title: A high-throughput single-cell analysis of human CD8⁺ T cell functions reveals discordance for cytokine secretion and cytolysis.
abstract: CD8+ T cells are a key component of the adaptive immune response to viral infection. An inadequate CD8+ T cell response is thought to be partly responsible for the persistent chronic infection that arises following infection with HIV. It is therefore critical to identify ways to define what constitutes an adequate or inadequate response. IFN-γ production has been used as a measure of T cell function, but the relationship between cytokine production and the ability of a cell to lyse virus-infected cells is not clear. Moreover, the ability to assess multiple CD8+ T cell functions with single-cell resolution using freshly isolated blood samples, and subsequently to recover these cells for further functional analyses, has not been achieved. As described here, to address this need, we have developed a high-throughput, automated assay in 125-pl microwells to simultaneously evaluate the ability of thousands of individual CD8+ T cells from HIV-infected patients to mediate lysis and to produce cytokines. This concurrent, direct analysis enabled us to investigate the correlation between immediate cytotoxic activity and short-term cytokine secretion. The majority of in vivo primed, circulating HIV-specific CD8+ T cells were discordant for cytolysis and cytokine secretion, notably IFN-γ, when encountering cognate antigen presented on defined numbers of cells. Our approach should facilitate determination of signatures of functional variance among individual effector CD8+ T cells, including those from mucosal samples and those induced by vaccines.
authors.affiliation: Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
authors.forename: Navin
authors.initials: N
authors.lastname: Varadarajan
authors.forename: Boris
authors.initials: B
authors.lastname: Julg
authors.forename: Yvonne J
authors.initials: YJ
authors.lastname: Yamanaka
authors.forename: Huabiao
authors.initials: H
authors.lastname: Chen
authors.forename: Adebola O
authors.initials: AO
authors.lastname: Ogunniyi
authors.forename: Elizabeth
authors.initials: E
authors.lastname: McAndrew
authors.forename: Lindsay C
authors.initials: LC
authors.lastname: Porter
authors.forename: Alicja
authors.initials: A
authors.lastname: Piechocka-Trocha
authors.forename: Brenna J
authors.initials: BJ
authors.lastname: Hill
authors.forename: Daniel C
authors.initials: DC
authors.lastname: Douek
authors.forename: Florencia
authors.initials: F
authors.lastname: Pereyra
authors.forename: Bruce D
authors.initials: BD
authors.lastname: Walker
authors.forename: J Christopher
authors.initials: JC
authors.lastname: Love
mesh_terms: D056704:Adaptive Immunity; D000938:Antigen-Presenting Cells / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D016207:Cytokines / Q000096:biosynthesis*; D003602:Cytotoxicity, Immunologic; D016655:HIV Core Protein p24 / Q000276:immunology; D015658:HIV Infections / Q000276:immunology; D020276:HIV Long-Term Survivors; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000096:biosynthesis; D059010:Single-Cell Analysis / Q000379:methods*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
chemical_list: D016207:Cytokines; D016655:HIV Core Protein p24; C514799:p24 protein, Human Immunodeficiency Virus Type 1; D007371:Interferon-gamma