RhoG-ELMO1-RAC1 is involved in phagocytosis suppressed by mono-butyl phthalate in TM4 cells

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 μM) 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.