Friberg D, Bryant J, Shannon W, Whiteside T L
Department of Pathology, University of Pittsburgh, Pennsylvania, USA.
Clin Diagn Lab Immunol. 1994 May;1(3):261-8. doi: 10.1128/cdli.1.3.261-268.1994.
Measurements of cytokine levels in serum may not adequately reflect the cytokine-producing potential of immune cells because of the short half-lives of cytokines and the presence of various inhibitors in human sera. In vitro cytokine production by peripheral blood mononuclear cells (PBMCs) can be an important and reliable measure of immunocompetence. 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 interferon (IFN-gamma) were established; the feasibility of using cryopreserved 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. Supernatants obtained from paired fresh and frozen PBMCs were quantitated 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 activated with lipopolysaccharide produced comparable levels of IL-1 beta. However, the mean levels of stimulated production of TNF-alpha, IFN-gamma, and IL-2 were significantly higher in cryopreserved versus fresh PBMCs (P < or = 0.0004). Correlations between the level of production of each cytokine by fresh versus cryopreserved in vitro-stimulated PBMCs were statistically significant, although of moderate magnitude. Spontaneous cytokine release by fresh versus cryopreserved cells was not significantly different.
由于细胞因子的半衰期较短，且人血清中存在各种抑制剂，因此血清中细胞因子水平的测量可能无法充分反映免疫细胞产生细胞因子的潜力。外周血单个核细胞（PBMC）的体外细胞因子产生可以是免疫能力的重要且可靠的指标。此外，PBMC体外自发释放细胞因子可作为其体内活化的指标。在本研究中，确定了PBMC体外产生白细胞介素-1β（IL-1β）、肿瘤坏死因子α（TNF-α）、IL-2和γ干扰素（IFN-γ）的正常范围；评估了使用冻存PBMC进行体外细胞因子产生测定的可行性；并比较了来自健康供体的新鲜和冻存PBMC自发（未刺激）与诱导产生细胞因子的情况。通过使用酶联免疫吸附测定法或针对世界卫生组织细胞因子标准标准化的放射免疫测定法，对从配对的新鲜和冷冻PBMC获得的上清液中的IL-1β、TNF-α、IL-2和IFN-γ进行定量。用脂多糖激活的新鲜或冻存PBMC产生的IL-1β水平相当。然而，冻存的PBMC与新鲜的PBMC相比，刺激产生的TNF-α、IFN-γ和IL-2的平均水平显著更高（P≤0.0004）。新鲜与冻存的体外刺激PBMC产生的每种细胞因子水平之间的相关性具有统计学意义，尽管程度适中。新鲜与冻存细胞的自发细胞因子释放没有显著差异。
