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Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes

Investigación publicada en Veterinary Immunology and Immunopathology

28 de julio de 2021

The gut associated lymphoid tissue (GALT) effector sites play a crucial role on the pathogenesis of many immune-mediated gastrointestinal diseases. The lymphocytes at these effector sites are principally T cells which present important morphological, phenotypical and functional differences. Flow cytometry (FC) is one of the most commonly used techniques to characterize intestinal lymphocytes in human and animal models. Published studies with a focus on dogs for intraepithelial lymphocytes (IEL) immunophenotyping exist in very limited numbers. Moreover, no lamina propria lymphocytes (LPL) isolation protocols in the canine species have been described for FC evaluation. In addition to immune intestinal dysregulation, imbalances in the peripheral blood immune system have been described in both human and animal gastrointestinal disorders. The aim of this study was to provide a protocol for canine IEL and LPL isolation for FC immunophenotyping of T cells subsets. Specifically, T helper, T cytotoxic, activated Th and Tc lymphocytes, regulatory, double negative, double positive, IFN-γ and IL-4 producing T cells, and to compare their respective populations between these effector sites and with the blood stream compartment in healthy dogs. The potential relationship of these cells distributions with age, sex and breed was also evaluated. This study included sixteen healthy dogs of different sexes and breeds with a mean age of 4.55 ± 2.93 years old. The selected protocols for the three immune compartments showed proper cell yield, purity, viability, and the absence of phenotypic and functional disturbances. Histologically, an adequate separation of the duodenal epithelium from the lamina propria was also observed. All the proposed T cells subsets were identified in the three immune compartments studied, showing some statistically significant differences in their distributions at these locations that highlight the importance of their individual evaluation. This study provides an adequate method for canine small intestine IEL and LPL isolation for FC immunophenotyping and is key for future studies on the gastrointestinal immune system associated with different canine diseases




Agulla B., Garcia-Sancho M., Sainz A., Rodriguez-Franco F., Diaz-Reganon D., Rodriguez-Bertos A. y Villaescusa A.




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Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes

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Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes


Participantes:

Universidad ComplutenseDepartamento de Medicina y Cirugía Animal. Facultad de Veterinaria. Universidad Complutense (UCM).

Universidad ComplutenseDepartamento de Sanidad Animal. Facultad de Veterinaria. Universidad Complutense (UCM).

Universidad ComplutenseServicio de Patología y Veterinaria Forense (SAP). Centro de Vigilancia Sanitaria Veterinaria (VISAVET). Universidad Complutense (UCM).







Veterinary Immunology and Immunopathology
FACTOR YEAR Q
2.046 2020

NLMID: 8002006

PMID: 34352607

ISSN: 0165-2427



TÍTULO: Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes


REVISTA: Vet. Immunol. Immunopathol.


NUMERACIÓN: 239:110305


AÑO: 2021


EDITORIAL: Elsevier Scientific


AUTORES: Agulla B., Garcia-Sancho M., Sainz A., Rodriguez-Franco F., Diaz-Reganon D., Rodriguez-Bertos A. and Villaescusa A.


PARTICIPANTES VISAVET


Antonio Manuel Rodríguez Bertos

DOI: https://doi.org/10.1016/j.vetimm.2021.110305


CITA ESTA PUBLICACIÓN:

Agulla B., Garcia-Sancho M., Sainz A., Rodriguez-Franco F., Diaz-Reganon D., Rodriguez-Bertos A. y Villaescusa A. Isolation and immunophenotyping by flow cytometry of canine peripheral blood and intraepithelial and lamina propria duodenal T lymphocytes. Veterinary Immunology and Immunopathology. 239:110305. 2021. (A). ISSN: 0165-2427. DOI: 10.1016/j.vetimm.2021.110305


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