Polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs) and new flame retardants (NFRs) are known thyroid hormone (TH) disruptors, but their disrupting mechanisms in humans are not completely understood. In this study, we aimed to explore the disrupting mechanisms of the aforementioned chemicals via examining TH-regulated proteins and gene expression in human serum. Adult participants from an e-waste dismantling (exposed group) and a control region (control group) in South China provided blood samples for the research. Some compounds of PCBs, PBDEs, and NFRs showed strong binding affinity to the thyroid-stimulating hormone (TSH), thyroglobulin, thyroxine-binding globulin (TBG), gene expression of TH receptor α (TRα) and β, and iodothyronine deiodinase I (ID1). The highly exposed individuals had lower levels of TBG, TSH, and expression of TRα, but higher expression of ID1 than those of the control group. The disruption of TH-regulated proteins and gene expression suggested the exertion of different and, at times, even contradictory effects on TH disruption. However, no statistically significant difference was found in the TH levels between the exposed and the control group, implying that the TH disruption induced by these chemicals depends on the combined influence of multiple mechanisms. Gene expression appears to be an effective approach for investigations of TH disruption and the potential health effects.