Serum biochemical profile and whole blood chemiluminescent (CL) responses in 8 Holstein cattle affected with leukocyte adhesion deficiency (LAD) were evaluated. Concentrations of sodium, chloride and calcium in serum from cattle affected with LAD were significantly (p<0.05) decreased as compared with controls. The characteristic changes in serum proteins were hypoalbuminemia and hyperglobulinemia, and the concentrations of albumin and gammaglobulin in serum from normal cattle and cattle affected with LAD were significantly (p<0.01) different. Significantly (p<0.01) diminished CL indices and prolonged peak time of CL responses in whole blood were detected in cattle affected with LAD. These findings indicate that the CL response associated with iC3b receptor mediated phagocytic activity is impaired in cattle affected with LAD. The whole blood CL assay appeared to be practical and useful for routine evaluation of blood samples from cattle affected with LAD.
Abstract Salp16, a 16‐kDa tick salivary gland protein, is known to be the molecule involved in the transmission of A naplasma phagocytophilum , an obligate intracellular pathogen causing zoonotic anaplasmosis, from its mammalian hosts to I xodes scapularis. Recently, the presence of A . phagocytophilum was documented in J apan and I xodes persulcatus was identified as one of its vectors. The purpose of this study was to identify S alp16 genes in I . persulcatus and characterize their function. Two cDNA clones encoding the S alp16 ‐like sequences were obtained from the salivary glands of fed female I . persulcatus ticks and designated S alp16 I per1 and I per2 . Gene expression analyses showed that the S alp16 I per genes were expressed specifically in the salivary glands and were up‐regulated by blood feeding. These proteins attenuated the oxidative burst of activated bovine neutrophils and inhibited their migration induced by the chemoattractant interleukin‐8 ( IL ‐8). These results demonstrate that S alp16 I per proteins contribute to the establishment of blood feeding as an immunosuppressant of neutrophil, an essential factor in innate host immunity. Further examination of the role of S alp16 I per in the transmission of pathogens, including A . phagocytophilum , will increase our understanding of the tick–host–pathogen interface.
In vivo responsiveness to epinephrine, expression of L‐selectin on neutrophils, changes in intracellular calcium ([Ca 2+ ] i ), sulfatide‐induced superoxide production and tyrosine phosphorylation in neutrophils were evaluated to elucidate the role of L‐selectin‐associated functions of normal and CD18‐deficient bovine neutrophils. The number of neutrophils in peripheral blood was significantly increased ( P < 0.05) in four normal calves at 5–20 min after in vivo administration of epinephrine; however, no significant increase of neutrophils was found in three calves with bovine leucocyte adhesion deficiency (BLAD). Expression of L‐selectin on neutrophils from three calves with BLAD was 61–77% of that of normal calves. Pretreatment of neutrophils with phorbol myristate acetate caused a marked decrease in the expression of L‐selectin on neutrophils from both normal and BLAD calves. The sulfatide‐induced sustained phase of [Ca 2+ ] i concentration in neutrophils from calves with BLAD was significantly ( P < 0.05) decreased. Following stimulation with aggregated IgG, the transient phase of [Ca 2+ ] i in neutrophils from normal and BLAD calves was increased; however, the sustained phase of [Ca 2+ ] i in BLAD neutrophils was significantly lower ( P < 0.05) than that of controls. Sulfatide‐induced O 2 − production and chemiluminescent response in neutrophils from calves with BLAD were 48–51% of those of normal calves and were inhibited by genistein and wortmannin, respectively, in a dose‐dependent manner. The amount of tyrosine phosphorylated 100 kDa protein in neutrophils from BLAD calves stimulated with sulfatides was 57% of that of controls. The degree of L‐selectin expression on neutrophils was correlated with the intracellular signalling events and the related superoxide production.
Two Holstein heifers with persistent and recurrent infections including ulcerative gingivitis, periodontitis, pneumonia, loss of teeth and stunted growth associated with marked neutrophilia were evaluated clinically and for neutrophil function, CD18 expression on neutrophils and CD18 genotype analysis by DNA-polymerase chain reaction (PCR) test. Adherence to nylon fibers and phagocytic activity of neutrophils from affected animals were significantly (p < 0.05) impaired as compared with those of controls. Neutrophils from affected heifers had decreased chemiluminescent (CL) responses when stimulated with opsonized zymosan, compared with those of controls. In contrast, neutrophils from affected heifers produced increased CL responses when stimulated with latex beads and phorbol myristate acetate compared with those of controls. The clinical findings, functional leukocyte abnormalities, deficiency in expression of CD18 on neutrophils, and the D128G mutation detected by DNA-PCR testing of affected heifers demonstrated that these heifers have bovine leukocyte adhesion deficiency (BLAD). Although both animals were confirmed to be homozygotes for BLAD by DNA-PCR test, they had differences in clinical, hematological and neutrophil functional characteristics.
The level of gelatinases in surrounding body fluids of actively remodelling tissue is indicative of basement membrane and extracellular matrix degradation under various physiological and pathological circumstances. To elucidate the association of gelatinase with mammary tissue remodelling during gradual or acute involution, in the first trial, goats milked twice daily (lactation) and goats receiving decreased milking frequency (involution) served to provide a total of 12 milk samples and 11 mammary secretion samples, respectively. In the second trial, 6 cows served to provide samples of dry secretion in 3 consecutive weeks immediately following milk stasis. Gelatin zymography was applied for gelatinase phenotyping and quantification on milk, plasma and the degranulation medium/lysate of milk somatic cells. Results indicated that the most prevalent gelatinase subtype switched from gelatinase A in milk to gelatinase B in involution secretion. Mammary secretion of goats during involution contained marginally higher protein level, significantly lower casein ratio and greater specific capacity of gelatinase B compared with those of milk during lactation. Specific capacities of gelatinases A and B in plasma of goats were similar during lactation and involution, while gelatinase B capacity in degranulation medium/lysates based on unit number of goat somatic cell was significantly higher during involution than during lactation. Milk stasis of cows induced a significant increase in specific capacity of gelatinase B, but not gelatinase A, of dry secretion up to the third week. Results of both trials agree that regional selective accretion of gelatinase B in milk might have played a role in mammary tissue remodelling during involution induced by either decreasing milking frequency or milk stasis. It is suggested that infiltrated polymorphonuclear neutrophils are one of the potential contributors responsible for the accumulation of gelatinase B during involution.
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