IN-VITRO CYTOKINE PRODUCTION BY NORMAL HUMAN PERIPHERAL-BLOOD MONONUCLEAR-CELLS AS A MEASURE OF IMMUNOCOMPETENCE OR THE STATE OF ACTIVATION

Measurements of cytokine levels in serum may not adequately reflect th e cytokine-producing potential of immune cells because of the short ha lf-lives of cytokines and the presence of various inhibitors in human sera. In vitro cytokine production by peripheral blood mononuclear cel ls (PBMCs) can be an important and reliable measure of immunocompetenc e. Also, spontaneous in vitro release of cytokines by PBMCs may serve as a measure of their activation in vivo. In the present study, normal ranges for the in vitro production by PBMCs of interleukin-1 beta (IL -1 beta), tumor necrosis factor alpha (TNF-alpha), IL-2, and gamma int erferon (IFN-gamma) were established; the feasibility of using cryopre served PBMCs for assays of in vitro cytokine production was evaluated; and spontaneous (unstimulated) versus induced production of cytokines by fresh and cryopreserved PBMCs from healthy donors was compared. Su pernatants obtained from paired fresh and frozen PBMCs were quantitate d for IL-1 beta, TNF-alpha, IL-2, and IFN-gamma by using enzyme-linked immunosorbent assay or a radioimmunoassay standardized against World Health Organization cytokine standards. Fresh or cryopreserved PBMCs a ctivated with lipopolysaccharide produced comparable levels of IL-1 be ta. However, the mean levels of stimulated production of TNF-alpha, IF N-gamma, and IL-2 were significantly higher in cryopreserved versus fr esh PBMCs (P less than or equal to 0.0004). Correlations between the l evel of production of each cytokined by fresh versus cryopreserved in vitro-stimulated PBMCs were statistically significant, although of mod erate magnitude. Spontaneous cytokine release by fresh versus cyropres erved cells was not significantly different. No correlation was observ ed between the TNF-alpha and IL-1 beta or the IL-2 and IFN-gamma level s produced by PBMCs from healthy donors, indicating that in vitro prod uction of each cytokine is independently regulated. Normal ranges for levels of all four cytokines were established by using PBMCs for the e valuation of spontaneous and induced cytokine production is more conve nient, economical, and reliable for the serial monitoring of clinical trials. The biologic and clinical usefulness of serial cytokine produc tion assays is illustrated in patients with advanced cancer who receiv ed therapy with biologic response modifiers.