Netea M G, Drenth J P, De Bont N, Hijmans A, Keuter M, Dharmana E, Demacker P N, van der Meer J W
Department of Medicine, University Hospital St. Radboud, Nijmegen, The Netherlands.
Cytokine. 1996 Sep;8(9):739-44. doi: 10.1006/cyto.1996.0098.
Whole blood cultures are used to study cytokine stimulation and release ex vivo. In the present study this method was compared with a more direct approach and a quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was used to assess mRNA expression for IL-1 beta and tumour necrosis factor alpha (TNF-alpha) and mRNA in whole blood. Stimulation of whole blood from normal donors with lipopolysaccharide (LPS) at various time intervals showed a parallel rise of immunogenic IL-1 beta and TNF-alpha as well as a rise of mRNA expression for IL-1 beta and TNF-alpha with peak levels for IL-1 beta after 4-6 h stimulation and for mRNA TNF-alpha expression after 2 h stimulation. These methods were used to explore cytokine production during the course of typhoid fever and after a 5 km run. In both conditions circulating cytokine concentrations were not influenced, but the TNF-alpha and IL-1 beta mRNA gene expression in circulating whole blood cells was increased in patients with typhoid fever. The LPS-stimulated production of TNF-alpha and IL-1 beta was decreased in both but there was no change for the mRNA content in whole blood for these cytokines. These findings demonstrate that RT-PCR is an attractive method to study the gene expression of cytokines in whole blood, an increased TNF-alpha and IL-1 beta gene expression is present in typhoid fever, and that the LPS stimulated downregulation of cytokines in exercise and typhoid fever may be mediated by post-transcriptional processes.
全血培养用于体外研究细胞因子的刺激和释放。在本研究中，将该方法与一种更直接的方法进行了比较，并使用定量逆转录聚合酶链反应（RT-PCR）来评估全血中白细胞介素-1β（IL-1β）和肿瘤坏死因子α（TNF-α）的mRNA表达以及mRNA含量。在不同时间间隔用脂多糖（LPS）刺激正常供体的全血，结果显示免疫原性IL-1β和TNF-α平行升高，以及IL-1β和TNF-α的mRNA表达升高，其中IL-1β在刺激4 - 6小时后达到峰值水平，TNF-α mRNA表达在刺激2小时后达到峰值水平。这些方法被用于探究伤寒热病程中和5公里跑步后细胞因子的产生情况。在这两种情况下，循环细胞因子浓度均未受影响，但伤寒热患者循环全血细胞中TNF-α和IL-1β mRNA基因表达增加。在这两种情况下，LPS刺激的TNF-α和IL-1β产生均减少，但这些细胞因子在全血中的mRNA含量没有变化。这些发现表明，RT-PCR是研究全血中细胞因子基因表达的一种有吸引力的方法，伤寒热中存在TNF-α和IL-1β基因表达增加的情况，并且运动和伤寒热中LPS刺激的细胞因子下调可能是由转录后过程介导 的。
