Noriaki Yamamoto

Local effects of transforming growth factor-β1 on rat calvaria: Changes depending on the dose and the injection site

Abstract: Transforming growth factor-β1 (TGF-β1) has opposite effects on osteoblastic cells in vitro, namely an inhibitory or stimulatory effect on cell differentiation. Because these effects are dependent on TGF-β1 concentration or culture condition, we investigated whether the in vivo effects of TGF-β1 on bone formation in infant rat calvaria were affected by the dose or the injection site. Human platelet-derived TGF-β1 was injected subcutaneously onto the periosteal surface of parietal bone of 4-week-old rats at doses of 5 or 20 ng/100 μl per animal for 14 days, and the local effect on bone formation was examined by bone histomorphometry. TGF-β1 treatment for 7 days decreased the mineral apposition rate, bone formation rate, and elongated mineralization lag time at the injection site. This change became more prominent when treatment continued for 14 days. These changes were restricted to the TGF-β1-exposed area. Multiple subcutaneous injections of a relatively high dose (200 ng/100 μl per animal) of TGF-β1 induced woven bone formation, in addition to marked inhibition of bone formation rate and prolongation of mineralization lag time. On the other hand, direct exposure of TGF-β1 in the subperiosteal layer induced woven bone with periosteal cell proliferation even at a single injection of a low dose (5 or 50 ng/25 μl) of TGF-β1. In conclusion, the in vivo effects of TGF-β1 on bone formation varied depending on its concentration and injection site. Also, subcutaneous injection of relatively low doses of TGF-β1 inhibited local lamellar bone formation.

Bone mineral density and bone histomorphometric assessments of postpregnancy osteoporosis: A report of five patients

Reports of five young women who developed vertebral fractures associated with pregnancy and lactation are presented (Fig. 1). Ages ranged from 24 to 37 (mean 30) years. All five patients have osteoporosis with two to nine vertebral fractures at presentation postpartum. Bone mineral density (BMD) was measured by single-photon absorptiometry, quantitative computer tomography, and dual-energy X-ray absorptiometry. BMD of the trabecular bone was less than normal values and it remained apparently low even several years after pregnancy. Histological findings of bone biopsy identified the bone loss with increasing bone resorption. Our present findings suggest that postpregnancy osteoporosis affects mainly the trabecular bone site, and the patients might have low peak bone mass and poor reversibility probably due to a low rate of remodeling.

Effects of intermittent administration of low dose human PTH(1–34) on cancellous and cortical bone of lumbar vertebral bodies in adult beagles

This study assessed the effect of low dose human parathyroid hormone [hPTH(1-34)] administration on cancellous and cortical bone of lumbar vertebrae in intact male beagles. 16 19-20-month-old beagle dogs were randomized into four groups: in group 1, the vehicle control group, saline was injected daily; in group 2, the sequential group, 0.375 microg/kg of PTH was injected daily for 4 weeks, then off 8 weeks, and this sequence was once repeated for another 4 and 8 weeks; in group 3, the same dose of PTH was injected once per week for 24 weeks; and, in group 4, PTH was injected three times per week for 24 weeks. Histomorphometric assessment on cancelllous and cortical bone (both ventral and dorsal shell) and two-dimensional node-strut analysis were done on the fifth lumbar vertebral bodies after calcein double bone labeling. In intact adult beagles, on the group treated with 0.375 microg/kg per day three times per week (group 4): (1) had a higher mean value in cancellous bone formation parameters [osteoid surface (+74%), osteoid volume (twofold), mineral apposition rate (+21%), and bone formation rate (twofold)]; (2) exhibited no effect on cortical thickness and porosity in both the ventral and dorsal shell; and (3) showed a lower mean value of node to termini (0.11 +/- 0.02 vs. 0.22 +/- 0.09) and a higher mean value of cortex to node (0.18 +/- 0.06 vs. 0.08 +/- 0.02), but not in trabeculae to trabeculae node, than age-related controls. In conclusion, we found that a low dose of PTH administration: (1) stimulated cancellous bone formation; (2) improved connectivity of trabeculae joined to the cortex; (3) did not decrease cortical thickness; and (4) did not increase cortical porosity in both ventral and dorsal cortexal shell of the lumbar vertebrae during this dosage and period in intact male beagles.

Maintenance of bone mass by physical exercise after discontinuation of intermittent hPTH(1–34) administration

Human PTH(1-34) has been recognized for its marked anabolic effect on bone, but that effect has been reported to be lost after cessation of PTH treatment. The objective of this study was to determine the fate of hPTH-stimulated bone and whether this anabolic effect of PTH could be maintained by daily exercise. Eleven-week-old Sprague-Dawley rats were ovariectomized (OVX) and human PTH(1-34) (30 micrograms/kg) was injected subcutaneously three times per week for 12 weeks beginning one week after surgery. After the cessation of PTH treatment, treadmill exercises were performed for 8 weeks (15.7 m/min, 1 h/day, 5 days/week). The results of histomorphometric assessment in the proximal tibial metaphysis demonstrated that hPTH treatment partially prevented OVX-induced cancellous bone loss. Eight weeks following the cessation of PTH treatment, PTH-stimulated bone mass went back to the OVX control level. Daily exercise did maintain PTH-stimulated bone mass; however, this exercise did not increase the bone mass in PTH-untreated OVX rats.

Weight gain in childhood and bone mass in female college students

The attainment of maximal peak bone mass early on in life is one of the most important strategies for the prevention of osteoporosis in women. The aim of this study was to clarify the correlation between gains in body size in all growth phases in childhood and adult bone mass in women. The subjects were 86 female first-year university students, aged 18–21 years. We measured the subjects’ bone mineral content (BMC) and bone mineral density (BMD) at the lumbar spine and the left hip, including the femoral neck, with dual energy X-ray absorptiometry. Each subject was measured for current height and weight. Height and weight at birth, and at 1.5, and 3 years were obtained from each maternity record book, and those between 6 and 18 years were obtained from their school health records. Other information, including physical activity and calcium intake, was obtained through an interview. Bivariate analysis showed that weight gains during the periods from birth to 1.5 years and from 9 to 12 years significantly correlated with both BMC and BMD values at any site. The stepwise method of multiple regression analysis showed that a weight gain during the period from birth to 1.5 years was significantly associated with BMC at the lumbar spine (P = 0.0001) and at the femoral neck (P = 0.0290) and with BMD at the lumbar spine (P = 0.0387). Birth weight was significantly associated with BMC at the lumbar spine (P = 0.0474) and the total hip (P = 0.0352), and weight gain during the period from 9 to 12 years was significantly associated with BMC at the femoral neck (P = 0.0376). In conclusion, birth weight and weight gain in infancy are important determinants of bone mass in young women. Additionally, a girl’s prepubertal growth spurt is likely to be a key phase for the acquisition of bone mass in relation to body weight. Our findings suggest that osteoporosis prevention programs may need to start very early in the life cycle.

Effects of human PTH(1-34) and bisphosphonate on the osteopenic rat model

It has been demonstrated that the intermittent administration of human parathyroid hormone (hPTH) is beneficial for restoration of bone mass in osteoporotic patients. The mechanisms of anabolic effects of hPTH have been determined by ovariectomized rat models and other larger remodeling animals. However, treatment with hPTH may increase the cancellous bone mass at the expense of cortical bone mass and cessation of the treatment results in rapid bone loss. Efforts have been made to maintain newly formed bone mass after withdrawal of the hPTH treatment. These issues are not well understood. In this article, the authors would like to represent previous studies of their own and others concerning these issues.

Discrepancy of response of hPTH administration and its withdrawal between trabecular and cortical bone sites in ovx rats

We demonstrated the differences of response between trabecular and cortical bone sites in the rat induced by OVX and hPTH(1-34) administration [subcutaneous injection, 30 micrograms/kg, 3 times/week, for 12 weeks] and its withdrawal [for 8 weeks]. We observed that hPTH(1-34) administration in OVX rats partially prevented OVX-induced cancellous bone loss in the proximal tiabial metaphysis and added cortical bone in the tibial shaft. After cessation of hPTH treatment, bone loss was observed both in trabecular and cortical bone; however, it was more dramatic on endocortical surfaces.

Low dose of human PTH(1-34) improved tibial subcortical bone mass without further cortical bone loss in adult intact beagles

Abstract: Cortical bone loss has not been observed in parathyroid hormone- (PTH-) treated rats, but clinical investigators suggest that it may augment cancellous bone mass at the expense of cortical bone in PTH-treated patients. In this study, the effects of PTH on cancellous and cortical bone mineral density of the tibia were studied in dogs with Haversian cortical bone remodeling by peripheral quantitative computed tomography (pQCT). Sixteen 19- to 20-month-old beagle dogs were randomized into four groups. In group 1, the vehicle control group, saline was injected daily throughout the experimental period. In group 2, the sequential group, 0.375 μg/kg of hPTH(1-34) was injected daily during the first 4 weeks, then stopped for 8 weeks, and this sequence was once repeated (on 4 weeks, off 8 weeks, and on 4 weeks, off 8 weeks). In group 3, the one-time group, the same dose of PTH was injected once per week for 24 weeks. In group 4, the three-time group, the same dose of PTH was given three times per week for 24 weeks. Peripheral QCT measurements were carried out 7 mm distal to the growth plate in the proximal metaphysis of the tibia. A voxel size of 0.295 mm and threshold for cortical and subcortical bone of 0.930 was chosen throughout the experiment. In adult beagles, group 4 had a significantly higher value in subcortical bone mineral density (SubCt.BMD) than control group (852 vs 771 mg/mm3; P < .05). Higher but non-significant increase occurred in the PTH-treated animals (groups 2, 3, 4) in the following parameters: total (Total.BMD), cancellous bone mineral density (Cn.BMD), cortical bone mineral density (Ct.BMD), and cortical thickness (Ct.Th). These findings show that a low dose of PTH improved subcortical bone mineral density without decreasing cortical bone mineral density during this dosage and period.

Human Parathyroid Hormone (1–34) Increases Cortical Bone Mass by Activating Bone Modeling in the Formation Mode in Ovariectomized Rats

The purpose of this study was to determine the efficacy of human parathyroid hormone, hPTH (1–34), in augmenting cortical bone mass in ovariectomized (OVX) growing rats. Thirty 11-week-old female Sprague-Dawley rats were divided into five groups of six animals each. Ovariectomy was performed on 18 rats and 12 rats were subjected to sham surgery. All rats were left untreated for the 4 weeks postsurgery. At the end of pretreatment period, groups of baseline sham (Group 1) and OVX (Group 2) rats were killed. The remaining rats were divided into three groups and each treated as follows. Sham-operated rats (Group 3) and ovariectomized rats (Group 4) were injected with saline vehicle. Ovariectomized rats (Group 5) were injected with hPTH (1–34), 30jLlg/kg, five times per week. All treatments were initiated at 4 weeks postsurgery for a 4-week period. To classify the effects of PTH on cortical bone modeling and remodeling during the treatment period, the PTH-treated group was injected subcutaneously with Oxytetracycline just prior to the initiation of the PTH treatment. All animals were double-labeled with subcutaneous injections of calcein on day 6 and day 2 before euthanization. Cross sections of the tibial diaphysis were subjected to quantitative bone histomorphometry. PTH treatment of OVX rats (Group 5) increased cortical bone area and width, tended to decrease the bone marrow area, and did not significantly increase the endocortical resorbing surface compared to the vehicle-treated OVX rats. PTH administration in the OVX rats increased cortical bone by the addition of new circumferential bone on both the periosteal and endocortical surfaces by stimulating osteoblastic recruitment; the newly formed bone was lamellar in nature. PTH administration activated cortical bone modeling in the formation mode (activation-formation) on either the periosteal or endocortical envelopes. The present data indicate that PTH stimulated cortical bone modeling in the formation mode and augmented cortical bone mass in the tibial diaphysis of OVX rats. These findings are in agreement with previous observations that dogs and humans respond similarly to PTH, suggesting that PTH administration may be a useful anabolic agent in the prevention and treatment of postmenopausal osteoporosis.

Maintenance of bone mineral density of femoral cortex in ovariectomized rats after withdrawal of concurrent administration of human parathyroid hormone (1-34) and incadronate disodium (YM175)

The aim of this study was to evaluate the potential use of a combination of human parathyroid hormone (1–34) [hPTH(1–34)] and bisphosphonate (incadronate disodium cycloheptylaminomethylenedisphosphonate monohydrate, YM175) as a therapy for osteoporosis. We examined the effects of concurrent administration of PTH and YM175 or single administration and the persistence of their therapeutic effect after withdrawal on bone mineral density (BMD) of the femur in ovariectomized rats with established osteopenia. One hundred and two 11-week-old Sprague-Dawley rats were divided into sham operation and ovariectomy (OVX) groups. OVX rats were untreated for the first 4 weeks post ovariectomy to allow for the development of moderate ovariectomy to allow for the development of moderate osteopenia. These animals were then subjected to various treatment regimens with either PTH, YM175, or both for 4 weeks. The animals were then killed at 4 or 12 weeks, after withdrawal of the treatment and the bone mineral density (BMD) of distal, middle, proximal part, and total area of the femur were determined by dual-energy X-ray absorptiometry (DXA). In the distal femur (cancellous bone-rich region), treatment with YM failed to restore BMD in OVX rats, while treatment with PTH alone (P<.01) or PTH + YM175 (P<.01) reversed BMD in OVX rats after 4 weeks of treatments. The restored distal BMD by PTH or PTH + YM175 treatments could be maintained thereafter until 12 weeks withdrawal. In midshaft of the femur (cortical bone-rich region), treatment with PTH (P<.05), YM175 (P<.05), and PTH + YM175 (P<.01) all could increase BMD after 4 weeks of treatments in the OVX rats, but only concurrent treatment with PTH + YM175 maintained the BMD of femoral midshaft for 12 weeks after withdrawal of the treatment. These results suggest that (1) concurrent treatment with PTH and YM175 could result in a bone gain not only in cancellous bone but also in cortical bone of the femur, and (2) the restored BMD could be maintained for 12 weeks after cessation of the treatment in cortical bone only by concurrent use of PTH + YM175 in immature ovariectomized rats