FREE SHIPPING over $40
OVER 10000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Canadian journal of veterinary research = Revue canadienne de recherche veterinaire1998; 62(2); 133-139;

Evaluation of an in vitro degranulation challenge procedure for equine pulmonary mast cells.

Abstract: Pulmonary mast cells (PMC) are important components of the inflammatory process in equine allergic lung diseases such as heaves. Very little, however, is known of the degranulation kinetics of these cells and thus, their pathophysiologic role remains largely speculative. The purpose of this study was to develop a repeatable protocol for in vitro equine PMC degranulation. Five mature horses (sex: 2 M, 3 F; age: 8.8 +/- 6.5 y), historically free of pulmonary disease and normal on clinical respiratory examination, arterial blood gas analysis, pulmonary mechanics testing and histamine inhalation challenge, were studied. Bronchoalveolar lavage was performed on 4 separate occasions, at least 2 d apart, in a different lung lobe on each occasion. The lavage fluid was concentrated by centrifugation. Cells were resuspended in modified HEPES/Tyrode, assessed for viability by Trypan blue exclusion, and PMC concentration determined. Cell inocula containing 30,000 PMC were incubated with 10(-8) to 6 x 10(-5) M A23187. Cells were then separated by centrifugation and histamine release (HR) was determined by fluorometric assay. The procedure was readily performed and yielded sufficient PMC for 30 to 60 inocula per lavage. Maximal HR (34.4 +/- 16.1%) was obtained with 10(-5) M A23187. The degranulation process was largely complete by 20 min but cell lysis was negligible. The challenge was repeatable within horse and produced a mean coefficient of variability of 23.0% following 20 min incubation with 10(-5) M A23187. We conclude that equine PMC degranulation can be repeatably performed in vitro and speculate that this protocol may be useful in further studies on the pathophysiology and treatment of equine allergic lung diseases.
Get Full Text
Publication Date: 1998-04-29 PubMed ID: 9553713PubMed Central: PMC1189460
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article
  • Research Support
  • Non-U.S. Gov't

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The study focuses on developing a reliable method to stimulate the degranulation of pulmonary mast cells (PMC) in horses in a controlled or in vitro environment, in order to better understand the role of these mast cells in equine allergic lung diseases.

Understanding the Research

  • The core part of the research revolves around the degranulation of pulmonary mast cells (PMC). These cells are integral to inflammation in horse lung diseases, including heaves, an allergic respiratory disease. Despite being key players in these diseases, there is limited understanding of their degranulation behaviour, and thus their precise pathophysiologic role remains largely speculative.
  • This study aims to provide some clarity by developing a repeatable in vitro (or laboratory-controlled) method for inciting and studying PMC degranulation.

Study Methodology

  • Researchers used five mature horses, all of which had no history of pulmonary disease. They underwent bronchoalveolar lavage—a procedure for sampling cells from the lung—on four separate occasions.
  • The cells harvested from this lung lavage were then concentrated by centrifugation and resuspended in a specialized solution for assessment of vitality through a procedure known as Trypan blue exclusion.
  • These cells were then stimulated, or ‘challenged’, with a compound named “A23187” in increasing concentrations. This substance induces the cells to degranulate or release inflammatory compounds. Post-challenge, the cells were separated by another round of centrifugation and the amount of histamine released, indicative of degranulation degree, was measured using a fluorometric assay.

Results and Conclusion

  • The study produced successful and repeatable results with the formulation of the in vitro degranulation challenge. It provided a sufficient quantity of PMC for multiple challenges.
  • Significant histamine release was achieved with a specific concentration of A23187, suggesting that the protocol was effective at inducing PMC degranulation. The process was largely complete within 20 minutes, with minimal cell lysis, thus maintaining the cells intact for further analysis.
  • Ultimately, the researchers conclude that in-vitro degranulation of equine PMCs is possible and repeatable. They suggest that this newfound technique will be helpful in future studies exploring the pathophysiology and potential treatments for equine allergic lung diseases.

Cite This Article

APA
Hare JE, Viel L, Conlon PD, Marshall JS. (1998). Evaluation of an in vitro degranulation challenge procedure for equine pulmonary mast cells. Can J Vet Res, 62(2), 133-139.

Publication

Canadian journal of veterinary research = Revue canadienne de recherche veterinaire
ISSN: 0830-9000
NlmUniqueID: 8607793
Country: Canada
Language: English
Volume: 62
Issue: 2
Pages: 133-139

Researcher Affiliations

Hare, J E
  • Department of Biomedical Sciences, Ontario Veterinary College, University of Guelph.
Viel, L
    Conlon, P D
      Marshall, J S

        MeSH Terms

        • Administration, Inhalation
        • Animals
        • Bronchoalveolar Lavage Fluid / cytology
        • Calcimycin / pharmacology
        • Carbon Dioxide / blood
        • Cytoplasmic Granules / physiology
        • Female
        • Histamine / administration & dosage
        • Histamine / pharmacology
        • Histamine Release
        • Horses
        • In Vitro Techniques
        • Lung / physiology
        • Male
        • Mast Cells / drug effects
        • Mast Cells / physiology
        • Mast Cells / ultrastructure
        • Oxygen / blood
        • Partial Pressure
        • Respiration / drug effects
        • Respiration / physiology
        • Respiratory Function Tests / veterinary

        References

        This article includes 28 references
        1. Wardlaw AJ, Cromwell O, Celestino D, Fitzharris P, Geddes DM, Collins JV, Kay AB. Morphological and secretory properties of bronchoalveolar lavage mast cells in respiratory diseases.. Clin Allergy 1986 Mar;16(2):163-73.
          pubmed: 2423272doi: 10.1111/j.1365-2222.1986.tb00760.xgoogle scholar: lookup
        2. Naylor JM, Clark EG, Clayton HM. Chronic obstructive pulmonary disease: Usefulness of clinical signs, bronchoalveolar lavage, and lung biopsy as diagnostic and prognostic aids.. Can Vet J 1992 Sep;33(9):591-8.
          pubmed: 17424075
        3. Winder NC, Grünig G, Hermann M, von Fellenberg R. Comparison of bronchoalveolar lavage and respiratory secretion cytology in horses with histologically diagnosed pulmonary disease.. Schweiz Arch Tierheilkd 1991;133(3):123-30.
          pubmed: 2063168
        4. McGorum BC, Dixon PM, Halliwell RE. Quantification of histamine in plasma and pulmonary fluids from horses with chronic obstructive pulmonary disease, before and after 'natural (hay and straw) challenges'.. Vet Immunol Immunopathol 1993 Apr;36(3):223-37.
          pubmed: 8506613doi: 10.1016/0165-2427(93)90021-ugoogle scholar: lookup
        5. Yamasaki Y, Shimamura O, Kizu A, Nakagawa M, Ijichi H. Modulation by alpha 2-adrenergic stimulation of IgE-mediated 14C-serotonin release from rat mast cells.. Agents Actions 1983 Jun;13(4):310-7.
          pubmed: 6137138doi: 10.1007/BF01971482google scholar: lookup
        6. Derksen FJ, Robinson NE, Armstrong PJ, Stick JA, Slocombe RF. Airway reactivity in ponies with recurrent airway obstruction (heaves).. J Appl Physiol (1985) 1985 Feb;58(2):598-604.
          pubmed: 3980362doi: 10.1152/jappl.1985.58.2.598google scholar: lookup
        7. Casolaro V, Galeone D, Giacummo A, Sanduzzi A, Melillo G, Marone G. Human basophil/mast cell releasability. V. Functional comparisons of cells obtained from peripheral blood, lung parenchyma, and bronchoalveolar lavage in asthmatics.. Am Rev Respir Dis 1989 Jun;139(6):1375-82.
          pubmed: 2471428doi: 10.1164/ajrccm/139.6.1375google scholar: lookup
        8. Wells E, Harper ST, Jackson CG, Mann J, Eady RP. Characterization of primate bronchoalveolar mast cells. I. IgE-dependent release of histamine, leukotrienes, and prostaglandins.. J Immunol 1986 Dec 15;137(12):3933-40.
          pubmed: 2431048
        9. Hare JE, Viel L, O'Byrne PM, Conlon PD. Effect of sodium cromoglycate on light racehorses with elevated metachromatic cell numbers on bronchoalveolar lavage and reduced exercise tolerance.. J Vet Pharmacol Ther 1994 Jun;17(3):237-44.
          pubmed: 7933063doi: 10.1111/j.1365-2885.1994.tb00239.xgoogle scholar: lookup
        10. Lindgren BR, Grundström N, Andersson RG. Comparison of the effects of clonidine and guanfacine on the histamine liberation from human mast cells and basophils and on the human bronchial smooth muscle activity.. Arzneimittelforschung 1987 May;37(5):551-3.
          pubmed: 2441713
        11. Kings MA, de Weck AL. Pharmacological and immunological aspects of histamine release from horse leucocytes.. Int Arch Allergy Appl Immunol 1980;62(4):397-408.
          pubmed: 6155351doi: 10.1159/000232542google scholar: lookup
        12. Magro AM, Rudofsky UH, Schrader WP, Prendergast J. Characterisation of IgE-mediated histamine release from equine basophils in vitro.. Equine Vet J 1988 Sep;20(5):352-6.
          pubmed: 2460333doi: 10.1111/j.2042-3306.1988.tb01544.xgoogle scholar: lookup
        13. Sommerhoff CP, Osborne ML, Gold WM, Lazarus SC. Functional and morphologic characterization of mast cells recovered by bronchoalveolar lavage from Basenji greyhound and mongrel dogs.. J Allergy Clin Immunol 1989 Feb;83(2 Pt 1):441-9.
          pubmed: 2465334doi: 10.1016/0091-6749(89)90131-0google scholar: lookup
        14. Schulman ES, Pollack RB, Post TJ, Peters SP. Histochemical heterogeneity of dispersed human lung mast cells.. J Immunol 1990 Jun 1;144(11):4195-201.
          pubmed: 1692857
        15. Soma LR, Beech J, Gerber NH Jr. Effects of cromolyn in horses with chronic obstructive pulmonary disease.. Vet Res Commun 1987;11(4):339-51.
          pubmed: 3118560doi: 10.1007/BF00346192google scholar: lookup
        16. Doucet MY, Jones TR, Ford-Hutchinson AW. Responses of equine trachealis and lung parenchyma to methacholine, histamine, serotonin, prostanoids, and leukotrienes in vitro.. Can J Physiol Pharmacol 1990 Mar;68(3):379-83.
          pubmed: 2322894doi: 10.1139/y90-053google scholar: lookup
        17. Tesarowski DB, Viel L, McDonell WN. Pulmonary function measurements during repeated environmental challenge of horses with recurrent airway obstruction (heaves).. Am J Vet Res 1996 Aug;57(8):1214-9.
          pubmed: 8836377
        18. Johansen T. Ethacrynic acid inhibition of histamine release from rat mast cells: effect on cellular ATP levels and thiol groups.. Eur J Pharmacol 1983 Sep 2;92(3-4):181-9.
          pubmed: 6195000doi: 10.1016/0014-2999(83)90285-6google scholar: lookup
        19. Flint KC, Leung KB, Pearce FL, Hudspith BN, Brostoff J, Johnson NM. Human mast cells recovered by bronchoalveolar lavage: their morphology, histamine release and the effects of sodium cromoglycate.. Clin Sci (Lond) 1985 Apr;68(4):427-32.
          pubmed: 2578914doi: 10.1042/cs0680427google scholar: lookup
        20. Chand N, Eyre P. Spasmolytic action of histamine in airway smooth muscle of horse.. Agents Actions 1978 Apr;8(3):191-8.
          pubmed: 665431doi: 10.1007/BF01966602google scholar: lookup
        21. Thomson JR, McPherson EA. Prophylactic effects of sodium cromoglycate on chronic obstructive pulmonary disease in the horse.. Equine Vet J 1981 Oct;13(4):243-6.
          pubmed: 6797824doi: 10.1111/j.2042-3306.1981.tb03507.xgoogle scholar: lookup
        22. SHORE PA, BURKHALTER A, COHN VH Jr. A method for the fluorometric assay of histamine in tissues.. J Pharmacol Exp Ther 1959 Nov;127:182-6.
          pubmed: 14446178
        23. Broadstone RV, LeBlanc PH, Derksen FJ, Robinson NE. In vitro responses of airway smooth muscle from horses with recurrent airway obstruction.. Pulm Pharmacol 1991;4(4):191-202.
          pubmed: 1806132doi: 10.1016/0952-0600(91)90011-qgoogle scholar: lookup
        24. Dirscherl P, Grabner A, Buschmann H. Responsiveness of basophil granulocytes of horses suffering from chronic obstructive pulmonary disease to various allergens.. Vet Immunol Immunopathol 1993 Oct;38(3-4):217-27.
          pubmed: 7507274doi: 10.1016/0165-2427(93)90082-fgoogle scholar: lookup
        25. Lowman MA, Rees PH, Benyon RC, Church MK. Human mast cell heterogeneity: histamine release from mast cells dispersed from skin, lung, adenoids, tonsils, and colon in response to IgE-dependent and nonimmunologic stimuli.. J Allergy Clin Immunol 1988 Mar;81(3):590-7.
          pubmed: 2450114
        26. Derksen FJ, Scott JS, Miller DC, Slocombe RF, Robinson NE. Bronchoalveolar lavage in ponies with recurrent airway obstruction (heaves).. Am Rev Respir Dis 1985 Nov;132(5):1066-70.
          pubmed: 4062037doi: 10.1164/arrd.1985.132.5.1066google scholar: lookup
        27. Viel L. Diagnostic procedures, prognosis and therapeutic approaches of chronic respiratory diseases in horses.. Can Vet J 1985 Jan;26(1):33-5.
          pubmed: 17422496
        28. Johansen T. Energy metabolism in rat mast cells in relation to histamine secretion.. Pharmacol Toxicol 1987;61 Suppl 2:1-20.
          pubmed: 2449683doi: 10.1111/j.1600-0773.1987.tb01597.xgoogle scholar: lookup

        Citations

        This article has been cited 1 times.
        1. White SJ, Couetil L, Richard EA, Marti E, Wilson PB. Microarray molecular mapping of horses with severe asthma. J Vet Intern Med 2024 Jan-Feb;38(1):477-484.
          doi: 10.1111/jvim.16951pubmed: 38071496google scholar: lookup
        Subscribe to our newsletter
        Join 99,850 horse owners like you who receive our email updates on horse health and nutrition.