RhoG-ELMO1-RAC1 is involved in phagocytosis suppressed by mono-butyl phthalate in TM4 cells
RhoG-ELMO1-RAC1 is involved in phagocytosis suppressed by mono-butyl phthalate in TM4 cells
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DOI：
10.1007/s11356-018-3503-z
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发表时间：
2018-12-01
影响因子：
5.8
通讯作者：
Wang, Yubang
中科院分区：
文献类型：
--
作者：
Gong, Pan;Chen, Shanshan;Wang, Yubang
邻苯二甲酸二丁酯（DBP）是最主要的邻苯二甲酸酯之一，在环境中普遍存在。DBP及其活性代谢产物邻苯二甲酸单丁酯（MBP）的雄性生殖毒性已在体内和体外研究中得到证实。本研究的目的是探讨RhoG-ELMO 1-RAC 1在MBP破坏TM 4细胞吞噬功能中的作用。小鼠Sertoli细胞系（TM 4细胞）被维持并用不同水平的MBP（1、10和100 μ M）处理24小时。然后，收获细胞用于进一步的实验。采用流式细胞术、免疫荧光法和油红O染色法检测TM 4细胞的吞噬能力。RAC 1 pull-down检测RAC 1活性（GTP-RAC 1）。分别通过qRT-PCR和Western印迹分析与吞噬作用相关的mRNA和蛋白质（包括ELMO 1、RhoG和RAC 1）的表达。MBP可明显抑制TM 4细胞的吞噬功能，下调GTP-RAC 1的表达和向膜的移动。此外，ELMO 1蛋白表达下调，在MBP处理后的剂量依赖性的方式。此外，与吞噬作用相关的蛋白质，包括RhoG和GTP-RAC 1的表达显著降低，但总RAC 1的表达保持不变。用ELMO 1或RhoG质粒转染TM 4细胞后，GTP-RAC 1表达显著增加，但在MBP和ELMO 1/RhoG质粒共同处理后，GTP-RAC 1表达恢复。本研究提示MBP可通过RhoG-ELMO 1-RAC 1途径降低支持细胞的吞噬功能。
Di-n-butyl phthalate (DBP) is one of the most dominant phthalate esters and is ubiquitous in the environment. Male reproductive toxicity of DBP and its active metabolite mono-butyl phthalate (MBP) has been demonstrated in in vivo and in vitro studies. The objective of this study was to explore the roles of RhoG-ELMO1-RAC1 in phagocytosis disrupted by MBP in TM4 cells. Mouse Sertoli cell lines (TM4 cells) were maintained and treated by various levels of MBP (1, 10, and 100 mu M) for 24 h. Then, cells were harvested for further experiments. Phagocytic capacity of TM4 cells was detected by flow cytometry, immunofluorescence, and oil red O staining. RAC1 activity (GTP-RAC1) was measured by RAC1 pull-down assay. Expression of mRNA and protein related to phagocytosis including ELMO1, RhoG, and RAC1 was analyzed by qRT-PCR and Western blots, respectively. MBP inhibited phagocytosis of TM4 cells and downregulated GTP-RAC1 expression and movement to membrane markedly. Furthermore, ELMO1 protein expression was downregulated in a dose-dependent manner after MBP treatments. Additionally, expression of proteins relating to phagocytosis, including RhoG and GTP-RAC1, was decreased significantly, but expression of total-RAC1 remained unchanged. GTP-RAC1 expression increased dramatically after TM4 cells were transfected with ELMO1 or RhoG plasmid, but restored under co-treatments with MBP and ELMO1/RhoG plasmid. This study suggests that MBP can reduce the phagocytosis of Sertoli cells through RhoG-ELMO1-RAC1 pathway.