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Objectives: This study aimed to elucidate whether the administration of parathyroid hormone (PTH) results in remodeling- or modeling-based bone formation in different regions of the murine femora, and whether the PTH-driven bone formation would facilitate osteoblastic differentiation into osteocytes., Methods: Six-week-old male C57BL/6J mice were employed to examine the distribution of alkaline phosphatase (ALP), PHOSPHO1, podoplanin, and calcein labeling in two distinct long bone regions: the metaphyseal trabeculae close to the chondro-osseous junction (COJ) and those distant from the COJ in three mouse groups, a control group receiving a vehicle (sham group) and groups receiving hPTH (1-34) twice a day (PTH BID group) or four times a day (PTH QID group) for two weeks., Results: The sham group showed PHOSPHO1-reactive mature osteoblasts localized primarily at the COJ, whereas the PTH BID/QID groups exhibited extended lines of PHOSPHO1-reactive osteoblasts even in regions distant from the COJ. The PTH QID group displayed fragmented calcein labeling in trabeculae close to the COJ, whereas continuous labeling was observed in trabeculae distant from the COJ. Osteoblasts tended to express podoplanin and PHOSPHO1 independently in the close and distant regions of the sham group, while osteoblasts in the PTH-administered groups showed immunoreactivity of podoplanin and PHOSPHO1 together in the close and distant regions., Conclusions: Administration of PTH may accelerate remodeling-based bone formation in regions close to the COJ while predominantly inducing modeling-based bone formation in distant regions. PTH appeared to simultaneously facilitate osteoblastic bone mineralization and differentiation into osteocytes in both remodeling- and modeling-based bone formation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Japanese Association for Oral Biology. Published by Elsevier B.V. All rights reserved.)

Teriparatide is a peptide derived from a parathyroid hormone (PTH) and an osteoporosis therapeutic drug with potent bone formation-promoting activity. To identify novel druggable genes that act downstream of PTH signaling and are potentially involved in bone formation, we screened PTH target genes in mouse osteoblast-like MC3T3-E1 cells. Here we show that Gprc5a, encoding an orphan G protein-coupled receptor, is a novel PTH-inducible gene and negatively regulates osteoblast proliferation and differentiation. PTH treatment induced Gprc5a expression in MC3T3-E1 cells, rat osteosarcoma ROS17/2.8 cells, and mouse femurs. Induction of Gprc5a expression by PTH occurred in the absence of protein synthesis and was mediated primarily via the cAMP pathway, suggesting that Gprc5a is a direct target of PTH signaling. Interestingly, Gprc5a expression was induced additively by co-treatment with PTH and 1α, 25-dihydroxyvitamin D3 (calcitriol), or retinoic acid in MC3T3-E1 cells. Reporter analysis of a 1 kb fragment of human GPRC5A promoter revealed that the promoter fragment showed responsiveness to PTH via the cAMP response element, suggesting that GPRC5A is also a PTH-inducible gene in humans. Gprc5a knockdown promoted cell viability and proliferation, as demonstrated by MTT and BrdU assays. Gprc5a knockdown also promoted osteoblast differentiation, as indicated by gene expression analysis and mineralization assay. Mechanistic studies showed that Gprc5a interacted with BMPR1A and suppressed BMP signaling induced by BMP-2 and constitutively active BMP receptors, ALK2 (ACVR1) Q207D and ALK3 (BMPR1A) Q233D. Thus, our results suggest that Gprc5a is a novel gene induced by PTH that acts in an inhibitory manner on both cell proliferation and osteoblast differentiation and is a candidate for drug targets for osteoporosis., (© 2024 Wiley Periodicals LLC.)

Chronic kidney disease-induced secondary hyperparathyroidism (CKD-SHPT) heightens fracture risk through impaired mineral homeostasis and elevated levels of uremic toxins (UTs), which in turn enhance bone remodeling. Etelcalcetide (Etel), a calcium-sensing receptor (CaSR) agonist, suppresses parathyroid hormone (PTH) in hyperparathyroidism to reduce excessive bone resorption, leading to increased bone mass. However, Etel's effect on bone quality, chemical composition, and strength is not well understood. To address these gaps, we established a CKD-SHPT rat model and administered Etel at a human-equivalent dose concurrently with disease induction. The effects on bone and mineral homeostasis were compared with a CKD-SHPT (vehicle-treated group) and a control group (rats without SHPT). Compared with vehicle-treated CKD-SHPT rats, Etel treatment improved renal function, reduced circulating UT levels, improved mineral homeostasis parameters, decreased PTH levels, and prevented mineralization defects. The upregulation of mineralization-promoting genes by Etel in CKD-SHPT rats might explain its ability to prevent mineralization defects. Etel preserved both trabecular and cortical bones with attendant suppression of osteoclast function, besides increasing mineralization. Etel maintained the number of viable osteocytes to the control level, which could also contribute to its beneficial effects on bone. CKD-SHPT rats displayed increased carbonate substitution of matrix and mineral, decreased crystallinity, mineral-to-matrix ratio, and collagen maturity, and these changes were mitigated by Etel. Further, Etel treatment prevented CKD-SHPT-induced deterioration in bone strength and mechanical behavior. Based on these findings, we conclude that in CKD-SHPT rats, Etel has multiscale beneficial effects on bone that involve remodeling suppression, mineralization gene upregulation, and preservation of osteocytes., Competing Interests: Declaration of competing interest There is no known conflict of interest related to this research work and no competing financial interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Glucocorticoids (GC) and parathyroid hormone (PTH) are widely used therapeutic endocrine hormones where their effects on bone and joint arise from actions on multiple skeletal cell types. In osteocytes, GC and PTH exert opposing effects on perilacunar canalicular remodeling (PLR). Suppressed PLR can impair bone quality and joint homeostasis, including in GC-induced osteonecrosis. However, combined effects of GC and PTH on PLR are unknown. Given the untapped potential to target osteocytes to improve skeletal health, this study sought to test the feasibility of therapeutically mitigating PLR suppression. Focusing on subchondral bone and joint homeostasis, we hypothesize that PTH(1-34), a PLR agonist, could rescue GC-suppressed PLR. The skeletal effects of GC and PTH(1-34), alone or combined, were examined in male and female mice by micro-computed tomography, mechanical testing, histology, and gene expression analysis. For each outcome, females were more responsive to GC and PTH(1-34) than males. GC and PTH(1-34) exerted regional differences, with GC increasing trabecular bone volume but reducing cortical bone thickness, stiffness, and ultimate force. Despite PTH(1-34)'s anabolic effects on trabecular bone, it did not rescue GC's catabolic effects on cortical bone. Likewise, cartilage integrity and subchondral bone apoptosis, tartrate-resistant acid phosphatase (TRAP) activity, and osteocyte lacunocanalicular networks showed no evidence that PTH(1-34) could offset GC-dependent effects. Rather, GC and PTH(1-34) each increased cortical bone gene expression implicated in bone resorption by osteoclasts and osteocytes, including Acp5, Mmp13, Atp6v0d2, Ctsk , differences maintained when GC and PTH(1-34) were combined. Since PTH(1-34) is insufficient to rescue GC's effects on young female mouse bone, future studies are needed to determine if osteocyte PLR suppression, due to GC, aging, or other factors, can be offset by a PLR agonist., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Yee, Meliadis, Kaya, Chang and Alliston.)

In this study, we used a bioinformatic approach to construct a miRNA-target gene interaction network potentially involved in the anabolic effect of parathyroid hormone analogue teriparatide [PTH (1-34)] on osteoblasts. We extracted a dataset of 26 microRNAs (miRNAs) from previously published studies and predicted miRNA target interactions (MTIs) using four software tools: DIANA, miRWalk, miRDB, and TargetScan. By constructing an interactome of PTH-regulated miRNAs and their predicted target genes, we elucidated signaling pathways regulating pluripotency of stem cells, the Hippo signaling pathway, and the TGF-beta signaling pathway as the most significant pathways in the effects of PTH on osteoblasts. Furthermore, we constructed intersection of MTI networks for these three pathways and added validated interactions. There are 8 genes present in all three selected pathways and a set of 18 miRNAs are predicted to target these genes, according to literature data. The most important genes in all three pathways were BMPR1A , BMPR2 and SMAD2 having the most interactions with miRNAs. Among these miRNAs, only miR-146a-5p and miR-346 have validated interactions in these pathways and were shown to be important regulators of these pathways. In addition, we also propose miR-551b-5p and miR-338-5p for further experimental validation, as they have been predicted to target important genes in these pathways but none of their target interactions have yet been verified. Our wet-lab experiment on miRNAs differentially expressed between PTH (1-34) treated and untreated mesenchymal stem cells supports miR-186-5p from the literature obtained data as another prominent miRNA. The meticulous selection of miRNAs outlined will significantly support and guide future research aimed at discovering and understanding the crucial pathways of osteoanabolic PTH-epigenetic effects on osteoblasts. Additionally, they hold potential for the discovery of new PTH target genes, innovative biomarkers for the effectiveness and safety of osteoporosis-affected treatment, as well as novel therapeutic targets., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Vrščaj, Marc and Ostanek.)

Introduction: Creating induced pluripotent stem cells (iPSCs) from somatic cells of patients with genetic diseases offers a pathway to generate disease-specific iPSCs carrying genetic markers. Differentiating these iPSCs into renal tubular cells can aid in understanding the pathophysiology of rare inherited renal tubular diseases through cellular experiments., Materials and Methods: Two Japanese patients with Pseudohypoparathyroidism (PHP), a 49-year-old woman and a 71-year-old man, were studied. iPSC-derived tubular cells were established from their peripheral blood mononuclear cells (PBMCs). We examined changes in intracellular and extracellular cyclic adenosine monophosphate (cAMP) levels in these cells in response to parathyroid hormone (PTH) stimulation., Results: Renal tubular cells, differentiated from iPSCs of a healthy control (648A1), showed a PTH-dependent increase in both intracellular and extracellular cAMP levels. However, the renal tubular cells derived from the PHP patients' iPSCs showed inconsistent changes in cAMP levels upon PTH exposure., Conclusion: We successfully created disease-specific iPSCs from PHP patients' PBMCs, differentiated them into tubular cells, and replicated the distinctive response of the disease to PTH in vitro. This approach could enhance our understanding of the pathophysiology of inherited renal tubular diseases and contribute to developing effective treatments., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Calcitriol and calcimimetics are used to treat hyperparathyroidism secondary to chronic kidney disease (CKD). Calcitriol administration and the subsequent increase in serum calcium concentration decrease parathyroid hormone (PTH) levels, which should reduce bone remodeling. We have previously reported that, when maintaining a given concentration of PTH, the addition of calcimimetics is associated with an increased bone cell activity. Whether calcitriol administration affects bone cell activity while PTH is maintained constant should be evaluated in an animal model of renal osteodystrophy. The aim of the present study was to compare in CKD PTH-clamped rats the bone effects of calcitriol and calcimimetic administration. The results show that the administration of calcitriol and calcimimetic at doses that induced a similar reduction in PTH secretion produced dissimilar effects on osteoblast activity in 5/6 nephrectomized (Nx) rats with secondary hyperparathyroidism and in Nx rats with clamped PTH. Remarkably, in both rat models, the administration of calcitriol decreased osteoblastic activity, whereas calcimimetic increased bone cell activity. In vitro, calcitriol supplementation inhibited nuclear translocation of β-catenin and reduced proliferation, osteogenesis, and mineralization in mesenchymal stem cells differentiated into osteoblasts. In conclusion, besides the action of calcitriol and calcimimetics at parathyroid level, these treatments have specific effects on bone cells that are independent of the PTH level., (© 2024 The Author(s). The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Objective: To investigate the effect of intermittent parathyroid hormone (iPTH) administration on pathological new bone formation during treatment of ankylosing spondylitis-related osteoporosis., Methods: Animal models with pathological bone formation caused by hypothetical AS pathogenesis received treatment with iPTH. We determined the effects of iPTH on bone loss and the formation of pathological new bone with micro-computed tomography (micro-CT) and histological examination. In addition, the tamoxifen-inducible conditional knockout mice (CAGGCre-ER TM ; PTH flox/flox , PTH-/-) was established to delete PTH and investigate the effect of endogenous PTH on pathological new bone formation., Results: iPTH treatment significantly improved trabecular bone mass in the modified collagen-induced arthritis (m-CIA) model and unbalanced mechanical loading models. Meanwhile, iPTH treatment did not enhance pathological new bone formation in all types of animal models. Endogenous PTH deficiency had no effects on pathological new bone formation in unbalanced mechanical loading models., Conclusion: Experimental animal models of AS treated with iPTH show improvement in trabecular bone density, but not entheseal pathological bone formation，indicating it may be a potential treatment for inflammatory bone loss does in AS., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Parathyroid hormone (PTH) serves dual roles in bone metabolism, exhibiting both anabolic and catabolic effects. The anabolic properties of PTH have been utilized in the treatment of osteoporosis with proven efficacy in preventing fractures. Despite these benefits, PTH can be administered therapeutically for up to 2 years, and its use in patients with underlying malignancies remains a subject of ongoing debate. These considerations underscore the need for a more comprehensive understanding of the underlying mechanisms. p21-activated kinase 4 (PAK4) is involved in bone resorption and cancer-associated osteolysis; however, its role in osteoblast function and PTH action remains unknown. Therefore, in this study, we aimed to clarify the role of PAK4 in osteoblast function and its effects on PTH-induced anabolic activity. PAK4 enhanced MC3T3-E1 osteoblast viability and proliferation and upregulated cyclin D1 expression. PAK4 also augmented osteoblast differentiation, as indicated by increased mineralization found by alkaline phosphatase and Alizarin Red staining. Treatment with PTH (1-34), an active PTH fragment, stimulated PAK4 expression and phosphorylation in a protein kinase A-dependent manner. In addition, bone morphogenetic protein-2 (which is known to promote bone formation) increased phosphorylated PAK4 (p-PAK4) and PAK4 levels. PAK4 regulated the expression of both phosphorylated and total β-catenin, which are critical for osteoblast proliferation and differentiation. Moreover, p-PAK4 directly interacted with β-catenin, and disruption of β-catenin's binding to T-cell factor impaired PAK4- and PTH-induced osteoblast differentiation. Our findings elucidate the effect of PAK4 on enhancing bone formation in osteoblasts and its pivotal role in the anabolic activity of PTH mediated through its interaction with β-catenin. These insights improve the understanding of the mechanisms underlying PTH activity and should inform the development of more effective and safer osteoporosis treatments., (© 2024 Wiley Periodicals LLC.)

Combined treatment with PTH(1-34) and mechanical loading confers increased structural benefits to bone than monotherapies. However, it remains unclear how this longitudinal adaptation affects the bone mechanics. This study quantified the individual and combined longitudinal effects of PTH(1-34) and mechanical loading on the bone stiffness and strength evaluated in vivo with validated micro-finite element (microFE) models. C57BL/6 mice were ovariectomised at 14-week-old and treated either with injections of PTH(1-34), compressive tibia loading or both interventions concurrently. Right tibiae were in vivo microCT-scanned every 2 weeks from 14 until 24-week-old. MicroCT images were rigidly registered to reference tibia and the cortical organ level (whole bone) and tissue level (midshaft) morphometric properties and bone mineral content were quantified. MicroCT images were converted into voxel-based homogeneous, linear elastic microFE models to estimate the bone stiffness and strength. This approach allowed us for the first time to quantify the longitudinal changes in mechanical properties induced by combined treatments in a model of accelerated bone resorption. Both changes of stiffness and strength were higher with co-treatment than with individual therapies, consistent with increased benefits with the tibia bone mineral content and cortical area, properties strongly associated with the tibia mechanics. The longitudinal data shows that the two bone anabolics, both individually and combined, had persistent benefit on estimated mechanical properties, and that benefits (increased stiffness and strength) remained after treatment was withdrawn., (© 2023 The Authors. Journal of Orthopaedic Research® published by Wiley Periodicals LLC on behalf of Orthopaedic Research Society.)

To clarify the cellular mechanism of cortical porosity induced by intermittent parathyroid hormone (PTH) administration, we examined the femoral cortical bone of mice that received 40 µg/kg/day (four times a day) human PTH (hPTH) (1-34). The PTH-driven cortical porosity initiated from the metaphyseal region and chronologically expanded toward the diaphysis. Alkaline phosphatase (ALP)-positive osteoblasts in the control mice covered the cortical surface, and endomucin-positive blood vessels were distant from these osteoblasts. In PTH-administered mice, endomucin-reactive blood vessels with TRAP-positive penetrated the ALP-positive osteoblast layer, invading the cortical bone. Statistically, the distance between endomucin-positive blood vessels and the cortical bone surface abated after PTH administration. Transmission electron microscopic observation demonstrated that vascular endothelial cells often pass through the flattened osteoblast layer and accompanied osteoclasts in the deep region of the cortical bone. The cell layers covering mature osteoblasts thickened with PTH administration and exhibited ALP, α-smooth muscle actin (αSMA), vascular cell adhesion molecule-1 (VCAM1), and receptor activator of NF-κB ligand (RANKL). Within these cell layers, osteoclasts were found near endomucin-reactive blood vessels. In PTH-administered femora, osteocytes secreted Dkk1, a Wnt inhibitor that affects angiogenesis, and blood vessels exhibited plasmalemma vesicle-associated protein, an angiogenic molecule. In summary, endomucin-positive blood vessels, when accompanied by osteoclasts in the ALP/αSMA/VCAM1/RANKL-reactive osteoblastic cell layers, invade the cortical bone, potentially due to the action of osteocyte-derived molecules such as DKK1., Competing Interests: Competing InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Osteoarthritis (OA) treatment is limited by the lack of effective nonsurgical interventions to slow disease progression. Here, we examined the contributions of the subchondral bone properties to OA development. We used parathyroid hormone (PTH) to modulate bone mass before OA initiation and alendronate (ALN) to inhibit bone remodeling during OA progression. We examined the spatiotemporal progression of joint damage by combining histopathological and transcriptomic analyses across joint tissues. The additive effect of PTH pretreatment before OA initiation and ALN treatment during OA progression most effectively attenuated load-induced OA pathology. Individually, PTH directly improved cartilage health and slowed the development of cartilage damage, whereas ALN primarily attenuated subchondral bone changes associated with OA progression. Joint damage reflected early transcriptomic changes. With both treatments, the structural changes were associated with early modulation of immunoregulation and immunoresponse pathways that may contribute to disease mechanisms. Overall, our results demonstrate the potential of subchondral bone-modifying therapies to slow the progression of OA.

In hyperparathyroidism (hyperPTH), excessive amounts of PTH are secreted, interfering with calcium regulation in the body. Several drugs can control the disease's side effects, but none of them is an alternative treatment to surgery. Therefore, new drug candidates are necessary. In this study, three computationally repositioned drugs, DG 041, IMD 0354, and cucurbitacin I, are evaluated in an in vitro model of hyperPTH. First, we integrated publicly available transcriptomics datasets to propose drug candidates. Using 3D spheroids derived from a single primary hyperPTH patient, we assessed their in vitro efficacy. None of the proposed drugs affected the viability of healthy cell control (HEK293) or overactive parathyroid cells at the level of toxicity. This behavior was attributed to the non-cancerous nature of the parathyroid cells, establishing the hyperPTH disease model. Cucurbitacin I and IMD 0354 exhibited a slight inverse relationship between increased drug concentrations and cell viability, whereas DG 041 increased viability. Based on these results, further studies are needed on the mechanism of action of the repurposed drugs, including determining the effects of these drugs on cellular PTH synthesis and secretion and on the metabolic pathways that regulate PTH secretion., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

This study investigated the efficacy of the two FDA-approved bone anabolic ligands of the parathyroid hormone receptor 1 (PTH1R), teriparatide or human parathyroid hormone 1-34 (PTH) and abaloparatide (ABL), to restoring skeletal health using a preclinical murine model of streptozotocin-induced T1-DM. Intermittent daily subcutaneous injections of equal molar doses (12 pmoles/g/day) of PTH (50 ng/g/day), ABL (47.5 ng/g/day), or vehicle, were administered for 28 days to 5-month-old C57Bl/6 J male mice with established T1-DM or control (C) mice. ABL was superior to PTH in increasing or restoring bone mass in control or T1-MD mice, respectively, which was associated with superior stimulation of trabecular and periosteal bone formation, upregulation of osteoclastic/osteoblastic gene expression, and increased circulating bone remodeling markers. Only ABL corrected the reduction in ultimate load, which is a measure of bone strength, induced by T1-DM, and it also increased energy to ultimate load. In addition, bones from T1-DM mice treated with PTH or ABL exhibited increased ultimate stress, a material index, compared to T1-DM mice administered with vehicle. And both PTH and ABL prevented the increased expression of the Wnt antagonist Sost/sclerostin displayed by T1-DM mice. Further, PTH and ABL increased to a similar extent the circulating bone resorption marker CTX and the bone formation marker P1NP in T1-DM after 2 weeks of treatment; however, only ABL sustained these increases after 4 weeks of treatment. We conclude that at equal molar doses, ABL is more effective than PTH in increasing bone mass and restoring the cortical and trabecular bone lost with T1-DM, due to higher and longer-lasting increases in bone remodeling., Competing Interests: Declaration of competing interest None, (Copyright © 2024 Elsevier Inc. All rights reserved.)

The intermittent administration of parathyroid hormone (PTH) exerts potent bone anabolic effects, which increase bone mineral density (BMD) and reduce fracture risk in osteoporotic patients. However, the underlying mechanisms remain unclear. Tmem119 has been proposed as a factor that is closely linked to the osteoblast phenotype, and we previously reported that PTH enhanced the expression of Tmem119 in mouse osteoblastic cells. However, roles of Tmem119 in the bone anabolic effects of PTH in vivo remain unknown. We herein investigated the roles of Tmem119 in bone anabolic effects of PTH using Tmem119-deficient mice. Tmem119 deficiency significantly reduced PTH-induced increases in trabecular bone volume and cortical BMD of femurs. Effects of Tmem119 deficiency on bone mass seemed predominant in female mice. Histomorphometric analyses with calcein labeling showed that Tmem119 deficiency significantly attenuated PTH-induced increases in the rates of bone formation and mineralization as well as numbers of osteoblasts. Moreover, Tmem119 deficiency significantly blunted PTH-induced decreases in phosphorylation of β-catenin and increases in alkaline phosphatase activity in osteoblasts. In conclusion, the present results indicate that Tmem119 is involved in bone anabolic effects of PTH through osteoblastic bone formation partly related to canonical Wnt-β-catenin signaling in mice., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest, financial or otherwise., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: To investigate the effect of recombinant human parathyroid hormone (rhPTH) biocomposite on bone-to-tendon interface (BTI) healing for surgical repair of a chronic rotator cuff tear (RCT) model of rabbit, focusing on genetic, histologic, biomechanical and micro-computed tomography (CT) evaluations., Methods: Sixty-four rabbits were equally assigned to the 4 groups: saline injection (group A), nanofiber sheet alone (group B), rhPTH-soaked nanofiber sheet (nanofiber sheet was soaked with rhPTH, group C), and rhPTH biocomposite (rhPTH permeated the nanofiber sheet by coaxial electrospinning, group D). The release kinetics of rhPTH (groups C and D) was examined for 6 weeks in vitro. Nanofiber scaffolds were implanted on the surface of the repair site 6 weeks after the induction of chronic RCT. Genetic and histologic analyses were conducted 4 weeks after surgery. Furthermore, genetic, histologic, biomechanical, micro-CT, and serologic analyses were performed 12 weeks after surgery., Results: In vivo, group D showed the highest collagen type I alpha 1 (COL1A1), collagen type III alpha 1 (COL3A1), and bone morphogenetic protein 2 (BMP-2) messenger RNA (mRNA) expression levels (all P < .001) 4 weeks after surgery; however, there were no differences between groups at 12 weeks postsurgery. After 12 weeks postsurgery, group D showed better collagen fiber continuity and orientation, denser collagen fibers, more mature bone-to-tendon junction, and greater fibrocartilage layer formation compared with the other groups (all P < .05). Furthermore, group D showed the highest load-to-failure rate (28.9 ± 2.0 N/kg for group A, 30.1 ± 3.3 N/kg for group B, 39.7 ± 2.7 N/kg for group C, and 48.2 ± 4.5 N/kg for group D, P < .001) and micro-CT outcomes, including bone and tissue mineral density, and bone volume/total volume rate (all P < .001) at 12 weeks postsurgery., Conclusions: In comparison to rhPTH-soaked nanofiber sheet and the other control groups, rhPTH biocomposite effectively accelerated BTI healing by enhancing the mRNA expression levels of COL1A1, COL3A1, and BMP-2 at an early stage and achieving tenogenesis, chondrogenesis, and osteogenesis at 12 weeks after surgical repair of a chronic RCT model of rabbit., Clinical Relevance: The present study might be a transitional study to demonstrate the efficacy of rhPTH biocomposites on BTI healing for surgical repair of chronic RCTs as an adaptable polymer biomaterial in humans., Competing Interests: Disclosure The authors report the following potential conflicts of interest or sources of funding: This research was supported by a grant from the Seoul National University Bundang Hospital research fund (No. 14-2020-0001; Effect of recombinant parathyroid hormone-mixed nanofiber sheet engineered with 3D printing for regeneration of tendon in a chronic rotator cuff tear model of rabbit) to J.H.O. All other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Full ICMJE author disclosure forms are available for this article online, as supplementary material., (Copyright © 2023 Arthroscopy Association of North America. Published by Elsevier Inc. All rights reserved.)

The autonomic nervous system plays a crucial role in regulating bone metabolism, with sympathetic activation stimulating bone resorption and inhibiting bone formation. We found that fractures lead to increased sympathetic tone, enhanced osteoclast resorption, decreased osteoblast formation, and thus hastened systemic bone loss in ovariectomized (OVX) mice. However, the combined administration of parathyroid hormone (PTH) and the β-receptor blocker propranolol dramatically promoted systemic bone formation and osteoporotic fracture healing in OVX mice. The effect of this treatment is superior to that of treatment with PTH or propranolol alone. In vitro, the sympathetic neurotransmitter norepinephrine (NE) suppressed PTH-induced osteoblast differentiation and mineralization, which was rescued by propranolol. Moreover, NE decreased the PTH-induced expression of Runx2 but enhanced the expression of Rankl and the effect of PTH-stimulated osteoblasts on osteoclastic differentiation, whereas these effects were reversed by propranolol. Furthermore, PTH increased the expression of the circadian clock gene Bmal1, which was inhibited by NE-βAR signaling. Bmal1 knockdown blocked the rescue effect of propranolol on the NE-induced decrease in PTH-stimulated osteoblast differentiation. Taken together, these results suggest that propranolol enhances the anabolic effect of PTH in preventing systemic bone loss following osteoporotic fracture by blocking the negative effects of sympathetic signaling on PTH anabolism., (© 2024. The Author(s).)

Parathyroid hormone 1 receptor (PTH1R) plays a key role in mediating calcium homeostasis and bone development, and aberrant PTH1R activity underlies several human diseases. Peptidic PTH1R antagonists and inverse agonists have therapeutic potential in treating these diseases, but their poor pharmacokinetics and pharmacodynamics undermine their in vivo efficacy. Herein, we report the use of a backbone-modification strategy to design a peptidic PTH1R inhibitor that displays prolonged activity as an antagonist of wild-type PTH1R and an inverse agonist of the constitutively active PTH1R-H223R mutant both in vitro and in vivo. This peptide may be of interest for the future development of therapeutic agents that ameliorate PTH1R malfunction.

In this study, we aimed to quantify the localised effects of mechanical loading (ML), low (20 μg/kg/day), moderate (40 μg/kg/day) or high (80 μg/kg/day) dosages of parathyroid hormone (PTH), and combined (PTHML) treatments on cortical bone adaptation in healthy 19-week old female C57BL/6 mice. To this end, we utilise a previously reported image analysis algorithm on μCT data of the mouse tibia published by Sugiyama et al. (2008) to measure changes in cortical area, marrow cavity area and local cortical thickness measures (ΔCt.Ar, ΔMa.Ar, ΔCt.Th respectively), evaluated at two cross-sections within the mouse tibia (proximal-middle (37 %) and middle (50 %)), and are compared to a superposed summation (P + M) of individual treatments to determine the effectiveness of combining treatments in vivo. ΔCt.Ar analysis revealed a non-linear, synergistic interactions between PTH and ML in the 37 % cross-section that saturates at higher PTH dosages, whereas the 50 % cross-section experiences an approximately linear, additive adaptation response. This coincided with an increase in ΔMa.Ar (indicating resorption of the endosteal surface), which was only counteracted by combined high dose PTH with ML in the middle cross-section. Regional analysis of ΔCt.Th changes reveal localised cortical thinning in response to low dose PTH treatment in the posteromedial region of the middle cross-section, signifying that PTH does not provide a homogeneous adaptation response around the cortical perimeter. We observe a synergistic response in the proximal-middle cross-section, with regions of compressive strain experiencing the greatest adaptation response to PTHML treatments, (peak ΔCt.Th of 189.32, 213.78 and 239.30 μm for low, moderate and high PTHML groups respectively). In contrast, PTHML treatments in the middle cross-section show a similar response to the superposed P + M group, with the exception of the combined high dose PTHML treatment which shows a synergistic interaction. These analyses suggest that, in mice, adding mechanical loading to PTH treatments leads to region specific bone responses; synergism of PTHML is only achieved in some regions experiencing high loading, while other regions respond additively to this combined treatment., Competing Interests: Declaration of competing interest The authors hereby declare that they have no conflict of interest in the submitted manuscript., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Objectives: Intermittent administration of parathyroid hormone (PTH) increases systemic bone mass. However, the effect of PTH on osseous and soft tissue healing around implants in osteoporosis patients remains unclear. This study aimed to investigate the effects of PTH on tissue healing around implants in ovariectomized rats and to compare systemic and intraoral administration routes., Material and Methods: Implants were placed at the healed sites of ovariectomized rats 3 weeks after maxillary first molar extraction. Rats were randomly divided into two groups that received either daily systemic subcutaneous or local intraoral PTH administration. Maxillae were dissected to examine bone architectures with micro-computed tomography images. Histomorphometric and immunohistochemical analyses were performed to evaluate osseous and soft tissue healing around the implants., Results: Regardless of the administration route, PTH significantly increased bone area and the numbers of osteoblasts, osteoclasts, and osteocytes in the first and second inside and outside areas of implant threads, in addition to decreasing the number of sclerostin + osteocytes. However, the intraoral PTH administration route was superior to the systemic route by significantly improving bone quality and promoting collagen production in the connective tissue around implants., Conclusions: Parathyroid hormone administration promoted both osseous and soft tissue healing around implants, irrespective of administration route. Interestingly, intraoral administration improved the evaluated parameters more than systemic administration. Thus, the intraoral route could become a useful treatment strategy for implant treatment in osteoporosis patients., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

We have previously shown that the small molecule hPTHR1 agonist PCO371 (1) orally and dose-dependently induces PTH-like calcemic and hypophostemic activity in thyroparathyroidectomized rats. Compound 2a, bearing a bicyclic aromatic ring, was identified as a novel hPTHR1 agonist during hit to lead modification. It showed moderate PTHR1 agonistic activity with an EC 20 value of 15 μM, and its metabolic stability in human liver microsome (hLM) as well as its solubility in phosphate buffer (PPb) and Fasted state simulated intestinal fluid (FaSSIF) were found to be poor. As results of the initial derivatization of 2a, we identified the indole derivatives as another scaffold. In this article, we report on the structure-activity relationship (SAR), structure-metabolism relationship (SMR), and structure-solubility relationship (SSR) of bicyclic aromatic derivatives, and the in vivo efficacy of 2j., (© 2024 Wiley-VCH GmbH.)

The intermittent injection of teriparatide, a recombinant fragment of human parathyroid hormone (PTH 1-34), activates anabolic activity on bone turnover. However, the PTH administration period is limited to 2 years. Thus, sequential therapy after discontinuation of PTH is required. Low-intensity pulsed ultrasound (LIPUS) has been widely used for bone fracture healing. In this study, we examined the effects of LIPUS on bone mass after PTH withdrawal in ovariectomized (OVX) model mice. The LIPUS-non-irradiated femoral trabecular bone mineral density (BMD) in the treated after PTH withdrawal was significantly decreased. Meanwhile, the femoral BMD in the OVX + PTH-LIPUS group was remarkably higher than that of the OVX group. Additionally, mRNA expression of Runx2, Osterix, Col1a1, and ALP increased significantly following LIPUS irradiation after PTH withdrawal. These results suggest that LIPUS protected against femoral trabecular BMD loss and up-regulated the osteogenic factors following PTH withdrawal in OVX mice., (© 2023 The Authors. Journal of Ultrasound in Medicine published by Wiley Periodicals LLC on behalf of American Institute of Ultrasound in Medicine.)

Stress fractures occur as a result of repeated mechanical stress on bone and are commonly found in the load-bearing lower extremities. Macrophages are key players in the immune system and play an important role in bone remodeling and fracture healing. However, the role of macrophages in stress fractures has not been adequately addressed. We hypothesize that macrophage infiltration into a stress fracture callus site promotes bone healing. To test this, a unilateral stress fracture induction model was employed in which the murine ulna of four-month-old, C57BL/6 J male mice was repeatedly loaded with a pre-determined force until the bone was displaced a distance below the threshold for complete fracture. Mice were treated daily with parathyroid hormone (PTH, 50 μg/kg/day) starting two days before injury and continued until 24 h before euthanasia either four or six days after injury, or treated with trabectedin (0.15 mg/kg) on the day of stress fracture and euthanized three or seven days after injury. These treatments were used due to their established effects on macrophages. While macrophages have been implicated in the anabolic effects of PTH, trabectedin, an FDA approved chemotherapeutic, compromises macrophage function and reduces bone mass. At three- and four-days post injury, callus macrophage numbers were analyzed histologically. There was a significant increase in macrophages with PTH treatment compared to vehicle in the callus site. By one week of healing, treatments differentially affected the bony callus as analyzed by microcomputed tomography. PTH enhanced callus bone volume. Conversely, callus bone volume was decreased with trabectedin treatment. Interestingly, concurrent treatment with PTH and trabectedin rescued the reduction observed in the callus with trabectedin treatment alone. This study reports on the key involvement of macrophages during stress fracture healing. Given these observed outcomes on macrophage physiology and bone healing, these findings may be important for patients actively receiving either of these FDA-approved therapeutics., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

The current study examined the gene expression profiles of anabolic and catabolic molecules after a single parathyroid hormone (PTH) injection in mice. No significant changes were observed in alkaline phosphatase area/tissue volume, tartrate-resistant acid phosphatase-positive osteoclasts, or static bone histomorphometry parameters. However, a sudden and significant decrease in Runx2 expression occurred at 1.5 h post-injection followed by immediate elevation, while sclerostin level was initially downregulated but gradually recovered. Meanwhile, Rankl expression initially increased and then returned to baseline. The prolonged elevation of anabolic molecules and transient increase in catabolic molecules may contribute to the anabolic effect of PTH treatment., Competing Interests: Conflict of interest None of the authors declares a conflict of interest., (Copyright © 2023 Japanese Association for Oral Biology. Published by Elsevier B.V. All rights reserved.)

Background and Objective: It is essential to know the quantitative interactions between biological tissues and external mechanical and chemical stimuli. This assists the physicians to better know the quantitative behavior of the tissue and plan more effective therapy. In the literature, the effect of the chemical and mechanical loading was investigated on the bone biological cell activities and some mechanical features, but a lack of prediction of bone injury under the chemical and mechanical factors was sensed. Therefore, the present study aims to investigate the effects of the application of major chemical factors involved in ossification, including RANKL 1 (Receptor Activator of Nuclear Factor Kappa Beta Ligand), PTH 2 (Parathyroid Hormone), and OPG 3 (Osteoprotegerin) on the mandibular bone biological osteoblast and osteoclast activities and mechanical properties. Moreover, the study assesses the bone injury possibility under uniform mastication pressure applied on the premolar tooth in terms of the mechanostat theory undergoing the effects of the chemical factors., Methods: A 3D geometry of the mandible-tooth assembly was generated from the CT image dataset. The geometry was next purified, solidified, and exported to FEM 4 (Finite Element Method) software to be meshed, where boundary conditions and loading were applied. Moreover, the mathematical system of differential equations to model the chemical factor effects on osteoblast and osteoclast activities as well as bone matrix volume fraction and elastic stiffness relations were applied. Next, the values of the equivalent strain were calculated to predict the injury states of the bone., Results: The results complied with the literature data. The results showed that RANKL and PTH increased the values of the equivalent strain from 450 με to 11500 με, while OPG reduced that from 450 με to 300 με., Conclusions: Therefore, RANKL and PTH doses of this study put the bone at risk of injury compared to the baseline of no dose applied, while OPG secured the bone from injury., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Osteocytes have recently been identified as a new regulator of bone remodeling, but the detailed mechanism of their differentiation from osteoblasts remains unclear. The purpose of this study is to identify cell cycle regulators involved in the differentiation of osteoblasts into osteocytes and determine their physiological significance. The study uses IDG-SW3 cells as a model for the differentiation from osteoblasts to osteocytes. Among the major cyclin-dependent kinases (Cdks), Cdk1 is most abundantly expressed in IDG-SW3 cells, and its expression is down-regulated during differentiation into osteocytes. Inhibition of CDK1 activity reduces IDG-SW3 cell proliferation and differentiation into osteocytes. Osteocyte and Osteoblast-specific Cdk1 knockout in mice (Dmp1-Cdk1 KO ) results in trabecular bone loss. Pthlh expression increases during differentiation, but inhibiting CDK1 activity reduces Pthlh expression. Parathyroid hormone-related protein concentration is reduced in the bone marrow of Dmp1-Cdk1 KO mice. Four weeks of Parathyroid hormone administration partially recovers the trabecular bone loss in Dmp1-Cdk1 KO mice. These results demonstrate that Cdk1 plays an essential role in the differentiation from osteoblast to osteocyte and the acquisition and maintenance of bone mass. The findings contribute to a better understanding of the mechanisms of bone mass regulation and can help develop efficient therapeutic strategies for osteoporosis treatment., (© 2023 Wiley-VCH GmbH.)

Background: Non-union formation still represents a major burden in trauma and orthopedic surgery. Moreover, aged patients are at an increased risk for bone healing failure. Parathyroid hormone (PTH) has been shown to accelerate fracture healing in young adult animals. However, there is no information whether PTH also stimulates bone regeneration in atrophic non-unions in the aged. Therefore, the aim of the present study was to analyze the effect of PTH on bone regeneration in an atrophic non-union model in aged CD-1 mice., Methods: After creation of a 1.8 mm segmental defect, mice femora were stabilized by pin-clip fixation. The animals were treated daily with either 200 mg/kg body weight PTH 1-34 (n = 17) or saline (control; n = 17) subcutaneously. Bone regeneration was analyzed by means of X-ray, biomechanics, micro-computed tomography (µCT) imaging as well as histological, immunohistochemical and Western blot analyses., Results: In PTH-treated animals bone formation was markedly improved when compared to controls. This was associated with an increased bending stiffness as well as a higher number of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts and CD31-positive microvessels within the callus tissue. Furthermore, PTH-treated aged animals showed a decreased inflammatory response, characterized by a lower number of MPO-positive granulocytes and CD68-positive macrophages within the bone defects when compared to controls. Additional Western blot analyses demonstrated a significantly higher expression of cyclooxygenase (COX)-2 and phosphoinositide 3-kinase (PI3K) in PTH-treated mice., Conclusion: Taken together, these findings indicate that PTH is an effective pharmacological compound for the treatment of non-union formation in aged animals., (© 2023. The Author(s).)

Low back pain (LBP) is one of the most prevalent diseases affecting quality of life, with no disease-modifying therapy. During aging and spinal degeneration, the balance between the normal endplate (EP) bilayers of cartilage and bone shifts to more bone. The aged/degenerated bony EP has increased porosity because of osteoclastic remodeling activity and may be a source of LBP due to aberrant sensory innervation within the pores. We used two mouse models of spinal degeneration to show that parathyroid hormone (PTH) treatment induced osteogenesis and angiogenesis and reduced the porosity of bony EPs. PTH increased the cartilaginous volume and improved the mechanical properties of EPs, which was accompanied by a reduction of the inflammatory factors cyclooxygenase-2 and prostaglandin E 2 . PTH treatment furthermore partially reversed the innervation of porous EPs and reversed LBP-related behaviors. Conditional knockout of PTH 1 receptors in the nucleus pulposus (NP) did not abolish the treatment effects of PTH, suggesting that the NP is not the primary source of LBP in our mouse models. Last, we showed that aged rhesus macaques with spontaneous spinal degeneration also had decreased EP porosity and sensory innervation when treated with PTH, demonstrating a similar mechanism of PTH action on EP sclerosis between mice and macaques. In summary, our results suggest that PTH treatment could partially reverse EP restructuring during spinal regeneration and support further investigation into this potentially disease-modifying treatment strategy for LBP.

Purpose: Recent studies have demonstrated the positive effects of parathyroid hormone (PTH) on bone healing, and findings support the use of PTH to accelerate bone healing following distraction osteogenesis. The goal of this review was to compile and discuss the mechanisms potentially underlying the effects of PTH on newly formed bone following a bone-lengthening procedure incorporating all relevant evidence in both animal and clinical studies., Methods: This review summarized all evidence from in vivo to clinical studies regarding the effects of PTH administration on a bone-lengthening model. In addition, a comprehensive evaluation of what is currently known regarding the potential mechanisms underlying the potential benefits of PTH in bone lengthening was presented. Some controversial findings regarding the optimal dosage and timing of administration of PTH in this model were also discussed., Results: The findings demonstrated that the potential mechanisms associated with the action of PTH on the acceleration of bone regeneration after distraction osteogenesis are involvement in mesenchymal cell proliferation and differentiation, endochondral bone formation, membranous bone formation, and callus remodeling., Conclusions: In the last 20 years, a number of animal and clinical studies have indicated that there is a prospective role for PTH treatment in human bone lengthening as an anabolic agent that accelerates the mineralization and strength of the regenerated bone. Therefore, PTH treatment can be viewed as a potential treatment to increase the amount of new calcified bone and the mechanical strength of the bone in order to shorten the consolidation stage after bone lengthening., (© 2023. International Osteoporosis Foundation and Bone Health and Osteoporosis Foundation.)

Bone defect repair remains a major clinical challenge that requires the construction of scaffolds that can regulate bone homeostasis. In this study, a photo-cured mesoporous bioactive glass (PMBG) precursor is developed as a tricalcium phosphate (TCP) agglomerant to obtain a double-phase PMBG/TCP scaffold via 3D printing. The scaffold exhibits multi-scale porous structures and large surface areas, making it a suitable carrier for the loading of parathyroid hormone (PTH) (1-34), which is used for the treatment of osteoporosis. In vitro and in vivo results demonstrate that PMBG/TCP scaffolds coordinated with PTH (1-34) can regulate bone homeostasis in a bidirectional manner to facilitate bone formation and inhibit bone resorption. Furthermore, bidirectional regulation of bone homeostasis by PTH (1-34) is achieved by inhibiting fibrogenic activation protein (FAP). Thus, PMBG/TCP scaffolds coordinated with PTH (1-34) are viable materials with considerable potential for application in the field of bone regeneration and provide an excellent solution for the design and development of clinical materials., (© 2023 Wiley-VCH GmbH.)

Introduction: Diabetes mellitus (DM) impairs fracture healing and is associated with susceptibility to infection, which further inhibits fracture healing. While intermittent parathyroid hormone (1-34) (iPTH) effectively improves fracture healing, it is unknown whether infection-associated impaired fracture healing can be rescued with PTH (teriparatide)., Methods: A chronic diet-induced type 2 diabetic mouse model was used to yield mice with decreased glucose tolerance and increased blood glucose levels compared to lean-fed controls. Methicillin-resistant Staphylococcus aureus (MRSA) was inoculated in a surgical tibia fracture model to simulate infected fracture, after which mice were treated with a combination of antibiotics and adjunctive teriparatide treatment. Fracture healing was assessed by Radiographic Union Scale in Tibial Fractures (RUST), micro-computed tomography (μCT), biomechanical testing, and histology., Results: RUST score was significantly poorer in diabetic mice compared to their lean nondiabetic counterparts. There were concomitant reductions in micro-computed tomography (μCT) parameters of callus architecture including bone volume/total volume, trabecular thickness, and total mineral density in type 2 diabetes mellitus (T2DM) mice. Biomechanicaltesting of fractured femora demonstrated diminished torsional rigidity, stiffness, and toughness to max torque. Adjuvant teriparatide treatment with systemic antibiotic therapy improved numerous parameters of bone microarchitecture bone volume, increased connectivity density, and increased trabecular number in both the lean and T2DM group. Despite the observation that poor fracture healing in T2DM mice was further impaired by MRSA infection, adjuvant iPTH treatment significantly improved fracture healing compared to antibiotic treatment alone in infected T2DM fractures., Discussion: Our results suggest that teriparatide may constitute a viable adjuvant therapeutic agent to improve bony union and bone microarchitecture to prevent the development of septic nonunion under diabetic conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Kwon, Cahill, Yu, Alder, Dussik, Jeong, Back and Lee.)

We reported accentuated lactational decreases in areal bone mineral density and only partial skeletal recovery after lactation in Ugandan women with HIV (WWH) initiated on tenofovir disoproxil fumarate-based antiretroviral therapy (TDF-based ART) during pregnancy compared to women without HIV (REF). WWH also had higher breast milk calcium in the first months of lactation. To investigate the mechanisms, we measured bone turnover markers (bone resorption: C-terminal telopeptide [CTX]; bone formation: procollagen type 1 N-terminal propeptide [P1NP], bone-specific and total alkaline phosphatase [BALP, TALP]), hormones (parathyroid hormone [PTH], intact fibroblast growth factor 23 [FGF23], 1,25-dihydroxyvitamin D [1,25(OH) 2 D]), vitamin D status (25-hydroxyvitamin D [25OHD]), and indices of mineral metabolism and renal function. Blood and urine samples collected at 36 weeks of gestation, 14 and 26 weeks of lactation, and 3-6 months after lactation were analyzed. Mean 25OHD was >50 nmol/L throughout. Both groups experienced similar biochemical changes during pregnancy and lactation to women in other settings, but within these patterns, the two groups differed significantly. Notably, WWH had higher PTH (+31%) and lower 1,25(OH) 2 D (-9%) and TmP/GFR (-9%) throughout, lower P1NP (-27%) and plasma phosphate (-10%) in pregnancy, higher CTX (+15%) and BALP (+19%), and lower eGFR (-4%) during and after lactation. P1NP/CTX ratio was lower in WWH than REF in pregnancy (-21%), less so in lactation (-15%), and similar after lactation. Additionally, WWH had lower plasma calcium (-5%), lower FGF23 (-16%) and fasting urinary calcium (-34%) at one or both lactation timepoints, and higher fasting urinary phosphate (+22%) at 26 weeks of lactation and after lactation. These differences resemble reported TDF effects, especially raised PTH, increased bone resorption, decreased bone formation, and decreased renal function, and may explain the observed differences in bone mineral density and breast milk calcium. Further studies are needed to determine whether HIV and TDF-based ART have long-term consequences for maternal bone health and offspring growth. © 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR)., (© 2023 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).)

Intermittent delivery of parathyroid hormone (PTH) could effectively promote bone regeneration, but the need for daily injection administration has limited its further clinical applications. Exposure to magnetic stimulation could regulate cell fate to promote osteogenesis. Herein, we developed a magnetized hydrogel with programmed PTH release and simultaneous magnetic actuation to promote osteogenic commitment. Ag dual-cross-linked hydrogel was formulated as GelMA - PVA (GP) biphasic reservoir with magnetic nanoparticles (GPM) and PTH (GPMP). Macroscopic and microscopic characterizations were performed to optimize the formulations. In vitro release assessment confirmed the programmable release of PTH with a pulsatile profile primed via magnetization in the first 4 days and a sustained release, controlled by an optimized GP matrix, for over a month. Stimulated by an alternating magnetic field, the hydrogels displayed a zigzag-shaped pulsatile release profile, and the cumulative release was enhanced by 8, 28, and 18% in In40, Ab40, and In20Ab20 (loading 40 μg PTH via incorporation, absorption, and their combination) formulations, respectively, compared with the same formulations without magnetic stimulation. An in vitro cytocompatibility test showed that all formulations were biocompatible and that PTH addition significantly promoted the proliferation of MC3T3-E1 pre-osteoblasts. In vivo studies presented enhanced new bone regeneration with significantly improved bone volume and bone mineral density in GPM and GPMP groups (increased by 120 and 251% compared with those of non-treated control), confirming their osteogenic effects and accelerated bone healing. This newly developed GPMP sample provides simultaneous osteogenesis effects via the programmed release of PTH and magnetically promoted bone regeneration and is promising in the facilitation of bone healing and treatment of various delayed/non-union conditions without the burden of daily injection.

Due to aging and long-term estrogen deficiency, postmenopausal women suffer muscle atrophy (MA), which is characterized by decreased muscle mass and muscle quality. Low-intensity pulsed ultrasound (LIPUS) is an acoustic wave inducing biological effects mainly by the mechanical stimulation and used as a non-invasive physical therapy for muscle repair. Parathyroid hormone (PTH) is an 84-amino-acid polypeptide, and its bioactive fragment [PTH (1-34)] has potential application in the treatment of MA. We speculate that the combination of physical therapy (i.e., the LIPUS) and regulatory hormone (i.e., the PTH) would be more effective in the treatment of MA. The objective of this study was to evaluate the individual and combined effects of LIPUS and PTH therapy on MA in estrogen deficiency mice. Seventy 8-week-old female C57BL/6J mice were used in this study and the MA model was induced by an intraperitoneal injection of 4-vinylcyclohexene diepoxide (VCD) for 20 consecutive days. The VCD-induced MA mice were randomly divided into MA, LIPUS, PTH and LIPUS + PTH (Combined) groups (n = 10/group). In the LIPUS group, the mice were treated by LIPUS in bilateral quadriceps muscles for 20 min, five times a week for 6 weeks. In the PTH group, the mice received subcutaneous injection of PTH (1-34) (80 ug/kg/d) five times a week, for 6 weeks. In the Combined group, the PTH was administrated 30 min before each LIPUS session. Hematoxylin-eosin (H&E) staining, serum biochemical analysis and quantitative real-time polymerase chain reaction (qRT-PCR) were applied to evaluate the therapeutic effects of related treatments. The results showed that the MA mice had a disordered estrus cycle, significantly decreased muscle mass and myofibers cross-sectional area (CSA). After treatments, LIPUS, PTH and Combined groups had a significantly increased CSA, compared with the MA mice without treatment. In addition, Combined group had a significantly increased mRNA expression of Pax7, MyoD and MyoG, compared with LIPUS and PTH monotherapy groups. Our findings indicated that the combination of LIPUS and PTH treatment improves muscle regeneration ability, which might have potential for treating MA in postmenopausal women., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Adaptation to different environments can be achieved by physiological shifts throughout development. Hormonal regulators shape the physiological and morphological traits of the evolving animals making them fit for the particular ecological surroundings. We hypothesized that the artificially induced hypersynthesis of calcitonin and parathyroid hormone mutually influencing calcium metabolism could affect bone formation during early ontogeny in fish imitating the heterochrony in craniofacial ossification in natural adaptive morphs. Conducting an experiment, we found that the long-standing treatment of salmonid juveniles with high doses of both hormones irreversibly shifts the corresponding hormone status for a period well beyond the time scale for total degradation of the injected hormone. The hormones program the ossification of the jaw suspension bones and neurocranial elements in a specific manner affecting the jaws position and pharingo-branchial area stretching. These morphological shifts resemble the adaptive variants found in sympatric pelagic and demersal morphs of salmonids. We conclude that solitary deviations in the regulators of calcium metabolism could determine functional morphological traits via transformations in skeletal development., (© 2023 Wiley Periodicals LLC.)

Background: β-Arrestin 2 (β-arr2) binds activated parathyroid hormone (PTH) receptors stimulating internalization. PTH stimulates both anabolic and catabolic effect on bone depending on the way it is administered. Intermittent PTH stimulation increases trabecular bone formation in mice, but this is decreased in mice lacking β-arr 2, suggesting a role for β-arr 2 in the anabolic effects of PTH. The role of β-arr 2 in the catabolic effects of continuous PTH (cPTH) treatment is not known., Objective: To assess the effects of cPTH administration on bone in mice lacking β-arr 2 compared to wild-type (WT)., Methods: Groups of male and female WT or β-arr2 knockout (KO) mice were administered either PTH or phosphate-buffered saline by osmotic pumps for 2 weeks. Following treatment, serum calcium and phosphate levels were measured, bone structure and mineral density were measured by microcomputed tomography, and bone cells measured by static and dynamic histomorphometry., Results: β-arr2 KO had no effects on skeletal development in mice of either sex. PTH treatment caused hypercalcemia and hypophosphatemia and decreased trabecular and cortical bone only in male WT mice. β-arr2 KO in male mice completely abrogated the effects of PTH on bone, while in female β-arr2 KO mice, PTH treatment increased trabecular bone with no effects on cortical bone., Conclusions: These results demonstrate a profound sex effect on skeletal responses to cPTH treatment, suggesting a protective effect of estrogen on bone loss. β-arr2 plays a role in restraining the anabolic effects of PTH in both male and female mice.

Parathyroid hormone (PTH) is one of the most important hormones for bone turnover and calcium homeostasis. It is unclear how the central nervous system regulates PTH. The subfornical organ (SFO) lies above the third ventricle and modulates body fluid homeostasis. Through retrograde tracing, electrophysiology, and in vivo calcium imaging, we identified the SFO as an important brain nucleus that responds to serum PTH changes in mice. Chemogenetic stimulation of GABAergic neurons in SFO induces decreased serum PTH followed by a decrease in trabecular bone mass. Conversely, stimulation of glutamatergic neurons in the SFO promoted serum PTH and bone mass. Moreover, we found that the blockage of different PTH receptors in the SFO affects peripheral PTH levels and the PTH's response to calcium stimulation. Furthermore, we identified a GABAergic projection from the SFO to the paraventricular nucleus, which modulates PTH and bone mass. These findings advance our understanding of the central neural regulation of PTH at cellular and circuit level., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)

The parathyroid hormone (PTH) 1 receptor (PTH1R) is a G protein-coupled receptor (GPCR) that regulates skeletal development and calcium homeostasis. Here, we describe cryo-EM structures of the PTH1R in complex with fragments of the two hormones, PTH and PTH-related protein, the drug abaloparatide, as well as the engineered tool compounds, long-acting PTH (LA-PTH) and the truncated peptide, M-PTH(1-14). We found that the critical N terminus of each agonist engages the transmembrane bundle in a topologically similar fashion, reflecting similarities in measures of Gαs activation. The full-length peptides induce subtly different extracellular domain (ECD) orientations relative to the transmembrane domain. In the structure bound to M-PTH, the ECD is unresolved, demonstrating that the ECD is highly dynamic when unconstrained by a peptide. High resolutions enabled identification of water molecules near peptide and G protein binding sites. Our results illuminate the action of orthosteric agonists of the PTH1R., Competing Interests: Declaration of interests P.M.S. is a co-founder and shareholder of Septerna Inc. D.W. is a shareholder of Septerna Inc., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

To repair systematically osteoporotic bone defects, it is important to make an effort to both diminish osteoporosis and enhance bone regeneration. Herein, a specifically monoporous microsphere (MPM) carrier encapsulating dosage-sensitive and short half-time parathyroid hormone (PTH) is constructed to tackle the issue. Compared with conventional microsphere carriers involving compact, porous, and mesoporous microspheres, the MPM is desirable to achieve precisely in situ delivery and to minimize topical accumulation. The findings show that the PTH loaded inside MPMs can be gradually released from the single hole of MPMs to improve the initial drug concentration. Also, the MPMs can self-shift with the daily movement of experimental animals to effectively reduce the topical aggregation of released drugs in vitro. In vivo evaluation further confirms that the implant of MPMs-PTH plays a dual role in stimulating the regenerative repair of the cranial defect and relieving osteoporosis in the whole body. Consequently, the current work develops a dynamically movable drug delivery system to achieve precisely in situ delivery, minimize topical accumulation, and systematically repair osteoporotic bone defects., (© 2022 Wiley-VCH GmbH.)

The classical paradigm of G protein-coupled receptor (GPCR) signaling via G proteins is grounded in a view that downstream responses are relatively transient and confined to the cell surface, but this notion has been revised in recent years following the identification of several receptors that engage in sustained signaling responses from subcellular compartments following internalization of the ligand-receptor complex. This phenomenon was initially discovered for the parathyroid hormone (PTH) type 1 receptor (PTH1R), a vital GPCR for maintaining normal calcium and phosphate levels in the body with the paradoxical ability to build or break down bone in response to PTH binding. The diverse biological processes regulated by this receptor are thought to depend on its capacity to mediate diverse modes of cyclic adenosine monophosphate (cAMP) signaling. These include transient signaling at the plasma membrane and sustained signaling from internalized PTH1R within early endosomes mediated by PTH. Here we discuss recent structural, cell signaling, and in vivo studies that unveil potential pharmacological outputs of the spatial versus temporal dimension of PTH1R signaling via cAMP. Notably, the combination of molecular dynamics simulations and elastic network model-based methods revealed how precise modulation of PTH signaling responses is achieved through structure-encoded allosteric coupling within the receptor and between the peptide hormone binding site and the G protein coupling interface. The implications of recent findings are now being explored for addressing key questions on how location bias in receptor signaling contributes to pharmacological functions, and how to drug a difficult target such as the PTH1R toward discovering nonpeptidic small molecule candidates for the treatment of metabolic bone and mineral diseases., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Beneficial effects of intermittent parathyroid hormone (PTH) on bone mass and architecture are described to either simply add to, or to synergise with those of mechanical loading. We evaluate whether interaction with in vivo loading is reinforced by PTH dosing regimen and exhibits compartment-specific sensitivities. Female 12-week-old C57Bl6 mice received daily (7/7) or interrupted 5 day/week (5/7) PTH for 3 weeks (two vehicle groups). All mice had six loading episodes (12N) applied to right tibia (left, non-loaded) for the last 2 weeks. Micro-CT scans were used to evaluate mass and architecture in almost the entire cortical and proximal trabecular regions. Epiphyseal cortical, trabecular and marrow space volumes, and bony growth-plate bridge incidence were evaluated. Statistical analyses employed a linear mixed-effects model at each percentile and 2-way ANOVA with post-hoc test for epiphyses and bridging. We found that daily PTH enhances cortical mass and modifies shape along almost the entire tibia and that these effects are partly mitigated by brief interruption in treatment. Mechanical loading alone augments cortical mass and modifies shape but only in a region proximal to the tibiofibular junction. The effect of combining load and daily PTH dosing is solely additive for cortical bone mass with no significant load: PTH interaction, but exhibits clear synergy with interrupted PTH treatment. Daily, not interrupted PTH stimulates trabecular bone gains, yet load:PTH interaction is present at limited regions with both daily and interrupted treatment. PTH treatment, but not loading, modifies epiphyseal bone but, in contrast, only loading modifies bridge number and areal density. Our findings demonstrate impressive local effects on tibial mass and shape of combined loading and PTH that are sensitive to dosing regimen and exert their effects modularly. These findings emphasise a need to clarify PTH dosing regimens and that advantages could be accrued by aligning treatment accordingly to patient requirements and life-style., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Central to the maintenance of calcium homeostasis is the regulated reabsorption of calcium along the nephron. To this end, parathyroid hormone (PTH) is released from the parathyroid gland in response to lowered plasma calcium levels. This hormone acts through the PTH 1 receptor along the nephron to increase urinary phosphate excretion and decrease urinary calcium excretion. In the proximal tubule, PTH inhibits phosphate reabsorption by reducing the abundance of sodium phosphate cotransporters in the apical membrane. PTH likely decreases calcium reabsorption from the proximal tubule, by reducing the reabsorption of sodium, an event necessary for the paracellular movement of calcium across this segment. In the thick ascending limb (TAL), PTH increases calcium permeability and may increase the electrical driving force thereby increasing calcium reabsorption in the TAL. Finally, in the distal convolution, PTH acts to increase transcellular calcium reabsorption by increasing the activity and abundance of the apically expressed calcium channel TRPV5., (© 2023 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

Objectives: To investigate the effects of intermittent parathyroid hormone on cementoblast-mediated periodontal repair in the context of orthodontic-induced root resorption., Materials and Methods: The rat model of orthodontic-induced root resorption was established. Sixty rats were randomly allocated into the experiment group (n = 30) and the control group (n = 30), either receiving a daily subcutaneous injection of recombinant human PTH or placebo vehicle. Enzyme-linked immunosorbent assay, Micro-computed tomography, hematoxylin and eosin staining, and immunohistochemistry staining were performed to detect the periodontal repair. In vitro, OCCM-30 cells were exposed to intermittent PTH (incubated with PTH for the first 6 h in each 24-h cycle). After three cycles, flow cytometry assay, alkaline phosphatase staining, and Alizarin red staining were performed. Quantitative real-time polymerase chain reaction and Western blotting were employed to further determine the effects of intermittent PTH., Results: Intermittent PTH-responsive repair enhancement was detected with the expression of bone sialoprotein, osteocalcin, collagen-1, and alkaline phosphatase significantly upregulated. Increased expressions of cementoblastic proteins were positively correlated to cycles of PTH administration. The proportion of cementoblasts in S and G2/M phases was increased; namely, intermittent PTH promoted cementoblast cell proliferation., Conclusions: Intermittent parathyroid hormone administration promotes cementoblast-mediated cementogenesis during periodontal repair in a time-dependent manner., (© 2022 Wiley Periodicals LLC.)

Hypoparathyroidism (HypoPT) is a disorder characterized by hypocalcemia, low or absent parathyroid hormone (PTH) levels, reduced bone remodeling, and high areal bone mineral density (aBMD). PTH is a therapeutic option, yet data on the prolonged clinical and skeletal effects of PTH treatment are limited. We tracked annual daily doses of calcium and active vitamin D supplements, calciotropic biochemistries, estimated glomerular filtration rate (eGFR), and aBMD measurements in 27 HypoPT patients (16 postsurgical, 11 nonsurgical) who were treated with recombinant human PTH(1-84) [rhPTH(1-84)] for at least 8 (n = 27) and up to 12 (n = 14) years. We also performed high-resolution-peripheral quantitative computed tomography (HRpQCT) imaging and report results at baseline, 5, 8, and 12 years of rhPTH(1-84) treatment. With prolonged use of rhPTH, reductions in the need for supplemental calcium and active vitamin D were maintained. The eGFR did not decline. Serum calcium was maintained within the lower limit of the normal range. aBMD by dual-energy X-ray absorptiometry (DXA) showed an increase at the lumbar spine and a decrease at the distal 1/3 radius. By HRpQCT, cortical volumetric BMD (vBMD) at the tibia decreased at year 5: -20.0% ± 1.5%. The magnitude of this reduction was mitigated in year 8: -8.5% ± 1.6% and in year 12: -10.3% ± 2.2% but all were significantly below the mean baseline value (p < 0.001). A similar pattern of decline was observed at the radius. Cortical porosity progressively increased at the tibia in year 5: 17.4% ± 10% (p < 0.05), year 8: 55.2% ± 11% (p < 0.001), and year 12: 83.5% ± 14% (p < 0.001). A similar pattern of increase was observed at the radius. Failure load, which was higher than normal at baseline, decreased but remained above normal at year 12. This is the longest experience, to date, with PTH therapy in HypoPT. These results demonstrate sustained biochemical stability but overall decreases in bone mass. © 2023 American Society for Bone and Mineral Research (ASBMR)., (© 2023 American Society for Bone and Mineral Research (ASBMR).)

Introduction: The intermittent use of recombinant human parathyroid hormone (iPTH) alters calcium metabolism and induces osteogenesis in experimental models. However, the real effects of iPTH in excitable cells and neurons that require membrane receptors to undergo membrane depolarization/repolarization (Na+K+ATPase) to generate ATP, voltage-gated calcium channel (calcium-IP3R-calponin) as well as GABAergic (GABAA) signaling remains unclear., Objectives: In this study, the expression of IP3R, Na+K+-ATPase, GABAA and calmodulin proteins were evaluated in histological sections of the cerebellum of rats following prolonged injection of iPTH., Methods: Twenty Wistar rats were used in this study and randomly assigned as either or control group. The test group were subcutaneously injected with 20 μg/kg of iPTH, 3×/week for 8 weeks, while the control group received 1 ml/kg of 0.9% saline solution. The rats were euthanized on the 60th day after the first administration, and their cerebellar vermis was removed and submitted to histological and immunohistochemical evaluation for detection of IP3R, Na+K+-ATPase, GABAA and calmodulin proteins. The expression of proteins was evaluated in the areas corresponding to the Purkinje cells as well as in neuropil of molecular layer of cerebellum. All results were transformed into a percentage for each area analyzed to verify significance between groups., Results: Rats that received iPTH demonstrated significant reduction of IP3R, calmodulin and GABAA in Purkinje cells and neuropil of molecular layer while the expression of Na+K+-ATPase was similar., Conclusion: It was concluded that iPTH decreased the expression of IP3R and calmodulin while it did not alter the expression of Na+K+-ATPase. These changes insinuate the ionic activity of calcium and sodium/potassium. Yet, the iPTH alters GABAergic signaling in Purkinje cells, suggesting neurotransmission activity changes in the cerebellum.

The administration of intermittent parathyroid hormone (iPTH) is anabolic to the skeleton. Recent studies with cultured osteoblasts have revealed that the expression of PHOSPHO1, a bone-specific phosphatase essential for the initiation of mineralisation, is regulated by PTH. Therefore, this study sought to determine whether the bone anabolic response to iPTH involves modulation of expression of Phospho1 and of other enzymes critical for bone matrix mineralisation. To mimic iPTH treatment, primary murine osteoblasts were challenged with 50 nM PTH for 6 h in every 48 h period for 8 days (4 cycles), 14 days (7 cycles) and 20 days (10 cycles) in total. The expression of both Phospho1 and Smpd3 was almost completely inhibited after 4 cycles, whereas 10 cycles were required to stimulate a similar response in Alpl expression. To explore the in vivo role of PHOSPHO1 in PTH-mediated osteogenesis, the effects of 14- and 28-day iPTH (80 µg/kg/day) administration was assessed in male wild-type (WT) and Phospho1 -/- mice. The expression of Phospho1, Alpl, Smpd3, Enpp1, Runx2 and Trps1 expression was enhanced in the femora of WT mice following iPTH administration but remained unchanged in the femora of Phospho1 -/- mice. After 28 days of iPTH administration, the anabolic response in the femora of WT was greater than that noted in Phospho1 -/- mice. Specifically, cortical and trabecular bone volume/total volume, as well as cortical thickness, were increased in femora of iPTH-treated WT but not in iPTH-treated Phospho1 -/- mice. Trabecular bone osteoblast number was also increased in iPTH-treated WT mice but not in iPTH-treated Phospho1 -/-  mice. The increased levels of Phospho1, Alpl, Enpp1 and Smpd3 in WT mice in response to iPTH administration is consistent with their contribution to the potent anabolic properties of iPTH in bone. Furthermore, as the anabolic response to iPTH was attenuated in mice deficient in PHOSPHO1, this suggests that the osteoanabolic effects of iPTH are at least partly mediated via bone mineralisation processes., (© 2022 The Authors. Cell Biochemistry and Function published by John Wiley & Sons Ltd.)

Aim: Two-pore channels (TPCs) constitute a small family of cation channels expressed in endo-lysosomal compartments. TPCs have been characterized as critical elements controlling Ca 2+ -mediated vesicular membrane fusion and thereby regulating endo-lysosomal vesicle trafficking. Exo- and endocytotic trafficking and lysosomal degradation are major mechanisms of adaption of epithelial transport. A prime example of highly regulated epithelial transport is the tubular system of the kidney. We therefore studied the localization of TPC protein 1 (TPC1) in the kidney and its functional role in the dynamic regulation of tubular transport., Methods: Immunohistochemistry in combination with tubular markers were used to investigate TPC1 expression in proximal and distal tubules. The excretion of phosphate and ammonium, as well as urine volume and pH were studied in vivo, in response to dynamic challenges induced by bolus injection of parathyroid hormone or acid-base transitions via consecutive infusion of NaCl, Na 2 CO 3 , and NH 4 Cl., Results: In TPC1-deficient mice, the PTH-induced rise in phosphate excretion was prolonged and exaggerated, and its recovery delayed in comparison with wildtype littermates. In the acid-base transition experiment, TPC1-deficient mice showed an identical rise in phosphate excretion in response to Na 2 CO 3 compared with wildtypes, but a delayed NH4Cl-induced recovery. Ammonium-excretion decreased with Na 2 CO 3 , and increased with NH 4 Cl, but without differences between genotypes., Conclusions: We conclude that TPC1 is expressed subapically in the proximal but not distal tubule and plays an important role in the dynamic adaptation of proximal tubular phosphate reabsorption towards enhanced, but not reduced absorption., (© 2023 The Authors. Acta Physiologica published by John Wiley & Sons Ltd on behalf of Scandinavian Physiological Society.)

PTH 1-34 (teriparatide) is approved by FDA for the treatment of postmenopausal osteoporosis. Iron overload is a major contributing factor for bone loss induced by unloading. Whether iron metabolism is involved in the regulation of PTH 1-34 on unloading-induced osteoporosis has not yet been reported. Here, we found that PTH 1-34 attenuated bone loss in unloading mice. PTH 1-34 regulated the disturbance of iron metabolism in unloading mice by activating Nrf2 and further promoting hepcidin expression in the liver. In addition, the Nrf2 inhibitor selectively blocked hepcidin expression in the liver of unloading mice, which neutralized the inhibitory effect of PTH 1-34 on bone loss and the recovery of iron metabolism in unloading mice. Finally, we found that PTH 1-34 promoted the differentiation and inhibited apoptosis of osteoblasts by regulating iron metabolism and maintaining redox balance under unloading conditions. Our results suggested that PTH 1-34 promoted bone formation by regulating iron metabolism under unloading conditions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Che, Ren, Chen, Wang, Zhang and Shang.)