This study examined the effect of 2 yr of treatment with the aminobisphosphonate alendronate (ALN) (0.05 or 0.25 mg/ kg i.v. ALN every 2 wk) on estrogen deficiency bone loss and bone strength changes in ovariectomized (OVX) baboons (n = 7 per group) and the ALN mode of action at the tissue level. Biochemical markers of bone turnover increased in OVX animals and were maintained by ALN treatment at non-OVX levels (low dose) or below (high dose). 2 yr of treatment produced no cumulative effects on bone turnover markers. Histomorphometry showed a marked increase in cancellous bone remodeling in OVX animals. Activation frequency increased from 0.48 to 0.86 per yr (L5 vertebra), and the osteoid surfaces from 9 to 13.5% (P < 0.05). No changes were observed in eroded and osteoclast surfaces. ALN treatment decreased activation frequency and indices of bone formation to control levels (low dose) or below (high dose), did not change indices of mineralization, and increased bone mineral density (BMD) in the lumbar vertebrae (L2-L4) by 15% at 0.25 mg/kg (P < 0.05), relative to vehicle-treated animals. The mean strength of cancellous bone (LA) increased by 44% (low ALN dose) and 100% (high dose), compared with vehicle. The strength of individual bones correlated with the square of the L2-L4 BMD (r = 0.91, P < 0.0034). In conclusion, ALN treatment reversed the effects of ovariectomy on cancellous bone turnover and increased bone mass and bone strength in baboons. (J. Clin. Invest. 1993. 92:2577-2586 Key words: osteoporosis * alendronate * histomorphometry-bone strength * bone mineral density

Bone morphogenetic proteins (BMPs) function during various aspects of embryonic development including skeletogenesis. However, their biological functions after birth are less understood. To investigate the role of BMPs during bone remodeling, we generated a postnatal osteoblast-specific disruption of Bmpr1a that encodes the type IA receptor for BMPs in mice. Mutant mice were smaller than controls up to 6 months after birth. Irregular calcification and low bone mass were observed, but there were normal numbers of osteoblasts. The ability of the mutant osteoblasts to form mineralized nodules in culture was severely reduced. Interestingly, bone mass was increased in aged mutant mice due to reduced bone resorption evidenced by reduced bone turnover. The mutant mice lost more bone after ovariectomy likely resulting from decreased osteoblast function which could not overcome ovariectomyinduced bone resorption. In organ culture of bones from aged mice, ablation of the Bmpr1a gene by adenoviral Cre recombinase abolished the stimulatory effects of BMP4 on the expression of lysosomal enzymes essential for osteoclastic bone resorption. These results demonstrate essential and age-dependent roles for BMP signaling mediated by BMPRIA (a type IA receptor for BMP) in osteoblasts for bone remodeling.Bone formation is a well characterized process; however, little is known about the molecular mechanisms that regulate bone remodeling, the physiological process through which bone mass is maintained constant. Remodeling consists of two distinct phases: initial bone resorption by the osteoclasts, followed by de novo bone formation by the osteoblasts (1). Differentiated osteoblasts are the only cells responsible for bone formation. Bone formation is thought to be regulated by hormones and by locally acting growth factors (2). Bone morphogenetic proteins (BMPs) 1 are secreted molecules and members of transforming growth factor-␤ superfamily (3, 4). They were discovered by their ectopic bone formation activity when implanted locally in soft tissues (5). Over the past decade, the phenotypes of mice with mutations in genes coding for this group of proteins and their receptors uncovered the essential roles for BMPs in wide variety of developmental processes, including skeletal development and patterning (6 -9). However, despite its powerful ability to induce ectopic osteogenesis, the essential role of BMPs in bone formation and bone metabolism in the adult skeleton has not been established (10) because of embryonic lethality resulting from mutations of genes encoding the most potent BMPs for bone formation, BMP2 and BMP4, and their receptors (11-13). We previously generated a null allele for Bmpr1a that encodes a type IA receptor for BMP (BMPRIA or ALK3). Mice homozygous for this null allele died by embryonic day 8.0 (E8.0) without mesoderm formation (13). Bmpr1a is expressed in most tissues throughout development and after birth (13,14). Expression of a dominant-negative form of BMPRIA in a cultured cell line or chick limb buds suggests ...

Estrogen deficiency in mammals is known to increase bone turnover and result in reduced bone mass. The bisphosphonate, 4-amino-1-hydroxybutylidene-1,1-bisphophonic acid disodium salt, alendronate (MK-217), is a potent inhibitor of bone resorption and was evaluated in this study for its ability to inhibit bone loss following ovariectomy in rats. Alendronate was administered sc in doses of 0.0, 0.056, 0.28, 1.40, and 7.0 mg P/kg/month, divided into two, four, or eight monthly subcutaneous injections for each dose, to female Sprague-Dawley rats (250-280 g) that underwent bilateral ovariectomy. Rats were sacrificed 12 weeks postovariectomy, the femora ashed, and the tibiae prepared for static and dynamic histomorphometric analyses. Femoral bone mass in vehicle-treated rats was reduced by 12% 12 weeks after ovariectomy compared to the nonovariectomized control group. In MK-217-treated rats femoral bone mass was significantly increased in a dose-dependent manner compared to either ovariectomized or nonovariectomized controls. Histomorphometric analysis showed significant increases in tibial trabecular bone volume with no decrease in osteoclast number. Doses delivered twice per month or eight times per month were equally effective in achieving the peak bone volume 12 weeks after ovariectomy. In conclusion, alendronate (MK-217) was effective in inhibiting bone loss due to estrogen deficiency in rats, and the magnitude of its effect was related primarily to the total amount of compound administered rather than the frequency of its administration.

To examine the effect of alendronate (4-amino-1-hydroxybutylidene bisphosphonate) on fracture repair, the drug was given to mature beagle dogs orogastrically at 2 mg/kg/day for 9 weeks preceding fracture. 16 weeks after fracture, or both before and after fracture (25 weeks). A transverse mid-diaphyseal fracture of the right radius was surgically induced and was stabilized by external coaptation splinting. Fracture healing and bone remodeling were evaluated by radiography, gross and histological examination, and bone histomorphometry. The mechanical properties of the fracture callus were determined by a four-point bending test. Radiographs and gross and microscopic examination demonstrated normal bone healing at the fracture site in all dogs. In dogs that received alendronate during the fracture healing period, at 16 weeks the calluses were approximately 2-3 times larger than those in dogs that received a placebo during the healing period. This is consistent with slower callus bone remodeling, an expected pharmacological effect of the compound. Bone histomorphometry demonstrated that treatment with alendronate did not inhibit bone formation or mineralization. Mechanical testing showed that the ultimate load at failure and the flexural rigidity of both the fractured and contralateral intact bone were unaffected by treatment with alendronate. Therefore, in this study, treatment with alendronate before or during fracture healing, or both, resulted in no adverse effects on the union, strength, or mineralization of bone in mature beagle dogs.