Potential Endocrine Disrupting Effects Of Short Chain Chlorinated Paraffins And Organophosphate Flame Retardants

Endocrine disrupting chemicals(EDCs)is an exogenous substance or mixture that interferes with the production,release,transport,metabolism,binding,action or elimination of natural hormones in the body responsible for the maintenance of homeostasis and the regulation of developmental processes,which has recently been the research hotspot in the domain of environmental science.Compared with other developed countries,China is in the primary stage of this research and there are still many deficiencies,especially high-throughput platform applied to screen the potential endocrine disrupting effects of chemicals.In this study,we adopted two in vitro models,dual-luciferase reporter gene assay and H295 R steroidogenesis assay,to investigate the endocrine disrupting effects of three short chain chlorinated paraffins(SCCPs)and nine organophosphate flame retardants(OPFRs),which were frenquently detected in the environment.Molecular docking was also used to study the interactions between chemicals and corresponding receptors.The main work and achievements of the research were shown as followed:(1)In the first study,we adopted two in vitro models,dual-luciferase reporter gene assay and H295 R steroidogenesis assay to investigate the potential estrogen,thyroid hormone and glucocorticoid effects of three SCCPs(C10-40.40%,C10-66.10%,and C11-43.20%).The results of reporter gene assay via estrogen receptor ?(ER?)revealed that all test chemicals induced estrogenic effects,in the following order: C11-43.20% > C10-66.10% > C10-40.40%,with REC20 values of 6.5×10-9 mol/L(M),2.7×10-8 M,and 2.8×10-7 M respectively.C10-40.40% and C10-66.10% also demonstrated remarkable anti-estrogenic activities,with RIC20 values of 1.1×10-7 M and 5.4×10-7 M.Only C11-43.20% showed glucocorticoid receptor-mediated(GR)antagonistic activity,with a RIC20 value of 2.6×10-8 M.Meanwhile,all test SCCPs stimulated the secretion of 17?-estradiol(E2),and C11-43.20% significantly increased the production of cortisol at a high level in H295 R cell line.The results of real-time polymerase chain reaction(RT-PCR)showed some steroidogenic genes,such as StAR,17?HSD,CYP11A1,CYP11B1,CYP19,and CYP21,were upregulated in a concentration-dependent manner by test chemicals.(2)In the second study,we employed the dual-luciferase reporter gene assay via glucocorticoid and mineralocorticoid receptors(GR/MR),H295 R steroidogenesis assay and molecular docking to evaluate the endocrine disrupting effects of nine OPFRs.In the reporter gene assay,none of OPFRs showed GR or MR agonistic activities.TMPP,TPHP,and TDBPP exhibited GR antagonistic activities,with RIC20 values of 1.2×10-6 M,2.6×10-6 M,and 1.1×10-6 M respectively.TNBP,TMPP,TPHP,TDCIPP,and TDBPP showed MR antagonistic properties,in the following order of TDBPP > TPHP > TMPP > TNBP > TDCIPP,with RIC20 values of 7.5×10-7 M,7.9×10-7 M,9.5×10-7 M,5.1×10-6 M,and 5.2×10-6 M respectively.In the H295 R steroidogenesis assay,the fold changes of eight steroidogenic genes,including St AR,HMGR,3?HSD2,CYP11A1,CYP11B1,CYP11B2,CYP17,and CYP21 in response to OPFRs,were further studied.CYP17,CYP21,and CYP11B1 expressions were significantly downregulated following TMPP,TPHP,or TDBPP exposure at a concentration of 2×10-6 M compared to vehicle.TMPP and TDBPP,which possessed the strongest potential endocrine disrupting effects,decreased the productions of cortisol and aldosterone,respectively.The results of molecular docking showed OPFRs could interact with corresponding receptors through ? bonds,hydrophobic bond and van der waals force.