鈴木 敦詞

Recent findings suggest that thyroid stimulating hormone (TSH) is a negative regulator of skeletal remodeling by reducing both differentiation of ostcoblasts and formation of osteoclasts. In addition, increased fracture risk in untreated hypothyroid patients has been reported to begin up to 8 years before diagnosis. The aim of the present study was to evaluate the effect of subclinical hypothyroidism on bone structure by using the heel QUS. Subjects were outpatients without any past or present history of thyroid disease. Among 2 10 postmenopausal women, 22 of 33 patients (Hypo), who had elevated serum TSH concentration (TSH >= 4 mu U/ml) with normal serum free thyroxine (FT4) concentration, agreed to join to this study. We also randomly selected 24 control subjects (Cont) from 176 postmenopausal women with normal thyroid status. Calcaneus osteo sono assessment indices (OSI) of right feet were measured using the ultrasound bone densitometry AOS-100. Serum TSH concentrations in Hypo patients (5.31 +/- 1.3 mu U/ml) were higher than those in Cont patients (2.05 +/- 1.1 mu U/ml), and there was significant difference of FT, concentrations (Cont 1.33 +/- 0.15 ng/dl; Hypo 1.19 +/- 0.17 ng/dl). OSI and its Z-score in Hypo subjects (OSI, 2.138 +/- 0.152; Z-Score -0.322 +/- 0.504 SD, Mean +/- SD) were significantly lower than those in Cont subjects (OSI, 2.347 +/- 0.243; Z-Score 0.322 +/- 0.91 SD, Mean +/- SD). Simple regression statistical analysis showed that OSI decreased according to the increase of serum TSH concentration (n = 47, P < 0.037). In addition, multiple regression analysis showed that the elevation of serum TSH concentration was associated with the decrease of OSI. These results suggest that the elevation of serum TSH concentration in subclinical hypothyroidism affects not bone turnover but bone structure as assessed by QUS.

We investigated the effect of arginine vasopressin (AVT) on inorganic phosphate (Pi) transport in A-10 rat aortic vascular smooth muscle cells (VSMCs). AVP time- and dose-dependently stimulated Na-dependent Pi transport in A-10 cells. This stimulatory effect of AVP on Pi transport was markedly suppressed by V I receptor antagonist. A protein kinase C (PKC) inhibitor calphostin C partially suppressed the stimulatory effect of AVP. The selective inhibitors of c-Jun-NH2-terminal mitogen-activated protein (MAP) kinase (Jun kinase) attenuated AVP-induced Pi transport, but Erk kinase or p38 MAP kinase inhibitors did not. Wortmannin, a phosphatidylinositol (PI) 3-kinase inhibitor, suppressed AVP-induced Pi transport. Rapamycin, a selective inhibitor of S, kinase, reduced this effect of AVP, while Akt kinase inhibitor did not. The combination of inhibitors for PKC, Jun kinase and PI 3-kinase completely suppressed the AVP-enhanced Pi transport. Furthermore, AVP rescued the VSMC from high phosphate-induced cell death and enhanced mineralization of these cells. In summary, these results suggest that AVP stimulates both Na-dependent Pi transport and mineralization in VSMCs. The mechanism is mediated by the activation of multiple signaling pathways including PKC, PI 3-kinase, S-6 kinase and Jun kinase.

25歳, 女性. 18歳頃より体重増加, 肥満精査治療目的に入院. 頸部と両腋窩部に黒色色素沈着を認め, 皮膚生検にて黒色表皮腫と診断された. HOMA-R指数14.3, グルコースクランプ法にてもM値1.74mg/kg/minと高度のインスリン抵抗性が示唆された. 食事療法及び塩酸メトホルミン750mg/日の投与により, 体重, 空腹時血清インスリン濃度が低下, 黒色表皮腫も消退した.

The prevalence of hypovitammosis D has been recently reevaluated, and diabetes is considered as a risk factor for osteoporosis. We studied the association of the prevalence of hypovitammosis D with the clinical features of diabetes. We conducted the observational study in 581 Japanese patients with type 2 diabetes mellitus and 51 normal subjects, and analyzed the relationship between serum 25-hydroxyvitamin D (25-OHD) concentration and the clinical features associated with type 2 diabetes. Mean serum 25-OHD concentration in type 2 diabetes patients was 17.0 +/- 7.1 ng/ml (Mean +/- SD) in winter, and was not statistically different from normal population (17.5 +/- 3.6 ng/ml). The prevalence of hypovitaminosis D (< 20ng/ml) was 70.6%. Serum concentrations of 25-OHD were associated with HbAlc (P= 0.013), age (P = 0.070) and serum albumin (P < 0.00 1), but were not related to BMI or the duration of diabetes. The levels of 25-OHD were significantly lower in the population with apparent microvascular complications, although serum creatinine levels were below 2.0 mg/dl. Serum 25-OHD concentrations in the group treated with insulin (15.4 +/- 6.5 ng/ml) was lower than those in the patients treated with diet alone (20.8 +/- 7.6 ng/ml) and with oral hypoglycemic agents (17.3 +/- 7.0 ng/ml). Furthermore, the highest incidence of osteoporotic fracture and/or back deformity was observed in insulin-treated patients with hypovitammosis D. In conclusion, these results suggest that microvascular complications and insulin treatment in type 2 diabetes patients are associated with the co-existence of hypovitaminosis D, and that hypovitammosis D in insulin-treated patients is possibly related to the risk of osteoporotic fracture.

Introduction: Bone morphogenetic proteins (BMPs) are produced by osteogenic cells and play an important role in bone formation. Inorganic phosphate (Pi) is a fundamental constituent of hydroxyapatite, and its transport by osteogenic cells is an important function for primary calcification of the bone matrix. In this study, we investigated the role of Pi transport in BMP-2-induced matrix mineralization. Materials and Methods: Confluent MC3T3-E1 osteoblast-like cells were exposed to BMP-2 for various time periods. Pi and alanine transport was determined using radiolabeled substrate, Pit-1 and Pit-2 expression by Northern blot analysis, cell differentiation by alkaline phosphatase activity, matrix mineralization by alizarin red staining, and the characteristics of mineral deposited in the matrix by transmission electron microscopy, electron diffraction analysis, and Fourier transformed infrared resolution (FTIR). Results: BMP-2 time- and dose-dependently stimulated Na-dependent Pi transport in MC3T3-E1 cells by increasing the V-max of the transport system. This effect was preceded by an increase in mRNA encoding Pit-1. but not Pit-2. BMP-2 also dose-dependently enhanced extracellular matrix mineralization, an effect blunted by either phosphonoformic acid or expression of antisense Pit-1. Enhanced Pi transport and matrix mineralization induced by BMP-2 were blunted by a specific inhibitor of the c-Jun-N-terminal kinase (JNK) pathway. Conclusions: Results presented in this study indicate that, in addition to its well-known effect on several markers of the differentiation of osteoblastic cells, BMP-2 also stimulates Pi transport activity through a selective increase in expression of type III Pi transporters Pit-1. In MC3T3-E1. cells, this effect is mediated by the JNK pathway and plays an essential role in bone matrix calcification induced by BMP-2.

We conducted an observational study in order to assess the prevalence of hypovitaminosis D and its seasonal changes, in the Tokai area (N35.3 E137.0), in 197 normal subjects in Japan. The mean serum 25-hydroxyvitamin D (25-OHD) level measured by direct radioimmunoassay (RIA) was lowest at the end of winter, and highest at the end of summer (15.1 +/- 7.1 ng/ml in March; 21.5 +/- 5.5 ng/ml in June; 31.6 +/- 5.6 ng/ml in September; 23.1 +/- 5.3 ng/ml in December; mean +/- SD). The prevalence of hypovitaminosis D ( < 20 ng/ml) was 86.7%, 33.4%, 1.0%, and 26.0% in March, June, September, and December, respectively. Mean plasma intact parathyroid hormone (iPTH) concentration was lowest at the end of summer and highest at the end of winter (28.2 +/- 9.3 pg/ml in March; 21.7 +/- 7.0 pg/ml in June; 19.8 +/- 6.9 pg/ml in September; and 25.7 +/- 9.2 pg/ml in December; mean +/- SD). Serum 25-OHD was inversely associated with iPTH ( coefficient, - 0.223; r = 0.251; P < 0.001). Serum 25-OHD levels were higher in men than in women. The serum 25-OHD level was positively associated with age, body weight, and body mass index, but not with body fat content. These results suggest a high prevalence of hypovitaminosis D associated with elevation of iPTH in Japan, in winter, even in a sunny area.

An accerelated polyol pathway in diabetes contributes to the development of diabetic complications. To elucidate diabetic nephropathy involving also renal tubular damage, we measured urinary sorbitol concentration concomitantly with urinary N-acetyl-D-glucosaminidase (NAG) excretion in WBN-kob diabetic rats. Twenty-four-hour urinary sorbitol concentrations increased in the diabetic rats in parallel with whole blood sorbitol concentrations. An increase in 24-h urinary NAG excretion coincided with the elevated urinary sorbitol levels in the diabetic rats. The administration of epalrestat, an aldose reductase inhibitor, reduced the increased whole blood and urinary sorbitol concentrations and urinary NAG excretion concomitantly with renal aldose reductase inhibition in the diabetic rats. These results indicate that diabetic nephropathy involves distorted cell function of renal tubules, and that treatment with epalrestat may prevent at least the progress of the nephropathy.

Prostaglandins are now recognized to be important regulators for both bone formation and resorption. Among them, prostaglandin E-1 (PGE(1)) has been reported to stimulate cAMP accumulation and to induce alkaline phosphatase (ALP) activity, a marker of differentiation, in osteoblast-like cells. Recently, we have shown that p38 mitogen-activated protein (MAP) kinase pathway regulates ALP activity in response to activation of Gi protein-coupled receptors in mouse osteoblast-like MC3T3-E1 cells (Suzuki et al., Endocrinology 140 (1999) 3177). In the present study, we investigated whether p38 MAP kinase is involved in ALP activation by PGE(1) in MC3T3-E1 osteoblast-like cells. PGE(1) dose-dependently enhanced ALP activities in the concentration range between 1 nM and 1 muM in MC3T3-E1 cells. SB203580, a specific inhibitor of p38 MAP kinase, blocked the increase in ALP activity induced by PGE(1). Further analysis with western blotting suggested that PGE(1) induced an increase in tyrosine (Tyr) phosphorylation of p38 MAP kinase. Both Bt(2)cAMP, a permeable analogue of cAMP, and forskolin, which directly activates adenylate cyclase, also induced an increase in Tyr phosphorylation of p38 MAP kinase. H-89, a potent inhibitor of protein kinase A (PKA), significantly suppressed PGE(1)-induced Tyr phosphorylation of p38 MAP kinase. The results of this study suggest that PGE(1) stimulates p38 MAP kinase through the activation of PKA, resulting in the enhancement of ALP activity. (C) 2003 Elsevier Ltd. All rights reserved.

Understanding the causes of diabetic vascular complications has become an increasingly important issue because of the rapidly rising prevalence of diabetes. Recently discovered vasoconstrictors and angiogenesis regulators, such as endothelin (ET) and vascular endothelial growth factor (VEGF), have been intensely studied for possible pathogenic roles in diabetic vascular complications. The present study was undertaken to clarify the effect of glycemic control on serum VEGF and plasma ET-1 concentrations in diabetic patients, and to identify other factors that may cause fluctuations of these substances. Plasma VEGF and ET-1 concentrations of 45 hospitalized diabetic patients and 54 control subjects were measured by enzyme immunoassay (EIA) and radioimmunoassay (RIA), respectively. Plasma VEGF was elevated in poorly controlled diabetic patients compared with healthy subjects and plasma VEGF concentrations declined after hospitalized treatment with either insulin or oral hypoglycemic agents in combination with diet. There was a significant correlation between plasma VEGF concentration and both fasting plasma glucose (FPG) and hemoglobin A(1c) (HbA(1c)). Plasma ET-1 in poorly controlled diabetic patients was higher than in healthy controls, but improved glycemic control did not affect plasma ET-1 concentrations. Thus, poor glycemic control causes increased levels of plasma VEGF, which may result in hypertension and vascular complications in diabetes. Short-term treatment resulting in good glycemic control can improve levels of VEGF and may provide beneficial effects on diabetic vascular complications. (C) 2004 Elsevier Inc. All rights reserved.

We investigated the effect of platelet-derived growth factor B homodimer (PDGF-BB) on inorganic phosphate (Pi) transport activity, which has been reported to be involved in the mechanism of atherosclerosis, in A-10 rat aortic vascular smooth muscle cells (VSMCs). PDGF-BB time- and dose-dependently stimulated Pi transport in A-10 cells. Using northern blot analysis, the PDGF-BB-enhanced Pi transporter (PiT) in A-10 cells was identified as Pit-1 (Glvr-1), a member of the type III Na-dependent PiT. An inhibitor of PDGF P-receptor tyrosine kinase suppressed PDGF-BB-induced Pi transport. Both a protein kinase C (PKC) inhibitor calphostin C and PKC down regulation suppressed the stimulatory effect of PDGF-BB on Pi transport. On the other hand, inhibition of mitogen-activated protein (MAP) kinases by selective inhibitors did Dot affect Pi transport. Ly294002, a phosphatidylinositol (PI) 3-kinase inhibitor, partially attenuated PDGF-BB-induced Pi transport. A selective inhibitor Of S-6 kinase, rapamycin, reduced this effect of PDGF-BB, while Akt kinase inhibitor did not. In summary, these results indicated that PDGF-BB is a potent and selective stimulator of Pi transport in VSMCs. The mechanism responsible for this effect is not mediated by MAP kinase, but involves activation of PKC, PI 3-kinase and S-6 kinase. (C) 2004 Published by Elsevier Ireland Ltd.

Signaling involved in osteoblastic cell differentiation remains largely unknown. This study further investigates mechanisms involved in BMP-2-induced osteoblastic cell differentiation. We report that BMP-2 can activate JNK and p38 in osteoblastic cells and provide evidences that these MAP kinases have distinct roles in regulating alkaline phosphatase and osteocalcin expression.

Endothelin-1 (ET-1) has been reported to modulate bone metabolism both in vivo and in vitro. In the present study, we investigated the effect of ET-1 on inorganic phosphate (Pi) transport in osteoblast-like cells, which is now considered to be important for the initiation of bone matrix calcification. ET-1 time- and dose-dependently stimulated Na-dependent Pi transport in mouse calvaria-derived osteoblast-like MC3T3-E1 cells, and this effect was dependent on transcriptional and translational process. Kinetic analysis indicated that the change in Pi transport activity induced by ET-1 was due to alteration in the number of the Pi transporter. BQ123, a selective antagonist for ETA receptor, suppressed the ET-1-induced Pi transport, but BQ788, a selective antagonist for ETB receptor, had no effect. The inhibition of phosphoinositide hydrolysis by phospholipase C (PLC) partially attenuated the Pi transport by ET-1. Propranolol, which inhibits phosphatidic acid phosphohydrolase, also suppressed ET-1-induced Pi transport. On the contrary, indomethacin did not affect the stimulatory effect of Pi transport by ET-1. Calphostin C, a protein kinase C (PKC) inhibitor, significantly blunted the stimulatory effect of ET-1 on Pi transport. Combined effect of PMA and ET-1 on Pi transport was not additive. Pi transport induced by ET-1 was also suppressed in PKC down-regulated cells. In conclusion, the results of the present study indicate that, in MC3T3-E1 osteoblast-like cells, ET-1 acting through ETA receptor links to a stimulation of Pi transport via activation of PKC through both phosphoinositide and phosphatidylcholine hydrolyses. J. Cell. Biochem. 83: 47-55, 2001. (C) 2001 Wiley-Liss, Inc.