1 Department of Immunology, Medical University of Białystok, Białystok, Poland.
2 Department of Regenerative Medicine and Immune Regulation, Medical University of Białystok, Białystok, Poland.
Hum Exp Toxicol. 2019 May;38(5):578-587. doi: 10.1177/0960327119828198. Epub 2019 Feb 6.
To investigate the mechanism of apoptosis dependent on the Fas/FasL (Fas ligand) complex in the presence of N-nitrosodimethylamine (NDMA) in human leukocytes.
Polymorphonuclear neutrophils (PMNs) and peripheral blood mononuclear cells (PBMCs) were isolated form whole blood by density centrifugation. The concentration of NDMA was assessed by cellular toxicity assay. Apoptotic cells were assessed with flow cytometry and the expression of pro- and antiapoptotic proteins was investigated by Western blotting in PMNs and PBMCs treated with NDMA and/or FasL.
PMNs showed a higher ratio of apoptotic cells than PBMCs after exposure to NDMA and/or FasL. Enhanced apoptosis was related to the increased expression of proapoptotic proteins in neutrophils following exposure to either NDMA or FasL. In PBMCs, the relation was observed after exposure to FasL only. PMNs and PBMCs incubated with NDMA and FasL simultaneously demonstrated the highest increase in protein expression.
NDMA shows a stronger proapoptotic effect with PMNs than with PBMCs. The Fas/FasL complex, along with other proapoptotic proteins of the receptor (Fas, FADD) and mitochondrial pathway (Noxa, Puma, Bim), plays a key role in the induction of neutrophil apoptosis. Synergic effects of NDMA and FasL which lead to higher induction of apoptosis in PMNs than in PBMCs indicates a multistage and varied regulation of apoptosis in different populations of leukocytes.
研究在人白细胞中存在 N-亚硝基二甲胺 (NDMA) 时 Fas/FasL（Fas 配体）复合物依赖性细胞凋亡的机制。
通过密度离心从全血中分离多形核粒细胞 (PMN) 和外周血单核细胞 (PBMC)。通过细胞毒性测定评估 NDMA 的浓度。通过流式细胞术评估凋亡细胞，并通过 Western 印迹法在 PMN 和 PBMC 中研究 NDMA 和/或 FasL 处理后促凋亡和抗凋亡蛋白的表达。
与单独暴露于 NDMA 或 FasL 相比，PMN 在暴露于 NDMA 和/或 FasL 后表现出更高比例的凋亡细胞。增强的凋亡与中性粒细胞暴露于 NDMA 或 FasL 后促凋亡蛋白表达增加有关。在 PBMC 中，仅在暴露于 FasL 后观察到这种关系。同时孵育 NDMA 和 FasL 的 PMN 和 PBMC 表现出蛋白表达的最大增加。
与 PBMC 相比，NDMA 对 PMN 表现出更强的促凋亡作用。Fas/FasL 复合物以及受体（Fas、FADD）和线粒体途径（Noxa、Puma、Bim）中的其他促凋亡蛋白在诱导中性粒细胞凋亡中起关键作用。NDMA 和 FasL 的协同作用导致 PMN 中凋亡的诱导高于 PBMC，这表明不同白细胞群体中凋亡的多阶段和多样化调节。
