Michiko Kanzawa

Plasma leptin concentrations are associated with bone mineral density and the presence of vertebral fractures in postmenopausal women

OBJECTIVE Although total fat body mass (FM) is considered to be one of the major determinants of bone mass, the mechanism by which FM and bone mass are positively correlated remains unclear. Leptin, the product of the obese (ob) gene, is secreted from adipocytes and its plasma levels are known to be positively correlated with %fat (FM divided by total body weight). There is recent evidence suggesting that leptin directly stimulates osteoblastic differentiation. Thus it is possible that the anabolic action of this hormone on bone may participate in the positive correlation between FM and bone mass. In this study, we analysed the relationships between either plasma leptin levels or %fat vs. bone mineral density (BMD) values as well as the presence of vertebral compression fractures, and evaluated whether or not plasma leptin levels were associated with BMD or bone fragility in a manner independent of FM.

Effect of high phosphate concentration on osteoclast differentiation as well as bone-resorbing activity.

Although high inorganic phosphate (Pi) concentration in culture media directly inhibits generation of new osteoclasts and also inhibits bone resorption by mature osteoclasts, its precise mechanism and the physiological role have not been elucidated. The present study was performed to investigate these issues. Increase in extracellular Pi concentration ([Pi]e) (2.5–4 mM) concentration dependently inhibited 1,25‐dihydroxyvitamin D3 [1,25(OH)2D3] or parathyroid hormone (PTH)‐(1‐34)‐induced osteoclast‐like cell formation from unfractionated bone cells in the presence of stromal cells. Increase in [Pi]e (2.5–4 mM) concentration dependently inhibited 1,25(OH)2D3‐, PTH‐(1‐34)‐, or receptor activator of NF‐κB ligand (RANKL) and macrophage colony‐stimulating factor (M‐CSF)‐induced osteoclast‐like cell formation from hemopoietic blast cells in the absence of stromal cells. Increase in [Pi]e (2.5–4 mM) dose dependently stimulated the expression of osteoprotegerin (OPG) mRNA and increased the expression of OPG mRNA suppressed by PTH‐(1‐34) or 1,25(OH)2D3 in unfractionated bone cells, while it did not affect RANKL mRNA. Increase in [Pi]e (2.5‐4 mM) concentration dependently inhibited the bone‐resorbing activity of isolated rabbit osteoclasts. Increase in [Pi]e (4 mM) induced the apoptosis of isolated rabbit osteoclasts while it did not affect the apoptosis of osteoclast precursor cells and mouse macrophage‐like cell line C7 cells that can differentiate into osteoclasts in the presence of RANKL and M‐CSF. These results indicate that increase in [Pi]e inhibits osteoclast differentiation both by up‐regulating OPG expression and by direct action on osteoclast precursor cells. It is also indicated that increase in [Pi]e inhibits osteoclastic activity at least in part by the direct induction of apoptosis of osteoclasts.

Determinants of bone mineral density and spinal fracture risk in postmenopausal Japanese women

Abstract: The present study analyzed the factors that determine bone mineral density (BMD) and predict spinal fracture risk in postmenopausal Japanese women. Two hundred and five postmenopausal Japanese women aged 48–84 years (mean age 64 years) were enrolled in the cross-sectional study. BMD of the lumbar spine, femoral neck and total body as well as body composition were measured by dual-energy X-ray absorptiometry (DXA). Mid-radial BMD was measured by single-photon absorptiometry. We also determined serum levels of insulin-like growth factor (IGF)-I, IGF binding protein-2, -3 and osteocalcin as well as urinary levels of pyridinoline (Pyr), deoxy-Pyr (D-Pyr) and growth hormone. Multiple regression analysis revealed that lean body mass (LBM) was positively correlated with BMD at all sites. In contrast, femoral neck BMD was highly related to fat mass as well as LBM, although fat mass was not an independent correlate of total body and mid-radial BMD. LBM and urinary D-Pyr were crucial determinants at all sites except the mid-radius in stepwise regression analysis. Fat mass and serum IGF-I were determinants of femoral neck and mid-radial BMD, respectively. In terms of reproductive history, parity affected lumbar BMD. Factors affecting BMD differed according to the site. On the other hand, lumbar BMD as well as serum levels of IGF-I and albumin were selected as predictors of spinal fracture risk in multiple logistic regression analysis. Lumbar BMD, serum IGF-I and LBM were selected in women with lumbar BMD above 0.727 g/cm2. In conclusion, the present study indicates that LBM is a more important determinant of BMD than fat mass at any site except the femoral neck. Age, serum IGF-I and urinary D-Pyr were also determinants of BMD, dependent on the regions measured. Lumbar BMD and LBM as well as serum levels of IGF-I and albumin were useful markers which predicted the risk of osteoporotic spinal fractures in postmenopausal Japanese women.

Thyroid hormone stimulates osteoclast differentiation by a mechanism independent of RANKL–RANK interaction

It is well known that thyroid hormone excess causes bone loss. However, the precise mechanism of bone loss by thyroid hormone still remains unclear. When T3 was added to unfractionated bone cells after degeneration of pre‐existent osteoclasts, T3 (1 pM–100 nM) dose‐dependently stimulated osteoclast‐like cell formation, irrespective of the presence of indomethacin and IL‐6 Ab. T3 increased the expression of osteoprotegerin (OPG) messenger RNA (mRNA), but not of receptor activator of nuclear factor κB ligand (RANKL) in unfractionated bone cells, suggesting that the stimulatory effect of T3 on osteoclast formation was not mediated by the RANKL/OPG system. We next examined the direct effect of T3 on osteoclast precursors in the absence of osteoblasts, using hemopoietic blast cells derived from spleen cells. T3 (1 pM–100 nM) dose‐dependently stimulated osteoclast‐like cell formation from osteoclast precursors. OPG did not inhibit T3‐induced osteoclast formation from osteoclast precursor cells. The polymerase chain reaction (PCR) product corresponding in size to the mouse T3 receptor α1 cDNA was detected in osteoclast precursors from mouse hemopoietic blast cells as well as mouse heart and mouse osteoblastic cell line MC3T3‐E1 cells, suggesting that T3 directly stimulated osteoclast‐like cell formation from osteoclast precursors in the absence of osteoblasts. Further, T3 increased the expression of c‐Fos mRNA at 15 min and 24 h and Fra‐1 mRNA at 2 and 6 h in osteoclast precursors. Consistent with the increased expression of c‐Fos mRNA observed by RT‐PCR, the activation of c‐Fos occurred in osteoclast precursor cells stimulated by T3, while the activation of neither NF‐κB nor MAPKs was observed by immunoblot analysis. Antisense oligodeoxynucleotides (as‐ODN) complementary to c‐Fos mRNA at 1 μM significantly inhibited T3‐induced osteoclast‐like cell formation from osteoclast precursors in the absence of stromal cells while sense‐ODN did not affect T3‐induced osteoclast‐like cell formation. These results indicate that T3 directly stimulates osteoclast differentiation at least in part by up‐regulation of c‐fos protein in osteoclast precursor cells. J. Cell. Physiol. 201: 17–25, 2004.

The Presence and Severity of Vertebral Fractures is Associated with the Presence of Esophageal Hiatal Hernia in Postmenopausal Women

Abstract: We examined the relationship between the presence of esophageal hiatal hernia (HH) assessed by endoscopy and the presence of vertebral fractures (VFs) in 87 Japanese postmenopausal women (age range 52–87 years). We found that 29 (63%) of 46 patients with HH (71.2 ± 6.1 years, mean ± SD) had one or more VFs, compared with 14 (34%) of 41 patients without HH (70.8 ± 6.8 years), which was a significant difference in the frequency of VFs (c2= 7.242; p= 0.0071). The average number of VFs per patient was significantly higher for the patients with HH than for those without HH (1.67 ± 1.75 vs 0.68 ± 1.21, p= 0.0032). There were no significant differences in absolute or age-matched bone mineral density (BMD) values at the lumbar spine (0.656 ± 0.131 vs 0.662 ± 0.148 g/cm2; Z-score, –0.35 ± 1.17 vs –0.26 ± 1.00) and there were no significant differences in biochemical parameters, age, years since menopause or body mass index (BMI) between the two groups. When patients were divided into those with reflux esophagitis (RE) (n= 30, 70.2 ± 7.3 years) and those without RE (n= 57, 71.4 ± 5.9 years), no significant differences were detected in any of the above parameters including the presence or number of VFs. The patients were further subdivided into four groups: those with ‘HH only’ (n= 23, 72.3 ± 4.6 years), with ‘RE only’ (n= 7, 70.9 ± 7.7 years), with ‘both’ (n= 23, 70.0 ± 7.3 years) and with ‘neither’ (n= 34, 70.8 ± 6.7 years). One or more VFs were found in 12 (52%), 1 (14%), 17 (74%), and 13 (38%) patients in each group, respectively, and the difference in frequency was significant (c2= 10.748; p= 0.0132). The average number of VFs per patient in each group was 1.57 ± 2.06, 0.14 ± 0.38, 1.78 ± 1.41 and 0.79 ± 1.30, respectively, and there were significant differences between the ‘both’ and ‘neither’ groups, and between the ‘both’ and ‘RE only’ groups (p<0.05). When univariate logistic regression analysis was performed with the presence of HH as a dependent variable and each of the presence of VFs, the number of VFs per patient, absolute or age-matched BMD values at the lumbar spine, BMI and plasma albumin as independent variables, the presence of VFs and the number of VFs per patient were selected as indices affecting the presence of HH (odds ratio: 3.29 and 1.59, 95% confidence interval: 1.36–7.94 and 1.14–2.23; p = 0.0080 and 0.0064, respectively). These results show that the presence and severity of VFs are associated with the presence of HH but not of RE in Japanese postmenopausal women, and suggest that kyphosis induced by multiple VFs might predispose elderly women to a complication with HH.

Association of Polymorphic Alleles of the Calcium-Sensing Receptor Gene with Parathyroid Hormone Secretion in Hemodialysis Patients

The present study was performed to investigate the association of calcium-sensing receptor (CaSR) genotypes with parathyroid hormone (PTH) secretion in hemodialysis patients. Subjects were 122 Japanese hemodialysis patients, including 39 patients with diabetes mellitus (DM). The CaSR polymorphisms tested were codon 990 in intracellular domain (A/A, A/G, and G/G groups) as well as T to C base change of intron 4 (T/T, T/C, and C/C groups). Statistical analysis of these polymorphisms revealed that the serum PTH level was significantly higher in the A/A group than in the G/G group in the former. In addition, the serum PTH level was also significantly higher in patients displaying C allele, as compared with the T/T group in the latter. This association of two polymorphisms with the serum PTH level was observed only in non-DM patients. Although two polymorphisms affected the PTH level independently, patients who possessed both genotypes (AAC+) had a markedly high level of PTH not only in the non-DM group but also in the DM group. The present findings indicate the possibility of the prediction for the extensive progression of secondary hyperparathyroidism through analyzing the CaSR polymorphisms in chronic hemodialysis patients.

Association of polymorphic alleles of the calcium‐sensing receptor gene with the clinical severity of primary hyperparathyroidism

OBJECTIVE Primary hyperparathyroidism (pHPT) is a heterogeneous disease in its clinical course and severity. Previous studies have suggested an association between the clinical severity of pHPT and the genotypes of vitamin D receptor, oestrogen receptors and PTH molecules. The Ca‐sensing receptor (CaR) is activated by an extracellular calcium ion and controls PTH secretion, and thus polymorphisms of CaR might be associated with the magnitude of PTH secretion and the clinical severity of pHPT. In this study, we examined the relationship between CaR polymorphisms and biochemical markers in pHPT patients.

Effect of recombinant human growth hormone in elderly osteoporotic women

Bone mineral density and growth hormone (GH) secretion rate both decline during normal human ageing. We evaluated the effects of recombinant human GH on markers of body composition and bone turnover in an open study in 8 elderly osteoporotic women aged 68–75 years (mean age 71 years).

Parathyroid hormone gene polymorphisms in primary hyperparathyroidism

Genetic contributions to bone mineral density (BMD) and bone turnover are well known. In the present study, we analysed the relationship between restriction fragment length polymorphisms of the PTH gene and the development of primary hyperparathyroidism (pHPT) as well as its severity.

Identification of an enhancer sequence in 5′-flanking region of 1A exon of mouse liver/bone/kidney-type alkaline phosphatase gene

The 5′‐flanking region of the 1A exon of the mouse liver/bone/kidney‐type alkaline phosphatase (L/B/K (ALP) gene was analyzed for the promoter activity. Luciferase assays revealed that a segment containing a 9 bp inverted repeat (nt‐1212 to ‐1179, designated as IRS for inverted repeat segment) had an enhancer activity. Though an E‐box at nt ‐234 did not have a prominent promoter activity, the effect of IRS required its presence. Both transcriptional activity and IRS binding to the nuclear proteins were similarly observed even in cells that did not express ALP. Bone morphogenetic protein‐2 (BMP‐2) induced the ALP expression in pluripotent C3H10T1/2 cells, but it did not increase the transcriptional activity of the promoter region. Though they do not determine cell specific or BMP‐2 inducible ALP expression, IRS and the E‐box at position ‐241 seem to be important for the basic transcriptional activity of ALP gene.