High-resolution characterization of cytokine-producing alloreactivity in naive and allograft-primed mice.
BACKGROUND
Whether alloreactive T cells in a naive host derive from naive or memory T cells remains unclear. It is also unclear whether graft rejection alters the phenotype of these T cells. Proliferation assays and cytokine enzyme-linked immunosorbent assays performed on culture supernatants do not differentiate primary T-cell alloreactivity from recall responses in allograft-primed mice, suggesting that these methods are inadequate measures of the alloreactive immune repertoire.


METHODS
To better characterize alloreactivity in naive and skin allograft-primed mice, we used a modified, high-resolution cytokine enzyme-linked immunosorbent spot assay capable of detecting cytokine production over short time periods.


RESULTS
Twenty-four-hour analysis of alloreactivity in mice that rejected fully MHC-disparate skin allografts revealed a high frequency of interferon (IFN)-gamma- and interleukin (IL)-4-producing, L-selectin-negative T cells, consistent with a memory phenotype. In contrast, 24-hr allostimulation of T cells from naive mice resulted in IL-2 production with minimal secretion of IFN-gamma or IL-4. The frequency of IL-2 producers was low and their phenotype was L-selectin positive, suggesting that they were naive and not memory T cells. When maintained in culture for 48 hr, however, the T cells from the primary mixed lymphocyte reaction began producing IFN-gamma, consistent with in vitro priming.


CONCLUSIONS
The primary alloresponse does not seem to involve clones that have been preprimed by environmental antigens, but instead behaves similarly to self-MHC-restricted immunity directed toward prototypic protein antigens: T cells with a naive phenotype are specifically induced to differentiate into high-frequency memory populations. These findings may have important implications for therapeutic induction of allograft tolerance.
Matesic
D
D
Department of Medicine, Cleveland Veterans Affairs Medical Center and the Institute of Pathology, Case Western Reserve University, Ohio 44106, USA.
Lehmann
P V
PV
Heeger
P S
PS
D000818:Animals; D016207:Cytokines / Q000096:biosynthesis* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006084:Graft Rejection / Q000276:immunology; D007156:Immunologic Memory; D016898:Interferon-alpha / Q000096:biosynthesis; D007376:Interleukin-2 / Q000096:biosynthesis; D015847:Interleukin-4 / Q000096:biosynthesis; D019041:L-Selectin / Q000276:immunology; D008198:Lymph Nodes / Q000276:immunology; D008213:Lymphocyte Activation / Q000276:immunology; D007959:Lymphocyte Culture Test, Mixed; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D010641:Phenotype; D016038:Skin Transplantation / Q000276:immunology*; D013154:Spleen / Q000276:immunology; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology*
D016207:Cytokines; D016898:Interferon-alpha; D007376:Interleukin-2; D019041:L-Selectin; D015847:Interleukin-4