate a dynamic change in a wide range of cytokines during early stages of pregnancy, with a critical period around 26-27 dpc. In addition, at 26-27 dpc, expression of the surface/adhesion molecules, CD5 and VCAM-1, is induced on vascular endothelium of the sheep endometrium, possibly as a direct consequence of the changed cytokine environment, and may be involved in directing the changes in leucocyte migration observed during pregnancy.
Asunto(s)
Citocinas/biosíntesis , Implantación del Embrión/inmunología , Endometrio/inmunología , Regulación de la Expresión Génica/inmunología , Preñez/inmunología , Molécula 1 de Adhesión Celular Vascular/biosíntesis , Animales , Movimiento Celular/inmunología , Femenino , Leucocitos/inmunología , Embarazo , Ovinos/inmunologíaRESUMEN
Escherichia coli is commonly isolated in canine pyometra, but little is known of the virulence factors that may be involved in the precipitation of this disease. The aim of this study was to compare the prevalence of uropathogenic virulence factor (UVF) genes in E. coli isolates from canine pyometra and from feces of healthy bitches to evaluate their role in the pathogenesis of pyometra. E. coli from 23 cases of canine pyometra and from the feces of 24 healthy bitches were analyzed, by polymerase chain reaction, for UVF genes associated with canine and human urinary tract infections (UTIs). The prevalences of UVFs in E. coli from canine pyometra were similar to that in canine and human uropathogenic E. coli. The prevalence of pap was greater (P=0.036) for E. coli from pyometra (52%) than for fecal isolates (21%), and the papGIII allele was present in all pap-containing isolates. The prevalences of genes for alpha-haemolysin and cytotoxic necrotising factor 1 were not significantly higher (P=0.075) in E. coli from pyometra than from feces. The proportion of pyometra strains with >or=3 UVFs was higher (P=0.039) than that of fecal strains, suggesting that possession of >or=3 UVF genes enhances the pathogenicity of the strain. Our findings demonstrate that E. coli associated with canine pyometra are similar to uropathogenic strains, and that operons that encode P fimbriae, alpha-haemolysin and cytotoxic necrotising factor 1 probably enhance the virulence and pathogenicity of the strain in the canine genital tract.
Asunto(s)
Enfermedades de los Perros/microbiología , Infecciones por Escherichia coli/veterinaria , Escherichia coli/patogenicidad , Enfermedades Uterinas/veterinaria , Factores de Virulencia/genética , Adhesinas de Escherichia coli/química , Adhesinas de Escherichia coli/genética , Animales , Toxinas Bacterianas/química , Toxinas Bacterianas/genética , Citotoxinas/química , Citotoxinas/genética , ADN Bacteriano/química , ADN Bacteriano/genética , Perros , Escherichia coli/genética , Infecciones por Escherichia coli/microbiología , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/genética , Heces/microbiología , Femenino , Proteínas Fimbrias/química , Proteínas Fimbrias/genética , Fimbrias Bacterianas/química , Fimbrias Bacterianas/genética , Hemaglutininas/química , Hemaglutininas/genética , Proteínas Hemolisinas/química , Proteínas Hemolisinas/genética , Reacción en Cadena de la Polimerasa/veterinaria , Análisis de Secuencia de ADN , Enfermedades Uterinas/microbiología , VirulenciaRESUMEN
Lactating animals are particularly susceptible to mastitis during the early stages of mammary gland involution following weaning. In this study we compared the phagocytic capacity of cells collected from sheep mammary secretions at different stages of involution. The ability of neutrophils and macrophages to ingest latex beads in an in vitro phagocytosis assay was found to be dependent on how heavily the phagocytes were loaded with milk constituents. There was a decline in the phagocytic capacity of neutrophils from 1 to 2 days after weaning, while macrophages collected from fully involuted glands were more effective phagocytes compared with earlier stages (7-15 days) of involution. In addition, dendritic cells present in fully involuted mammary gland secretions (30 days after weaning) were highly phagocytic. These studies demonstrate that neutrophils and macrophages in sheep mammary secretions at early stages of involution are incapacitated, and as such may compromise the immune status of the mammary gland.
Asunto(s)
Leucocitos/inmunología , Glándulas Mamarias Animales/metabolismo , Fagocitosis/fisiología , Ovinos/fisiología , Destete , Animales , Células Dendríticas/inmunología , Femenino , Citometría de Flujo , Lactancia/inmunología , Macrófagos/inmunología , Glándulas Mamarias Animales/inmunología , Microscopía Electrónica , Neutrófilos/inmunologíaRESUMEN
PROBLEM: Previous studies have shown that the proportion of gammadeltaTCR+ large granulated lymphocytes (LGLs) increased markedly during pregnancy and declined dramatically by 2 days after parturition in sheep interplacentomal uterine epithelium. In the present study, the distribution, dynamics and fate of these cells, just before, during and immediately after parturition are described. METHODS OF STUDY: Interplacentomal tissues were collected at 140 days postcoitus (dpc), 148 dpc, during parturition, 1-2 hr postpartum, 1 day postpartum (dpp) and 3 dpp, and were studied using light and electron microscopy, and immuno histochemistry. Uterine washings were collected at 148 dpc and examined for the presence of LGLs. Semi-thin Araldite sections taken at different stages were used to quantify the intraepithelial LGLs, non-granulated lymphocytes (NGLs) and apoptotic cells, whereas frozen sections were used to quantify CD45R+, CD8+ and gammadeltaTCR+ intraepithelial lymphocytes (IELs). RESULTS: A dramatic decline in the proportion of IELs in the luminal epithelium during parturition was observed, mainly because of the decline in CD45R+, CD8+ and gammadeltaTCR+ IELs. There was also a significant decline in the number of granules/ LGL at parturition. This was accompanied by the presence of apoptotic cells of which some were LGLs. The proportions of IELs, LGLs and apoptotic cells markedly increased at 3 dpp. LGLs were found both in uterine washings at 148 dpc and in the uterine lumen at 3 dpp. Apoptosis of glandular epithelial cells was also evident at parturition and markedly increased at 1 dpp. CONCLUSIONS: Our results demonstrated that the dramatic decline in the proportion of gammadeltaTCR+ LGLs at parturition was because of de-granulation, apoptosis and migration of these cells into the uterine lumen.