title: Tuberculin-induced lymphocyte proliferation in whole blood: an antigen specific method for assessing immunosuppressive agents.
abstract: Antigen-stimulated whole blood cultures have not been used to study the effects of immunosuppressive drugs. The aim of this study was to assess the potential usefulness of tuberculin purified protein derivative (PPD)-stimulated lymphocyte proliferation in whole blood for studying the effects of T cell inhibitory agents. We have investigated whether PPD causes antigen specific T cell proliferation, and the role of the major histocompatibility complex class II (MHC class II), co-stimulation and IL-2 in the development of this response. We have also studied the effects of prednisolone and cyclosporin on lymphocyte proliferation. Heparinised blood from healthy volunteers was diluted in culture medium and incubated with PPD. Cell proliferation, measured by liquid scintillation counting, was maximal using 1000 units/ml PPD incubated in 10% whole blood for 6-7 days. A population of large CD4+ lymphocytes appeared in cultures incubated with PPD, suggesting that the major responding population was composed of T lymphocytes. There was no significant response to the negative control antigen KLH, indicating that proliferation was antigen specific. Monoclonal antibodies (mAbs) against MHC, CD2, CD26, CD28 and IL-2 inhibited proliferation. Prednisolone was more potent than cyclosporin in this assay (IC50 values; prednisolone 20 nmol, cyclosporin 278 nmol). For the first time, this report shows that the PPD causes antigen specific lymphocyte proliferation in whole blood, which is dependent on antigen presentation via MHC class II, co-stimulation and IL-2 production. Because the proliferative response is dependent on the major interactions that lead to T cell activation, this simple assay can be used to assess the effects of novel immunomodulators.
authors.affiliation: North West Lung Research Centre, South Manchester University Hospitals Trust, Wythenshawe, M23 9LT, Manchester, UK. dave_sd@hotmail.com
authors.forename: S D
authors.initials: SD
authors.lastname: Singh
authors.forename: C G
authors.initials: CG
authors.lastname: Booth
mesh_terms: D001773:Blood Cells / Q000187:drug effects / Q000276:immunology; D002455:Cell Division / Q000187:drug effects; D016903:Drug Monitoring / Q000379:methods*; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D066298:In Vitro Techniques; D008213:Lymphocyte Activation / Q000187:drug effects*; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology; D014373:Tuberculin / Q000276:immunology / Q000494:pharmacology
chemical_list: D007166:Immunosuppressive Agents; D014373:Tuberculin