Effects of Levothyroxine Treatment on Fertility and Pregnancy Outcomes in Subclinical Hypothyroidism: A Systematic Review and Meta-Analysis of Randomized Controlled Trials.

Background: Subclinical hypothyroidism, defined by elevated thyrotropin (TSH) and normal free thyroxine levels, is associated with adverse pregnancy outcomes, including preterm birth, pre-eclampsia, and small for gestational age. Despite the uncertainty regarding the effectiveness of levothyroxine (LT4) treatment on pregnancy outcomes in subclinical hypothyroidism, LT4 is widely administered with a pre-treatment threshold TSH level of 2.5 mU/L. The aim of this study is to investigate the efficacy of periconceptional LT4 treatment for subclinical hypothyroidism, including TSH levels >2.5 mU/L, and identify the characteristics of subclinical hypothyroidism that can benefit from LT4 treatment. Methods: We conducted a systematic review and meta-analysis of randomized controlled trials from inception to February 2023. We analyzed the pooled effects of LT4 on subclinical hypothyroidism before and during pregnancy. The main outcomes before pregnancy were live birth, pregnancy, and miscarriage. The main outcomes during pregnancy were live birth, miscarriage, and preterm birth. We conducted subgroup analyses to compare the effects of LT4 on subclinical hypothyroidism with TSH levels of 2.5-4.0 and >4.0 mU/L. Results: Of the 888 studies identified, 27 full-text articles were screened for eligibility. Five studies on pre-conception treatment with 768 participants and eight studies on treatment during early pregnancy with 2622 participants were analyzed. One of the two studies on pre-conception treatment in subclinical hypothyroidism with TSH >4.0 mU/L had high risk of bias and the other was composed of 64 participants. Pre-conception LT4 treatment had no significant effect in improving rates of live births and pregnancies, or reducing miscarriages (risk ratio [RR], 95% confidence interval): 1.41 (0.84-2.36), 1.73 (0.88-3.39), and 0.46 (0.11-2.00), respectively. LT4 treatment during pregnancy was not significantly associated with higher rates of live births (RR 1.03, 0.98-1.09) nor decreased miscarriage rates (RR 1.01, 0.66-1.53). The effect of LT4 treatment on preterm birth during pregnancy was significantly different depending on the TSH values (p = 0.04); a positive effect was shown in the subclinical hypothyroidism subgroup with TSH >4.0 mU/L (RR 0.47, 0.20-1.10), while no significant effect was observed in the subgroup with TSH 2.5-4.0 mU/L (RR 1.35, 0.79-2.31). Conclusions: Pre-conceptional LT4 treatment for subclinical hypothyroidism does not improve fertility or decrease the incidence of miscarriages. However, further well-designed studies are needed for pre-conceptional treatment, especially in TSH >4.0 mU/L. LT4 treatment during pregnancy had a positive effect on preterm birth; nevertheless, this was only applicable to subclinical hypothyroidism with TSH >4.0 mU/L.

Thyroglobulin regulates the expression and localization of the novel iodide transporter solute carrier family 26 member 7 (SLC26A7) in thyrocytes.

Solute carrier family 26 member 7 (SLC26A7), identified as a causative gene for congenital hypothyroidism, was found to be a novel iodide transporter expressed on the apical side of the follicular epithelium of the thyroid. We recently showed that TSH suppressed the expression of SLC26A7 and induces its localization to the plasma membrane, where it functions. We also showed that the ability of TSH to induce thyroid hormone synthesis is completely reversed by an autocrine negative-feedback action of thyroglobulin (Tg) stored in the follicular lumen. In the present study, we investigated the potential effect of follicular Tg on SLC26A7 expression and found that follicular Tg significantly suppressed the promoter activity, mRNA level, and protein level of SLC26A7 in rat thyroid FRTL-5 cells. In addition, follicular Tg inhibited the ability of TSH to induce the membrane localization of SLC26A7. In rat thyroid sections, the expression of SLC26A7 was weaker in follicles with a higher concentration of Tg, as evidenced by immunofluorescence staining. These results indicate that Tg stored in the follicular lumen is a feedback suppressor of the expression and membrane localization of SLC26A7, thereby downregulating the transport of iodide into the follicular lumen.

Thyroid stimulating hormone suppresses the expression and activity of cytosolic sulfotransferase 1a1 in thyrocytes.

Sulfonation is an important step in the metabolism of dopamine, estrogens, dehydroepiandrosterone, as well as thyroid hormones. However, the regulation of cytosolic sulfotransferases in the thyroid is not well understood. In a DNA microarray analysis of rat thyroid FRTL-5 cells, we found that the mRNA expression of 10 of 48 sulfotransferases was significantly altered by thyroid stimulating hormone (TSH), with that of sulfotransferase family 1A member 1 (SULT1A1) being the most significantly affected. Real-time PCR and Western blot analyses revealed that TSH, forskolin and dibutyryl cyclic AMP significantly suppressed SULT1A1 mRNA and protein levels in a time- and concentration-dependent manner. Moreover, immunofluorescence staining of FRTL-5 cells showed that SULT1A1 is localized in the perinuclear area in the absence of TSH but is spread throughout the cytoplasm with reduced fluorescence intensity in the presence of TSH. Sulfotransferase activity in FRTL-5 cells, measured using 3'-phosphoadenosine-5'-phosphosulfate as a donner and p-nitrophenol as an acceptor substrate, was significantly reduced by TSH. These findings suggest that the expression and activity of SULT1A1 are modulated by TSH in thyrocytes.

Inhibitory effects of methimazole and propylthiouracil on iodotyrosine deiodinase 1 in thyrocytes.

Methimazole (MMI) and propylthiouracil (PTU) are commonly used for the treatment of Graves' disease. They share similar inhibitory effects on thyroid hormone biosynthesis by interfering with thyroid peroxidase (TPO)-mediated oxidation and organification of iodine. However, their potential effects on other thyroid functional molecules have not been explored in depth. To identify novel effects of MMI and PTU, DNA microarray analysis, real-time PCR, Western blotting, immunofluorescence staining and confocal laser scanning microscopy were performed using FRTL-5 rat thyroid cells. DNA microarray analysis indicated that both MMI and PTU suppress iodotyrosine deiodinase 1 (Iyd, Dehal1) mRNA levels. Further studies revealed that Dehal1 mRNA levels was stimulated by TSH, insulin and serum, while it was suppressed by iodine and a follicular concentration of thyroglobulin. MMI and PTU significantly suppressed Dehal1 expression induced by TSH, insulin and serum. On the other hand, although MMI suppressed Dehal1 expression in the absence of TSH, PTU only weakly suppressed Dehal1 without TSH. These results suggest that PTU and MMI may use different mechanisms to regulate Dehal1 expression, and TSH may play essential and differential roles in mediating PTU and MMI signals in thyrocytes. The drugs also inhibited re-distribution of Dehal1 protein into newly formed lysosomes following thyroglobulin endocytosis. These findings imply complex and multifaceted regulation of Dehal1 in the thyroid and suggest that MMI and PTU modulate Dehal1 expression and distribution of the protein in thyrocytes to exert their effect.

Follicular thyroglobulin induces cathepsin H expression and activity in thyrocytes.

Thyroglobulin (Tg) stored in thyroid follicles exerts a potent negative-feedback effect on each step of pre-hormone biosynthesis, including Tg gene transcription and iodine uptake and organification, by suppressing the expression of specific transcription factors that regulate these steps. Pre-hormones are stored in the follicular colloid before being reabsorbed. Following lysosomal proteolysis of its precursor, thyroid hormone (TH) is released from thyroid follicles. Although the suppressive effects of follicular Tg on each step of pre-hormone biosynthesis have been extensively characterized, whether follicular Tg accumulation also affects hormone reabsorption, proteolysis, and secretion is unclear. In this study we explored whether follicular Tg can regulate the expression and function of the lysosomal endopeptidases cathepsins. We found that in the rat thyroid cell line FRTL-5 follicular Tg induced cathepsin H mRNA and protein expression, as well as cathepsin H enzyme activity. Double immunofluorescence staining showed that Tg endocytosis promoted cathepsin H translocalization into lysosomes where it co-localized with internalized Tg. These results suggest that cathepsin H is an active participant in lysosome-mediated pre-hormone degradation, and that follicular Tg stimulates mobilization of pre-hormones by activating cathepsin H-associated proteolysis pathways.

Excessive Cytosolic DNA Fragments as a Potential Trigger of Graves' Disease: An Encrypted Message Sent by Animal Models.

Graves' hyperthyroidism is caused by autoantibodies directed against the thyroid-stimulating hormone receptor (TSHR) that mimic the action of TSH. The establishment of Graves' hyperthyroidism in experimental animals has proven to be an important approach to dissect the mechanisms of self-tolerance breakdown that lead to the production of thyroid-stimulating TSHR autoantibodies (TSAbs). "Shimojo's model" was the first successful Graves' animal model, wherein immunization with fibroblasts cells expressing TSHR and a major histocompatibility complex (MHC) class II molecule, but not either alone, induced TSAb production in AKR/N (H-2k) mice. This model highlights the importance of coincident MHC class II expression on TSHR-expressing cells in the development of Graves' hyperthyroidism. These data are also in agreement with the observation that Graves' thyrocytes often aberrantly express MHC class II antigens via mechanisms that remain unclear. Our group demonstrated that cytosolic self-genomic DNA fragments derived from sterile injured cells can induce aberrant MHC class II expression and production of multiple inflammatory cytokines and chemokines in thyrocytes in vitro, suggesting that severe cell injury may initiate immune responses in a way that is relevant to thyroid autoimmunity mediated by cytosolic DNA signaling. Furthermore, more recent successful Graves' animal models were primarily established by immunizing mice with TSHR-expressing plasmids or adenovirus. In these models, double-stranded DNA vaccine contents presumably exert similar immune-activating effect in cells at inoculation sites and thus might pave the way toward successful Graves' animal models. This review focuses on evidence suggesting that cell injury-derived self-DNA fragments could act as Graves' disease triggers.

A Novel Role for Flotillin-Containing Lipid Rafts in Negative-Feedback Regulation of Thyroid-Specific Gene Expression by Thyroglobulin.

BACKGROUND: Thyroglobulin (Tg) stored in thyroid follicles regulates follicular function in thyroid hormone (TH) synthesis by suppressing thyroid-specific gene expression in a concentration-dependent manner. Thus, Tg is an intrinsic negative-feedback regulator that can restrain the effect of thyrotropin (TSH) in the follicle. However, the underlying mechanisms by which Tg exerts its prominent autoregulatory effect following recognition by thyrocytes remains unclear. METHODS: In order to identify potential proteins that recognize and interact with Tg, mass spectrometry was used to analyze immunoprecipitated Tg-bound proteins derived from Tg-treated rat thyroid FRTL-5 cells. RESULTS: Flotillin 1 and flotillin 2, two homologs that are integral membrane proteins in lipid rafts, were identified as novel Tg-binding proteins with high confidence. Further studies revealed that flotillins physically interact with endocytosed Tg, and together these proteins redistribute from the cell membrane to cytoplasmic vesicles. Treatment with the lipid raft disrupter methyl-ß-cyclodextrin abolished both the endocytosis and the negative-feedback effect of Tg on thyroid-specific gene expression. Meanwhile, siRNA-mediated knockdown of flotillin 1 or flotillin 2 also significantly inhibited Tg effects on gene expression. CONCLUSION: Together these results indicate that flotillin-containing lipid rafts are essential for follicular Tg to be recognized by thyrocytes and exert its negative-feedback effects in the thyroid.

Follicular thyroglobulin enhances gene expression necessary for thyroid hormone secretion.

We have previously shown that follicular thyroglobulin (Tg) has an unexpected function as an autocrine negative-feedback regulator of thyroid hormone (TH) biosynthesis. Tg significantly suppressed the expression of genes necessary for iodide transport and TH synthesis by counteracting stimulation by TSH. However, whether follicular Tg also regulates intracellular TH transport and its secretion from thyrocytes is not known. In the present study, we examined the potential effect of follicular Tg on TH transport and secretion by quantifying the expression of two TH transporters: monocarboxylate transporter 8 (MCT8) and µ-crystallin (CRYM). Our results showed that follicular Tg at physiologic concentrations enhanced both the mRNA and protein expression levels of MCT8 and CRYM in a time- and dose-dependent manner in rat thyroid FRTL-5 cells. Although both the sodium/iodide symporter (NIS), an essential transporter of iodide from blood into the thyroid, and MCT8, a transporter of synthesized TH from the gland, were co-localized on the basolateral membrane of rat thyrocytes in vivo, Tg decreased NIS expression and increased the expression of MCT8 by counteracting TSH action. Thus, the effect of Tg on TH secretion opposed its previously described negative-feedback suppression of TH synthesis. Our results indicate that Tg mediates a complex intrinsic regulation of gene expression that is necessary to balance two opposing vectorial transport systems: the inflow of newly synthesized TH and the outflow of TH by external secretion.

Rapid preparation of high-purity nuclear proteins from a small number of cultured cells for use in electrophoretic mobility shift assays.

BACKGROUND: Highly purified nuclear protein is required when using an electrophoretic mobility shift assay (EMSA) to study transcription factors, e.g. nuclear factor-κB (NF-κB), a major transcription factor that regulates both innate and adaptive immune responses following infection. Although many protocols have been developed for nuclear protein extraction, they are not necessarily optimized for use in EMSA, often require a large number of cells and long processing times, and do not always result in complete separation of the nuclear and cytoplasmic fractions. RESULTS: We have developed a simple, rapid and cost-effective method to prepare highly purified nuclear proteins from a small number of both suspended and adherent cultured cells that yields nuclear proteins comparable to those prepared by a standard large-scale method. The efficiency of the method was demonstrated by using EMSA to show the successful detection, in multilple concurrent samples, of NF-κB activation upon tetradecanoyl phorbol acetate (TPA) stimulation. CONCLUSIONS: This method requires only a small number of cells and no specialized equipment. The steps have been simplified, resulting in a short processing time, which allows researchers to process multiple samples simultaneously and quickly. This method is especially optimized for use in EMSA, and may be useful for other applications that include proteomic analysis.

Iodine excess as an environmental risk factor for autoimmune thyroid disease.

The global effort to prevent iodine deficiency disorders through iodine supplementation, such as universal salt iodization, has achieved impressive progress during the last few decades. However, iodine excess, due to extensive environmental iodine exposure in addition to poor monitoring, is currently a more frequent occurrence than iodine deficiency. Iodine excess is a precipitating environmental factor in the development of autoimmune thyroid disease. Excessive amounts of iodide have been linked to the development of autoimmune thyroiditis in humans and animals, while intrathyroidal depletion of iodine prevents disease in animal strains susceptible to severe thyroiditis. Although the mechanisms by which iodide induces thyroiditis are still unclear, several mechanisms have been proposed: (1) excess iodine induces the production of cytokines and chemokines that can recruit immunocompetent cells to the thyroid; (2) processing excess iodine in thyroid epithelial cells may result in elevated levels of oxidative stress, leading to harmful lipid oxidation and thyroid tissue injuries; and (3) iodine incorporation in the protein chain of thyroglobulin may augment the antigenicity of this molecule. This review will summarize the current knowledge regarding excess iodide as an environmental toxicant and relate it to the development of autoimmune thyroid disease.

Thyroglobulin increases thyroid cell proliferation via the suppression of specific microRNAs.

Thyroglobulin (Tg), stored in the follicular lumen, has also been shown recently to perform two unexpected roles: as an autocrine negative-feedback suppressor of thyroid function in the presence of TSH and as a potent inducer of thyroid cell growth in the absence of TSH. However, the underlying molecular mechanism(s) remain unclear. To elucidate a molecular pathway linking Tg to increased cell proliferation, we examined the regulation of microRNAs (miRNAs) by Tg using an miRNA microarray. We identified 21 miRNAs whose expression was significantly suppressed by Tg in rat thyroid FRTL-5 cells. Using specific miRNA analogs, we determined that miR-16, miR-24, and miR-195 mediate the induction of thyroid cell growth by Tg. The expression of miR-16 and miR-195 target genes, Mapk8, Ccne1, and Cdc6, which were previously shown to be essential for TSH-stimulated thyroid cell growth, were also induced by Tg. Moreover, the Tg-induced expression of these genes was reduced by overexpression of miR-16 and miR-195. Similarly, the induction of c-Myc by Tg was reduced by miR-24 overexpression. These results suggest that Tg could alter thyroid cell proliferation by increasing the expression of cell division-related genes such as Mapk8, Ccne1, Cdc6, and c-Myc through its suppression of specific microRNAs (miR-16, miR-24, and miR-195). In addition, we identified phosphatidylinositol 3-kinase as a key signaling pathway, linking Tg with cell proliferation. The present data support an important role for miRNAs as effectors for the effect of Tg on cell proliferation and perhaps other functions of Tg in the thyroid cell.

A case of severe acute necrotizing pancreatitis after administration of sitagliptin.

A 55-year-old Japanese man with a 3-year history of type 2 diabetes mellitus was admitted to our hospital for upper abdominal pain. Control of diabetes mellitus was good with voglibose and metformin, with sitagliptin added to this regimen 8 months prior. His pancreatic enzyme levels were elevated, and abdominal computed tomography (CT) showed diffuse pancreatic swelling with fluid accumulation and ascites of CT grade 3. The patient was diagnosed with severe acute pancreatitis. There were no obvious causes for pancreatitis except the recently administered sitagliptin. Since incretin-related drugs entered the market, the number of incretin-related drugs prescriptions rapidly increased and so did the incidence of pancreatitis. There are several reports suggesting the correlation between incretin-related drugs and pancreatitis, such as a report based on data obtained from the United States Food and Drug Administration (FDA) which revealed a significant correlation between the administration of exenatide or sitagliptin and pancreatitis. However, there also is a report that denied the evidence for such in a large cohort study. The relation between incretin based drugs and pancreatitis is still controversial.

Demonstration of innate immune responses in the thyroid gland: potential to sense danger and a possible trigger for autoimmune reactions.

BACKGROUND: Autoimmune thyroid disease is an archetypal organ-specific autoimmune disorder that is characterized by the production of thyroid autoantibodies and lymphocytic infiltration into the thyroid. However, the underlying mechanisms by which specific thyroid antibodies are produced are largely unknown. Recent studies have shown that innate immune responses affect both the phenotype and the severity of autoimmune reactions. Moreover, it appears that even non-immune cells, including thyroid cells, have an ability to launch such responses. The aim of this study was to conduct a more detailed analysis of innate immune responses of the thyroid upon stimulation with various "non-self" and "self" factors that might contribute to the initiation of autoimmune reactions. METHODS: We used rat thyroid FRTL-5 cells, human thyroid cells, and mice to investigate the effects of various pathogen-associated molecular patterns (PAMPs), danger-associated molecular patterns (DAMPs), and iodide on gene expression and function that were related to innate immune responses. RESULTS: RT-PCR analysis showed that both rat and human thyroid cells expressed mRNAs for Toll-like receptors (TLRs) that sensed PAMPs. Stimulation of thyrocytes with TLR ligands resulted in activation of the interferon-beta (IFN-ß) promoter and the nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB)-dependent promoter. As a result, pro-inflammatory cytokines, chemokines, and type I interferons were produced. Similar activation was observed when thyroid cells were stimulated with double-stranded DNA, one of the typical DAMPs. In addition to these PAMPs and DAMPs, treatment of thyroid cells with high concentrations of iodide increased mRNA expression of various cytokines. CONCLUSION: We show that thyroid cells express functional sensors for exogenous and endogenous dangers, and that they are capable of launching innate immune responses without the assistance of immune cells. Such responses may relate to the development of thyroiditis, which in turn may trigger autoimmune reactions.

Propylthiouracil increases sodium/iodide symporter gene expression and iodide uptake in rat thyroid cells in the absence of TSH.

BACKGROUND: Propylthiouracil (PTU) and methimazole (MMI) are drugs that are widely used to treat Graves' disease. Although both exert an antithyroid effect primarily by blocking thyroid peroxidase activity, their molecular structure and other actions are different. We hypothesized that PTU and MMI may have differential effects on thyroid-specific gene expression and function. METHODS: The effects of PTU and MMI on thyroid-specific gene expression and function were examined in rat thyroid FRTL-5 cells using DNA microarray, reverse transcriptase (RT)-polymerase chain reaction (PCR), real-time PCR, Western blot, immunohistochemistry, and radioiodine uptake studies. RESULTS: DNA microarray analysis showed a marked increase in sodium/iodide symporter (NIS) gene expression after PTU treatment, whereas MMI had no effect. RT-PCR and real-time PCR analysis revealed that PTU-induced NIS mRNA levels were comparable to those elicited by thyroid-stimulating hormone (TSH). PTU increased 5'-1880-bp and 5'-1052-bp activity of the rat NIS promoter. While PTU treatment also increased NIS protein levels, the size of the induced protein was smaller than that induced by TSH, and the protein localized predominantly in the cytoplasm rather than the plasma membrane. Accumulation of (125)I in FRTL-5 cells was increased by PTU stimulation, but this effect was weaker than that produced by TSH. CONCLUSIONS: We found that PTU induces NIS expression and iodide uptake in rat thyroid FRTL-5 cells in the absence of TSH. Although PTU and MMI share similar antithyroid activity, their effects on other thyroid functions appear to be quite different, which could affect their therapeutic effectiveness.

Regulation of dual oxidase expression and H2O2 production by thyroglobulin.

BACKGROUND: Thyroglobulin (Tg) is a macromolecular precursor in thyroid hormone synthesis to which iodine is stably bound. Tg, which is stored in the follicular space, is also a potent negative feedback regulator of follicular function, and this is achieved by suppressing mRNA levels of thyroid-specific genes such as the sodium/iodide symporter (Slc5a5), Tg, and thyroid peroxidase. Dual oxidase 1 (DUOX1) and DUOX2, originally identified in the thyroid, are nicotinamide adenine dinucleotide phosphate (NADPH) oxidases that are necessary to produce the H2O2 required for thyroid hormone biosynthesis. Since follicular Tg regulates the expression of genes that are essential for thyroid hormone synthesis, we hypothesized that Tg might also regulate DUOX expression and H2O2 production. METHODS: Rat thyroid FRTL-5 cells were treated with Tg, and the mRNA expression of Duox1 and Duox2 and their corresponding maturation factors Duoxa1 and Duoxa2 were evaluated by DNA microarray and real-time PCR. Duox2 promoter activity was examined by luciferase reporter gene assay. Protein levels of DUOX2 were also examined by Western blot analysis. Intracellular H2O2 generation was quantified by a fluorescent dye, 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate, and acetyl ester (CM-H2DCFDA). RESULTS: mRNA levels of Duox2 and its activation factor Duoxa2 (but not Duox1 or Duoxa1) were significantly suppressed by Tg in a dose-dependent manner and a time-dependent fashion in rat thyroid FRTL-5 cells. DUOX2 promoter activity was significantly suppressed by Tg in a dose-dependent manner. Protein levels of DUOX2 and H2O2 generation in cells were also reduced by Tg treatment. CONCLUSIONS: We show that physiological concentrations of Tg suppressed the expression and function of DUOX2 in thyroid cells. These results suggest that Tg is a strong suppressor of the expression and the activity of DUOX2/DUOXA2, thereby regulating iodide organification and hormone synthesis in the thyroid. The evidence supports a reported model in which accumulated Tg in thyroid follicles plays important roles in autoregulating the function of individual follicles, which produces the basis of follicular heterogeneity.

Essential role of hormone-sensitive lipase (HSL) in the maintenance of lipid storage in Mycobacterium leprae-infected macrophages.

Mycobacterium leprae (M. leprae), the causative agent of leprosy, parasitizes within the foamy or enlarged phagosome of macrophages where rich lipids accumulate. Although the mechanisms for lipid accumulation in the phagosome have been clarified, it is still unclear how such large amounts of lipids escape degradation. To further explore underlying mechanisms involved in lipid catabolism in M. leprae-infected host cells, we examined the expression of hormone-sensitive lipase (HSL), a key enzyme in fatty acid mobilization and lipolysis, in human macrophage THP-1 cells. We found that infection by live M. leprae significantly suppressed HSL expression levels. This suppression was not observed with dead M. leprae or latex beads. Macrophage activation by peptidoglycan (PGN), the ligand for toll-like receptor 2 (TLR2), increased HSL expression; however, live M. leprae suppressed this increase. HSL expression was abolished in the slit-skin smear specimens from patients with lepromatous and borderline leprosy. In addition, the recovery of HSL expression was observed in patients who experienced a lepra reaction, which is a cell-mediated, delayed-type hypersensitivity immune response, or in patients who were successfully treated with multi-drug therapy. These results suggest that M. leprae suppresses lipid degradation through inhibition of HSL expression, and that the monitoring of HSL mRNA levels in slit-skin smear specimens may be a useful indicator of patient prognosis.

Thyroglobulin (Tg) activates MAPK pathway to induce thyroid cell growth in the absence of TSH, insulin and serum.

The growth of thyroid cells is tightly regulated by thyroid stimulating hormone (TSH) through the cyclic adenosine 3', 5'-monophosphate (cAMP) signaling pathway by potentiating the mitogenic activity of insulin and insulin-like growth factors (IGFs). However, we recently reported that thyroglobulin (Tg), a major product of the thyroid, also induces the growth of thyroid cells cultured in 0.2% serum in the absence of TSH and insulin. In this report, we demonstrate that Tg induced phosphorylation of molecules of the c-Raf/MEK/ERK pathway of the mitogen-activated protein kinase (MAPK). The MEK-1/2 inhibitor PD98059 suppressed Tg-induced phosphorylation of ERK1/2 and reduced bromodeoxyuridine (BrdU) incorporation. Tg also induced expression of the essential transcriptional factors c-Myc, c-Fos and c-Jun and phosphorylation of the retinoblastoma (Rb) protein. The present results, together with the previous report, suggest that Tg utilizes multiple signaling cascades to induce thyroid cell growth independent of TSH/cAMP stimulation.

Identification of microRNAs that mediate thyroid cell growth induced by TSH.

TSH is a major regulator of thyroid cell growth and endocrine function. It is known that cAMP and phosphatidylinositol 3-kinase (PI3K) are responsible for mediating the action of TSH. Activation of these signals results in the induction of a series of transcription factors and cell cycle regulating proteins, which induce cell proliferation. In addition to such canonical transcriptional regulation, it was recently shown that microRNA (miRNA or miR) constitutes another key mechanism for the regulation of gene expression. However, whether TSH action is mediated by miRNA in the thyroid is unknown. In this study, we have performed miRNA microarray analysis and demonstrated that TSH significantly decreases expression of 47 miRNA in thyroid cells. Among these, we have shown, using their specific agonists, that overexpression of miR-16 and miR-195 suppressed cell cycle progression and DNA synthesis that was induced by TSH. In silico analysis predicted that Mapk8, Ccne1, and Cdc6, the expression of which was up-regulated by TSH, are potential target genes for these miRNA, and overexpression of miR-16 and miR-195 suppressed expression of these target genes. The decrease of miR-16 and miR-195 expression by TSH was reproduced by forskolin and N(6),2'-O-dibutyryladenosine cAMP and reversed by the protein kinase A inhibitor H89 and the PI3K inhibitor LY294002. These results suggest that TSH activates cAMP/protein kinase A and PI3K cascades to decrease miR-16 and miR-195, which induce Mapk8, Ccne1, and Cdc6 to activate cell proliferation.

Current status of leprosy: epidemiology, basic science and clinical perspectives.

Leprosy has affected humans for millennia and remains an important health problem worldwide, as evidenced by nearly 250 000 new cases detected every year. It is a chronic infectious disorder, caused by Mycobacterium leprae, that primarily affects the skin and peripheral nerves. Recent advances in basic science have improved our knowledge of the disease. Variation in the cellular immune response is the basis of a range of clinical manifestations. The introduction of multidrug therapy has significantly contributed to a decrease in the prevalence of the disease. However, leprosy control activities, including monitoring and prevention programs, must be maintained.

Innate immune activation and thyroid autoimmunity.

CONTEXT: Autoimmune thyroid disease (AITD) is the archetypal organ-specific autoimmune disorder and is characterized by the production of thyroid autoantibodies. However, the underlying mechanisms by which specific antibodies against thyroid proteins are produced are largely unknown. EVIDENCE ACQUISITION: Published peer-reviewed basic and clinical literatures on immunology and autoimmune diseases were identified through searches of PubMed for articles published from January 1971 to May 2011. Articles resulting from these searches and relevant references cited in those articles were reviewed. All the relevant articles were written in English. EVIDENCE SYNTHESIS: Recent studies have indicated that innate immune responses induced by both exogenous and endogenous factors affect the phenotype and severity of autoimmune reactions. One of the recent topics is the effect of self-genomic DNA fragments on immune activation. Expression of major histocompatibility complex class II on the autoimmune target cells seems to play an important role in the presentation of endogenous antigens. Accumulated evidence from animal models has generated new insights into the pathogenesis of AITD. CONCLUSION: AITD develops by a combination of genetic susceptibility and environmental factors. Innate immune responses are associated with thyroid dysfunction, tissue destruction, and the likely development and perpetuation of AITD. In addition to the other factors, cell injury may contribute to the activation of innate immune response and the development of AITD.