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ABSTRACT: Generation of inflammatory mediators and leukocyte recruitment to infection at an epithelial surface were studied during Escherichia coli-induced mastitis. One uninfected gland of each of eight midlactation cows was challenged with only 30 CFU of E. coli McDonald strain 487, a serum-resistant isolate from a cow with mastitis. Bacteria grew logarithmically during the first 10 to 12 h after challenge, reaching concentrations of more than 10(5) CFU/ml with no detectable host response during this time. An intense inflammatory reaction began approximately 12 h after the challenge and was characterized by a breakdown in the blood-milk permeability barrier followed by pyrexia and a pronounced leukocytic influx. Coincident with the onset of mammary inflammation was the appearance of neutrophil chemotactic activity in the milk from infected glands. Factors able to upregulate CD18 expression on peripheral blood neutrophils also appeared in milk at this time. The lack of appearance of chemotactic and CD18-upregulating activities until 12 h after challenge indicated that delays in neutrophil recruitment resulted from an initial lack of bacterial recognition and inflammatory mediator production. Production of complement fragment C5a, tumor necrosis factor, and interleukin-1 (IL-1) occurred earlier than production of IL-6 or IL-8. The early and intense production of C5a indicates that this chemoattractant may be more important than IL-8 during the initial recruitment and activation of neutrophils to a developing E. coli infection.
Infection and Immunity 09/1997; 65(8):3286-92. · 4.16 Impact Factor
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ABSTRACT: To define causes of increased susceptibility to coliform mastitis after parturition.
12 healthy Holstein cows assigned to 2 groups. Group-1 cows (n = 6) had calved between 6 and 10 days earlier. Group-2 cows (n = 6) were in midlactation.
Cows from each group were paired and challenge exposed with Escherichia coli in 1 mammary gland. Mastitis severity was determined by bacterial concentration in milk, pyrexia, and milk production. Measures of host defense were neutrophil chemotaxis, adhesion molecule expression, leukocyte recruitment, and cytokine production.
After challenge exposure, group-1 cows had more rapid E coli growth, higher peak bacterial concentration, and higher fever. Leukocyte recruitment was poor in 1 group-1 cow that had peracute mastitis. In contrast, leukocyte recruitment in 5 other group-1 cows began sooner than that in group-2 cows. In these group-1 cows, prechallenge-exposure milk somatic cell counts (SCC) were significantly lower than those in group-2 cows. Prechallenge-exposure SCC were correlated to stimulated CD18 expression (R2 = 0.79), and both measures correlated inversely with bacterial growth rate (R2 = -0.75). Values for tumor necrosis factor alpha, interleukin 1, and interleukin 8 in group-1 cows after challenge exposure were greater than or equal to those in group-2 cows.
Weak leukocyte recruitment to the mammary gland is associated with increased severity of coliform mastitis. Impaired production of cytokines measured is not a cause of increased susceptibility to coliform mastitis in early lactation.
Low milk SCC after calving may increase susceptibility to severe coliform mastitis.
American Journal of Veterinary Research 12/1996; 57(11):1569-75. · 1.27 Impact Factor
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ABSTRACT: Production of inflammatory cytokines and concentrations of growth hormone and insulin-like growth factor-I (IGF-I) were studied during experimental Escherichia coli mastitis to determine their potential involvement in reduced animal performance during infectious disease. During the first 10 to 14 hr after intramammary infusion of E. coli, bacteria multiplied to maximum levels of 10(4)-10(9) cfu/ml of milk with no clinical signs of mastitis. A rapid and intense inflammatory response, characterized by udder swelling, increased bovine serum albumin (BSA) and somatic cell count (SCC) in milk of infected glands, and elevated rectal temperature and serum cortisol concentration, began at approximately 12 hr after challenge. Lactational performance was reduced greatly at 24 hr, and the maximal decrease averaged 76% and 63% among infected and uninfected glands, respectively, of challenged cows; three cows became temporarily agalactic in all glands. By 6 days, all cows had nearly or completely eliminated the E. coli, and milk production had partially recovered. Milk composition showed an initial decrease in fat percentage followed by an increase thereafter. Protein percentage was increased and lactose content was reduced during most of the mastitic episode. High concentrations of tumor necrosis factor (TNF) and interleukin-1 (IL-1) were detected in milk of infected glands, and their appearance preceded or coincided with development of the mammary inflammation, systemic reaction, and hypogalactia. Serum growth hormone concentration was higher among challenged cows, whereas serum IGF-I concentrations changed little during the mastitic episode. Concentrations of IGF-I in milk whey increased from 5.0 to 12.2 ng/ml among infected glands and from 4.4 to 8.5 ng/ml among contralateral, uninfected glands; IGF binding proteins also increased in the milk of infected glands. These data demonstrate that (i) reduced lactational performance is not caused by reduced concentrations of growth hormone or IGF-I and (ii) inflammatory cytokines are produced at a time consistent with a possible role in the inhibition of milk synthesis.
Proceedings of The Society for Experimental Biology and Medicine 12/1995; 210(2):140-9.
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Science 10/1995; 269(5230):1590-1. · 31.20 Impact Factor
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ABSTRACT: The role of interleukin 1 (IL-1) as an inflammatory mediator during mastitis and the therapeutic effect of recombinant human IL-1 receptor antagonist (IL-1ra) for bovine mastitis was studied. Cows were intramammarily infused with lipopolysaccharide (25 micrograms) in 1 mammary gland. Half the cows also received infusions of 5 mg of IL-1ra into the same mammary gland just prior to endotoxin infusion and 4, 8, and 12 hours later. After endotoxin infusion, tumor necrosis factor and high IL-1 bioactivity were detected in whey from infused glands. Vascular permeability changes and neutrophil accumulation in milk paralleled the appearance of cytokines. A systemic reaction, characterized by pyrexia and an increase in blood cortisol concentration, also were observed. Milk yield was inhibited and milk composition was altered in infused and noninfused glands. The increase in IL-1 bioactivity in milk after endotoxin infusion was almost completely prevented in glands receiving IL-1ra. However, IL-1ra had no effect on local inflammation, systemic reaction, or impairment in productive performance. These results indicate that IL-1 does not mediate its effect within the milk compartment, and suggest either that IL-1 is not critical to the mastitic response or that intramammary infusion of IL-1ra does not place the antagonist where IL-1 interacts with its receptor.
American Journal of Veterinary Research 04/1995; 56(3):313-20. · 1.27 Impact Factor
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ABSTRACT: Milk somatic cells play a protective role against infectious disease in the bovine mammary gland. Many genetic and environmental factors affect the number and kinds of leukocytes that account for the vast majority of somatic cells in milk. Neutrophils constitute the vast majority of somatic cells in mammary glands that are infected with mastitis pathogens. The recruitment of neutrophils into the infected mammary gland is a normal part of the cow's defense mechanisms that is very effective for eradicating the majority of infections that occur. For many reasons, milk production and milk quality are negatively impacted by the presence of inflammation in infected glands. Because of the negative effects of high SCC in milk, various approaches are needed to reduce milk SCC. In the future, genetic gains for milk quality and mastitis resistance may be made by removing bulls from breeding programs when their daughters are predisposed to high SCC.
Journal of Dairy Science 03/1994; 77(2):619-27. · 2.56 Impact Factor
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ABSTRACT: The cDNA clone encoding for the bovine peripheral lymph node homing receptor (L-selectin) was isolated and sequenced. The predicted amino acid sequence of bovine L-selectin showed an overall high identity with that of human and murine L-selectin. However, the cytoplasmic tail of bovine L-selectin showed little similarity to that of human and murine L-selectin. The monoclonal antibody DREG-56, which recognizes human L-selectin, blocked the binding of bovine peripheral blood lymphocytes to high-walled endothelial venules in murine peripheral lymph nodes. Surface expression of bovine L-selectin was high in lymphocytes isolated from peripheral lymph nodes and low in lymphocytes isolated from Peyer's patches. This evidence strongly suggests that bovine L-selectin is a peripheral lymphocyte homing receptor.
Veterinary Immunology and Immunopathology 09/1993; 37(3-4):201-15. · 2.08 Impact Factor
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ABSTRACT: In Holstein cattle, an inherited disease has been recognized recently in which leukocytes lack surface glycoproteins termed beta 2 integrins, which are important in cell adhesion processes. This disease is the homologue of leukocyte adhesion deficiency in human beings and has been termed bovine leukocyte adhesion deficiency. The molecular basis of this disease is failure to produce normal CD18. The gene encoding bovine CD18 and its abnormal mutation have been sequenced, allowing specific diagnosis of the condition by DNA amplification by polymerase chain reaction followed by specific endonuclease digestion. This test was applied to formalin-fixed archival tissues from 18 cattle that had been admitted to the veterinary medical teaching hospital between 1975 and 1991 and that had had persistent and severe neutrophilia. Blood samples were collected from 2 additional cattle, and leukocytes from these samples also were tested. Fourteen cattle were confirmed to have been homozygous for the bovine leukocyte adhesion deficiency gene. Cattle with this condition had ranged in age from 2 weeks to 8 months at admission. They typically had had chronic bacterial infections that had failed to respond to or had recurred after conventional treatment. Consistent findings in these cattle included signs of bronchopneumonia, gingivitis, periodontitis, and peripheral lymphadenopathy. Severe neutrophilia, usually without a left shift, was a hallmark of the disease; consistent clinical biochemical findings included hypoalbuminemia, hyperglobulinemia, and hypoglycemia. This disease is important because it mimics common calfhood diseases such as pneumonia and diarrhea, but is ultimately consistently fatal before adulthood.
Journal of the American Veterinary Medical Association 03/1993; 202(3):445-9. · 1.79 Impact Factor
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ABSTRACT: The role of interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor alpha during endotoxin-induced mastitis in cows was characterized. Six cows had 10 micrograms of Escherichia coli lipopolysaccharide infused into 1 mammary gland. Three other cows served as nontreated controls. Within 1.5 to 2.5 hours after infusion, endotoxin caused obvious edema of the mammary gland and increased serum albumin concentration in milk of infused glands 6 times. Milk somatic cell count began to increase 3 to 5 hours after infusion in all treated glands. At 7 hours after infusion, somatic cell counts were increased > 10 times, compared with counts in milk from control cows. Pyrexia of > 1 C developed in only 1 cow, but all treated cows had serum cortisol concentrations > 50 ng/ml in response to endotoxin treatment. High concentrations of IL-1 (10 to 600 U/ml) and IL-6 (2 to 22 U/ml) were detected in milk of infused glands beginning 2.5 to 4 hours after infusion. Endotoxin did not induce detectable amounts of tumor necrosis factor activity in milk or serum. Swelling and mammary gland permeability changes preceded any detectable increase in IL-1 and IL-6 activity, indicating that these clinical signs of inflammation were not mediated by these cytokines. Systemic responses and the leukocytic influx into endotoxin-infused glands developed after or concurrently with initial increases in IL-1 and IL-6 activities in milk. These results suggested that IL-1 and IL-6 may have a role in mammary gland defenses and in the pathophysiologic changes during endotoxin-induced mastitis.
American Journal of Veterinary Research 01/1993; 54(1):80-5. · 1.27 Impact Factor
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ABSTRACT: Two point mutations were identified within the gene encoding bovine CD18 in a Holstein calf afflicted with leukocyte adhesion deficiency (LAD). One mutation causes an aspartic acid to glycine substitution at amino acid 128 (D128G) in the highly conserved extracellular region of this adhesion glycoprotein, a region where several mutations have been found to cause human LAD. The other mutation is silent. Twenty calves with clinical symptoms of LAD were tested, and all were homozygous for the D128G allele. In addition, two calves homozygous for the D128G allele were identified during widespread DNA testing, and both were subsequently found to exhibit symptoms of LAD. The carrier frequency for the D128G allele among Holstein cattle in the United States is approximately 15% among bulls and 6% among cows. This mutation is also prevalent among Holstein cattle throughout the world, placing this disorder among the most common genetic diseases known in animal agriculture. All cattle with the mutant allele are related to one bull, who through the use of artificial insemination sired many calves in the 1950s and 1960s. The organization of the dairy industry and the diagnostic test described herein will enable nearly complete eradication of bovine LAD within 1 year. These results also demonstrate that bovine LAD is genetically homologous and phenotypically similar to human LAD, thus providing a useful animal model for studies of LAD and beta 2 integrin function.
Proceedings of the National Academy of Sciences 11/1992; 89(19):9225-9. · 9.68 Impact Factor
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American Journal Of Pathology 07/1992; 140(6):1489-92. · 4.89 Impact Factor
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ABSTRACT: The bovine cDNA (CD18) encoding CD18, a cell-surface glycoprotein involved in multiple leukocyte functions, was sequenced and compared with the human and murine sequences. Portions of the 5'- and 3'-untranslated regions of the nucleotide sequences are conserved among the three species, including a 3' A+T-rich region believed to regulate mRNA stability and translational efficiency. The 2833-bp bovine sequence coded for a protein of 769 amino acids (aa). Overall, the deduced aa sequences were greater than 80% identical among the three species. The aa 96-389 and those in the cytoplasmic domain were very highly conserved with approx. 95% aa identity. All Cys residues and potential Asn-glycosylation sites present in the bovine sequence were also present in the human and murine sequences. The aa identity was also found in those regions where mutations were found to cause the genetic disease, leukocyte adhesion deficiency. These data identify functionally important regions of the CD18 mRNA and protein.
Gene 06/1992; 114(2):267-71. · 2.34 Impact Factor
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The Cornell veterinarian 05/1992; 82(2):103-9.
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ABSTRACT: A series of experiments was conducted to define the role of cortisol in the hypogalactia during endotoxin-induced mastitis. In the first experiment, three of six nonmastitic cows were given a continuous infusion of trilostane, a 3 beta-hydroxysteroid dehydrogenase inhibitor that blocks enhanced cortisol synthesis. Trilostane had no effect in these cows. In the second experiment, six midlactation cows were given 10 micrograms of endotoxin in each of two homolateral quarters to induce mastitis. Three of these cows also received trilostane. Increased serum cortisol following endotoxin infusion was blocked by trilostane treatment, whereas serum glucose and rectal temperatures were unaffected. Preventing the cortisol increase failed to reduce hypogalactia in endotoxin-infused or uninfused quarters. Decreases in milk production and increases in measures of mammary inflammation were greater in trilostane-treated cows, indicating that endogenous cortisol may moderate the cow's inflammatory response. In the third experiment, three of six nonmastitic cows were injected intramuscularly with 150 IU of ACTH. Serum cortisol concentration exceeded 70 ng/ml for at least 3 h in cows receiving ACTH. This cortisol concentration, comparable with concentrations found during endotoxin mastitis, did not inhibit milk production. Together, these data demonstrate that the acute cortisol increase does not mediate the hypogalactia associated with endotoxin-induced mastitis.
Journal of Dairy Science 04/1992; 75(3):739-46. · 2.56 Impact Factor
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ABSTRACT: Male Holtzman rats were given subcutaneous doses of a purified Pasteurella multocida group D heat-labile toxin on alternate days for up to 22 days. Rats were necropsied at 18 days or 36 days (14 days after last dose of toxin) or when moribund, and testicles were taken for histologic and ultrastructural examination. Other selected tissues, including liver and spleen, were taken for histologic examination. Histologically, testicular and splenic lesions occurred more consistently and at much smaller doses when compared with lesions in other target organs such as liver. Testicular and splenic lesions were present in all rats (6/6) given 0.8 micrograms/kg toxin and were seen in some rats (1/6) given as little as 0.2 micrograms/kg toxin. Only 3/6 rats given 0.8 micrograms/kg toxin had hepatic lesions; no hepatic lesions were seen at doses of 0.2 micrograms/kg. Testicles from toxin-treated rats were smaller and weighed less than controls. Seminiferous tubules were moderately dilated and lined by polygonal sertoli cells. The normal spermatogenic maturation sequence and mature spermatids were absent, and many tubules contained multinucleate spermatocytes. Severely affected tubules were necrotic and mineralized. Ultrastructurally, there was necrosis of adluminal spermatocytes, multinucleate cell formation, and spaces between Sertoli cell plasma membranes. Testicular lesions were similar to those described for vitamin D-deficient rats, vitamin A-deficient rats, vasectomized rats, and rats given intravenous tumor necrosis factor; however, rats given lethal doses of toxin did not have elevated levels of TNF alpha activity.
Toxicologic Pathology 02/1992; 20(1):103-11. · 1.91 Impact Factor
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ABSTRACT: Healthy, midlactation cows were given intramammary infusions of 10 micrograms of endotoxin in two homolateral quarters. Productive, inflammatory, and systemic responses were studied to investigate the pathophysiological effects of mastitis on lactational performance. Endotoxin suppressed milk yield in all quarters of treated cows. A more severe and prolonged suppression occurred in infused quarters compared with uninfused quarters. The fat percentage of milk from all quarters was increased with a greater increase occurring in infused quarters. The protein composition of milk was elevated, and the lactose concentration was depressed in infused quarters. Mammary inflammation--as measured by milk SCC, NAGase, serum albumin, and lactoferrin--was limited to infused quarters. Changes in milk NAGase closely paralleled changes in milk SCC. Daily feed intake was unaffected, and serum glucose levels did not decline following infusion. The lactose concentration of urine increased rapidly after infusion. Reduction in milk yield in all quarters, but varying changes in milk composition in infused versus uninfused quarters suggest that mastitic hypogalactia is mediated by multiple pathophysiological events and is not solely due to inflammatory damage to the mammary epithelium. Part of the reduced lactational performance may result from escape of milk components from the udder into the circulation.
Journal of Dairy Science 12/1991; 74(11):3763-74. · 2.56 Impact Factor
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ABSTRACT: Midlactation cows were infused with 10 micrograms endotoxin in the same two homolateral quarters after each of several consecutive milkings to study the effect of prolonged, endotoxin-induced mastitis on lactational performance. The initial infusion induced an acute response with systemic involvement. Inflammation developed in infused quarters, and milk production declined and milk composition was altered in all quarters. Subsequent infusions failed to induce systemic responses. Furthermore, milk yield and composition in uninfused quarters returned to pre-treatment levels despite further infusions. In infused quarters, milk yield, protein percentage and serum albumin concentration showed partial recovery during the endotoxin infusion period. In contrast, decreased lactose concentration, and increased cell count, N-acetyl-beta-D-glucosaminidase and lactoferrin levels persisted throughout the infusion period. After infusions were stopped, all measurements returned to near pretreatment levels. These data demonstrate that systemic, but not local, responses become refractory to multiple intramammary endotoxin infusions, and that multiple infusions have continued but little progressive or permanent, inhibitory effects on lactational performance despite a persistent mammary leucocytosis.
Research in Veterinary Science 12/1991; 51(3):272-7. · 1.65 Impact Factor
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ABSTRACT: Nonpregnant lactating cows were given 100 micrograms of endotoxin via the jugular vein to determine effects of intravenous endotoxin administration on mammary inflammation and lactational performance. At the first milking (11 h) posttreatment, milk yield was reduced 33%. Milk fat percentage was elevated at this time, but lactose concentration was decreased. Milk yield and composition returned to pretreatment levels within 2 d. Clinical mastitis was not induced by endotoxin treatment, but milk SCC, NAGase, serum albumin, and lactoferrin were increased by 50%. This increase was small compared with increases during mastitis and may have resulted from lower milk volume. These results support the hypothesis that part of the reduced lactational performance during endotoxin mastitis is mediated by systemic pathophysiological responses and indicate that intravenous endotoxin administration may be a useful model to study adverse effects of infectious disease on lactational performance.
Journal of Dairy Science 11/1991; 74(10):3407-11. · 2.56 Impact Factor
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ABSTRACT: As part of a project to identify the pathophysiological cause or causes of mastitic hypogalactia, midlactation cows were infused in two homolateral quarters with 10 micrograms of endotoxin while being milked four times daily to resolve better the temporal changes in mammary synthetic activity during endotoxin mastitis. Milk fat was decreased by the first milking (5 h) postinfusion and then recovered rapidly. In contrast, milk yield and the yields of protein and lactose were not significantly inhibited until the second milking, and these yields recovered slowly thereafter. The decline in milk yield by infused quarters was only 20% greater than the decline by uninfused quarters in this experiment. Mammary inflammation developed rapidly in infused quarters as milk serum albumin concentration was maximal at the first milking. Milk SCC and NAGase were also elevated at this time, and maximal levels occurred at milkings 2 to 4. Increased temperature, increased cortisol, and a mild anorexia were apparent at the first milking only. Endotoxin treatment had no effect on serum prolactin or glucose. These data suggest that the delayed hypogalactia is consequent to the mammary inflammation and systemic responses following endotoxin infusion. The results indicate that different pathophysiological events may inhibit synthesis of the different milk components.
Journal of Dairy Science 06/1991; 74(5):1527-38. · 2.56 Impact Factor
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ABSTRACT: Twenty-one, middle to late lactation Holstein cows were assigned to one of three treatments in a completely randomized design to examine physiological changes associated with intramammary or intravenous administration of Escherichia coli endotoxin. Treatments were 1) Hank's balanced salt solution infusion in two contralateral quarters (control), 2) E. coli endotoxin infusion in two contralateral quarters, and 3) intravenous infusion of E. coli endotoxin. Blood was sampled and rectal temperature was measured at 30-min intervals. Endotoxin treatment was at 0900 h and sampling continued until 1700 h. Serum prolactin, cortisol, and 13,14-dihydro-15-keto-prostaglandin F2 alpha were measured. A pyretic response was observed in intravenous and intramammary treatment groups after endotoxin treatment. Response peak was higher (41.1 vs. 40.3 degrees C) and occurred later (6 vs. 4.5 h posttreatment) in the intramammary than the intravenous treatment group. Significant prolactin peaks were observed also in intravenous and intramammary endotoxin treatment groups. Prolactin peaked higher (288 vs. 112 ng/ml) and occurred sooner (1 vs. 4 h posttreatment) in the intravenous than in the intramammary treatment group. Cortisol followed a trend similar to prolactin. Cortisol peaked higher (100 vs. 82 ng/ml) and sooner (2.5 vs. 4.5 h posttreatment) in the intravenous than in the intramammary treatment group. Concentrations of 13,14-dihydro-15-keto-prostaglandin F2 alpha increased rapidly posttreatment in the intravenous group only.
Journal of Dairy Science 04/1990; 73(3):627-32. · 2.56 Impact Factor
