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Flow cytometric detection of activation-induced cell death (apoptosis) in peripheral blood lymphocyte subpopulations from healthy cats


Holznagel, Edgar; Hofmann-Lehmann, Regina; Allenspach, Karin; Hüttner, Silke; Willett, Brian; Groscurth, Peter; Niederer, Eva; Lutz, Hans (1996). Flow cytometric detection of activation-induced cell death (apoptosis) in peripheral blood lymphocyte subpopulations from healthy cats. Veterinary Immunology and Immunopathology, 52(1-2):1-14.

Abstract

Human peripheral blood lymphocytes (PBLs) from healthy individuals are resistant to in vitro-induced apoptosis, but activated human lymphocytes can readily undergo apoptosis. The activation of human lymphocytes is accompanied by the upregulation of a cell surface antigen, the major histocompatibility complex (MHC) class II-antigen. Only a minority of PBLs are usually MHC class II-antigen-positive in healthy humans. In contrast, in healthy cats the majority of feline PBLs are MHC class II-antigen-positive. We have, therefore, investigated the sensitivity of feline peripheral blood lymphocytes obtained from specified pathogen free (SPF) cats to the induction of apoptosis. Feline PBLs from SPF cats (n = 16) and human PBLs from healthy donors (n = 2) were isolated. After short-term culture, cells were examined for the presence of fragmented DNA as a result of apoptosis by a DNA agarose gel electrophoresis method and for the presence of DNA double strand breaks by in situ 3' end labeling. In addition, relative DNA content per cell was flow cytometrically determined using propidium iodide (PI) or 7-actinomycin-D (7-AAD) and apoptotic cells were identified on the basis of a reduced DNA content. Cell surface antigens and cellular DNA were analyzed simultaneously by dual-color flow cytometric analyses in order to study lymphocyte subsets. Single- and dual-color analysis revealed that, in contrast to human lymphocytes, feline lymphocytes rapidly underwent apoptosis when cultured overnight in medium. Furthermore, the majority of apoptotic cells was found within the MHC II-positive cell subject.

Abstract

Human peripheral blood lymphocytes (PBLs) from healthy individuals are resistant to in vitro-induced apoptosis, but activated human lymphocytes can readily undergo apoptosis. The activation of human lymphocytes is accompanied by the upregulation of a cell surface antigen, the major histocompatibility complex (MHC) class II-antigen. Only a minority of PBLs are usually MHC class II-antigen-positive in healthy humans. In contrast, in healthy cats the majority of feline PBLs are MHC class II-antigen-positive. We have, therefore, investigated the sensitivity of feline peripheral blood lymphocytes obtained from specified pathogen free (SPF) cats to the induction of apoptosis. Feline PBLs from SPF cats (n = 16) and human PBLs from healthy donors (n = 2) were isolated. After short-term culture, cells were examined for the presence of fragmented DNA as a result of apoptosis by a DNA agarose gel electrophoresis method and for the presence of DNA double strand breaks by in situ 3' end labeling. In addition, relative DNA content per cell was flow cytometrically determined using propidium iodide (PI) or 7-actinomycin-D (7-AAD) and apoptotic cells were identified on the basis of a reduced DNA content. Cell surface antigens and cellular DNA were analyzed simultaneously by dual-color flow cytometric analyses in order to study lymphocyte subsets. Single- and dual-color analysis revealed that, in contrast to human lymphocytes, feline lymphocytes rapidly underwent apoptosis when cultured overnight in medium. Furthermore, the majority of apoptotic cells was found within the MHC II-positive cell subject.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:05 Vetsuisse Faculty > Veterinary Clinic > Department of Farm Animals
Dewey Decimal Classification:570 Life sciences; biology
630 Agriculture
Uncontrolled Keywords:Apoptosis; Cat; CD4; CD8; Flow cytometry; MHC II
Language:English
Date:15 June 1996
Deposited On:28 Dec 2016 10:24
Last Modified:08 Dec 2017 21:40
Publisher:Elsevier
ISSN:0165-2427
Publisher DOI:https://doi.org/10.1016/0165-2427(95)05541-X
PubMed ID:8807772

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