Hörig H, Spagnoli G C, Filgueira L, Babst R, Gallati H, Harder F, Juretic A, Heberer M
Department of Surgery, University of Basel, Switzerland.
J Cell Biochem. 1993 Dec;53(4):343-51. doi: 10.1002/jcb.240530412.
Glutamine is required for the proliferation of lymphocytes, but quantitative effects on discrete steps of activation remain unknown to date. Therefore the influence of glutamine (range: 0 mM-1 mM) on the in vitro response of human peripheral blood mononuclear cells (PBMC) to a mitogenic anti-CD3 monoclonal antibody (mAb) was investigated. Expression of surface activation markers by flow cytometry, presence of mRNA of cytokine genes by polymerase chain reaction, release of cytokines by ELISA, and entering into the cell cycle by flow cytometry were sequentially analyzed. Proliferation was measured by a 3H-thymidine incorporation assay. mRNA coding for IL-2, IL-2 receptor, IL-4, IL-5, GM-CSF, and IFN-gamma was detectable independently from exogenous glutamine provision; expression of the cell surface activation marker CD69 was also glutamine independent. In contrast, later activation events including the expression of the surface activation markers CD25, CD45RO, and CD71 as well as the production of IFN-gamma were found to require exogenous glutamine supply. In contrast, production of TNF-alpha could be observed in the absence of glutamine and was increased to a limited extent by exogenous glutamine. The overall lymphocyte response as reflected by entering into the cell cycle and proliferation was directly correlated with the glutamine concentration of the culture medium. Efficient progression through the cell cycle was found to require at least 0.5 mM glutamine and an increase in glutamine concentration from 0.1 mM to 1 mM enhanced proliferation by 50%. These results were supported by data obtained following anti-CD3 stimulation of a CD4+ T cell clone.(ABSTRACT TRUNCATED AT 250 WORDS)
谷氨酰胺是淋巴细胞增殖所必需的，但迄今为止，其对激活离散步骤的定量影响尚不清楚。因此，研究了谷氨酰胺（范围：0 mM - 1 mM）对人外周血单个核细胞（PBMC）体外对促有丝分裂抗CD3单克隆抗体（mAb）反应的影响。通过流式细胞术依次分析表面激活标志物的表达、通过聚合酶链反应检测细胞因子基因的mRNA、通过酶联免疫吸附测定法检测细胞因子的释放以及通过流式细胞术检测进入细胞周期的情况。通过3H-胸腺嘧啶核苷掺入试验测量增殖。编码白细胞介素-2（IL-2）、IL-2受体、IL-4、IL-5、粒细胞-巨噬细胞集落刺激因子（GM-CSF）和干扰素-γ（IFN-γ）的mRNA的检测与外源性谷氨酰胺的供应无关；细胞表面激活标志物CD69的表达也不依赖于谷氨酰胺。相反，发现包括表面激活标志物CD25、CD45RO和CD71的表达以及IFN-γ的产生在内的后期激活事件需要外源性谷氨酰胺供应。相比之下，在没有谷氨酰胺的情况下可以观察到肿瘤坏死因子-α（TNF-α）的产生，并且外源性谷氨酰胺使其在有限程度上增加。进入细胞周期和增殖所反映的总体淋巴细胞反应与培养基中的谷氨酰胺浓度直接相关。发现细胞周期的有效进展至少需要0.5 mM谷氨酰胺，并且谷氨酰胺浓度从0.1 mM增加到1 mM可使增殖增加50%。抗CD3刺激CD4 + T细胞克隆后获得的数据支持了这些结果。（摘要截短于250字）
