Secretory granules in endocrine cells selectively store bioactive peptide hormones and amines, which are secreted in a regulated manner upon appropriate stimulation. In addition to bioactive substances, various proteins and lipids characteristic of secretory granules are likely recruited to a restricted space at the trans-Golgi Network (TGN), and the space then matures to the secretory granule. Although experimental findings so far have strongly suggested that aggregation- and receptor-mediated processes are essential for the formation of secretory granules, the putative link between these two processes remains to be clarified. Recently, secretogranin III (SgIII) has been identified as a specific binding protein for chromogranin A (CgA), a representative constituent of the core aggregate within secretory granules, and it was later revealed that SgIII can also bind to the cholesterol-rich membrane domain at the TGN. Based on its multifaceted binding properties, SgIII may act as a central player in the formation of cholesterol-rich membrane platforms. Upon these platforms, essential processes for secretory granule biogenesis coordinately occur; that is, selective recruitment of prohormones, processing and modifying of prohormones, and condensation of mature hormones as an aggregate. This review summarizes the findings and theoretical concepts on the issue to date and then focuses on the putative role of SgIII in secretory granule biogenesis in endocrine cells.

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Patients with unresectable parathyroid carcinoma develop severe hypercalcemia, bone fractures and renal failure, and become unresponsive to conventional treatments. It has been shown that successful induction of anti-parathyroid hormone (PTH) antibodies, using PTH peptide fragments for immunisation, normalized serum levels of calcium as well as improved clinical symptoms. Here, we report our experience of PTH immunization in a Japanese female suffering from refractory hypercalcemia and renal failure caused by unresectable metastatic parathyroid carcinoma. Upon immunization, there were apparent clinical responses including reduction of serum levels of Ca along with anti-PTH antibodies induction. Therefore, we concluded that PTH immunization was an effective treatment against hypercalcemia caused by metastatic parathyroid carcinomas that are unresponsive to conventional treatments.

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The association of the FTO gene polymorphism, rs9939609, with obesity was examined using the population of the Takahata study (n (M/F): 2,639 (1,168 / 1,470); age: 63.0 ± 10.2 years), a Japanese community-based study. The effects of lifestyle-related factors, including nutritional intake and physical activities, on the association were also examined. Body mass index (BMI) was significantly associated with the FTO gene polymorphism (p<0.001). A case-control association study of the FTO gene polymorphism with obesity using multiple logistic regression analysis showed a significant association of the genotype AA (odds ratio, 1.53 [95% confidential interval, 1.04-2.24]) after adjustment for age and gender. Analysis to examine the differences in lifestyle-related factors among the genotype groups showed a significant difference in the energy expenditure for moderate to high-intensity physical activity (PA) (≥ 3.0 METs) (p=0.012) with a significant decrease toward the genotype AA (p=0.027). The effect of energy expenditure for moderate to high-intensity PA on the association of the polymorphism with obesity was then examined using study groups stratified based on the energy expenditure for moderate to high-intensity PA (Low-PA and High-PA). The BMI was significantly higher in the genotype AA in the Low-PA group (p=0.016) but not in the High-PA group (p=0.103). Furthermore, the genotype AA was significantly associated with obesity (odds ratio, 2.39 [95% confidential interval, 1.19-4.80]) in the Low-PA group but not in the High- PA group (p=0.650). The FTO gene, rs9939609, was associated with obesity, and the association was evident in subjects with low-PA, suggesting a PA-dependent association.

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Oxidative stress has been implicated as a causal role in atherosclerosis, microvascular complications of diabetes as well as in beta cell failure in type 2 diabetes. PPARγ agonists not only improve insulin sensitivity but also eliminate oxidative stress. In mouse, catalase, a major antioxidant enzyme, is directly regulated by PPARγ through two PPARγ binding elements in its promoter. This study examined the regulatory mechanisms of catalase expression in human. Expression of catalase was significantly upregulated in human primary adipocytes upon treatment with a PPARγ agonist. However, the mouse PPARγ response elements are not functionally conserved in human catalase promoter. In luciferase reporter assay containing human catalase promoter, PPARγ /RXRα, in combination of a PPARγ agonist significantly transactivated 19 kb of promoter and this was mediated via a novel PPARγ response element (PPRE) at -12 kb from transcription initiation site of human catalase gene. Electrophoretic mobility shift assay showed direct binding of PPARγ to this PPRE. Together, our results indicate that PPARγ regulates the expression of catalase gene in human through a PPRE distinct from that of mouse, and could explain, at least in part, the observed inhibitory effects of PPARγ on oxidative stress in human.

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We examined the inhibitory effect of thyroid blocking antibody (TBAb) on the thyroid stimulating activity of human chorionic gonadotropin (HCG) and equine CG (ECG). Five TBAb positive sera obtained from patients who had been hypothyroid but were currently on T4 treatment. The TSH binding inhibitory immunoglobulin (TBII) activities of the sera were 60-160 IU/L. Inhibition of TSH binding to the TSH receptor (TSHR) [TSH binding inhibition (TBI) activity] of HCG or ECG, and inhibition of TBAb on HCG or ECG-stimulated cAMP production were examined. Both HCG and ECG preparations showed weak TBI activity in the presence of small amounts of protein [bovine serum albumin (BSA)] but were negative in the presence of large amounts of protein [normal human serum (NHS) or BSA]. Four thousand IU/mL of HCG and ECG preparation caused cAMP production similar to 100μU/mL of bovine (b) TSH. The inhibitory effect of TBAb on cAMP production by this amount of HCG or ECG was then examined. The inhibitory effect of TBAb on cAMP production by HCG and ECG was similar to bTSH, and TBAb positive sera with more than 40 IU/L TBII activity completely blocked cAMP production by HCG, ECG and bTSH. This suggests that common alpha -subunit of both HCG and TSH are involved in the inhibitory effect of TBAb. Previous reports demonstrated that the thyroid stimulating activity of thyroid stimulating antibody (TSAb) was blocked by deglycosyalted HCG (competitive antagonist of TSH binding to TSHR). The fact and our present study suggest that TSH, HCG ECG, TSAb and TBAb have a similar binding site (alpha-subunitmimicking binding site) on the TSH receptor.

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Both glucocorticoid and insulin are known to have an anabolic effect on lipogenesis. Acetyl-CoA, an intermediate product of glycolysis, is supplied for fatty acid synthesis when carbohydrate intake is sufficient. Acetyl-CoA carboxylase (ACC), consisting of two isoenzymes ACC1 and ACC2, mediates the conversion from acetyl-CoA to malonyl-CoA, and thus plays a key role for the regulation of lipogenesis. In this study, we surveyed the effects of glucocorticoid and insulin on the transcriptional activity of the alternative promoters of ACCs (PI-PIII for ACC1, and PI and PII for ACC2) using the HepG2 human hepatocyte cell line in vitro. We also examined the roles of the insulin and/or glucose-regulated transcriptional factor(s) such as SREBP1c, LXRα/β, and ChREBP on each promoter of the ACC genes. We found that both insulin and glucocorticoid had potent positive effects on all the promoters examined, and additive effects of both hormones were recognized in ACC1 PI and ACC2 PI. Furthermore, a representative insulin-responsive transcription factor SREBP1c showed significant stimulatory effects on all the promoters of ACC genes, among which those on ACC1 PIII and ACC2 PI were most prominent. On the other hand, the effect of LXRα was rather selective; it showed a marked stimulatory effect only on ACC1 PII. LXRβ and ChREBP had minimal, if any, effects on some of the promoters. Altogether, our data suggest that insulin and glucocorticoid have positive effects on both ACC1 and ACC2 gene transcription. SREBP1c might be a master regulator of the expression of both genes regardless of the promoter utilized, whereas LXRα seems to play a promoter-specific role. Since ACC1 facilitates lipogenesis by stimulating fatty acid synthesis and ACC2 inhibits lipolysis, both insulin and glucocorticoid seem to play an important role in the pathogenesis of obesity and/or hepatic steatosis.

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Hypoglycemia is reported to be one of the manifestations of a patient with hypothalamic sarcoid infiltrates due to impaired counter-regulation of glucose. But, without hypothalamic lesion, patients with sarcoidosis would not be expected to have hypoglycemia. We recently identified a patient with an isolated sarcoidosis of the spleen who had experienced frequent fasting hypoglycemia which completely disappeared after splenectomy. During hypoglycemia, serum insulin was undetectable. Endocrinological examination revealed no abnormality. The objective was to investigate whether the patient’s hypoglycemia was due to ectopic secretion of an insulin-mimetic factor by the splenic sarcoidosis. Serum insulin-like growth factor-I (IGF-I) and IGF-II were measured by RIA. Serum visfatin and free IGF-I were by ELISA. A high molecular weight form of IGF-II, termed “big” IGF-II, was identified by Western blotting. Tissue IGF-I was quantified by real time RT-PCR after RNA extraction. Before operation, total and free serum IGF-I, serum IGF-II and serum visfatin were within reference range. Big IGF-II was not detected in patient’s serum extract. After operation, hypoglycemia did not recur and serum insulin returned to normal, while serum IGF-I decreased by half the preoperative level. RT-PCR revealed that mRNA level of IGF-I in the sarcoidosis tissue was about 1.8-fold greater than that in the normal spleen tissue. These data suggest that ectopic secretion of IGF-I by the splenic sarcoidosis and its direct access to the liver via the portal vein might cause fasting hypoglycemia mainly by suppressing hepatic gluconeogenesis.

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As a screening test for Cushing’s syndrome, the evaluation of late-night cortisol levels is indispensable. We evaluated the usefulness and accuracy of plasma, urinary, and salivary cortisol levels measured late at night for the diagnosis of Cushing’s syndrome. High cortisol levels (> 5 μg/dL) during the night are indicative of Cushing’s syndrome, although night plasma cortisol levels are not readily reproducible because of the stressful situation. There was no correlation between plasma and urinary cortisol levels late at night, and late-night urinary cortisol levels provided weak information for the diagnosis of Cushing’s syndrome. By contrast, late-night plasma and salivary cortisol levels showed a positive correlation, and salivary cortisol sampling was found to be useful for the diagnosis of Cushing’s syndrome, because more than 0.4 μg/dL of late-night salivary cortisol levels gave a sensitivity of 86% and a specificity of 100% in our hospital. This method is also useful for the diagnosis of early or mild stage Cushing’s syndrome, so-called subclinical Cushing’s syndrome. Inherent differences between assays make it difficult to define optimal diagnostic criteria. However, the relative levels of salivary cortisol ratio, which is presented as a relative level, compared with the mean levels of healthy subjects in each institute, is useful for the screening of Cushing’s syndrome as the cut-off level of 1.5 shows both high sensitivity and specificity in subclinical and overt Cushing’s syndrome. Late-night salivary cortisol measurement is therefore a primary method of choice in the screening of patients suspected of having Cushing’s syndrome.

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The effect of stress associated with acute weight reduction on adipocytokine production is incompletely understood. In the present study, we have investigated the changes in circulating adipocytokine concentrations and urinary concentrations of stress markers in male collegiate wrestlers during acute weight reduction for a competition. Twenty healthy Japanese male wrestlers (18-22 years of age) who participated in the national collegiate wrestling tournament were studied. Body weight, body fat amount, serum testosterone, serum leptin, serum adiponectin, urinary 8-hydroxy-2’- deoxyguanosine (8-OHdG) and urinary biopyrrins were analyzed during acute weight reduction for the competition. Body weight, body fat amount and the serum concentrations of testosterone, leptin and adiponectin significantly decreased on the day of weigh-in compared with the levels 12 days before weigh-in. In contrast, urinary concentrations of 8-OHdG and biopyrrins significantly increased on the day of weigh-in compared with the concentrations 12 days before weigh-in. A positive correlation was observed between the serum concentrations of adiponectin and testosterone, and a negative correlation was observed between the concentrations of serum adiponectin and urinary biopyrrins. The present results suggest that rapid weight reduction increases the urinary concentrations of stress markers, which is associated with a decrease in serum concentrations of adiponectin.

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A 19-year-old girl presented at our emergency room with hypokalemic periodic paralysis. She had a thyrotoxic goiter and had experienced three paralytic attacks during the previous 2 years on occasions when she stopped taking antithyroid drugs. In addition to thyrotoxic periodic paralysis (TPP), she had metabolic acidosis, urinary potassium loss, polyuria and polydipsia. Her reduced ability to acidify urine during spontaneous metabolic acidosis was confirmed by detection of coexisting distal renal tubular acidosis (RTA). The polyuria and polydipsia were caused by nephrogenic diabetes insipidus, which was diagnosed using the water deprivation test and vasopressin administration. Her recurrent and frequent paralytic attacks may have been the combined effects of thyrotoxicosis and RTA. Although the paralytic attack did not recur after improving the thyroid function, mild acidosis and nephrogenic DI have been remained subsequently. Patients with TPP, especially females with atypical metabolic features, should be investigated for possible precipitating factors.

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Recently, mutations in nuclear genes encoding two mitochondrial complex II subunit proteins, Succinate dehydrogenase D (SDHD) and SDHB, have been found to be associated with the development of familial pheochromocytomas and paragangliomas (hereditary pheochromocytoma/paraganglioma syndrome: HPPS). Growing evidence suggests that the mutation of SDHB is highly associated with abdominal paraganglioma and the following distant metastasis (malignant paraganglioma). In the present study, we used multiplex ligation dependent probe amplification (MLPA) analysis to identify a large heterozygous SDHB gene deletion encompassing sequences corresponding to the promoter region, in addition to exon 1 and exon 2 malignant paraganglioma patient in whom previously characterized SDHB mutations were undetectable. This is the first Japanese case report of malignant paraganglioma, with a large SDHB deletions. Our present findings strongly support the notion that large deletions in the SDHB gene should be considered in patients lacking characterized SDHB mutations.

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Table 3 contains a typographical error.

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