Histocultures allow studying intercellular interactions within human tissues, and they can be employed to model host-pathogen interactions under controlled laboratory conditions. Ex vivo infection of human tissues with human immunodeficiency virus (HIV), among other viruses, has been successfully used to investigate early disease pathogenesis, as well as a platform to test the efficacy and toxicity of antiviral drugs. In the present protocol, we explain how to process and infect with HIV-1 tissue explants from human tonsils and cervical mucosae, and maintain them in culture on top of gelatin sponges at the liquid-air interface for about two weeks. This non-polarized culture setting maximizes access to nutrients in culture medium and oxygen, although progressive loss of tissue integrity and functional architectures remains its main limitation. This method allows monitoring HIV-1 replication and pathogenesis using several techniques, including immunoassays, qPCR, and flow cytometry. Of importance, the physiologic variability between tissue donors, as well as between explants from different areas of the same specimen, may significantly affect experimental results. To ensure result reproducibility, it is critical to use an adequate number of explants, technical replicates, and donor-matched control conditions to normalize the results of the experimental treatments when compiling data from multiple experiments (i.e., conducted using tissue from different donors) for statistical analysis.

Ex vivo infection of human lymphoid tissue and female genital mucosa with human immunodeficiency virus 1 and histoculture / A. Introini, C. Vanpouille, W. Fitzgerald, K. Broliden, L. Margolis. - In: JOURNAL OF VISUALIZED EXPERIMENTS. - ISSN 1940-087X. - 2018:140(2018), pp. e57013.1-e57013.6. [10.3791/57013]

Ex vivo infection of human lymphoid tissue and female genital mucosa with human immunodeficiency virus 1 and histoculture

A. Introini
Primo
;
2018

Abstract

Histocultures allow studying intercellular interactions within human tissues, and they can be employed to model host-pathogen interactions under controlled laboratory conditions. Ex vivo infection of human tissues with human immunodeficiency virus (HIV), among other viruses, has been successfully used to investigate early disease pathogenesis, as well as a platform to test the efficacy and toxicity of antiviral drugs. In the present protocol, we explain how to process and infect with HIV-1 tissue explants from human tonsils and cervical mucosae, and maintain them in culture on top of gelatin sponges at the liquid-air interface for about two weeks. This non-polarized culture setting maximizes access to nutrients in culture medium and oxygen, although progressive loss of tissue integrity and functional architectures remains its main limitation. This method allows monitoring HIV-1 replication and pathogenesis using several techniques, including immunoassays, qPCR, and flow cytometry. Of importance, the physiologic variability between tissue donors, as well as between explants from different areas of the same specimen, may significantly affect experimental results. To ensure result reproducibility, it is critical to use an adequate number of explants, technical replicates, and donor-matched control conditions to normalize the results of the experimental treatments when compiling data from multiple experiments (i.e., conducted using tissue from different donors) for statistical analysis.
Cervical mucosa; Ex vivo; Female reproductive tract; Histoculture; HIV; Human immunodeficiency virus 1; Immunology and infection; Infection; Issue 140; Lymphoid tissue; Pathogenesis; Tissue explants; Tonsils
Settore MED/04 - Patologia Generale
Settore MED/50 - Scienze Tecniche Mediche Applicate
Settore BIO/13 - Biologia Applicata
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/871587
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