Nitric oxide-repressed Forkhead factor FoxE1 expression is involved in the inhibition of TSH-induced thyroid peroxidase levels.

Thyroid peroxidase (TPO) is essential for thyroid hormone synthesis mediating the covalent incorporation of iodine into tyrosine residues of thyroglobulin process known as organification. Thyroid-stimulating hormone (TSH) via cAMP signaling is the main hormonal regulator of TPO gene expression. In thyroid cells, TSH-stimulated nitric oxide (NO) production inhibits TSH-induced thyroid-specific gene expression, suggesting a potential autocrine role of NO in modulating thyroid function. Indeed, NO donors downregulate TSH-induced iodide accumulation and organification in thyroid cells. Here, using FRTL-5 thyroid cells as model, we obtained insights into the molecular mechanism underlying the inhibitory effects of NO on iodide organification. We demonstrated that NO donors inhibited TSH-stimulated TPO expression by inducing a cyclic guanosine monophosphate-dependent protein kinase-mediated transcriptional repression of the TPO gene. Moreover, we characterized the FoxE1 binding site Z as mediator of the NO-inhibited TPO expression. Mechanistically, we demonstrated that NO decreases TSH-induced FoxE1 expression, thus repressing the transcripcional activation of TPO gene. Taken together, we provide novel evidence reinforcing the inhibitory role of NO on thyroid cell function, an observation of potential pathophysiological relevance associated with human thyroid pathologies that come along with changes in the NO production.

S-Nitrosylation of NF-κB p65 Inhibits TSH-Induced Na(+)/I(-) Symporter Expression.

Nitric oxide (NO) is a ubiquitous signaling molecule involved in a wide variety of cellular physiological processes. In thyroid cells, NO-synthase III-endogenously produced NO reduces TSH-stimulated thyroid-specific gene expression, suggesting a potential autocrine role of NO in modulating thyroid function. Further studies indicate that NO induces thyroid dedifferentiation, because NO donors repress TSH-stimulated iodide (I(-)) uptake. Here, we investigated the molecular mechanism underlying the NO-inhibited Na(+)/I(-) symporter (NIS)-mediated I(-) uptake in thyroid cells. We showed that NO donors reduce I(-) uptake in a concentration-dependent manner, which correlates with decreased NIS protein expression. NO-reduced I(-) uptake results from transcriptional repression of NIS gene rather than posttranslational modifications reducing functional NIS expression at the plasma membrane. We observed that NO donors repress TSH-induced NIS gene expression by reducing the transcriptional activity of the nuclear factor-κB subunit p65. NO-promoted p65 S-nitrosylation reduces p65-mediated transactivation of the NIS promoter in response to TSH stimulation. Overall, our data are consistent with the notion that NO plays a role as an inhibitory signal to counterbalance TSH-stimulated nuclear factor-κB activation, thus modulating thyroid hormone biosynthesis.

Functional toll-like receptor 4 conferring lipopolysaccharide responsiveness is expressed in thyroid cells.

Lipopolysaccharide (LPS), a glycolipid found in the cell wall of Gram-negative bacteria, exerts pleiotropic biological effects in different cell types. LPS is mainly recognized by the Toll-like receptor (TLR) 4/MD2/Cluster of differentiation 14 complex (CD14). We previously demonstrated that LPS produced a direct action on thyroid cells, including up-regulation of thyroglobulin gene expression. This work aimed to study further the effect of LPS on thyroid function and to elucidate the mechanism by which LPS is recognized by the thyroid cell. We could detect the transcript and protein expression of TLR4, MD2, and CD14 in thyroid cells, and that these proteins are localized at the plasma membrane. The sodium iodide symporter (NIS) is the transporter involved in the iodide uptake, the first step in thyroid hormonogenesis. We demonstrated that LPS increases the TSH-induced iodide uptake and NIS protein expression. The LPS agonist lipid A reproduced LPS effect, whereas the LPS antagonist, polymyxin B, abrogated it. By the use of anti-TLR4 blocking antibodies and the transient expression of TLR4 dominant-negative forms, we evidenced the involvement of TLR4 in the LPS action. The enrichment of TLR4 expressing Fisher rat thyroid cell line-5 (FRTL-5) cells confirmed that TLR4 confers LPS responsiveness to thyroid cells. In conclusion, we revealed for the first time that all the components of the LPS receptor complex are expressed in thyroid cells. Evidence that the effects of LPS on rodent thyroid function involve TLR4-induced signaling was obtained. The fact that thyroid cells are able to recognize and respond to LPS supports a role of the endotoxin as a potential modifier of thyroid function.

Nitric oxide/cGMP signaling inhibits TSH-stimulated iodide uptake and expression of thyroid peroxidase and thyroglobulin mRNA in FRTL-5 thyroid cells.

OBJECTIVE: Nitric oxide (NO) induces morphological and functional alterations in primary cultured thyroid cells. The aim of this paper was to analyze the direct influence of a long-term exposition to NO on parameters of thyroid hormone biosynthesis in FRTL-5 cells. DESIGN: Cells were treated with the NO donor sodium nitroprusside (SNP) for 24-72 h. MAIN OUTCOME: SNP (50-500 micromol/L) reduced iodide uptake in a concentration-dependent manner. The inhibition of iodide uptake increased progressively with time and matched nitrite accumulation. SNP inhibited thyroperoxidase (TPO) and thyroglobulin (TG) mRNA expression in a concentration-dependent manner. SNP enhanced 3',5'-cyclic guanosine monophosphate (cGMP) production. 3',5'-cyclic adenosine phosphate (cAMP) generation was reduced by a high SNP concentration after 48 h. 8-Bromoguanosine 3',5'-cyclic monophosphate (8-Br-cGMP), a cGMP analog, inhibited iodide uptake as well as TPO and TG mRNA expression. The cGMP-dependent protein kinase (cGK) inhibitor KT-5823 reversed SNP or 8-Br-cGMP-inhibited iodide uptake. Thyroid-stimulating hormone pretreatment for 24-48 h prevented SNP-reduced iodide uptake although nitrite levels remained unaffected. CONCLUSION: These findings favor a long-term inhibitory role of the NO/cGMP pathway on parameters of thyroid hormone biosynthesis. A novel property of NO to inhibit TPO and TG mRNA expression is supported. The NO action on iodide uptake could involve cGK mediation. The long-term inhibition of steps of thyroid hormonogenesis by NO could be of interest in thyroid pathophysiology.