PTHrP Regulates Fatty Acid Metabolism via Novel lncRNA in Breast Cancer Initiation and Progression Models.

Parathyroid hormone-related peptide (PTHrP) is the primary cause of malignancy-associated hypercalcemia (MAH). We previously showed that PTHrP ablation, in the MMTV-PyMT murine model of breast cancer (BC) progression, can dramatically prolong tumor latency, slow tumor growth, and prevent metastatic spread. However, the signaling mechanisms using lineage tracing have not yet been carefully analyzed. Here, we generated Pthrpflox/flox; Cre+ mT/mG mice (KO) and Pthrpwt/wt; Cre+ mT/mG tumor mice (WT) to examine the signaling pathways under the control of PTHrP from the early to late stages of tumorigenesis. GFP+ mammary epithelial cells were further enriched for subsequent RNA sequencing (RNAseq) analyses. We observed significant upregulation of cell cycle signaling and fatty acid metabolism in PTHrP WT tumors, which are linked to tumor initiation and progression. Next, we observed that the expression levels of a novel lncRNA, GM50337, along with stearoyl-Coenzyme A desaturase 1 (Scd1) are significantly upregulated in PTHrP WT but not in KO tumors. We further validated a potential human orthologue lncRNA, OLMALINC, together with SCD1 that can be regulated via PTHrP in human BC cell lines. In conclusion, these novel findings could be used to develop targeted strategies for the treatment of BC and its metastatic complications.

SCF-SKP2 E3 ubiquitin ligase links mTORC1/ER stress/ISR with YAP activation in murine renal cystogenesis.

The Hippo pathway nuclear effector Yes-associated protein (YAP) potentiates the progression of polycystic kidney disease (PKD) arising from ciliopathies. The mechanisms underlying the increase in YAP expression and transcriptional activity in PKD remain obscure. We observed that in kidneys from mice with juvenile cystic kidney (jck) ciliopathy, the aberrant hyperactivity of mechanistic target of rapamycin complex 1 (mTORC1), driven by ERK1/2 and PI3K/AKT cascades, induced ER proteotoxic stress. To reduce this stress by reprogramming translation, the protein kinase R-like ER kinase-eukaryotic initiation factor 2α (PERK/eIF2α) arm of the integrated stress response (ISR) was activated. PERK-mediated phosphorylation of eIF2α drove the selective translation of activating transcription factor 4 (ATF4), potentiating YAP expression. In parallel, YAP underwent K63-linked polyubiquitination by SCF S-phase kinase-associated protein 2 (SKP2) E3 ubiquitin ligase, a Hippo-independent, nonproteolytic ubiquitination that enhances YAP nuclear trafficking and transcriptional activity in cancer cells. Defective ISR cellular adaptation to ER stress in eIF2α phosphorylation-deficient jck mice further augmented YAP-mediated transcriptional activity and renal cyst growth. Conversely, pharmacological tuning down of ER stress/ISR activity and SKP2 expression in jck mice by administration of tauroursodeoxycholic acid (TUDCA) or tolvaptan impeded these processes. Restoring ER homeostasis and/or interfering with the SKP2-YAP interaction represent potential therapeutic avenues for stemming the progression of renal cystogenesis.

Burosumab Treatment for Autosomal Recessive Hypophosphatemic Rickets Type 1 (ARHR1).

CONTEXT: Autosomal recessive hypophosphatemic rickets (ARHR) are rare, heritable renal phosphate-wasting disorders that arise from overexpression of the bone-derived phosphaturic hormone fibroblast growth factor 23 (FGF23) leading to impaired bone mineralization (rickets and osteomalacia). Inactivating mutations of Dentin matrix protein 1 (DMP1) give rise to ARHR type 1 (ARHR1). Short stature, prominent bowing of the legs, fractures/pseudofractures, and severe enthesopathy are prominent in this patient population. Traditionally, treatment consists of oral phosphate replacement and the addition of calcitriol but this approach is limited by modest efficacy and potential renal and gastrointestinal side effects. OBJECTIVE: The advent of burosumab (Crysvita), a fully humanized monoclonal antibody to FGF23 for the treatment of X-linked hypophosphatemia and tumor-induced osteomalacia, offers a unique opportunity to evaluate its safety and efficacy in patients with ARHR1. RESULTS: Monthly administration of burosumab to 2 brothers afflicted with the disorder resulted in normalization of serum phosphate, healing of pseudofracture, diminished fatigue, less bone pain, and reduced incapacity arising from the extensive enthesopathy and soft tissue fibrosis/calcification that characterizes this disorder. No adverse effects were reported following burosumab administration. CONCLUSION: The present report highlights the beneficial biochemical and clinical outcomes associated with the use of burosumab in patients with ARHR1.

Role of PTHrP nuclear localization and carboxyl terminus sequences in postnatal spinal cord development.

Parathyroid hormone-related peptide (PTHrP) acts under physiological conditions to regulate normal development of several tissues and organs. The role of PTHrP in spinal cord development has not been characterized. Pthrp knock in (Pthrp KI) mice were genetically modified to produce PTHrP in which there is a deficiency of the nuclear localization sequence (NLS) and C-terminus. Using this genetically modified mouse model, we have characterized its effect on spinal cord development early postnatally. The spinal cords from Pthrp KI mice displayed a significant reduction in its length, weight, and cross-sectional area compared to wild-type controls. Histologically, there was a decreased development of neurons and glial cells that caused decreased cell proliferation and increased apoptosis. The neural stem cells (NSCs) cultures also revealed decreased cell proliferation and differentiation and increased apoptosis. The proposed mechanism of delayed spinal cord development in Pthrp KI mice may be due to alteration in associated pathways in regulation of cell-division cycles and apoptosis. There was significant downregulation of Bmi-1 and upregulation of cyclin-dependent kinase inhibitors p27, p21, and p16 in Pthrp KI animals. We conclude that NLS and C-terminus peptide segments of PTHrP play an important role in inhibiting cell apoptosis and stimulation of cellular proliferation necessary for normal spinal cord development.

Parathyroid Hormone-Related Protein (PTHrP): An Emerging Target in Cancer Progression and Metastasis.

PTHrP was first discovered as the most common mediator of malignancy-associated hypercalcemia. Subsequently, the discovery of its ubiquitous expression in normal tissues unraveled its role as a physiological autocrine/paracrine regulator. The significance of PTHrP in cancer is not confined to malignancy-associated hypercalcemia, and sufficient evidence now also supports its role in skeletal metastasis through its modulation of bone turnover. Furthermore, our own studies have recently shown the critical role of PTHrP in breast cancer initiation, growth, and metastasis. More recently, we have provided new evidence that overexpression of PTHrP is associated with higher incidence of brain metastasis and decreased overall survival in triple-negative breast cancer patients. Further mechanistic studies in human and mouse model are necessary to fully understand the role of PTHrP in tumor progression and metastasis.

DNA damage checkpoint pathway modulates the regulation of skeletal growth and osteoblastic bone formation by parathyroid hormone-related peptide.

We previously demonstrated that parathyroid hormone-related peptide (PTHrP) 1-84 knockin (Pthrp KI) mice, which lacked a PTHrP nuclear localization sequence (NLS) and C-terminus, displayed early senescence, defective osteoblastic bone formation, and skeletal growth retardation. However, the mechanism of action of the PTHrP NLS and C-terminus in regulating development of skeleton is still unclear. In this study, we examined alterations of oxidative stress and DNA damage response-related molecules in Pthrp KI skeletal tissue. We found that ROS levels, protein expression levels of γ-H2AX, a DNA damage marker, and the DNA damage response markers p-Chk2 and p53 were up-regulated, whereas gene expression levels of anti-oxidative enzymes were down-regulated significantly. We therefore further disrupted the DNA damage response pathway by deleting the Chk2 in Pthrp KI (Chk2-/-KI) mice and did comparison with WT, Chk2-/- and Pthrp KI littermates. The Pthrp KI mice with Chk2 deletion exhibited a longer lifespan, improvement in osteoblastic bone formation and skeletal growth including width of growth plates and length of long bones, trabecular and epiphyseal bone volume, BMD, osteoblast numbers, type I collagen and ALP positive bone areas, the numbers of total colony-forming unit fibroblasts (CFU-f), ALP+ CFU-f and the expression levels of osteogenic genes. In addition, the genes associated with anti-oxidative enzymes were up-regulated significantly, whereas the tumor suppressor genes related to senescence were down-regulated in Chk2-/- KI mice compared to Pthrp KI mice. Our results suggest that Chk2 deletion in Pthrp KI mice can somewhat rescue defects in osteoblastic bone formation and skeletal growth by enhancing endochondral bone formation and osteogenesis. These studies therefore indicate that the DNA damage checkpoint pathway may be a target for the nuclear action of PTHrP to regulate skeletal development and growth.

Defective interplay between mTORC1 activity and endoplasmic reticulum stress-unfolded protein response in uremic vascular calcification.

Vascular calcification increases the risk of cardiovascular disease and death in patients with chronic kidney disease (CKD). Increased activity of mammalian target of rapamycin complex 1 (mTORC1) and endoplasmic reticulum (ER) stress-unfolded protein response (UPR) are independently reported to partake in the pathogenesis of vascular calcification in CKD. However, the association between mTORC1 activity and ER stress-UPR remains unknown. We report here that components of the uremic state [activation of the receptor for advanced glycation end products (RAGE) and hyperphosphatemia] potentiate vascular smooth muscle cell (VSMC) calcification by inducing persistent and exaggerated activity of mTORC1. This gives rise to prolonged and excessive ER stress-UPR as well as attenuated levels of sestrin 1 ( Sesn1) and Sesn3 feeding back to inhibit mTORC1 activity. Activating transcription factor 4 arising from the UPR mediates cell death via expression of CCAAT/enhancer-binding protein (c/EBP) homologous protein (CHOP), impairs the generation of pyrophosphate, a potent inhibitor of mineralization, and potentiates VSMC transdifferentiation to the osteochondrocytic phenotype. Short-term treatment of CKD mice with rapamycin, an inhibitor of mTORC1, or tauroursodeoxycholic acid, a bile acid that restores ER homeostasis, normalized mTORC1 activity, molecular markers of UPR, and calcium content of aortas. Collectively, these data highlight that increased and/or protracted mTORC1 activity arising from the uremic state leads to dysregulated ER stress-UPR and VSMC calcification. Manipulation of the mTORC1-ER stress-UPR pathway opens up new therapeutic strategies for the prevention and treatment of vascular calcification in CKD.

Romosozumab or Alendronate for Fracture Prevention in Women with Osteoporosis.

BACKGROUND: Romosozumab is a monoclonal antibody that binds to and inhibits sclerostin, increases bone formation, and decreases bone resorption. METHODS: We enrolled 4093 postmenopausal women with osteoporosis and a fragility fracture and randomly assigned them in a 1:1 ratio to receive monthly subcutaneous romosozumab (210 mg) or weekly oral alendronate (70 mg) in a blinded fashion for 12 months, followed by open-label alendronate in both groups. The primary end points were the cumulative incidence of new vertebral fracture at 24 months and the cumulative incidence of clinical fracture (nonvertebral and symptomatic vertebral fracture) at the time of the primary analysis (after clinical fractures had been confirmed in ≥330 patients). Secondary end points included the incidences of nonvertebral and hip fracture at the time of the primary analysis. Serious cardiovascular adverse events, osteonecrosis of the jaw, and atypical femoral fractures were adjudicated. RESULTS: Over a period of 24 months, a 48% lower risk of new vertebral fractures was observed in the romosozumab-to-alendronate group (6.2% [127 of 2046 patients]) than in the alendronate-to-alendronate group (11.9% [243 of 2047 patients]) (P<0.001). Clinical fractures occurred in 198 of 2046 patients (9.7%) in the romosozumab-to-alendronate group versus 266 of 2047 patients (13.0%) in the alendronate-to-alendronate group, representing a 27% lower risk with romosozumab (P<0.001). The risk of nonvertebral fractures was lower by 19% in the romosozumab-to-alendronate group than in the alendronate-to-alendronate group (178 of 2046 patients [8.7%] vs. 217 of 2047 patients [10.6%]; P=0.04), and the risk of hip fracture was lower by 38% (41 of 2046 patients [2.0%] vs. 66 of 2047 patients [3.2%]; P=0.02). Overall adverse events and serious adverse events were balanced between the two groups. During year 1, positively adjudicated serious cardiovascular adverse events were observed more often with romosozumab than with alendronate (50 of 2040 patients [2.5%] vs. 38 of 2014 patients [1.9%]). During the open-label alendronate period, adjudicated events of osteonecrosis of the jaw (1 event each in the romosozumab-to-alendronate and alendronate-to-alendronate groups) and atypical femoral fracture (2 events and 4 events, respectively) were observed. CONCLUSIONS: In postmenopausal women with osteoporosis who were at high risk for fracture, romosozumab treatment for 12 months followed by alendronate resulted in a significantly lower risk of fracture than alendronate alone. (Funded by Amgen and others; ARCH ClinicalTrials.gov number, NCT01631214 .).

The parathyroid hormone regulates skin tumour susceptibility in mice.

Using a forward genetics approach to map loci in a mouse skin cancer model, we previously identified a genetic locus, Skin tumour modifier of MSM 1 (Stmm1) on chromosome 7, conferring strong tumour resistance. Sub-congenic mapping localized Parathyroid hormone (Pth) in Stmm1b. Here, we report that serum intact-PTH (iPTH) and a genetic polymorphism in Pth are important for skin tumour resistance. We identified higher iPTH levels in sera from cancer-resistant MSM/Ms mice compared with susceptible FVB/NJ mice. Therefore, we performed skin carcinogenesis experiments with MSM-BAC transgenic mice (Pth MSM-Tg) and Pth knockout heterozygous mice (Pth +/-). As a result, the higher amounts of iPTH in sera conferred stronger resistance to skin tumours. Furthermore, we found that the coding SNP (rs51104087, Val28Met) localizes in the mouse Pro-PTH encoding region, which is linked to processing efficacy and increased PTH secretion. Finally, we report that PTH increases intracellular calcium in keratinocytes and promotes their terminal differentiation. Taken together, our data suggest that Pth is one of the genes responsible for Stmm1, and serum iPTH could serve as a prevention marker of skin cancer and a target for new therapies.

A Case of Male Osteoporosis: A 37-Year-Old Man with Multiple Vertebral Compression Fractures.

While the contributing role of testosterone to bone health is rather modest compared to other factors such as estradiol levels, male hypogonadism is associated with low bone mass and fragility fractures. Along with stimulating physical puberty by achieving virilization and a normal muscle mass and improving psychosocial wellbeing, the goals of testosterone replacement therapy in male hypogonadism also include attainment of age-specific bone mineral density. We report on a 37-year-old man who presented with multiple vertebral compression fractures several years following termination of testosterone replacement therapy for presumed constitutional delay in growth and puberty. Here, we discuss the management of congenital hypogonadotropic hypogonadism with hyposmia (Kallmann syndrome), with which the patient was ultimately diagnosed, the role of androgens in the acquisition of bone mass during puberty and its maintenance thereafter, and outline specific management strategies for patients with hypogonadism and high risk for fragility fractures.

Fibroblast Growth Factor 23 Regulation by Systemic and Local Osteoblast-Synthesized 1,25-Dihydroxyvitamin D.

Circulating levels of fibroblast growth factor 23 (FGF23) increase during the early stages of kidney disease, but the underlying mechanism remains incompletely characterized. We investigated the role of vitamin D metabolites in regulating intact FGF23 production in genetically modified mice without and with adenine-induced uremia. Exogenous calcitriol (1,25-dihydroxyvitamin D) and high circulating levels of calcidiol (25-hydroxyvitamin D) each increased serum FGF23 levels in wild-type mice and in mice with global deficiency of the Cyp27b1 gene encoding 25-hydroxyvitamin D 1-α-hydroxylase, which produces 1,25-hydroxyvitamin D. Compared with wild-type mice, normal, or uremic mice lacking Cyp27b1 had lower levels of serum FGF23, despite having high concentrations of parathyroid hormone, but administration of exogenous 1,25-dihydroxyvitamin D increased FGF23 levels. Furthermore, raising serum calcium levels in Cyp27b1-depleted mice directly increased FGF23 levels and indirectly enhanced the action of ambient vitamin D metabolites via the vitamin D receptor. In chromatin immunoprecipitation assays, 25-hydroxyvitamin D promoted binding of the vitamin D receptor and retinoid X receptor to the promoters of osteoblastic target genes. Conditional osteoblastic deletion of Cyp27b1 caused lower serum FGF23 levels, despite normal circulating levels of vitamin D metabolites. In adenine-induced uremia, only a modest increase in serum FGF23 levels occurred in mice with osteoblastic deletion of Cyp27b1 (12-fold) compared with a large increase (58-fold) in wild-type mice. Therefore, in addition to the direct effect of high circulating concentrations of 25-hydroxyvitamin D, local osteoblastic conversion of 25-hydroxyvitamin D to 1,25-dihydroxyvitamin D appears to be an important positive regulator of FGF23 production, particularly in uremia.

FGF23 Is Not Required to Regulate Fetal Phosphorus Metabolism but Exerts Effects Within 12 Hours After Birth.

Loss of fibroblast growth factor-23 (FGF23) causes hyperphosphatemia, extraskeletal calcifications, and early mortality; excess FGF23 causes hypophosphatemia with rickets or osteomalacia. However, FGF23 may not be important during fetal development. FGF23 deficiency (Fgf23 null) and FGF23 excess (Phex null) did not alter fetal phosphorus or skeletal parameters. In this study, we further tested our hypothesis that FGF23 is not essential for fetal phosphorus regulation but becomes important after birth. Although coreceptor Klotho null adults have extremely high FGF23 concentrations, intact FGF23 was normal in Klotho null fetuses, as were fetal phosphorus and skeletal parameters and placental and renal expression of FGF23 target genes. Pth/Fgf23 double mutants had the same elevation in serum phosphorus as Pth null fetuses, as compared with normal serum phosphorus in Fgf23 nulls. We examined the postnatal time courses of Fgf23 null, Klotho null, and Phex null mice. Fgf23 nulls and Klotho nulls were normal at birth, but developed hyperphosphatemia, increased renal expression of NaPi2a and NaPi2c, and reduced renal phosphorus excretion between 5 and 7 days after birth. Parathyroid hormone remained normal. In contrast, excess FGF23 exerted effects in Phex null males within 12 hours after birth, with the development of hypophosphatemia, reduced renal expression of NaPi2a and NaPi2c, and increased renal phosphorus excretion. In conclusion, although FGF23 is present in the fetal circulation at levels that may equal adult values, and there is robust expression of FGF23 target genes in placenta and fetal kidneys, FGF23 itself is not an important regulator of fetal phosphorous metabolism.

The Role of Parathyroid Hormone-Related Protein (PTHrP) in Osteoblast Response to Microgravity: Mechanistic Implications for Osteoporosis Development.

Prolonged skeletal unloading through bedrest results in bone loss similar to that observed in elderly osteoporotic patients, but with an accelerated timeframe. This rapid effect on weight-bearing bones is also observed in astronauts who can lose up to 2% of their bone mass per month spent in Space. Despite the important implications for Spaceflight travelers and bedridden patients, the exact mechanisms involved in disuse osteoporosis have not been elucidated. Parathyroid hormone-related protein (PTHrP) regulates many physiological processes including skeletal development, and has been proposed as a mechanosensor. To investigate the role of PTHrP in microgravity-induced bone loss, trabecular and calvarial osteoblasts (TOs and COs) from Pthrp +/+ and -/- mice were subjected to actual Spaceflight for 6 days (Foton M3 satellite). Pthrp +/+, +/- and -/- osteoblasts were also exposed to simulated microgravity for periods varying from 6 days to 6 weeks. While COs displayed little change in viability in 0g, viability of all TOs rapidly decreased in inverse proportion to PTHrP expression levels. Furthermore, Pthrp+/+ TOs displayed a sharp viability decline after 2 weeks at 0g. Microarray analysis of Pthrp+/+ TOs after 6 days in simulated 0g revealed expression changes in genes encoding prolactins, apoptosis/survival molecules, bone metabolism and extra-cellular matrix composition proteins, chemokines, insulin-like growth factor family members and Wnt-related signalling molecules. 88% of 0g-induced expression changes in Pthrp+/+ cells overlapped those caused by Pthrp ablation in normal gravity, and pulsatile treatment with PTHrP1-36 not only reversed a large proportion of 0g-induced effects in Pthrp+/+ TOs but maintained viability over 6-week exposure to microgravity. Our results confirm PTHrP efficacy as an anabolic agent to prevent microgravity-induced cell death in TOs.

Overexpression of Bmi1 in Lymphocytes Stimulates Skeletogenesis by Improving the Osteogenic Microenvironment.

To investigate whether overexpression of Bmi1 in lymphocytes can stimulate skeletogenesis by improving the osteogenic microenvironment, we examined the skeletal phenotype of EµBmi1 transgenic mice with overexpression of Bmi1 in lymphocytes. The size of the skeleton, trabecular bone volume and osteoblast number, indices of proliferation and differentiation of bone marrow mesenchymal stem cells (BM-MSCs) were increased significantly, ROS levels were reduced and antioxidative capacity was enhanced in EµBmi1 mice compared to WT mice. In PTHrP1-84 knockin (Pthrp(KI/KI)) mice, the expression levels of Bmi1 are reduced and potentially can mediate the premature osteoporosis observed. We therefore generated a Pthrp(KI/KI) mice overexpressing Bmi1 in lymphocytes and compared them with Pthrp(KI/KI) and WT littermates. Overexpression of Bmi1 in Pthrp(KI/KI) mice resulted in a longer lifespan, increased body weight and improvement in skeletal growth and parameters of osteoblastic bone formation with reduced ROS levels and DNA damage response parameters. Our results demonstrate that overexpression of Bmi1 in lymphocytes can stimulate osteogenesis in vivo and partially rescue defects in skeletal growth and osteogenesis in Pthrp(KI/KI) mice. These studies therefore indicate that overexpression of Bmi1 in lymphocytes can stimulate skeletogenesis by inhibiting oxidative stress and improving the osteogenic microenvironment.

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.

CYP24 inhibition as a therapeutic target in FGF23-mediated renal phosphate wasting disorders.

CYP24A1 (hereafter referred to as CYP24) enzymatic activity is pivotal in the inactivation of vitamin D metabolites. Basal renal and extrarenal CYP24 is usually low but is highly induced by its substrate 1,25-dihydroxyvitamin D. Unbalanced high and/or long-lasting CYP24 expression has been proposed to underlie diseases like chronic kidney disease, cancers, and psoriasis that otherwise should favorably respond to supplemental vitamin D. Using genetically modified mice, we have shown that renal phosphate wasting hypophosphatemic states arising from high levels of fibroblast growth factor 23 (FGF23) are also associated with increased renal Cyp24 expression, suggesting that elevated CYP24 activity is pivotal to the pathophysiology of these disorders. We therefore crossed 2 mouse strains, each with distinct etiology for high levels of circulating FGF23, onto a Cyp24-null background. Specifically, we evaluated Cyp24 deficiency in Hyp mice, the murine homolog of X-linked dominant hypophosphatemic rickets, and transgenic mice that overexpress a mutant FGF23 (FGF23R176Q) that is associated with the autosomal dominant form of hypophosphatemic rickets. Loss of Cyp24 in these murine models of human disease resulted in near-complete recovery of rachitic/osteomalacic bony abnormalities in the absence of any improvement in the serum biochemical profile. Moreover, treatment of Hyp and FGF23R1760-transgenic mice with the CYP24 inhibitor CTA102 also ameliorated their rachitic bones. Our results link CYP24 activity to the pathophysiology of FGF23-dependent renal phosphate wasting states and implicate pharmacologic CYP24 inhibition as a therapeutic adjunct for their treatment.

Deficiency of the parathyroid hormone-related peptide nuclear localization and carboxyl terminal sequences leads to premature skin ageing partially mediated by the upregulation of p27.

We previously reported that deficiency of the PTHrP nuclear localization sequence (NLS) and C-terminus in PTHrP knockin (PTHrP KI) mice resulted in premature ageing of skin. P27, a cyclin-dependent kinase inhibitor, was upregulated in PTHrP KI mice and acted as a downstream target of the PTHrP NLS to regulate the proliferation of vascular smooth muscle cells. To determine the effects of p27 deficiency on premature skin ageing of PTHrP KI mice, we compared the skin phenotypes of PTHrP KI mice to those of p27 knockout (p27(-/-) ) mice and to those of double homozygous p27-deficient and PTHrP KI (p27(-/-) PTHrP KI) mice at 2 weeks age. Compared with wild-type littermates, PTHrP KI mice displayed thinner skin and decreased subcutaneous fat and collagen fibres, decreased skin cell proliferation and increased apoptosis, higher expression of p27, p19 and p53 and lower expression of cyclin E and CDK2, and increased reactive oxygen species levels and decreased antioxidant capacity. Deficiency of p27 in the PTHrP KI mice at least in part corrected the skin premature ageing phenotype resulting in thicker skin and increased subcutaneous fat and collagen. These alternations were associated with higher expression of CDK2 and cyclin E, lower expression of p19 and p53, and enhanced antioxidant capacity with increased skin cell proliferation and inhibition of apoptosis. Our results indicate that the NLS and C-terminus of PTHrP play a critical role in preventing skin from premature ageing that is partially mediated by p27.

Bone Is a Major Target of PTH/PTHrP Receptor Signaling in Regulation of Fetal Blood Calcium Homeostasis.

The blood calcium concentration during fetal life is tightly regulated within a narrow range by highly interactive homeostatic mechanisms that include transport of calcium across the placenta and fluxes in and out of bone; the mechanisms of this regulation are poorly understood. Our findings that endochondral bone-specific PTH/PTHrP receptor (PPR) knockout (KO) mice showed significant reduction of fetal blood calcium concentration compared with that of control littermates at embryonic day 18.5 led us to focus on bone as a possibly major determinant of fetal calcium homeostasis. We found that the fetal calcium concentration of Runx2 KO mice was significantly higher than that of control littermates, suggesting that calcium flux into bone had a considerable influence on the circulating calcium concentration. Moreover, Runx2:PTH double mutant fetuses showed calcium levels similar to those of Runx2 KO mice, suggesting that part of the fetal hypocalcemia in PTH KO mice was caused by the increment of the mineralized bone mass allowed by the formation of osteoblasts. Finally, Rank:PTH double mutant mice had a blood calcium concentration even lower than that of the either Rank KO or PTH KO mice alone at embryonic day 18.5. These observations in our genetic models suggest that PTH/PTHrP receptor signaling in bones has a significant role of the regulation of fetal blood calcium concentration and that both placental transport and osteoclast activation contribute to PTH's hypercalcemic action. They also show that PTH-independent deposition of calcium in bone is the major controller of fetal blood calcium level.