Loss of µ-crystallin causes PPARγ activation and obesity in high-fat diet-fed mice.

The thyroid hormone-binding protein µ-crystallin (CRYM) mediates thyroid hormone action by sequestering triiodothyronine in the cytoplasm and regulating the intracellular concentration of thyroid hormone. As thyroid hormone action is closely associated with glycolipid metabolism, it has been proposed that CRYM may contribute to this process by reserving or releasing triiodothyronine in the cytoplasm. We aimed to clarify the relationship between CRYM and glycolipid metabolism by comparing wild-type and CRYM knockout mice fed a high-fat diet. Each group was provided a high-fat diet for 10 weeks, and then their body weight and fasting blood glucose levels were measured. Although no difference in body weight was observed between the two groups with normal diet, the treatment with a high-fat diet was found to induce obesity in the knockout mice. The knockout group displayed increased dietary intake, white adipose tissue, fat cell hypertrophy, and hyperglycemia in the intraperitoneal glucose tolerance test. In CRYM knockout mice, liver fat deposits were more pronounced than in the control group. Enhanced levels of PPARγ, which is known to cause fatty liver, and ACC1, which is a target gene for thyroid hormone and is involved in the fat synthesis, were also detected in the livers of CRYM knockout mice. These observations suggest that CRYM deficiency leads to obesity and lipogenesis, possibly in part through increasing the food intake of mice fed a high-fat diet.

Implications of thyroid autoimmunity in infertile women with subclinical hypothyroidism in the absence of both goiter and anti-thyroid antibodies: lessons from three cases.

There is a great deal of research interest regarding the underlying causes of slightly elevated TSH values in patients with subclinical hypothyroidism (SH) without abnormal findings on ultrasonography or anti-thyroid antibodies. Twelve infertile women with thyroglobulin antibody (TGAb) and thyroid peroxidase antibody (TPOAb)-negative nongoitrous SH were referred to our department of endocrinology between September 2007 and September 2015. None had been diagnosed with autoimmune thyroid disease or had any possible causes of SH. In all cases, LT4 was prescribed to bring TSH value below 2.5 mIU/L. Among those with infertility treatments, six (50%) became pregnant and gave birth to infants. Here, we report three of these six women who successfully became pregnant with infertility treatments and were found to have thyroid autoimmunity on data obtained during the postpartum period. Two developed postpartum thyroiditis, and the remaining one woman was temporarily weakly positive for TPOAb at 9 months postpartum. We describe three infertile subclinically hypothyroid women without goiter or anti-thyroid antibodies with potential thyroid autoimmunity. Thyroid autoimmunity is one of the most important issues for management of pregnant women, and thus, our findings are noteworthy for the care of infertile women with SH. This report provides valuable insights into the presence of autoimmunity in nongoitrous thyroid-associated antibody-negative SH patients.

Cytosolic T3-binding protein modulates dynamic alteration of T3-mediated gene expression in cells.

µ-Crystallin (CRYM) is also known as NADPH-dependent cytosolic T3-binding protein. A study using CRYM-null mice suggested that CRYM stores triiodothyronine (T3) in tissues. We previously established CRYM-expressing cells derived from parental GH3 cells. To examine the precise regulation of T3-responsive genes in the presence of CRYM, we evaluated serial alterations of T3-responsive gene expression by changing pericellular T3 concentrations in the media. We estimated the constitutive expression of three T3-responsive genes, growth hormone (GH), deiodinase 1 (Dio1), and deiodinase 2 (Dio2), in two cell lines. Subsequently, we measured the responsiveness of these three genes at 4, 8, 16, and 24 h after adding various concentrations of T3. We also estimated the levels of these mRNAs 24 and 48 h after removing T3. The levels of constitutive expression of GH and Dio1 were low and high in C8 cells, respectively, while Dio2 expression was not significantly different between GH3 and C8 cells. When treated with T3, Dio2 expression was significantly enhanced in C8 cells, while there were no differences in GH or Dio1 expression between GH3 and C8 cell lines. In contrast, removal of T3 retained the mRNA expression of GH and Dio2 in C8 cells. These results suggest that CRYM expression increases and sustains the T3 responsiveness of genes in cells, especially with alteration of the pericellular T3 concentration. The heterogeneity of T3-related gene expression is dependent on cellular CRYM expression in cases of dynamic changes in pericellular T3 concentration.

A new experimental model of ATP-sensitive K⁺ channel-independent insulinotropic action of glucose: a permissive role of cAMP for triggering of insulin release from rat pancreatic ß-cells.

In pancreatic ß-cells, glucose metabolism leads to closure of ATP sensitive K⁺ channels (K(ATP) channel) and Ca²âº influx, which is regarded as a required step for triggering of insulin release. Here, we demonstrate that glucose triggers rapid insulin release independent from its action on K(ATP) channels given the cellular cAMP is elevated. We measured insulin release from rat pancreatic islets by static and perifusion experiments. Changes in cytosolic free Ca²âº concentration ([Ca²âº]i) were monitored using fura-2 loaded rat pancreatic ß-cells. Glucose-induced insulin release was abolished when Ca²âº influx was inhibited by a combination of 250 µM diazoxide, an opener of K(ATP) channel, and 10 µM nifedipine, a blocker of L-type voltage-dependent Ca²âº channels. However, with both nifedipine and diazoxide, glucose induced a 5-fold increase in insulin release in the presence of 10 µM forskolin, an activator of adenylyl cyclase. In the presence of diazoxide, nifedipine, and forskolin, 22 mM glucose sharply increased the rate of insulin release within 2 min which peaked at 6 min: this was followed by a further gradual increase in insulin release. In contrast, it lowered [Ca(2+)]i with a nadir at 2-3 min followed by a gradual increase in [Ca²âº]i. The glucose effect was mimicked by 20 mM α-ketoisocaproic acid, a mitochondrial fuel, and it was nullified by 2 mM sodium azide, an inhibitor of mitochondrial electron transport. Cerulenin, an inhibitor of protein acylation, decreased the glucose effect. In conclusion, a rise in [Ca²âº]i through voltage-dependent Ca²âº channels is not mandatory for glucose-induced triggering of insulin release.

Humoral hypercalcemia associated with gastric carcinoma secreting parathyroid hormone: a case report and review of the literature.

Hypercalcemia with concomitant elevation of serum parathyroid hormone (PTH) and PTH-related protein (PTHrP) levels was found in a patient with advanced gastric carcinoma and multiple liver metastases. The most common features are hypercalcemia associated with hypersecretion of PTHrP and physiological suppression of PTH secretion in the syndrome of humoral hypercalcemia of malignancy (HHM). Although we initially made a diagnosis of primary hyperparathyroidism concomitant with HHM due to gastric cancer, diagnostic imaging studies, such as echography, CT, sestamibi scintigraphy, and autopsy findings, did not reveal evidence of any parathyroid tumors or ectopic parathyroid glands in the mediastinum. Both primary and metastatic tumor cells showed positive staining with PTH-specific antibody as well as PTHrP-specific antibody on immunohistochemical examination. PTH concentration in the cytosolic fraction of the metastatic tumor was elevated compared to that from a control patient with no calcium metabolic disorders in vitro. These findings indicated that PTH secreted ectopically by gastric cancer cells, not by parathyroid glands, caused hypercalcemia in this patient. To our knowledge, this is the first case report of PTH-secreting gastric carcinoma cells. We report the case and a review of the previous reported PTH-secreting non-parathyroid tumors along with the mechanisms of secretion.

Glucose-incretin interaction revisited.

Pancreatic beta cell dysfunction is pivotal to the development of diabetes, and restoration of insulin action is of primary importance. Here, we present a review of the mechanism of insulin secretion by pancreatic beta cells and discuss the mutual interaction of signaling pathways in stimulus-secretion coupling to better understand the scientific basis of pharmacological treatment for insulin secretion deficiency. Glucose stimulates insulin secretion via membrane depolarization by closure of ATP-sensitive K(+) channels (K(ATP) channels) and opening of L-type voltage-dependent Ca(2+) channels. The resultant elevation of cytosolic free Ca(2+) triggers insulin exocytosis. This is termed the "K(ATP)-dependent pathway" and is shared by sulfonylurea, which closes K(ATP) channels. Glucose also stimulates insulin release independent of its action on K(ATP) channels. This is referred to as the "K(ATP)-independent pathway," the molecular basis of which remains elusive. In the pancreatic beta cell, incretin hormones increase cAMP level, which enhances glucose-stimulated insulin release by protein kinase A-dependent and -independent mechanisms. Importantly, cAMP does not directly augment Ca(2+)-stimulated insulin release per se. The stimulatory level of ambient glucose is an absolute requirement for incretin to enhance insulin release. Therefore, incretin/cAMP enhances K(ATP)-independent insulinotropic action of glucose. The robust glucose-lowering effect of DPP4 inhibitor add-on in diabetic patients with sulfonylurea secondary failure is intriguing. With the clinical availability of DPP4 inhibitor and GLP-1 mimetics, the importance of the interactions between cAMP signaling and K(ATP) channel-independent actions of glucose is reappraised.

Fetal goitrous hypothyroidism due to maternal thyroid stimulation-blocking antibody: a case report.

Most fetal goitrous hypothyroidisms are reportedly caused by the maternal use of an antithyroid drug or fetal dyshormonogenesis. However, fetal goitrous hypothyroidism due to the transplacental passage of maternal thyroid stimulation-blocking antibody (TSBAb) is extremely rare. A woman at 28 weeks of gestation was found to have a fetal goiter by ultrasonography. Because the maternal serum showed hypothyroidism with an elevated titer of TSBAb, levothyroxine sodium was administered. The patient delivered a male infant, 3,412 g, with a goiter at term. Umbilical blood revealed primary hypothyroidism with increased TSBAb, and the infant was given levothyroxine sodium. After a month, neonatal thyroid function and TSBAb levels became normal. Attention should be paid to possible fetal hypothyroidism when a fetal goiter is observed to avoid impaired mental development of the neonate.

OTX5 regulates pineal expression of the zebrafish REV-ERB alpha through a new DNA binding site.

The pineal gland plays a central role in the photoneuroendocrine system and acts as a photosensory organ in lower vertebrates. The orphan nuclear receptor Rev-erbalpha (NR1D1) has previously been shown to be expressed in the pineal and to be regulated with a robust circadian rhythm during zebrafish embryogenesis. This early pineal expression is under the control of the transcription factor Orthodenticle homeobox 5 (Otx5). In this paper, we show that Otx5 regulates the second zfRev-erbalpha promoter, ZfP2. Despite the absence of a classical Otx-binding site within ZfP2, this regulation depends on the integrity of the Otx5 homeodomain. Mapping experiments as well as EMSAs show that this interaction between Otx5 and ZfP2 depends on a noncanonical bipartite Otx-binding site (GANNCTTA and TAAA) that we called pineal expression related element (PERE). We showed that PERE is necessary for pineal expression in vivo by injecting zebrafish embryos with wild type and mutated versions of zfRev-erbalpha promoter fused to green fluorescent protein. Interestingly, PERE is found upstream of other genes expressed in the pineal gland, suggesting that it may play an important role in governing pineal expression. Our data establish that PERE is a novel cis-acting element contributing to pineal-specific gene expression and to Otx target gene regulation.

Repression of insulin gene transcription by indirect genomic signaling via the estrogen receptor in pancreatic beta cells.

The mechanism whereby 17ß-estradiol (E2) mediates insulin gene transcription has not been fully elucidated. In this study, exposure of hamster insulinoma (HIT-T15) cells to 5 × 10-9 to 1 × 10-7 M E2 led to a concentration-dependent decrease of insulin mRNA levels. Transient expression of the estrogen receptor (ER) in HIT-T15 cells revealed that estrogen receptor α (ERα) repressed transcription of the rat insulin II promoter in both ligand-dependent and ligand-independent manners. The N-terminal A/B domain of ERα was not required for either activity. However, the repression was absent with mutated ER lacking the DNA-binding domain. Moreover, introducing mutations in the D-box and P-box of the zinc finger of ER (C227S, C202L) also abolished the repression. Deletion of the insulin promoter region revealed that nucleotide positions - 238 to - 144 (relative to the transcriptional start site) were needed for ER repression of the rat insulin II gene. PDX1- and BETA2-binding sites were required for the repression, but an estrogen response element-like sequence or an AP1 site in the promoter was not involved. In conclusion, we found that estrogen repressed insulin mRNA expression in a beta cell line. In addition, the ER suppressed insulin gene transcription in a ligand-independent matter. These observations suggest ER may regulate insulin transcription by indirect genomic signaling.

ACTH-producing thymic neuroendocrine tumor initially presenting as psychosis: A case report and literature review.

A 32-year-old woman was referred to our hospital because of severe psychosis and was found to have an ectopic ACTH-producing thymic neuroendocrine tumor. Laboratory data revealed an elevated serum cortisol and plasma ACTH level, hypokalemia, and metabolic alkalosis. Chest computed tomography (CT) revealed an anterior mediastinal mass and multiple pulmonary nodules. As the patient was unable to communicate because of her consciousness disturbance, she was managed with artificial ventilation and deep sedation. Metyrapone and potassium supplementation were administered, and steroid psychosis gradually improved. Thoracic surgery was performed and the histopathological diagnosis was thymic neuroendocrine tumor with positive anti-ACTH immunohistochemical staining. Here we present details of the case and review the literature.