Laboratory of Molecular Medicine, Department of Clinical Immunology Section 7631, Faculty of Health and Medical Sciences, University Hospital of Copenhagen, Denmark.
National Heart, Lung and Blood Institute, National Institute of Health, Bethesda, MD, 20814, USA.
Immunobiology. 2019 Mar;224(2):307-315. doi: 10.1016/j.imbio.2018.12.004. Epub 2018 Dec 27.
Complement activation is traditionally thought to occur in the extracellular space. However, it has been suggested that complement proteins are activated and function at additional locations. T cells contain intracellular stores of C3 and C5 that can be cleaved into C3a and C5a and bind to intracellular receptors, which have been shown to be of vital importance for the differentiation and function of these cells. However, whether the origin of the complement proteins located within T cells is derived from endogenous produced complement or from an uptake dependent mechanism is unknown. The presence of intracellular C3 in T cells from normal donors was investigated by fluorescence microscopy and flow cytometry. Moreover, mRNA expression levels of several genes encoding for complement proteins with primary focus on C3, C3aR, C5 and C5aR1 during resting state and upon activation of CD4 T cells were investigated by a quantitative PCR technique. Furthermore, the gene expression level was evaluated at different time points. We confirmed the presence of intracellular C3 protein in normal T-cells. However, we could not see any increase in mRNA levels using any activation strategy tested. On the contrary, we observed a slight increase in C3 and C5aR1 mRNA only in the non-activated T-cells compared to the activated T cells, and a decrease in the activated T-cells at different incubation time points. Our results show that there is a baseline intracellular expression of the complement C3, C5, C3aR and C5aR1 genes in normal CD4 T cells, but that expression is not increased during T-cell activation, but rather down regulated. Thus, the pool of intracellular complement in CD4 T cells may either be due to accumulated complement due low-grade expression or arise from the circulation from an uptake dependent mechanism, but these possibilities are not mutually exclusive.
补体激活传统上被认为发生在细胞外空间。然而，有人提出补体蛋白在其他位置被激活和发挥作用。T 细胞含有可被切割为 C3a 和 C5a 并与细胞内受体结合的细胞内 C3 和 C5 储备，这些受体已被证明对这些细胞的分化和功能至关重要。然而，位于 T 细胞内的补体蛋白的来源是来自内源性产生的补体还是依赖摄取的机制尚不清楚。通过荧光显微镜和流式细胞术研究了正常供体 T 细胞中细胞内 C3 的存在。此外，通过定量 PCR 技术研究了几种补体蛋白编码基因的 mRNA 表达水平，主要关注 C3、C3aR、C5 和 C5aR1，在 CD4 T 细胞静息状态和激活时。此外，还评估了不同时间点的基因表达水平。我们证实了正常 T 细胞中存在细胞内 C3 蛋白。然而，我们没有看到使用任何测试的激活策略时 mRNA 水平的任何增加。相反，与激活的 T 细胞相比，我们仅在未激活的 T 细胞中观察到 C3 和 C5aR1 mRNA 水平略有增加，并且在不同孵育时间点的激活 T 细胞中观察到减少。我们的结果表明，正常 CD4 T 细胞中存在补体 C3、C5、C3aR 和 C5aR1 基因的基线细胞内表达，但在 T 细胞激活过程中表达不会增加，而是下调。因此，CD4 T 细胞中的细胞内补体池可能是由于低水平表达导致的补体积累，或者是由于摄取依赖机制从循环中产生的，但这些可能性并不相互排斥。
