Hiroyuki Tanaka

Altered Biochemical Markers of Bone Turnover in Humans During 120 Days of Bed Rest

Microgravity induces significant and progressive bone loss in both humans and animals. This is the consequence of disturbed bone remodeling. We performed a bed rest experiment to simulate microgravity and tried to clarify bone metabolism by measuring biochemical markers of bone turnover. Six healthy volunteers participated in 120 days of bed rest. The parameters of calcium homeostasis, calcitropic hormones, and biochemical markers of bone turnover were examined. After ambulatory control evaluation, all subjects underwent 120 days of bed rest. Metabolic evaluation was performed in a baseline period, and on days 7, 16, 50, 72, 92, and 108 during bed rest, and on days 10 and 25 during a recovery period. Bed rest induced an increase in urinary calcium (Ca) excretion and serum Ca and bone resorption markers. Urine pyridinoline, deoxypyridinoline, and type I collagen cross-linked N-telopeptide increased more rapidly than urinary Ca excretion and serum Ca. Tartrate-resistant acid phosphatase (TRAP) increased even in the recovery period. Carboxy-terminal propeptide of type I collagen, a bone formation marker, significantly decreased on days 50, 92, and 108 of bed rest. These changes of biochemical markers of bone metabolism, except for TRAP, rapidly returned toward control levels in the recovery period. Immunoreactive parathyroid hormone showed a modest decrease during bed rest and a significant increase in the recovery period. Insulin-like growth factor I (IGF-I) and its binding protein, insulin-like growth factor binding protein-3, increased during bed rest, indicating the possibility of resistance to IGF-I in bones under reduced mechanical stress and strain. Bone loss from unloading results from the combination of acceleration of bone resorption and subsequent retardation of bone formation.

Direct action of 1,25-dihydroxyvitamin D on bone: VDRKO bone shows excessive bone formation in normal mineral condition☆

In the present study, the direct role of Vitamin D in bone metabolism was investigated. Vitamin D has been suggested to be an important hormone for bone metabolism, but there has been little evidence that Vitamin D actively participates in this process. Here, we show the direct action of Vitamin D by transplanting the bone of the Vitamin D receptor null mutant mice (VDR-/-) to the wild-type mouse. This procedure allowed us to investigate the changes in the bone without VDR in the normal humoral environment. Unexpectedly, the volume and the density of the VDR-/- bone transplanted to the wild-type mouse were significantly increased compared with the control (wild-type bone transplanted to the wild-type mouse). We show that Vitamin D has key roles in bone metabolism negatively.

Growth Hormone Therapy in Achondroplasia

Achondroplasia is one of the most common causes of severe rhizomelic dwarfism. We have previously reported the growth-promoting effect of growth hormone (GH) in this disorder. In this expanded clinical study, dose dependency and the long-term effect of GH were also investigated. Prepubertal children with achondroplasia (82 males and 63 females) were randomly divided into 2 groups. Patients were treated with 0.5 IU/kg per week or 1.0 IU/kg per week subcutaneous recombinant human GH. Of 75 patients, the mutational analysis of fibroblast growth factor receptor-3 revealed that G1138A was detected in 70 and G1138C was found in 2. GH increased growth rate and height z score in a dose-dependent manner. GH also increased serum insulin-like growth factor (IGF)-I, IGF-binding protein-3 and osteocalcin. No adverse effects were observed in either group. We conclude that GH therapy is a useful method for improvement of severe growth retardation of achondroplasia.

The Effects of Bone Marrow Transplantation on X-linked Hypophosphatemic Mice

The genes responsible for X‐linked hypophosphatemic (XLH) vitamin D‐resistant rickets and the murine homolog, hypophosphatemic mice (Hyp), were identified as PHEX and Phex (phosphate‐regulating gene with homology to endopeptidases on the X chromosome), respectively. However, the mechanism by which inactivating mutations of PHEX cause XLH remains unknown. We investigated the mechanisms by syngeneic bone marrow transplantation (BMT) from wild mice to Hyp mice. The expression of the Phex gene was detected in mouse BM cells. BMT introduced a chimerism in recipient Hyp mice and a significant increase in the serum phosphorus level. The renal sodium phosphate cotransporter gene expression was significantly increased. The effect of BMT on the serum phosphorus level depended on engraftment efficiencies, which represent the dosage of normal gene. Similarly, the serum alkaline phosphatase (ALP) activity was decreased and bone mineral density was increased. Furthermore, the renal expression of 25‐hydroxyvitamin D3 24‐hydroxylase, which is a key enzyme in the catabolic pathway and is increased in XLH/Hyp, was improved. From these results, we conclude that transplantation of normal BM cells improved abnormal bone mineral metabolism and deranged vitamin D metabolism in Hyp by replacing defective gene product(s) with normal gene product(s). This result may provide strong evidence for clinical application of BMT in metabolic bone disorders.

PTHrP rescues ATDC5 cells from apoptosis induced by FGF receptor 3 mutation.

An activation mutation in the FGFR3 gene causes ACH. The effects of the FGFR3 mutants on apoptosis were analyzed in a chondrogenic cell line. ACH chondrocytes exhibited marked apoptotic with downregulation of PTHrP expression. Rescue of these cells by PTHrP replacement implies a potential therapy for this disorder.

Osteoclastogenesis inhibitory factor/osteoprotegerin reduced bone loss induced by mechanical unloading

Skeletal unloading resulting from space flight and prolonged immobilization causes bone loss. Such bone loss ostensibly results from a rapid increase in bone resorption and subsequent sustained reduction in bone formation, but this mechanism remains unclear. Osteoclastogenesis inhibitory factor/osteoprotegerin (OCIF/OPG) is a recently identified potent inhibitor of osteoclast formation. We studied effects of OPG administration on tail-suspended growing rats to explore the therapeutic potential of OPG in the treatment and prevention of bone loss during mechanical unloading, such as that which occurs during space flight. Treatment with OPG in tail suspension increased the total bone mineral content (BMC g) of the tibia and femur and the total bone mineral density (BMD g/cm2) of the tibia. Moreover, treatment with OPG prevented reduction not only of BMC and BMD, but also of bone strength occurring through femoral diaphysis. Treatment with OPG in tail-suspended rats improved BMC, BMD and bone strength to levels of normally loaded rats treated with vehicle. Treatment with OPG in normally loaded rats significantly decreased urinary excretion of deoxypyridinoline, but the effect of OPG in tail suspension was unclear. These results indicate that OPG may be useful in inhibiting bone loss-engendered mechanical unloading.

Molecular Basis for the Treatment of Achondroplasia

Achondroplasia (ACH), the most common form of short-limbed dwarfism, and its related disorders are caused by constitutively activated point-mutated fibroblast growth factor receptor 3 (FGFR3). Recent studies have provided a large body of evidence to prove chondrocyte proliferation and differentiation in these disorders. However, little is known about the possible effects of the FGFR3 mutants on apoptosis of chondrocytes. In the present study, we analyzed apoptosis using a chondrogenic cell line, ATDC5, expressing the FGFR3 mutants causing ACH and thanatophoric dysplasia, which is a more severe neonatal lethal form comprising type I and type II. We found that the introduction of these mutated FGFR3s into ATDC5 cells decreased mRNA expression of parathyroid hormone-related peptide (PTHrP) and induced apoptosis. Importantly, replacement of PTHrP prevented the apoptotic changes in ATDC5 cells expressing ACH mutant. Insulin-like growth factor (IGF)-I, which is an important mediator of growth hormone (GH), also reduced apoptosis in ATDC5 cells expressing ACH mutant. IGF-I prevented apoptosis through the phosphatidylinositol 3-kinase and mitogen-activated protein kinase pathways, indicating the mechanisms by which GH treatment improves disturbed bone growth in ACH.

Osteoporosis in male and female leprosy patients.

Abstract. We measured the bone mineral density (BMD) of 353 leprosy patients (197 males 50–89 years old, average age 70.2; and 156 females 53–90 years old, average age 72.9) and serum levels of free testosterone (FT) in 81 males. The BMD of the lumbar vertebrae (L2–L4), diaphysis of the radius (1/3 radius), and the neck of the femur (neck) was measured using DXA (QDR 4500). The BMD of −2.5 SD YAM (young adult mean) in Japanese men and women was used as the cutoff value for osteoporosis in the respective genders: BMD of L2–L4, 0.751 g/cm2 (male), 0.747 g/cm2 (female); 1/3 radius, 0.655 g/cm2 (male), 0.550 g/cm2 (female); neck, 0.581 g/cm2 (female). The percentages of males with osteoporosis were 31.3% in the 50th, 32.9% in the 60th, 44.9% in the 70th, and 40.7% in the 80th decade at L2–L4. Similarly, the percentages were 33.3%, 58.3%, 74.3%, and 75.0%, respectively, at 1/3 radius. Among females, the percentages were 22.2%, 41.3%, 44.9%, and 68.8%, respectively, at L2–L4; 0%, 42.9%, 89.5%, and 78.6%, respectively, at 1/3 radius; and 11.1%, 38.6%, 67.7%, and 84.6% respectively, at neck. FT in men ranged from almost 0 to normal at each decade and BMD levels were significantly correlated with FT in all three regions of the skeleton (P < 0.0001). More than 30% of osteoporosis was found at each decade and FT may be one of the main factors affecting BMD in male leprosy patients.

Bone assessment of female long-distance runners

The objective of the present study was to investigate the effects of long-distance training on the bones in different growth stages by evaluating the bones of female high school athletes and female adult athletes who engage in long-distance training. Thirteen female high school athletes (aged 15–17 years) and 7 female adult athletes (aged 21–25 years) were enrolled in the present study. Bone mineral density (BMD) and bone mineral content (BMC) were measured using dual X-ray absorptiometry (DXA). Cortical bone status was also evaluated by measuring the tibial speed of sound (t-SOS). The bone metabolic status was evaluated by osteocalcin (OC) and tartrate-resistant acid phosphatase (TRAP). OC (P < 0.01) and lumbar BMD (P < 0.05) were significantly higher in high school athletes who were in the modeling period than in the adult athletes. Leg BMD (P < 0.01) and t-SOS (P < 0.05) were significantly higher in adult athletes who were in the remodeling period than in high school athletes. When lumbar BMD and leg BMD were compared according to menstrual conditions, the high school athletes with regular menstruation showed a significantly higher level of bone density than those having irregular or absent menstruation. These results indicate that the effect of long-distance training on bone metabolism is influenced strongly by sex hormones during late puberty. We concluded that the effect of long-distance training on bone metabolism in the female high school athletes (i.e., during puberty) was different from that in the adult athletes.

Analysis of linear growth in survivors of childhood acute lymphoblastic leukemia.

Abstract.u2003Therapy for childhood acute lymphoblastic leukemia (ALL) is entering a new era in terms of quality-of-life. In the current study, 21 patients with childhood-onset ALL were assessed for linear growth, bone mineral density (BMD), and endocrinological status, focusing especially on longitudinal analysis of the growth of each patient. Linear growth was uniformly attenuated during therapy in all patients. In contrast, after the cessation of therapy, the growth of each patient varied widely from attenuated to dramatic catch-up growth. In pubertal survivors who had received chemotherapy and cranial irradiation during prepuberty, the degree of growth after the cessation of therapy was negatively correlated with changes in height Z scores during therapy (r = −0.76, P = 0.004). One of the factors involved in catch-up growth, urinary N-telopeptide/creatinine (U-NTx/Cr), was significantly higher in patients whose Z scores decreased after cessation of therapy (P = 0.01), despite normal pubertal development and normal endocrinological assessments. The present study revealed individual differences in linear growth after the cessation of therapy and suggests the importance of catch-up growth during puberty.