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Flow Cytometric Measurement of Bovine Milk Neutrophil Phagocytosis

Flödescytometrisk bestämning av fagocytos hos neutrofila leukocyter isolerade från mjölk

Acta Veterinaria Scandinavica volume 28, pages 333–342 (1987)Cite this article

Abstract

A flow cytometric method for the evaluation of the phagocytic capacity of bovine milk polymorphonuclear neutrophils (PMN) is described. Milk PMN were isolated from stripping milk collected from udder quarters fitted with abraded intramammary devices (AIMD). A significant increase in the milk somatic cell count was observed in the stripping milk after the insertion of AIMD (308×103 and 1447×l03 cells/ml milk before respectively after the insertion of the AIMD, p < 0.001).

PMN were also isolated from blood by a discontinous gradient of Percoli. Blood and milk PMN were incubated for 15 min with FITC-labeled bacteria in a ratio of 1 PMN:20 bacteria and a final serum concentration of 10 %. The number of extracellular bacteria and the percentage of phagocytic cells were measured by a flow cytometer.

Percentage of phagocytized bacteria by milk PMN was significantly lower than that by blood PMN (p < 0.05). A smaller number of active phagocytes was present among cells isolated from milk than among cells isolated from blood. The phagocytic capacity of milk PMN reflects that of blood PMN in the same animal. A large variation in the phagocytic capacity of blood and milk PMN among animals was observed.

Sammanfattning

I arbetet beskrivs en flödescytometrisk metod för bestämning av fagocytoskapaciteten hos neutrofila leukocyter (PMN) i komjölk. PMN isolerades från eftermjölksprov från juverfjärdedelar i vars juvercisterner spiraler gjorda av plasttråd med skrovlig yta (AIMD) hade placerats. Cellkoncentrationen i mjölken ökade från 308 × 103 til 1447 × 103 celler/ml efter införandet av plastspiralerna i juverfjärdedelarna. Ökningen var signifikant (p < 0,001).

För isolering av PMN från blod användes en diskontinuerlig Percollgradient.

PMN från blod och mjölk inkuberades i 10 % serum med FITC-märkta bakterier i proportionen 1 PMN:20 bakterier. Inkubationstid, 15 min. Antalet extracellulära bakterier och frekvensen fagocyterande celler mättes med en flödescytometer.

PMN isolerade från mjölk fagocyterade signifikant lägre andel av bakterierna än PMN isolerade från blod (p < 0,05). En lägre andel av fagocyterna i mjölken var aktiva jämfört med i blodet.

Fagocytoskapaciteten hos PMN isolerade från mjölken speglar den hos blodet från samma djur. Variationen mellen djur är stor.

References

  • Brunsting A, Mullaney PF: Differential light scattering from spherical mammalian cells. Biophys. J. 1974,74, 439–453.

    Article  Google Scholar 

  • Bryant RE, DesPrez RM, VanWay MH, Rogers DE: Studies on human leukocyte motility. I. Effects of alterations in pH, electrolyte concentration and phagocytosis on leukocytes migration, adhensiveness and aggregation. J. exp. Med. 1966,124, 483–499.

    Article  CAS  Google Scholar 

  • Hageltorn M, Saad AM: Flow cytometric characterization of bovine blood and milk leukocytes. Amer. J. vet. Res. 1986,47, 2012–2016.

    CAS  PubMed  Google Scholar 

  • Henricks PAJ, Van Der Toi ME, Verhof J: Aggregation of human polymorphonuclear leucocytes during phagocytosis of bacteria. Immunology, 1984,52,671–678.

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kent GM, Newbould FHS: Phagocytosis and related phenomena in polymorphonuclear leukocytes from cow’s milk. Canad. J. comp. Med. 1969,55,214–219.

    Google Scholar 

  • Lee CS, Wooding FBP, Kemp P: Identification properties and differential counts of cell populations using electron microscopy of dry cows secretions, colostrum and milk from normal cows. J. Dairy Res. 1980,47, 39–50.

    Article  CAS  Google Scholar 

  • Newbould FHS: Enhancement of phagocytosis in bovine milk leukocytes in vitro. Canadl J. comp. Med. 1970,34, 261–264.

    CAS  Google Scholar 

  • Newbould FHS: The effect of added serum and glucose, and some inherent factors, on the phagocytosis in vitro by milk leukocytes from several cows. Canad. J. comp. Med. 1973,57, 189–194.

    Google Scholar 

  • Paape M J, Pearson RE, Wergin WP, Guidry AJ: Enhancement of chemotactic response of polymorphonuclear leukocytes into the mammary gland and isolation from milk. J. Dairy Sci. 1976,60, 53–62.

    Article  Google Scholar 

  • Paape MJ, Guidry AJ: Effect of fat and casein on intracellular killing of Staphylococcus aureus by milk leukocytes. Proc. Soc. exp. Med. 1977,755, 588–593.

    Article  Google Scholar 

  • Paape MJ, Pearson RE, Schultze WD: Variation among cows in the ability of milk to support phagocytosis and in the ability of polymorphonuclear leukocytes to phagocytose Staphylococcus aureus. Amer. J. vet. Res. 1978,59, 1907–1910.

    Google Scholar 

  • Paape MJ, Schultze WD, Guidry AJ, Kortum WM, Weinland BT: Effect of an intramam-mary polyethylene device on the concentration of leukocytes and immunoglobulins in milk and on the leukocyte response to Escherichia coli endotoxin and challenge exposure with Staphylococcus aureus. Amer. J. vet. Res. 1981,42,774–783.

    CAS  PubMed  Google Scholar 

  • Paape MJ, Norman JC: Intensification of milk somatic cell response to intramammary device. Amer. J. vet. Res. 1984,45, 1572–1575.

    CAS  PubMed  Google Scholar 

  • Paape M J, Acton JC, Weinland BT: Effect of IMD on milk yield, tissue damage and cellular composition. J. Dairy Sci. 1985, 68 (Supplement), 193.

    Google Scholar 

  • Pickering LK, Cleary TG, Kohl S, Getz S: Polymorphonuclear leukocytes of human colostrum. I. Oxidative metabolism and kinetics of killing of radiolabeled S. aureus. J. infect. Dis. 1980,742, 685–693.

    Article  Google Scholar 

  • Russell MW, Reiter Β: Phagocytic deficiency of bovine milk leukocytes: An effect of casein. J. Reticuloendothelial Soc. 1975,18, 1–13.

    CAS  Google Scholar 

  • Saad AM, Hageltorn M: Flow cytometric characterization of bovine blood neutrophil phagocytosis of fluorescent bacteria and zymosan particles. Acta vet. scand. 1985,26, 289–307.

    Article  CAS  Google Scholar 

  • Williams MR, Hibbitt KG, Field TR, Bunch KJ: Further studies on the variation among cows, bulls and calves in the ability of their blood polymorphonuclear leukocytes to kill Staphylococcus aureus. Brit. vet. J. 1984,140, 307–313.

    Article  CAS  Google Scholar 

  • Wisniowski J, Ronanikowa K, Grojewski H: Phagocytosis phenomenon in the mammary gland of cows. 1. The opsonizing factor and the phagocytic activity of leukocytes in milk and blood. Bull. Vet. Inst. Pulway 1965,9, 140–144.

    Google Scholar 

  • Young MD, Paape MJ, Guidry AJ: Ability of skimmed milk from quarters fitted with abraded intramammary device (AIMD) to support phagocytosis. J. Dairy Sci. 1984, 67 (Supplement), 172.

    Google Scholar 

Download references

Acknowledgment

Prof. Göran Åström is cordially thanked for helpful discussion and advices. Mrs. Gudrun Mellberg-Wieslander is thanked for typewriting the manuscript. This study was supported by grants from the Swedish Council for Forestry and Agricultural Research.

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Authors and Affiliations

  1. Department of Obstetrics and Gynaecology, Swedish University of Agricultural Sciences, S-750 07, Uppsala, Sweden

    Alaa M. Saad

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Saad, A.M. Flow Cytometric Measurement of Bovine Milk Neutrophil Phagocytosis. Acta Vet Scand 28, 333–342 (1987). https://doi.org/10.1186/BF03548601

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  • DOI: https://doi.org/10.1186/BF03548601

Keywords

  • intramammary device
  • milk cells
  • blood PMN

Acta Veterinaria Scandinavica

ISSN: 1751-0147