Thyrocytes Express a Functional Toll-Like Receptor 3: Overexpression Can Be Induced by Viral Infection and Reversed by Phenylmethimazole and Is Associated with Hashimoto’s Autoimmune Thyroiditis

Abstract

Toll-like receptors (TLRs) initiate an innate immune response. TLR3 on dendritic cells recognize double-stranded (ds) RNA and then signal increases in cytokines and recognition molecules important for immune cell interactions. In this report, we demonstrate TLR3 mRNA and protein are expressed on Fisher rat thyroid cell line-5 (FRTL-5) thyroid cells and are functional because incubating cells with polyinosine-polycytidylic acid causes 1) transcriptional activation of both the nuclear factor kappaB (NF-kappaB)/Elk1 and interferon (IFN) regulatory factor-3/IFN-beta signal paths, 2) posttranscriptional activation of NF-kappaB and ERK1/2, and 3) increased IFN-beta mRNA. TLR3 can be overexpressed, along with dsRNA-dependent protein kinase, major histocompatibility complex-I or II, and IFN regulatory factor-1, by transfecting dsRNA into the cells, infection with Influenza A virus, or incubation with IFN-beta, but not by incubation with dsRNA or IFNgamma, or by dsDNA transfection. A methimazole (MMI) derivative, phenylmethimazole, to a significantly greater degree than MMI, prevents overexpression by inhibiting increased transcriptional activation of IRF-3 and of IFN-stimulated response elements, phosphorylation of signal transducers and activation of transcription (STAT-1), but not NF-kappaB activation. TLR3 can be functionally overexpressed in cultured human thyrocytes by dsRNA transfection or IFN-beta treatment. Immunohistochemical studies show that TLR3 protein is overexpressed in human thyrocytes surrounded by immune cells in 100% of patients with Hashimoto's thyroiditis examined, but not in normal or Graves' thyrocytes. We conclude that functional TLR3 are present on thyrocytes; TLR3 downstream signals can be overexpressed by pathogen-related stimuli; overexpression can be reversed by phenylmethimazole to a significantly greater extent than MMI by inhibiting only the IFN regulatory factor-3/IFN-beta/signal transducers and activation of transcription arm of the TLR3 signal system; and TLR3 overexpression can induce an innate immune response in thyrocytes, which may be important in the pathogenesis of Hashimoto's thyroiditis and in the immune cell infiltrates.

Evading innate immunity in nonviral mRNA delivery: don’t shoot the messenger

“…Courreges et al describe the molecular basis for this inhibition, which seems to be a prevention of dsRNA-induced IRF3 translocation and homodimerization [105]. The observation that C10 blocks IRF3 transactivation is consistent with prior studies which demonstrate C10-mediated inhibition of the TLR3-regulated IRF3/IFN-β/STAT signal pathway [106,107]. In the same way, establishment of a cellular anti-RNA state can be prevented through inhibition of IRF7.…”

Proinflammatory response of human leukemic cells to dsRNA transfection linked to activation of dendritic cells

“…[15][16][17] In spite of the shared endpoints, the outcome of dsRNA treatment is dependent on cell type 15,30 and the method of exposure. 18,19,21,31 Indeed, in our study, we demonstrate that it is necessary to transfect dsRNA into the cytoplasm of AML cells in order to exert its immunostimulatory effects. These data corroborate the previously published data on dsRNA treatment of thyrocytes or murine non-plasmacytoid DC.…”

Increased expression of Toll-like receptor 3 in intrahepatic biliary epithelial cells at sites of ductular reaction in diseased livers

“…In addition, cultured RA synovial fibroblasts were activated by the TLR3 ligand polyI:C and by RNA released from necrotic synovial fluid cells, suggesting that necrotic cells may act as an endogenous TLR3 ligand leading to the stimulation of proinflammatory gene expression and autoimmunity [22][23][24]. The overexpression of TLR3 in thyrocytes is associated with the development of Hashimoto's autoimmune thyroiditis [25]. TLR3 activation can drastically enhance susceptibility to immune destruction of solid organs, as seen in autoimmune hepatitis [26].…”