KRN568 is a calcimimetic compound which acts on the calcium sensing receptors (CaR) on the parathyroid gland to suppress secretion of PTH. A recent report has demonstrated that CaRs are expressed on cultured human antral gastrin cells and that gastrin secretion is stimulated by an increase in extracellular calcium level. However, the effect of KRN568 on serum gastrin levels has yet to be clinically assessed. We therefore studied the effect of this calcimimetic on gastrin secretion in healthy subjects enrolled in the phase 1 study for KRN568 currently male volunteers at fasting and after meal. One subject proved to be a poor metabolizer (PM) for this compound and showed more than 10-fold high concentrations of plasma KRN568 (fasting Cmax 90.8 and non-fasting 83.8ng/ml) compared to the other 5 individuals (Cmax 6.5±2.2 and 7.4±1.6ng/ml, respectively). Plasma gastrin levels showed mild but apparent increase (from 30 to 125pg/ml) in this particular subject, while there were no significant increases in the other five people (from 34±6 to 63±3pg/ml) after oral administration of 400mg KRN568 at fasting. In the PM, administration of KRN568 resulted in extraordinarily high serum drug levels associated with transient increase of gastrin levels. This observation suggested that calcium-induced stimulation of gastrin secretion in human was mediated by a mechanism involving CaR. Potential side effects related to the increased gastrin secretion may be warranted in the practical use of this compound.
The present study has been designed to quantify and compare right and left carotid intima-media thicknesses (IMT) in type 2 diabetics and healthy controls. It was also intended to investigate the effects of various risk factors on the carotid IMT in these subjects. A total of 122 subjects; 70 patients with type 2 diabetes and 52 non-diabetic subjects as controls, were recruited for the study. Right and left common carotid artery stiffness indices were assessed with ultrasonography in both groups. Age, body mass index (BMI), duration of diabetes, cigarette smoking, lipid profile including lipoprotein a, Chlamydia pneumonia seropositivity, glycemic indices, fasting insulin levels, serum fibrinogen levels and presence of hypertension, coronary artery disease, degenerative complications of diabetes mellitus were all assessed in order to define their role as determinants of carotid artery IMT. The difference between the groups regarding mean carotid IMT was statistically significant for the left carotid arteries (p=0.028) and borderline significance was found for the right carotid arteries (p=0.055). Age has a very strong association with carotid IMT in diabetic patients (p<0.0001) with univariate analysis. According to the results of multivariate analysis, age and BMI were found to be the most important independent determinants of carotid IMT for both sides. When age was excluded from the model, BMI and coronary artery disease were found to have strong association with IMT on the right (p=0.0036 and 0.0249) and BMI was the only significant determinant for the left side (p=0.0025). This study shows that carotid IMT is greater in diabetic subjects compared with healthy controls. For the diabetic subjects, age, BMI and presence of coronary heart disease have a strong influence on the atherosclerotic process of the carotid arteries.
The correlation between various factors obtained during oral glucose tolerance tests (OGTT) and insulin sensitivity tests (steady state plasma glucose, SSPG method) were analyzed in non-diabetic subjects by Pearson's simple correlation. Similar data were obtained for normal as well as normal and impaired glucose tolerance (IGT) subjects. Three factors (AUCI1200AUCG1200, AUCI1200 and AUCI1200, area of under curve (AUC), insulin (I), glucose (G)) were found to have significant correlation (r=0.47-0.54) with SSPG, thus indicating insulin resistance. Those with hyperinsulinemia were noted to have insulin resistance with high precision (98%), but insulin resistant subjects without hyperinsulinemia were all judged to have normal sensitivity. These three factors all had high specificity (98%), but low sensitivity (21-25%) with false negative results being obtained in 75-78%, and false positive results in less than 2%. It was difficult to distinguish positive and negative for the insulin resistance by receiver operating characteristic (ROC) curve of I0G0. The sensitivity of which was 12.8% and specificity 93.8%. By ROC curve, the specificities and sensitivities for the proposed factors, including homeostatic model assessment (HOMA) product (I0G0, I0-120, G0-120, insulin and glucose at 0-120' during OGTT) ranged from 60 to 70%, which is far from an acceptable level of 90% or more. Therefore, these factors were useful to correlate insulin sensitivity with OGTT results in hyperinsulinemic subjects or in general non-diabetic subjects, but less useful when it came to identifying individual cases with insulin resistance. These parameters should be used only in non-diabetic subjects and their significance is thought to be restricted to cases with hyperinsulinemia.
The human ACTH receptor (ACTH-R) gene consists of two exons; the first encodes the 5'-untranslated region (UTR) and the second encodes part of the 5'-UTR, the entire coding region, and the whole of the 3'-UTR. While investigating the transcription initiation site of the human ACTH-R gene by 5'-rapid amplification of cDNA ends, we found a 157-bp alternate exon in the 5'-UTR. This newly identified exon was irrelevant to the alternate exon in the mouse ACTH-R gene which we reported previously.
Whether GH secretion in women varies over the menstrual cycle is uncertain. Previous investigations have led to conflicting conclusions; some studies suggested that there is an estrogen mediated rise in GH during the periovulatory (PO) and luteal (L) phases whereas others indicated no change in GH axis over the cycle. Differences in conclusions could relate to heterogeneity of the study populations, GH sampling paradigms or sensitivity of the GH assays used. In order to investigate whether GH secretion varied over the cycle, 24-h GH profiles using every 10-min sampling were obtained in 6 ovulatory women during the early follicular (EF), PO and L phases of the cycle. The TSH response to TRH, GH response to GRH and fasting plasma IGF-I were measured on each occasion. There was a trend toward higher integrated GH concentration (IGHC) during the PO phase, although this difference was not statistically significant (3284±721 vs 4542±872 vs 4071 ±699μg/min/L; EF vs PO vs L; p=0.09). Similarly, deconvolution estimated GH secretion did not vary over the cycle (p=0.56). There were no differences in GH pulse amplitude or frequency. There were no correlations between IGHC and sex steroids. Serum IGF-I was constant over the cycle (272±38 vs 277±31 vs 265±38μg/L; p=0.89). The TSH response to TRH and GH response to GRH did not vary over the cycle. We concluded that the effect of changes in the ovarian steroid milieu on the GH axis during spontaneous menstrual cycles is minimal.
In this study, we analyzed plasma lipid and lipoprotein levels before and after treatment with 1-desamino-8-D-arginine vasopressin (DDAVP) in subjects with partial and complete central diabetes insipidus (DI) in order to determine how a shortage and supplement of this hormone affect plasma lipid metabolism. The subjects consisted of 6 patients with partial and 6 with complete central DI. After treatment with DDAVP through nasal cavity, plasma total cholesterol (TC) level did not decrease either in complete or partial form. Plasma triglyceride (TG) levels decreased from 306±175mg/dl to 198±91(35% decrease, p=0.027) in complete form, while TG did not change significantly in partial form. A detailed investigation of plasma lipoprotein metabolism during treatment with DDAVP was carried out in 3 of the 6 subjects with complete form of DI. Lipoprotein lipase activity and mass in post-heparin plasma from those three subjects tended to increase after treatment with DDAVP, along with the complete disappearance of an unusual lipoprotein between low density lipoprotein (LDL) and very low density lipoprotein (VLDL) as analyzed by polyacrylamide gel electrophoresis. These results suggest that the DDAVP treatment has a favorable effect on lipid and lipoprotein metabolism, especially triglyceride-rich lipoproteins, either directly or through modifying factors contributing to lipid metabolism.In this study, we analyzed plasma lipid and lipoprotein levels before and after treatment with 1-desamino-8-D-arginine vasopressin (DDAVP) in subjects with partial and complete central diabetes insipidus (DI) in order to determine how a shortage and supplement of this hormone affect plasma lipid metabolism. The subjects consisted of 6 patients with partial and 6 with complete central DI. After treatment with DDAVP through nasal cavity, plasma total cholesterol (TC) level did not decrease either in complete or partial form. Plasma triglyceride (TG) levels decreased from 306±175mg/dl to 198±91(35% decrease, p=0.027) in complete form, while TG did not change significantly in partial form. A detailed investigation of plasma lipoprotein metabolism during treatment with DDAVP was carried out in 3 of the 6 subjects with complete form of DI. Lipoprotein lipase activity and mass in post-heparin plasma from those three subjects tended to increase after treatment with DDAVP, along with the complete disappearance of an unusual lipoprotein between low density lipoprotein (LDL) and very low density lipoprotein (VLDL) as analyzed by polyacrylamide gel electrophoresis. These results suggest that the DDAVP treatment has a favorable effect on lipid and lipoprotein metabolism, especially triglyceride-rich lipoproteins, either directly or through modifying factors contributing to lipid metabolism.
Circulating human growth hormone (GH) consists of several molecular isoforms. Increased proportion of circulating non-22K hGH and 20K hGH was reported in active acromegaly. In this study, we studied the release of 20K and 22K hGH from cultured GH-producing human pituitary adenoma cells in vitro. Pituitary adenoma cells obtained from 6 acromegalic patients were cultured and submitted to perifusion experiments. Concentrations of 20K and 22K hGH in the serum and the perifusion effluent were determined by specific enzyme-linked immunosorbent assays recently developed. The %20K value varied in a wide range from 3.58 to 8.72% in vitro and was lower than in the serum (mean±SD: 6.57±1.88% vs 9.08±2.12%, P<0.05). There was no correlation between the %20K values in vitro and in vivo (r=0.31, P>0.05). The in vitro secretions of 20K and 22K hGH were in parallel and strongly correlated (r=0.953, P<0.001). These findings suggest that different GH-producing pituitary adenoma cells secrete 20K hGH in variable amounts and that the proportion of 20K hGH in the serum might be affected by metabolic clearance of hGH isoforms. It was also suggested that 20K and 22K hGH might be secreted in toto from GH-producing human pituitary adenoma cells.
The high frequency of cutaneous manifestations in patients with multiple endocrine neoplasia type 1 (MEN 1) has recently been reported. Since prevalence of some cutaneous diseases varies among different ethnic groups, we examined the frequency of facial angiofibromas in Japanese patients with familial MEN 1. Among 27 patients with germline MENI gene mutation and one asymptomatic gene carrier, angiofibromas were identified in 43% (12/28) of the subjects. This frequency was significantly lower than that of Caucasian patients, but nonetheless almost equaled those of pituitary tumors and pancreas endocrine tumors. Angiofibromas should be considered as one of major manifestations in MEN 1 regardless of patients' ethnic origin, and clinicians should pay careful attention to the cutaneous lesions in patients with endocrine tumors.
Subclinical thyroiditis or thyroid dysfunction is relatively common in the elderly. To estimate the effectiveness of measurement of serum levels of anti-thyroglobulin and anti-microsomal or thyroid peroxidase antibodies for detecting focal lymphocytic thyroiditis (FLT) in the elderly, we examined the relationships between antibody titer and postmortem histological finding of the thyroid gland in 180 consecutive autopsies (69 women and 111 men) over 60years of age without any overt clinical thyroid or collagen diseases. FLT was found in 25 cases (13.9%) with female predominance (21.7% in female vs. 9.0% in male). Measurements of serum levels of anti-thyroglobulin and anti-thyroid peroxidase antibodies by radioimmunoassay (TgAb and TPOAb, respectively) were compared with the measurements of anti-thyroglobulin and anti-microsomal antibodies by a hemagglutination technique (TGHA and MCHA, respectively), using sera from 25 patients with FLT and age- and sex-matched 51 patients without FLT. Among 25 cases with FLT, TgAb and TPOAb were positive in 17 (68%) and 12 (48%), respectively. There was a close relationship between degree of FLT and serum level of TgAb or TPOAb (P<0.0001). On the other hand, TGHA and MCHA were positive only in 8 (32%) and 10 (40%), respectively. TgAb and TPOAb were more sensitive than TGHA (68% vs. 32%, P<0.05) and MCHA (48% vs. 40%) to detect FLT. Positive findings in either TgAb or TPOAb significantly improved sensitivity (76%) compared with that of TGHA or MCHA (44%) (P<0.05). Specificities of combined measurements of TgAb and TPOAb (90%) were not significantly different from those of TGHA and MCHA (100%). These findings indicate that TgAb is a more sensitive method for detecting FLT and that its diagnostic sensitivity for FLT increases by using it in combination with TPOAb. Therefore, in the elderly without clinically or biochemically overt thyroid dysfunction, positive TgAb and/or TPOAb could imply presence of FLT, and their titers might reflect degree of inflammation.
Human chorionic gonadotropin (hCG) is a member of a family of heterodimeric glycoprotein hormones that contain a common α-subunit but differ in their hormone-specific β-subunits. Both subunits have five and six disulfide bonds, respectively, which consist of cystine knot structure. It is evident from numerous studies that the structure of β-subunits is rigid, whereas that of α-subunit is flexible and can be molded by a β-subunit. Previously, we reported that secreted forms of α mutants where either cysteine residue in the disulfide bond 7-31 or 59-87 was converted to alanine contained a disulfide-linked homodimer in addition to a monomer. To study whether the hCGβ-subunit affects the conformations of α mutants, α-subunits lacking either the 7-31 or 59-87 disulfide bond were expressed with wild-type (WT) hCGβ in Chinese hamster ovary cells, and homodimer formation and glycosylation of dimerized α-subunit were assessed by continuous labeling with [35S]methionine/cysteine, immunoprecipitation with anti-α or -hCGβ serum, digestion with endoglycosidase-H or -F, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in a non-reducing condition. Our data showed that α homodimer was not observed in the half-Cys mutants except one, where cysteine at position 7 was converted to alanine, in the presence of β-subunit. This finding indicated that hCGβ-subunit rescued the α half-Cys mutants from the formation of intermolecular disulfide-linked homodimer by preferentially combining with the α mutants. In both free WT and all mutants treated with endoglycosidase-H, no or faint bands were recognized as the same migration as seen in endoglycosidase-F treatment. Even in the endoglycosidase-H sensitive cases, the amount of sensitive α-subunits was less than 5% of total α-subunits. In contrast to free α-subunits, distinct endoglycosidase-H sensitive bands were seen in both WT and mutants, although the ratio was various. We concluded that hCGβ-subunit affects the folding and glycosylation of the α-subunit mutants.
The management of occult small thyroid nodules found by ultrasonography is sometimes problematic. Self-palpation may be utilized in the follow-up of these nodules. We developed a method of thyroid self-examination (TSE). We examined the size of the nodules palpable by this method by ultrasonography. In 78 patients examined, all 38 tumors more than 2cm in size, whereas only one of 14 tumors less than or equal to 1cm in size, were palpable. These results suggest the clinical usefulness of TSE in the follow-up of thyroid tumors, which are very common and slow-growing.
Familial male-limited precocious puberty (MPP) is described in a 10year old patient with typical symptoms of the disease. Sequence analysis of genomic DNA clearly demonstrated a heterozygous T1193C transition in exon 11 of the LH receptor (LHR) gene, which results in M398T substitution in the second transmembrane helix of the protein product of this gene. The same mutation was found in the patient's mother and in her brother. The grandmother and the relatives of the patient's father were free of the mutation. The boy was successfully treated with inhibitors of steroid biosynthesis and androgen antagonists. It is suggested that this mutation caused constitutive activation of the LHR, which results in excessive formation of androgens in Leydig cells and is responsible for the symptoms of precocious puberty in this patient. This is the second case of the familial form of MPP that was maternally inherited.
To clarify the role of nitric oxide (NO) in the ontogeny of arginine vasopressin (AVP) and oxytocin (OXT) neurons in the hypothalamo-neurohypophysial system (HNS), we observed the coexpression pattern of NADPH-diaphorase (NADPH-d) activity and AVP- or OXT-immunoreactivity (IR) in the rat hypothalamus and posterior pituitary during the postnatal period. The enzymatic activity of NADPH-d was observed in the supraoptic nucleus (SON), paraventricular nucleus (PVN), median eminence (ME) and posterior pituitary throughout the postnatal development. AVP-containing neurons were clearly observed from postnatal day 1 in both the SON and PVN, while OXT-containing neurons were recognized from postnatal day 14. The coexistence of NADPH-d and AVP or OXT was detected in the SON from postnatal day 14. At postnatal day 21, the coexpression pattern was approximately the same as that of the SON and PVN in adult rats. Our findings indicated that the expression of NADPH-d and OXT was observed from almost the same postnatal period in both the SON and PVN. In addition, the pattern of increased numbers of NADPH-d positive fibers was similar to that of OXT-immunoreactive fibers in both the inner layer of the ME and the posterior pituitary. A good correlation was thus obtained between OXT expression and NADPH-d activity in the HNS during postnatal development. The present study suggests that NO is more closely involved in the expression and regulation of secretion of OXT than AVP.
It remains controversial whether or not a correlation exists between serum leptin levels and insulin resistance, and, if such a correlation does exist, whether it is independent of adiposity. To investigate the possible existence of an independent correlation, we have assessed serum leptin levels and insulin resistance in Japanese diabetic and non-diabetic subjects by means of Homeostatic Model Assessment (HOMA-R). Sixty-four Japanese patients with Type 2 diabetes mellitus (DM) (33 men and 31 women) and 53 sex-, age-, and body mass index (BMI)-matched non-diabetic adults (29 men and 24 women) were enrolled. The fasting plasma level of glucose (FPG) and the fasting serum levels of immunoreactive insulin (FIRI) and leptin were determined. Multiple linear regression analysis demonstrated that, in both male diabetic and male non-diabetic subjects, HOMA-R and BMI were independently correlated with serum leptin levels. In females, BMI, but not HOMA-R, was correlated to the serum levels of leptin in both groups. There was no statistically significant difference in the partial regression coefficients between male diabetic and male non-diabetic subjects. These results suggest that the correlation of HOMA-R to the serum levels of leptin in females is dependent on BMI. In males, the relationship between serum leptin levels and the insulin resistance was not affected by the extent of glucose intolerance.
It is known that measurement of accommodation is useful to evaluate the sympathetic activity of intraocular muscles. To find if sympathetic overactivity is involved in eyelid retraction in euthyroid Graves' disease, we measured accommodation in two patients with this disease, whose serum concentrations of free T3, free T4 and TSH were within reference ranges. Accommodation was measured with a computer-assisted infrared optometer with an iriscoder, and the results were expressed as the change in the eye's refractive power (in diopters) in response to the movement of a target beam. In patient 1, the accommodation amplitude was low, indicating sympathetic overactivity. This amplitude rose to near the reference range when timolol maleate drops were used, and the eyelid retraction disappeared when guanethidine drops were given. During the use of guanethidine drops, accommodation remained normal. In patient 2, who had normal accommodation, eyelid retraction did not change with guanethidine administration, but improved with intravenous methylprednisolone pulse therapy. These two cases suggested that even in euthyroid Graves' disease, eyelid retraction is caused by sympathetic overactivity, and pulse therapy with methylprednisolone may be effective for eyelid retraction when guanethidine drops are not effective.
Congenital lipoid adrenal hyperplasia (CLAH) is an autosomal recessive disorder characterized by impaired synthesis of adrenal and gonadal steroids. It was demonstrated that loss-of-function mutations in the steroidogenic acute regulatory protein (StAR) gene cause CLAH and that 46, XX patients with CLAH develop spontaneous puberty. We had reported that three 46, XX patients with CLAH had presented spontaneous puberty and one of the patients had developed life-threatening ovarian cysts, before the etiology of CLAH had been clarified. In the present study, we analyzed their StAR gene and demonstrated mutations. Endocrinological examinations of the patients revealed that serum LH and FSH levels and their responses to the LHRH stimulation were not exaggerated before the onset of puberty. Serum LH levels and its response to LHRH were increased during puberty, whereas serum FSH levels remained within the normal range. Serum estradiol increased after the administration of human menopausal gonadotropins in the pubertal patient, suggesting that the ovary might have another system than StAR to facilitate cholesterol transport into the mitochondria. Although the patients had menstrual cycles, they remained anovulatory, and the resultant increased secretion of LH was speculated to be responsible for the development of ovarian cysts.
Octreotide usually induces long-acting inhibition of GH secretion without rebound hypersecretion. We report a patient with acromegaly who showed rebound elevation of GH and insulin-like growth factor-1(IGF-1) after octreotide withdrawal. The patient responded remarkably to octreotide therapy with normalization of GH and IGF-1 and shrinkage of pituitary macroadenoma. Octreotide therapy was stopped when he developed liver dysfunction. GH and IGF-1 were markedly elevated for two weeks after cessation of octreotide.
The insulin secretory pattern as a phenotype of type 2 diabetes is an impairment in the rapid, pulsatile secretion of insulin in response to a rise in blood glucose after meal-intake. The restoration of endogenous rapid insulin secretion after oral glucose load was established for the first time by using nateglinide, which is a newly developed insulin secretagogue, in obese patients with type 2 diabetes mellitus. It was clearly demonstrated that with the nateglinide, serum insulin levels were quickly raised, and glycemic response curves were almost normalized with the same amount of insulin secretion during 180min. Therefore, the lack of rapid, pulsatile secretion of insulin in response to glycemic rise after oral glucose load, rather than insulin resistance, is responsible for postprandial glycemic response in obese type 2 diabetes patients.