Bisphosphonates for treatment of osteoporosis: expected benefits, potential harms, and drug holidays.

OBJECTIVE: To outline the efficacy and risks of bisphosphonate therapy for the management of osteoporosis and describe which patients might be eligible for bisphosphonate "drug holiday." QUALITY OF EVIDENCE: MEDLINE (PubMed, through December 31, 2012) was used to identify relevant publications for inclusion. Most of the evidence cited is level II evidence (non-randomized, cohort, and other comparisons trials). MAIN MESSAGE: The antifracture efficacy of approved first-line bisphosphonates has been proven in randomized controlled clinical trials. However, with more extensive and prolonged clinical use of bisphosphonates, associations have been reported between their administration and the occurrence of rare, but serious, adverse events. Osteonecrosis of the jaw and atypical subtrochanteric and diaphyseal femur fractures might be related to the use of bisphosphonates in osteoporosis, but they are exceedingly rare and they often occur with other comorbidities or concomitant medication use. Drug holidays should only be considered in low-risk patients and in select patients at moderate risk of fracture after 3 to 5 years of therapy. CONCLUSION: When bisphosphonates are prescribed to patients at high risk of fracture, their antifracture benefits considerably outweigh their potential for harm. For patients taking bisphosphonates for 3 to 5 years, reassess the need for ongoing therapy.

Alterations in phosphorus, calcium and PTHrP contribute to defects in dental and dental alveolar bone formation in calcium-sensing receptor-deficient mice.

To determine whether the calcium-sensing receptor (CaR) participates in tooth formation and dental alveolar bone development in mandibles in vivo, we examined these processes, as well as mineralization, in 2-week-old CaR-knockout (CaR(-/-)) mice. We also attempted to rescue the phenotype of CaR(-/-) mice by genetic means, in mice doubly homozygous for CaR and 25-hydroxyvitamin D 1alpha-hydroxylase [1alpha(OH)ase] or parathyroid hormone (Pth). In CaR(-/-) mice, which exhibited hypercalcemia, hypophosphatemia and increased serum PTH, the volumes of teeth and of dental alveolar bone were decreased dramatically, whereas the ratio of the area of predentin to total dentin and the number and surface of osteoblasts in dental alveolar bone were increased significantly, as compared with wild-type littermates. The normocalcemia present in CaR(-/-);1alpha(OH)ase(-/-) mice only slightly improved the defects in dental and alveolar bone formation observed in the hypercalcemic CaR(-/-) mice. However, these defects were completely rescued by the additional elimination of hypophosphatemia and by an increase in parathyroid hormone-related protein (PTHrP) expression in the apical pulp, Hertwig's epithelial root sheath and mandibular tissue in CaR(-/-); Pth(-/-) mice. Therefore, alterations in calcium, phosphorus and PTHrP contribute to defects in the formation of teeth and alveolar bone in CaR-deficient mice. This study indicates that CaR participates in the formation of teeth and in the development of dental alveolar bone in mandibles in vivo, although it appears to do so largely indirectly.

Role of 1,25-dihydroxyvitamin D in alleviating alveolar bone loss and gingival inflammation in ligature-induced periodontitis.

OBJECTIVES: The goal of this study was to assess if endogenous 1,25(OH)2D deficiency enhanced, whereas exogenous 1,25(OH)2D3 supplementation alleviated alveolar bone loss and gingival inflammation induced by ligature-induced periodontitis. METHODS: A model of ligature-induced experimental periodontitis was generated in wild-type (WT) and Cyp27b1-knockout (KO) mice on a rescue diet (RD), and un-ligated genotype-matched littermates as control, or in WT mice on a normal diet (ND) with vehicle treatment or 1,25(OH)2D3 treatment, and un-ligated WT littermates as control. Alveolar bone mass and turnover, T cell infiltration and inflammatory cytokines in gingival tissues were examined. RESULTS: In WT mice, ligature-induced alveolar bone loss occurred by inhibiting alveolar bone formation. This was characterized by reduction of osteoblast numbers, alkaline phosphatase activity and type I collagen synthesis, as well as by augmentation of osteoclastic alveolar bone resorption and gingival inflammation, including increases of osteoclast numbers, inflammatory positive cells and up-regulation of mRNA expression levels of inflammatory cytokines. Alveolar bone destruction and gingival inflammation were more severe in diet-matched Cyp27b1-KO mice than in WT littermates on RD. Supplementation of exogenous 1,25(OH)2D3 alleviated alveolar bone loss and gingival inflammation in ligated WT mice on ND, but those parameters did not reach levels observed in un-ligated WT ones. CONCLUSIONS: Endogenous 1,25(OH)2D deficiency enhanced, whereas exogenous 1,25(OH)2D3 supplementation alleviated alveolar bone loss and gingival inflammation induced by ligature-induced periodontitis.

Fibroblast growth factor 23 overexpression impacts negatively on dentin mineralization and dentinogenesis in mice.

1. Though previous studies have shown that fibroblast growth factor 23 (FGF23) mRNA expression localizes in ameloblasts and odontoblasts in teeth, it is unclear what effect FGF23 overexpression has on dentin mineralization and dentinogenesis. Toward this end, the phenotypes of mandibles and teeth were compared between 6-week-old FGF23 transgenic mice and their wild-type littermates by radiography, microcomputed tomography scanning, histology, histochemistry and immunohistochemistry. 2. The mineral density was reduced in all teeth, including molars and incisors, and in the mandible, and the mineralized tooth volume in incisor and molars, and the mineralized cortical and alveolar bone volume in mandibles were decreased in FGF23 transgenic mice compared with their wild-type littermates. The dental volume, reparative dentin area, the expression of dentin sialoprotein in dentin, and the deposition of type I collagen and osteocalcin in the dental matrix were significantly reduced. However, the predentin volume and the expression of biglycan in dentin were increased in FGF23 transgenic mice compared with their wild-type littermates. 3. The results of the present study show that FGF23 overexpression plays a negative regulatory role on dentin mineralization and dentinogenesis.

Probing the Scope and Mechanisms of Calcitriol Actions Using Genetically Modified Mouse Models.

Genetically modified mice have provided novel insights into the mechanisms of activation and inactivation of vitamin D, and in the process have provided phenocopies of acquired human disease such as rickets and osteomalacia and inherited diseases such as pseudovitamin D deficiency rickets, hereditary vitamin D resistant rickets, and idiopathic infantile hypercalcemia. Both global and tissue-specific deletion studies leading to decreases of the active form of vitamin D, calcitriol [1,25(OH)2D], and/or of the vitamin D receptor (VDR), have demonstrated the primary role of calcitriol and VDR in bone, cartilage and tooth development and in the regulation of mineral metabolism and of parathyroid hormone (PTH) and FGF23, which modulate calcium and phosphate fluxes. They have also, however, extended the spectrum of actions of calcitriol and the VDR to include, among others: modulation, jointly and independently, of skin metabolism; joint regulation of adipose tissue metabolism; cardiovascular function; and immune function. Genetic studies in older mice have also shed light on the molecular mechanisms underlying the important role of the calcitriol/VDR pathway in diseases of aging such as osteoporosis and cancer. In the course of these studies in diverse tissues, important upstream and downstream, often tissue-selective, pathways have been illuminated, and intracrine, as well as endocrine actions have been described. Human studies to date have focused on acquired or genetic deficiencies of the prohormone vitamin D or the (generally inactive) precursor metabolite 25-hyrodxyvitamin D, but have yet to probe the pleiotropic aspects of deficiency of the active form of vitamin D, calcitriol, in human disease. © 2020 American Society for Bone and Mineral Research © 2020 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

SIRT1/FOXO3a axis plays an important role in the prevention of mandibular bone loss induced by 1,25(OH)2D deficiency.

It has been reported that 1,25 dihydroxyvitamin D [1,25(OH)2D] deficiency leads to the loss of mandibular bone, however the mechanism is unclear. We investigated whether the Sirt1/FOXO3a signaling pathway is involved in this process. Using a 1,25(OH)2D deficiency model induced by genetic deletion in mice of 25-hydroxyvitamin D-1α hydroxylase [1α(OH)ase-/- mice]. We first documented a sharp reduction of expression levels of Sirt1 in the 1α(OH)ase-/- mice in vivo. Next, we demonstrated dose-dependent upregulation of Sirt1 by treatment with exogenous 1,25(OH)2D3in vitro. We then identified a functional VDR binding site in the Sirt1 promoter. By crossing Prx1-Sirt1 transgenic mice with 1α(OH)ase-/- mice we demonstrated that the overexpression of Sirt1 in mesenchymal stem cells (MSCs) greatly improved the 1α(OH)ase-/- mandibular bone loss phenotype by increasing osteoblastic bone formation and reducing osteoclastic bone resorption. In mechanistic studies, we showed, in 1α(OH)ase-/- mice, decreases of Sirt1 and FoxO3a, an increase in oxidative stress as reflected by a reduction of the antioxidant enzymes peroxiredoxin1 (Prdx1), SOD1 and SOD2 expression, and an increase of markers for osteocyte senescence and senescence associated secretory phenotypes (SASP), including ß-galactosidase (ß-gal), p16, p53 and p21. The targeted overexpression of Sirt1 in the 1α(OH)ase-/- mice restored the expression levels of these molecules. Finally, we demonstrated that a Sirt1 agonist can upregulate FOXO3a activity by increasing deacetylation and nuclear translocation. Overall, results from this study support the concept that targeted increases in Sirt1/FOXO3a signaling levels can greatly improve the bone loss caused by 1,25(OH)2D deficiency.

Evolution of Our Understanding of the Hyperparathyroid Syndromes: A Historical Perspective.

We review advancing and overlapping stages for our understanding of the expressions of six hyperparathyroid (HPT) syndromes: multiple endocrine neoplasia type 1 (MEN1) or type 4, multiple endocrine neoplasia type 2A (MEN2A), hyperparathyroidism-jaw tumor syndrome, familial hypocalciuric hypercalcemia, neonatal severe primary hyperparathyroidism, and familial isolated hyperparathyroidism. During stage 1 (1903 to 1967), the introduction of robust measurement of serum calcium was a milestone that uncovered hypercalcemia as the first sign of dysfunction in many HPT subjects, and inheritability was reported in each syndrome. The earliest reports of HPT syndromes were biased toward severe or striking manifestations. During stage 2 (1959 to 1985), the early formulations of a syndrome were improved. Radioimmunoassays (parathyroid hormone [PTH], gastrin, insulin, prolactin, calcitonin) were breakthroughs. They could identify a syndrome carrier, indicate an emerging tumor, characterize a tumor, or monitor a tumor. During stage 3 (1981 to 2006), the assembly of many cases enabled recognition of further details. For example, hormone non-secreting skin lesions were discovered in MEN1 and MEN2A. During stage 4 (1985 to the present), new genomic tools were a revolution for gene identification. Four principal genes ("principal" implies mutated or deleted in 50% or more probands for its syndrome) (MEN1, RET, CASR, CDC73) were identified for five syndromes. During stage 5 (1993 to the present), seven syndromal genes other than a principal gene were identified (CDKN1B, CDKN2B, CDKN2C, CDKN1A, GNA11, AP2S1, GCM2). Identification of AP2S1 and GCM2 became possible because of whole-exome sequencing. During stages 4 and 5, the newly identified genes enabled many studies, including robust assignment of the carriers and non-carriers of a mutation. Furthermore, molecular pathways of RET and the calcium-sensing receptor were elaborated, thereby facilitating developments in pharmacotherapy. Current findings hold the promise that more genes for HPT syndromes will be identified and studied in the near future. © 2018 American Society for Bone and Mineral Research.

1,25-dihydroxyvitamin D deficiency accelerates alveolar bone loss independent of aging and extracellular calcium and phosphorus.

BACKGROUND: Vitamin D is critical for bone homeostasis and immunomodulation. We therefore assessed whether 1,25-dihydroxyvitamin D (1,25(OH)2 D) deficiency in mice with targeted deletion of the gene encoding 25-hydroxyvitamin D-1α-hydroxylase (1α(OH)ase [1αOH)ase-/- mice]) results in alveolar bone loss and periodontal inflammation in vivo. METHODS: Ten-week-old and 12-month-old 1α(OH)ase-/- mice and wild-type littermates were fed a normal diet or a rescue diet, and the phenotype of the periodontium was then analyzed using microcomputed tomography, histology, immunohistochemistry, and real-time Reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Alveolar bone loss was increased and maxillary bone mineral density (BMD), osteoblast numbers, and the number of osterix-positive cells were decreased significantly in 1α(OH)ase-/- mice compared with wild-type mice. Although aging from 10 weeks to 12 months accentuated these changes, and a rescue diet reduced them, the alterations in the 1α(OH)ase-/- mice exceeded the effects of aging and diet change. Nuclear factor kappa light-chain-enhancer of activated B cells (NF-кB) p65 and CD3 positive cells, and the gene expression levels of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, matrix metalloproteinase (MMP)-3 and -8 were all increased significantly in periodontal tissues of 1α(OH)ase-/- mice compared with wild-type mice. Aging from 10 weeks to 12 months also accentuated these changes, and a rescue diet reduced them, however, the alterations in the 1α(OH)ase-/- mice exceeded the effects of aging and diet change. CONCLUSION: 1,25(OH)2 D deficiency in the 1α(OH)ase-/- mice accelerated alveolar bone loss by inhibiting osteoblastic bone formation and enhancing periodontal tissue degeneration in a calcium- and phosphorus- as well as an age-independent manner.

PTHrP Nuclear Localization and Carboxyl Terminus Sequences Modulate Dental and Mandibular Development in Part via the Action of p27.

To determine whether the action of the PTHrP nuclear localization sequence and C terminus is mediated through p27 in modulating dental and mandibular development, compound mutant mice, which are homozygous for both p27 deletion and the PTHrP1-84 knock-in mutation (p27(-/-)Pthrp(KI/KI)), were generated. Their teeth and mandibular phenotypes were compared with those of p27(-/-), Pthrp(KI/KI), and wild-type mice. At 2 weeks of age, the mandibular mineral density, alveolar bone volume, osteoblast numbers, and dental volume, dentin sialoprotein-immunopositive areas in the first molar were increased significantly in p27(-/-) mice and decreased dramatically in both Pthrp(KI/KI) and p27(-/-) Pthrp(KI/KI) mice compared with wild-type mice; however, these parameters were partly rescued in p27(-/-) Pthrp(KI/KI) mice compared with Pthrp(KI/KI) mice. These data demonstrate that the deletion of p27 in Pthrp(KI/KI) mice can partially rescue defects in dental and mandibular development. Furthermore, we found that deletion of p27 in Pthrp(KI/KI) mice partially corrected the dental and mandibular phenotype by modulating cell cyclin-regulating molecules and antioxidant enzymes. This study therefore indicates that the p27 pathway may function downstream in the action of PTHrP nuclear localization sequence to regulate dental and mandibular development.

Partial rescue of the Hyp phenotype by osteoblast-targeted PHEX (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) expression.

Inactivating mutations and/or deletions of PHEX/Phex (phosphate-regulating gene with homologies to endopeptidases on the X chromosome) are responsible for X-linked hypophosphatemic rickets in humans and in the murine homolog Hyp. The predominant osteoblastic expression of Phex has implicated a primary metabolic osteoblast defect in the pathophysiology of this disorder. By targeting PHEX expression to osteoblasts in the Hyp genetic background, we aimed to correct the corresponding biochemical and morphological abnormalities and obtain information on their pathogenetic mechanism. When transgene Phex expression, driven by a mouse pro-alpha1(I) collagen gene promoter, was crossed into the Hyp background, it improved the defective mineralization of bone and teeth but failed to correct the hypophosphatemia and altered vitamin D metabolism associated with the disorder. Ex vivo bone marrow cultures confirmed the amelioration in the Hyp-associated matrix mineralization defect after Phex expression. These findings suggest that while the Hyp bone and teeth abnormalities partially correct after PHEX gene transfer, additional factors and/or sites of PHEX expression are likely critical for the elaboration of the appropriate molecular signals that alter renal phosphate handling and vitamin D metabolism in this disorder.

1,25(OH)2D deficiency induces temporomandibular joint osteoarthritis via secretion of senescence-associated inflammatory cytokines.

1,25-Dihydroxyvitamin D [1,25(OH)(2)D] insufficiency appears to be associated with several age-related diseases. Insufficient levels of serum 25-hydroxyvitamin D has been shown to lead to the progression of osteoarthritis (OA) while underlying biological mechanisms remain largely unknown. In this study, we sought to determine whether 1,25(OH)(2)D deficiency has a direct effect on the process of murine temporomandibular joint (TMJ) OA in 25-hydroxyvitamin D 1α-hydroxylase knockout [1α(OH)ase(-/-)] mice that had been fed a rescue diet (high calcium, phosphate, and lactose) from weaning until 6 or 18 months of age. Our results showed that the bone mineral density and subchondral bone volume were reduced in mandibular condyles, articular surfaces were collapsed, the thickness of articular cartilage and cartilage matrix protein abundance were progressively decreased and eventually led to an erosion of articular cartilage of mandibular condyles. We also found that DNA damage, cellular senescence and the production of senescence-associated inflammatory cytokines were increased significantly in 1α(OH)ase(-/-) mice. This study demonstrates that 1,25(OH)(2)D deficiency causes an erosive TMJ OA phenotype by inducing DNA damage, cellular senescence and the production of senescence-associated inflammatory cytokines. Our results indicate that 1,25(OH)(2)D plays an important role in preventing the development and progression of OA.

An efficient paradigm for genetic epidemiology cohort creation.

Development of novel methodologies to efficiently create large genetic epidemiology cohorts is needed. Here we describe a rapid, precise and cost-efficient method for collection of DNA from cases previously experiencing an osteoporotic fracture by identifying cases using and administrative health-care databases. Over the course of 14 months we collected DNA from 1,130 women experiencing an osteoporotic fracture, at a cost of $54 per sample. This cohort is among the larger DNA osteoporotic fracture collections in the world. The novel method described addresses a major unmet health care research need and is widely applicable to any disease that can be identified accurately through administrative data.

Bone formation regulates circulating concentrations of fibroblast growth factor 23.

We examined the role of bone remodeling in the regulation of circulating concentrations of FGF23 using mouse models manifesting differing degrees of coupled and uncoupled bone turnover. Administration of the antiresorptive agent osteoprotegerin produced a profound reduction in bone resorption and formation in male and oophorectomized female mice, accompanied by an increase in serum levels of fibroblast growth factor 23 (FGF23) and a reduction in circulating 1,25-dihydroxyvitamin D [1,25(OH)(2)D]. In contrast, exogenous PTH(1-34) administration increased bone turnover and reduced circulating FGF23. In 1,25(OH)(2)D-deficient, 25-hydroxyvitamin D 1alpha-hydroxylase null mice on a high-calcium diet, endogenous PTH was elevated, bone formation but not resorption was increased, and serum FGF23 was virtually undetectable; on a rescue diet, serum calcium was normalized, PTH levels were reduced, bone formation was reduced, and serum FGF23 levels increased. After PTH treatment of wild-type mice, gene expression of dentin matrix protein 1 (DMP1) in bone was increased, whereas gene expression of FGF23 was reduced. In vitro studies in the osteoblastic cell line UMR-106 showed that externally added DMP1 could inhibit FGF23 gene expression and production stimulated by 1,25(OH)(2)D(3). The results show that osteoblastic bone formation is a potent modulator of FGF23 production and release into the circulation, suggest that the biological consequences on mineral homeostasis of circulating FGF23 may also be dependent on the prevailing rate of bone turnover, and provide evidence that DMP1 may be a direct negative regulator of FGF23 production in osteoblastic cells.

Distinctive anabolic roles of 1,25-dihydroxyvitamin D(3) and parathyroid hormone in teeth and mandible versus long bones.

To assess the roles of 1,25-dihydroxyvitamin D (1,25(OH)(2)D) and parathyroid hormone (PTH) in hard tissue formation in oro-facial tissues, we examined the effect of either 1,25(OH)(2)D or PTH deficiency on dentin and dental alveolar bone formation and mineralization in the mandibles, and osteoblastic bone formation in long bones of 1alpha-hydroxylase knockout (1alpha(OH)ase(-/-)) mice. Compared with wild-type mice, the mineral density was decreased in the teeth and mandibles, and unmineralized dentin (predentin and biglycan immunopositive dentin) and unmineralized bone matrix in the dental alveolar bone were increased in 1alpha(OH)ase(-/-) mice. The dental volume, reparative dentin volume, and dentin sialoprotein immunopositive areas were reduced in 1alpha(OH)ase(-/-) mice. The cortical thickness, dental alveolar bone volume, and osteoblast number were all decreased significantly in the mandibles; in contrast, the osteoblast number and surface were increased in the trabecular bone of the tibiae in 1alpha(OH)ase(-/-) mice consistent with their secondary hyperparathyroidism. The expression of PTH receptor and IGF1 was reduced slightly in mandibles, but enhanced significantly in the long bones in the 1alpha(OH)ase(-/-) mice. To control for the role of secondary hyperparathyroidism, we also examined teeth and mandibles in 6-week-old PTH(-/-) mice. In these animals, dental and bone volumes in mandibles were not altered when compared with their wild-type littermates. These results suggest that 1,25(OH)(2)D(3) plays an anabolic role in both dentin and dental alveolar bone as it does in long bones, whereas PTH acts predominantly in long bones rather than mandibular bone.