Human P2X7 receptor variants Gly150Arg and Arg276His polymorphisms have differential effects on risk association and cellular functions in pancreatic cancer.

BACKGROUND: The purinergic P2X7 receptor (P2X7R) plays an important role in the crosstalk between pancreatic stellate cells (PSCs) and cancer cells, thus promoting progression of pancreatic ductal adenocarcinoma (PDAC). Single nucleotide polymorphisms (SNPs) in the P2X7R have been reported for several cancers, but have not been explored in PDAC. MATERIALS AND METHODS: Blood samples from PDAC patients and controls were genotyped for 11 non-synonymous SNPs in P2X7R and a risk analysis was performed. Relevant P2X7R-SNP GFP variants were expressed in PSCs and cancer cells and their function was assayed in the following tests. Responses in Ca2+ were studied with Fura-2 and dye uptake with YO-PRO-1. Cell migration was monitored by fluorescence microscopy. Released cytokines were measured with MSD assay. RESULTS: Risk analysis showed that two SNPs 474G>A and 853G>A (rs28360447, rs7958316), that lead to the Gly150Arg and Arg276His variants, had a significant but opposite risk association with PDAC development, protecting against and predisposing to the disease, respectively. In vitro experiments performed on cancer cells and PSCs expressing the Gly150Arg variant showed reduced intracellular Ca2+ response, fluorescent dye uptake, and cell migration, while the Arg276His variant reduced dye uptake but displayed WT-like Ca2+ responses. As predicted, P2X7R was involved in cytokine release (IL-6, IL-1ß, IL-8, TNF-α), but the P2X7R inhibitors displayed varied effects. CONCLUSION: In conclusion, we provide evidence for the P2X7R SNPs association with PDAC and propose that they could be considered as potential biomarkers.

Differential expression of the inflammatory ciita gene may be accompanied by altered bone properties in intact sex steroid-deficient female rats.

OBJECTIVE: The class II transactivator (CIITA), encoded by the CIITA gene, controls expression of immune response regulators, which affect bone homeostasis. Previously, we investigated a functional CIITA polymorphism in elderly women. Women carrying the allele associated with lower CIITA levels displayed higher bone mineral density (BMD), but also higher bone loss. The present exploratory study in a rat model sought to investigate effects of differential expression of Ciita on bone structural integrity and strength. Two strains DA (normal-to-high expression) and DA.VRA4 (lower expression) underwent ovariectomy (OVX) or sham-surgery at ~ 14-weeks of age (DA OVX n = 8, sham n = 4; DA.VRA4 OVX n = 10, sham n = 2). After 16-weeks, femoral BMD and bone mineral content (BMC) were measured and morphometry and biomechanical testing performed. RESULTS: In DA.VRA4 rats, BMD/BMC, cross-sectional area and biomechanical properties were lower. Ciita expression was accompanied by OVX-induced changes to cross-sectional area and femoral shaft strength; DA rats had lower maximum load-to-fracture. Thus, while lower Ciita expression associated with lower bone mass, OVX induced changes to structural and mechanical bone properties were less pronounced. CONCLUSION: The data tentatively suggests association between Ciita expression and structural and mechanical bone properties, and a possible role in bone changes resulting from estrogen deficiency.

Vascular Endothelial Growth Factor and Mesenchymal Stem Cells Revealed Similar Bone Formation to Allograft in a Sheep Model.

INTRODUCTION: Mesenchymal stem cells (MSCs) and vascular endothelial growth factor (VEGF) are key factors in bone regeneration. Further stimulation should establish an enhanced cell environment optimal for vessel evolvement and hereby being able to attract bone-forming cells. The aim of this study was to generate new bone by using MSCs and VEGF, being able to stimulate growth equal to allograft. METHODS: Eight Texel/Gotland sheep had four titanium implants in a size of 10 × 12 mm inserted into bilateral distal femurs, containing a 2 mm gap. In the gap, autologous 3 × 106 MSCs seeded on hydroxyapatite (HA) granules in combination with 10 ng, 100 ng, and 500 ng VEGF release/day were added. After 12 weeks, the implant-bone blocks were harvested, embedded, and sectioned for histomorphometric analysis. Bone formation and mechanical fixation were evaluated. Blood samples were collected for the determination of bone-related biomarkers and VEGF in serum at weeks 0, 1, 4, 8, and 12. RESULTS: The combination of 3 × 106 MSCs with 10 ng, 100 ng, and 500 ng VEGF release/day exhibited similar amount of bone formation within the gap as allograft (P > 0.05). Moreover, no difference in mechanical fixation was observed between the groups (P > 0.05). Serum biomarkers showed no significant difference compared to baseline (all P > 0.05). CONCLUSION: MSCs and VEGF exhibit significant bone regeneration, and their bone properties equal to allograft, with no systemic increase in osteogenic markers or VEGF with no visible side effects. This study indicates a possible new approach into solving the problem of insufficient allograft, in larger bone defects.

Extracellular purines and bone homeostasis.

Maintenance of a healthy skeleton is highly dependent on an intricate regulation of bone metabolism, as changes in the balance between bone formation and bone resorption leads to bone loss, bone fragility and ultimately bone fractures. During the last three decades it has become increasingly evident that physiological release of purines in the extracellular space is imperative for bone homeostasis and is orchestrated via the network of purinoceptors. Adenosine derivatives are released locally in the skeleton either by the bone forming osteoblasts or the bone degrading osteoclasts actioned directly by processes like mechanical loading and indirectly by systemic hormones. Adenosine derivatives directly affect the bone cells by their action on the membranal receptors or have co-stimulatory actions with bone active hormones such as parathyroid hormone or the gut hormones. Any deviations leading to increased levels of extracellular adenosine derivatives in the bone tissue such as in pathologic situations, trigger complex pathways with opposing effects on tissue health as presented by studies involving a range of model organisms. Pathological conditions where skeletal purinergic signaling is affected are following tissue injury like microdamage and macroscopic fractures; and during inflammatory processes where nucleotides and nucleosides play an important part in the pathophysiological skeletal response. Moreover, adenosine derivatives also play an important role in the interaction between malignant cells and bone cells in several types of cancers involving the skeleton, such as but not limited to multiple myeloma and bone osteolysis. Much knowledge has been gained over the last decades. The net- resulting phenotype of adenosine derivatives in bone (including the ratio of ATP to Adenosine) is highly dependent on CD39 and CD73 enzymes together with the expression and activity of the specific receptors. Thus, each component is important in the physiological and pathophysiological processes in bone. Promising perspectives await in the future in treating skeletal disorders with medications targeting the individual components of the purinergic signaling pathway.

Analytical considerations and plans to standardize or harmonize assays for the reference bone turnover markers PINP and ß-CTX in blood.

Procollagen type I N-propeptide (PINP) and the C-terminal telopeptide of type I collagen (ß-CTX) in blood have been designated as reference bone turnover markers in osteoporosis by the International Osteoporosis Foundation (IOF) and International Federation of Clinical Chemistry and Laboratory Medicine (IFCC). The IFCC Committee on Bone Metabolism (C-BM) has examined current commercial assays and performed a multicentre study to examine the agreement between assays for PINP and ß-CTX in serum and plasma. The results of these studies will inform our work towards the harmonization of PINP assays and the standardization of ß-CTX assays in blood, with the development of common calibrators and reference measurement procedures in collaboration with the reagent manufacturing industry. Successful achievement of these goals will help develop universally acceptable practice guidelines for the management of osteoporosis with the inclusion of common reference intervals and treatment targets for PINP and ß-CTX.

Fibroblast growth factor-23 is associated with imaging markers of diabetic cardiomyopathy and anti-diabetic therapeutics.

BACKGROUND: The biomarker fibroblast growth factor-23 (FGF-23) has been associated with increased cardiovascular morbidity and mortality in both patients with and without type 2 diabetes. The aim of this study was to evaluate the relationship between FGF-23 and cardiac structure, function and perfusion in patients with type 2 diabetes and normal or mildly impaired kidney function. Furthermore, to investigate the association between FGF-23, anti-diabetes therapy and the classic complications and risk factors associated with type 2 diabetes. METHODS: In this cross-sectional study, 246 patients with type 2 diabetes underwent echocardiography and advanced cardiac magnetic resonance imaging to assess left ventricular (LV) structure and function. In addition, myocardial blood flow (MBF) during rest and pharmacological stress (adenosine 140 µg/kg/min) were evaluated in 183 of the patients. Patients with eGFR < 60 ml/min/1.73 m2 were excluded. RESULTS: Median (Q1-Q3) FGF-23 was 74 (58-91) ng/L. Patients with FGF-23 above the median had lower MBF during stress (2.3 ± 0.9 vs. 2.7 ± 0.9 ml/min/g, P = 0.001) and lower overall myocardial perfusion reserve (MPR) (2.7 ± 0.8 vs. 3.3 ± 1.1, P < 0.001). LV mass (143 ± 40 vs. 138 ± 36 g, P = 0.04) and E/e* (8.5 ± 3.2 vs. 7.6 ± 2.7, P = 0.04) were higher in patients with FGF-23 above the median. In a linear model adjusted for age, sex, eGFR and hypertension, increasing FGF-23 was associated with decreased MPR (P < 0.01, R2 = 0.11) and increased E/e* (P < 0.01, R2 = 0.07). FGF-23 was lower in patients receiving glucagon like peptide-1 (GLP-1) analogues (71 (57-86) vs. 80 (60-98) ng/L, P = 0.01) than in those who did not receive GLP-1 analogues. CONCLUSIONS: In patients with type 2 diabetes and normal or mildly impaired kidney function, increased levels of FGF-23 are associated with impaired cardiac diastolic function and decreased MPR, caused by a decrease in maximal MBF during stress. Use of GLP-1 analogues is associated with decreased levels of FGF-23. Clinical trial registration https://www.clinicaltrials.gov . Unique identifier: NCT02684331. Date of registration: February 18, 2016.

Human P2X7 Receptor Causes Cycle Arrest in RPMI-8226 Myeloma Cells to Alter the Interaction with Osteoblasts and Osteoclasts.

Multiple myeloma is a malignant expansion of plasma cells and aggressively affects bone health. We show that P2X7 receptor altered myeloma growth, which affects primary bone cells in vitro. Expression on six human myeloma cell lines confirmed the heterogeneity associated with P2X7 receptor. Pharmacology with 2'(3')-O-(4-benzoylbenzoyl) adenosine 5'-triphosphate (BzATP) as agonist showed dose-dependent membranal pores on RPMI-8226 (p = 0.0027) and blockade with P2X7 receptor antagonists. Ca2+ influx with increasing doses of BzATP (p = 0.0040) was also inhibited with antagonists. Chronic P2X7 receptor activation reduced RPMI-8226 viability (p = 0.0208). No apoptosis or RPMI-8226 death was observed by annexin V/propidium iodide (PI) labeling and caspase-3 cleavage, respectively. However, bromodeoxyuridine (BrdU) labelling showed an accumulation of RPMI-8226 in the S phase of cell cycle progression (61.5%, p = 0.0114) with significant decline in G0/G1 (5.2%, p = 0.0086) and G2/M (23.5%, p = 0.0015) phases. As myeloma pathology depends on a positive and proximal interaction with bone, we show that P2X7 receptor on RPMI-8226 inhibited the myeloma-induced suppression on mineralization (p = 0.0286) and reversed the excessive osteoclastic resorption. Our results demonstrate a view of how myeloma cell growth is halted by P2X7 receptor and the consequences on myeloma-osteoblast and myeloma-osteoclast interaction in vitro.

Glucose-Dependent Insulinotropic Polypeptide (GIP) Reduces Bone Resorption in Patients With Type 2 Diabetes.

CONTEXT: In healthy individuals, glucose-dependent insulinotropic polypeptide (GIP) enhances insulin secretion and reduces bone resorption by up to 25% estimated by absolute placebo-corrected changes in carboxy-terminal type 1 collagen crosslinks (CTX) during GIP and glucose administration. In patients with type 2 diabetes (T2D), GIP's insulinotropic effect is impaired and effects on bone may be reduced. OBJECTIVE: To investigate GIP's effect on bone biomarkers in patients with T2D. DESIGN: Randomized, double-blinded, crossover study investigating 6 interventions. PATIENTS: Twelve male patients with T2D. INTERVENTIONS: A primed continuous 90-minute GIP infusion (2 pmol/kg/min) or matching placebo (saline) administered at 3 plasma glucose (PG) levels (i.e., paired days with "insulin-induced hypoglycemia" (PG lowered to 3 mmol/L), "fasting hyperglycemia" (mean PG ~8 mmol/L), or "aggravated hyperglycemia" (mean PG ~12 mmol/L). MAIN OUTCOME MEASURES: Bone biomarkers: CTX, procollagen type 1 N-terminal propeptide (P1NP) and PTH. RESULTS: On days with insulin-induced hypoglycemia, CTX was suppressed by up to 40 ±â€…15% during GIP administration compared with 12 ±â€…11% during placebo infusion (P < 0.0001). On days with fasting hyperglycemia, CTX was suppressed by up to 36 ±â€…15% during GIP administration, compared with 0 ±â€…9% during placebo infusion (P < 0.0001). On days with aggravated hyperglycemia, CTX was suppressed by up to 47 ±â€…23% during GIP administration compared with 10 ±â€…9% during placebo infusion (P = 0.0005). At all glycemic levels, P1NP and PTH concentrations were similar between paired days after 90 minutes. CONCLUSIONS: Short-term GIP infusions reduce bone resorption by more than one-third (estimated by absolute placebo-corrected CTX reductions) in patients with T2DM, suggesting preserved bone effects of GIP in these patients. PRÉCIS: Short-term GIP infusions reduce the bone resorption marker CTX by one-third in patients with type 2 diabetes independent of glycemic levels.

Hypophosphatemic Hypovitaminosis D Induces Osteomalacia in the Adult Female Rat.

Osteomalacia is a bone-demineralizing disease of adulthood, often caused by hypovitaminosis D. Current animal models of the disease mimic osteomalacia as a consequence of gastric bypass or toxic exposure to metals, but a relevant model of diet-induced osteomalacia is lacking. For that purpose, 7-month-old female Sprague Dawley rats were randomly assigned into 2 weight-stratified groups and maintained for 4 months on synthetic diets containing negligible or normal levels of vitamin D. The dietary regimen resulted in vitamin D deficiency as measured by 25-hydroxyvitamin D serum levels; however, hypovitaminosis D per se did not affect biomarkers of calcium metabolism and bone turnover, nor did it result in increased osteoid. Thus, vitamin D depletion through the diet was found to be insufficient to induce an osteomalacia-like phenotype in the adult rat. After 4 months, the phosphate content of the vitamin D-depleted diet had decreased to 0.16% (calcium:phosphorus ratio of 5.85), resulting in an osteomalacic-like condition (trabecular osteoid surface/bone surface constituted 33%; CI, 26-40). The diet change also affected both metabolic and bone turnover biomarkers, including significantly suppressing serum fibroblast growth factor 23. Furthermore, decreased dietary phosphate in a vitamin D-depleted diet led to microarchitectural changes of trabecular and cortical bone, lower bone mass density, lower bone mass content and decreased bone strength, all indicating reduced bone quality. Taken together, our results show that osteomalacia can be induced in the adult female rat by depleting vitamin D and lowering phosphate content in the diet.

GIP's effect on bone metabolism is reduced by the selective GIP receptor antagonist GIP(3-30)NH2.

Infusion of the incretin hormone glucose-dependent insulinotropic polypeptide (GIP) suppresses the bone resorption marker carboxy-terminal type 1 collagen crosslinks (CTX). Using separate and combined infusions of the selective GIP receptor (GIPR) antagonist, GIP(3-30)NH2, and GIP, we investigated how GIPR inhibition affects bone turnover markers. Ten healthy men (median age 22.5 years (range 21-25), BMI 21.3kg/m2 (19.9-24.7)) participated in a randomized, doubled blinded, placebo-controlled, crossover study with four 1h 12mmol/l-hyperglycemic clamps on four separate study days with concomitant infusions of GIP, GIP+GIP(3-30)NH2, GIP(3-30)NH2, and placebo, respectively, separated by a period of at least one week. GIP was infused at 1.5pmol/kg/min and GIP(3-30)NH2 at 800pmol/kg/min. Plasma glucose was clamped at 12.0±1.2mmol/l and plasma levels of GIP and GIP(3-30)NH2 amounted to ∼80pmol/l and ∼50nmol/l, respectively. GIP suppressed CTX more than placebo (baseline-subtracted AUC -6,811±1,260 vs. -3,012±3,018ng/l×min, P= 0.002) and resulted in CTX values of 53 ± 6.9% (GIP) versus 81 ± 10% of baseline (placebo), respectively (P = 0.0006), at the end of the hyperglycemic clamp. Co-infusion of GIP and GIP(3-30)NH2 attenuated the GIP-induced CTX suppression by 51±33% (P = 0.01). The peak value of the bone formation marker N-terminal propeptide of type 1 procollagen (P1NP) peaked at higher levels during GIP (109±6.7% of baseline) than during GIP(3-30)NH2 infusion (101±8.9%) (P = 0.049) and GIP suppressed PTH levels compared to GIP(3-30)NH2 alone (P = 0.0158). In conclusion, blockade of the GIPR with GIP(3-30)NH2 diminished GIP-induced CTX and P1NP responses, showing that these effects are GIPR-mediated and that GIPR antagonism might interfere with bone resorption.

Bone turnover markers in children and adolescents with type 1 diabetes-A systematic review.

Type 1 diabetes (T1D) is associated with impaired bone health and both osteocalcin (OCN) and procollagen type 1 amino terminal propetide (P1NP) (markers of bone formation) and C-terminal cross-linked telopeptide (CTX) (marker of bone resorption) are decreased in adult patients with T1D. We review the existing literature characterizing these bone turnover markers in children and adolescents with T1D and by meta-analysis examine whether alterations in OCN, P1NP, and CTX are evident and if potential changes correlate to the metabolic control (hemoglobin A1c, HbA1c). Systematic searches at MEDLINE and EMBASE were conducted in January 2018 identifying all studies describing OCN, P1NP, or CTX in children and adolescents with T1D. A total of 26 studies were included, representing data from more than 1000 patients with T1D. Pooled analyses of standard mean difference and summary effects analysis were performed when sufficient data were available. Pooled analysis revealed mean OCN to be significantly lower in children and adolescents with T1D compared to healthy controls (standard mean difference: -1.87, 95% confidence interval, CI: -2.83; -0.91) whereas both P1NP and CTX did not differ from the controls. Only data on OCN was sufficient to make pooled correlation analysis revealing a negative correlation between OCN and HbA1c (-0.31 95% CI: -0.45; -0.16). In conclusion, OCN is decreased in children and adolescents with T1D, whether CTX and P1NP are affected as well is unclear, due to very limited data available. New and large studies including OCN, P1NP, and CTX (preferably as z-scores adjusting for age variability) is needed to further elucidate the status of bone turnover in children and adolescents with T1D.

Metformin-induced changes of the gut microbiota in healthy young men: results of a non-blinded, one-armed intervention study.

AIMS/HYPOTHESIS: Individuals with type 2 diabetes have an altered bacterial composition of their gut microbiota compared with non-diabetic individuals. However, these alterations may be confounded by medication, notably the blood-glucose-lowering biguanide, metformin. We undertook a clinical trial in healthy and previously drug-free men with the primary aim of investigating metformin-induced compositional changes in the non-diabetic state. A secondary aim was to examine whether the pre-treatment gut microbiota was related to gastrointestinal adverse effects during metformin treatment. METHODS: Twenty-seven healthy young Danish men were included in an 18-week one-armed crossover trial consisting of a pre-intervention period, an intervention period and a post-intervention period, each period lasting 6 weeks. Inclusion criteria were men of age 18-35 years, BMI between 18.5 kg/m2 and 27.5 kg/m2, HbA1c < 39 mmol/mol (5.7%) and plasma creatinine within the normal range. No prescribed medication, including antibiotics, for 2 months prior to recruitment were allowed and no previous gastrointestinal surgery, discounting appendectomy or chronic illness requiring medical treatment. During the intervention the participants were given metformin up to 1 g twice daily. Participants were examined five times in the fasting state with blood sampling and recording of gastrointestinal symptoms. Examinations took place at Frederiksberg Hospital, Denmark before and after the pre-intervention period, halfway through and immediately after the end of intervention and after the wash-out period. Faecal samples were collected at nine evenly distributed time points, and bacterial DNA was extracted and subjected to 16S rRNA gene amplicon sequencing in order to evaluate gut microbiota composition. Subjective gastrointestinal symptoms were reported at each visit. RESULTS: Data from participants who completed visit 1 (n=23) are included in analyses. For the primary outcome the relative abundance of 11 bacterial genera significantly changed during the intervention but returned to baseline levels after treatment cessation. In line with previous reports, we observed a reduced abundance of Intestinibacter spp. and Clostridium spp., as well as an increased abundance of Escherichia/Shigella spp. and Bilophila wadsworthia. The relative abundance at baseline of 12 bacterial genera predicted self-reported gastrointestinal adverse effects. CONCLUSIONS/INTERPRETATION: Intake of metformin changes the gut microbiota composition in normoglycaemic young men. The microbiota changes induced by metformin extend and validate previous reports in individuals with type 2 diabetes. Secondary analyses suggest that pre-treatment gut microbiota composition may be a determinant for development of gastrointestinal adverse effects following metformin intake. These results require further investigation and replication in larger prospective studies. TRIAL REGISTRATION: Clinicaltrialsregister.eu 2015-000199-86 and ClinicalTrials.gov NCT02546050 FUNDING: This project was funded by Danish Diabetes Association and The Novo Nordisk Foundation.

Osteogenic impact of football training in 55- to 70-year-old women and men with prediabetes.

The effects of football training on bone health were examined in 55- to 70-year-old sedentary women and men with prediabetes. Patients (n = 50) with prediabetes (age; 61 ± 9 years, BMI 29.7 ± 0.6 kg/m2 , body fat content; 37 ± 1%, VO2max ; 22.7 ± 0.8 mL/min/kg and mean arterial pressure; 104 ± 3 mm Hg) were randomized into a football training group (FTG; n = 27, 14 women) and a control group (CON; n = 23, 11 women). At baseline, 73% and 24% were diagnosed with femur osteopenia and osteoporosis, respectively. FTG performed football training twice weekly 30-60-minute sessions in 16 weeks, and both FTG and CON received professional dietary advice. Pre- and post-intervention whole-body and regional bone mineral content (BMC) and density (BMD) were determined with DXA-scans, and venous blood samples were drawn and analyzed for plasma bone turnover markers. Change scores were greater (P < 0.05) in FTG compared to CON in leg BMD (0.023 ± 0.005 vs -0.004 ± 0.001 g/cm2 ) and in leg BMC (32 ± 8 vs -4 ± 6 g). Between-group changes in favor of FTG (P < 0.05) also occurred in the femur neck BMD (3.2%) and femur shaft BMD (2.5%). Whole-body BMC and BMD were unchanged in both groups during the intervention. In FTG, resting plasma osteocalcin, P1NP, and CTX-1 rose (P < 0.05) by 23 ± 8, 52 ± 9 and 38 ± 7%, with greater change scores (P < 0.05) than in CON. Finally, P1NP (formation)/CTX-1 (resorption) ratio increased (P < 0.05) in FTG (127 ± 15 vs 150 ± 11) from pre- to post-intervention, with no change in CON (124 ± 12 and 123 ± 12). In conclusion, football training provides a powerful osteogenic stimulus and improves bone health in 55- to 70-year-old women and men diagnosed with prediabetes.

Bone Turnover Markers in Patients With Nonalcoholic Fatty Liver Disease and/or Type 2 Diabetes During Oral Glucose and Isoglycemic Intravenous Glucose.

Context: Nonalcoholic fatty liver disease (NAFLD) is associated with type 2 diabetes (T2D) and vice versa, and both conditions are associated with an increased risk of fractures and altered bone turnover. Although patients with NAFLD typically suffer from decreased bone mineral density (BMD), T2D is associated with normal to high BMD. The pathophysiology is uncertain but may involve the gut-bone axis. Objective: We investigated the influence of the gut on glucose-induced changes in plasma bone turnover markers in healthy controls and patients with T2D and/or biopsy-verified NAFLD. Design: Cross-sectional cohort study. Patients: Patients with NAFLD with normal glucose tolerance, patients with NAFLD and T2D, patients with T2D without liver disease, and healthy controls. Interventions: Four-hour 50-g oral glucose tolerance test (OGTT) and an isoglycemic intravenous glucose infusion (IIGI). Main Outcome Measures: Collagen type 1 C-telopeptide (CTX), osteocalcin, procollagen type 1 N-terminal propeptide (P1NP), and parathyroid hormone. Results: Plasma glucose levels achieved during OGTTs were successfully matched on corresponding IIGI days. Patients with NAFLD and T2D exhibited similar CTX suppression during the two glucose challenges (P = 0.46) and pronounced suppression of P1NP during IIGI compared with OGTT. Conversely, remaining groups showed greater (P < 0.05) CTX suppression during OGTT and similar suppression of bone formation markers during IIGI and OGTT. Conclusions: OGTT-induced CTX suppression seems to be impaired in patients with NAFLD and T2D, but preserved in patients with either NAFLD or T2D, suggesting that coexistence of T2D and NAFLD may affect gut-bone axis.

Whole-body Magnetic Resonance Imaging in Axial Spondyloarthritis: Reduction of Sacroiliac, Spinal, and Entheseal Inflammation in a Placebo-controlled Trial of Adalimumab.

OBJECTIVE: To investigate whether adalimumab (ADA) reduces whole-body (WB-) magnetic resonance imaging (MRI) indices for inflammation in the entheses, peripheral joints, sacroiliac joints, spine, and the entire body in patients with axial spondyloarthritis (axSpA). METHODS: An investigator-initiated, randomized, placebo-controlled, double-blinded 48-week followup trial included 49 patients with axSpA, who had Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) ≥ 4.0 despite treatment with nonsteroidal antiinflammatory drugs and a clinical indication for tumor necrosis factor inhibitor treatment. Patients were randomized to subcutaneous ADA 40 mg or placebo every other week for 6 weeks; thereafter, all patients received ADA. Conventional MRI and WBMRI were performed at weeks 0, 6, 24, and 48. The primary WBMRI endpoint was the proportion of patients with an improvement in WBMRI total inflammation index above the smallest detectable change (SDC) at Week 6. RESULTS: The primary WBMRI endpoint (improvement of SDC > 2.3) was met in 11 (44%) patients in the ADA group and 3 (13%) patients in the placebo group (p = 0.025, Fisher's exact test). The primary conventional MRI endpoint, the minimally important change in Spondyloarthritis Research Consortium of Canada Spine MRI Inflammation Index at Week 6, was achieved by 9 (36%) patients in the ADA group and 4 (17%) patients in the placebo group (p = 0.20). The primary clinical endpoint, BASDAI reduction > 50% or 2.0 at Week 24, was attained by 32 (65%) patients. CONCLUSION: ADA provided significant reductions in WBMRI indices of peripheral, axial, and whole-body inflammation in patients with axSpA. WBMRI is promising for objective assessment and monitoring of peripheral and axial disease activity in future clinical trials.

Bone turnover, calcium homeostasis, and vitamin D status in Danish vegans.

BACKGROUND/OBJECTIVES: A vegan diet has been associated with increased bone fracture risk, but the physiology linking nutritional exposure to bone metabolism has only been partially elucidated. This study investigated whether a vegan diet is associated with increased bone turnover and altered calcium homeostasis due to insufficient intake of calcium and vitamin D. SUBJECTS/METHODS: Fractionated and total 25-hydroxyvitamin D (25(OH)-D), parathyroid hormone (PTH), calcium, and four bone turnover markers (osteocalcin, N-terminal propeptide of type I procollagen (PINP), bone-specific alkaline phosphatase (BAP), and C-terminal telopeptide of type I collagen (CTX)) were measured in serum from 78 vegans and 77 omnivores. RESULTS: When adjusting for seasonality and constitutional covariates (age, sex, and body fat percentage) vegans had higher concentrations of PINP (32 [95% CI: 7, 64]%, P = 0.01) and BAP (58 [95% CI: 27, 97]%, P < 0.001) compared to omnivores, whereas CTX (30 [95% CI: -1, 72]%, P = 0.06) and osteocalcin (21.8 [95% CI: -9.3, 63.7]%, P = 0.2) concentrations did not differ between the two groups. Vegans had higher serum PTH concentration (38 [95% CI: 19, 60]%; P < 0.001) and lower 25(OH)-D serum concentration (-33 [95% CI: -45, -19]%; P < 0.001), but similar serum calcium concentration (-1 [95% CI: -3, 1]%, P = 0.18 compared to omnivores. CONCLUSIONS: Vegans have higher levels of circulating bone turnover markers compared to omnivores, which may in the long-term lead to poorer bone health. Differences in dietary habits including intake of vitamin D and calcium may, at least partly, explain the observed differences.

Glucose-Dependent Insulinotropic Polypeptide (GIP) Inhibits Bone Resorption Independently of Insulin and Glycemia.

Context: The gut hormone glucose-dependent insulinotropic polypeptide (GIP) causes postprandial insulin release and inhibits bone resorption assessed by carboxy-terminal collagen crosslinks (CTX). Objective: To study if GIP affects bone homeostasis biomarkers independently of insulin release and glycemic level. Design: Randomized, double-blinded, crossover study with 5 study days. Patients: Ten male C-peptide-negative patients with type 1 diabetes. Interventions: On 3 matched days with "low glycemia" (plasma glucose in the interval 3 to 7 mmol/L for 120 minutes), we administered intravenous (IV) GIP (4 pmol × kg-1 × min-1), glucagon-like peptide 1 (1 pmol × kg-1 × min-1), or placebo (saline), and on 2 matched days with "high glycemia" (plasma glucose 12 mmol/L for 90 minutes), we administered either GIP or saline. Main Outcome Measures: CTX, procollagen type 1 N-terminal propeptide (P1NP), and parathyroid hormone (PTH). Results: During low glycemia: GIP progressively suppressed CTX from baseline by up to 59 ± 18% compared with 24 ± 10% during saline infusion (P < 0.0001). Absolute values of P1NP and PTH did not differ between days. During high glycemia: GIP suppressed CTX from baseline by up to 59 ± 19% compared with 7 ± 9% during saline infusion (P < 0.0001). P1NP did not differ between days. GIP suppressed PTH after 60 minutes compared with saline (P < 0.01), but this difference disappeared after 90 minutes. Conclusions: Short-term GIP infusions robustly reduce bone resorption independently of endogenous insulin secretion and during both elevated and low plasma glucose, but have no effect on P1NP or PTH after 90 minutes.

Optimizing combination of vascular endothelial growth factor and mesenchymal stem cells on ectopic bone formation in SCID mice.

INTRODUCTION: Insufficient blood supply may limit bone regeneration in bone defects. Vascular endothelial growth factor (VEGF) promotes angiogenesis by increasing endothelial migration. This outcome, however, could depend on time of application. Sheep mesenchymal stem cells (MSCs) in severe combined immunodeficient (SCID) mice were used in this study to evaluate optimal time points for VEGF stimulation to increase bone formation. METHODS: Twenty-eight SCID (NOD.CB17-Prkdcscid /J) mice had hydroxyapatite granules seeded with 5 × 105 MSCs inserted subcutaneous. Pellets released VEGF on days 1-7, days 1-14, days 1-21, days 1-42, days 7-14, and days 21-42. After 8 weeks, the implant-bone-blocks were harvested, paraffin embedded, sectioned, and stained with both hematoxylin and eosin (HE) and immunohistochemistry for human vimentin (hVim) staining. Blood samples were collected for determination of bone-related biomarkers in serum. RESULTS: The groups with 5 × 105 MSCs and VEGF stimulation on days 1-14 and days 1-21 showed more bone formation when compared to the control group of 5 × 105 MSCs alone (p < 0.01). Serum biomarkers had no significant values. The hVim staining confirmed the ovine origin of the observed ectopic bone formation. CONCLUSION: Optimal bone formation of MSCs was reached when stimulating with VEGF during the first 14 or 21 days after surgery. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3326-3332, 2017.

Effect of GLP-1 receptor agonist treatment on body weight in obese antipsychotic-treated patients with schizophrenia: a randomized, placebo-controlled trial.

AIMS: Schizophrenia is associated with cardiovascular co-morbidity and a reduced life-expectancy of up to 20 years. Antipsychotics are dopamine D2 receptor antagonists and are the standard of medical care in schizophrenia, but the drugs are associated with severe metabolic side effects such as obesity and diabetes. Glucagon-like peptide-1 receptor agonists (GLP-1RAs) are registered for treatment of both obesity and type 2 diabetes. We investigated metabolic effects of the GLP-1RA, exenatide once-weekly, in non-diabetic, antipsychotic-treated, obese patients with schizophrenia. MATERIAL AND METHODS: Antipsychotic-treated, obese, non-diabetic, schizophrenia spectrum patients were randomized to double-blinded adjunctive treatment with once-weekly subcutaneous exenatide (n = 23) or placebo (n = 22) injections for 3 months. The primary outcome was loss of body weight after treatment and repeated measures analysis of variance was used as statistical analysis. RESULTS: Between March 2013 and June 2015, 40 patients completed the trial. At baseline, mean body weight was 118.3 ± 16.0 kg in the exenatide group and 111.7 ± 18.0 kg in the placebo group, with no group differences ( P = .23). The exenatide and placebo groups experienced significant ( P = .004), however similar ( P = .98), weight losses of 2.24 ± 3.3 and 2.23 ± 4.4 kg, respectively, after 3 months of treatment. CONCLUSIONS: Treatment with exenatide once-weekly did not promote weight loss in obese, antipsychotic-treated patients with schizophrenia compared to placebo. Our results could suggest that the body weight-lowering effect of GLP-1RAs involves dopaminergic signaling, but blockade of other receptor systems may also play a role. Nevertheless, anti-obesity regimens effective in the general population may not be readily implemented in antipsychotic-treated patients with schizophrenia.

Football training in men with prostate cancer undergoing androgen deprivation therapy: activity profile and short-term skeletal and postural balance adaptations.

PURPOSE: To investigate the activity profile of football training and its short-term effects on bone mass, bone turnover markers (BTMs) and postural balance in men with prostate cancer (PCa) undergoing androgen deprivation therapy (ADT). METHODS: This was a randomised 12-week study in which men with PCa undergoing ADT were assigned to a football intervention group [FTG, n = 29, 67 ± 7 (±SD) years] training 2‒3 times per week for 45‒60 min or to a control group (n = 28, 66 ± 5 years). The activity profile was measured using a 5-Hz GPS. The outcomes were total body and leg bone mineral content (BMC) and density, BTMs and postural balance. RESULTS: In the last part of the 12 weeks, FTG performed 194 ± 41 accelerations and 296 ± 65 decelerations at >0.6 m/s/s and covered a distance of 905 ± 297 m at speeds >6 km/h and 2646 ± 705 m per training session. Analysis of baseline-to-12-week change scores showed between-group differences in favour of FTG in total body BMC [26.4 g, 95 % confidence interval (CI): 5.8-46.9 g, p = 0.013], leg BMC (13.8 g, 95 % CI: 7.0‒20.5 g, p < 0.001) and markers of bone formation: P1NP (36.6 µg/L, 95 % CI: 10.4‒62.8 µg/L, p = 0.008) and osteocalcin (8.6 µg/L, 95 % CI: 3.3‒13.8 µg/L, p < 0.01). The number of decelerations correlated to the increase in leg BMC (r = 0.65, p = 0.012). No between-group differences were observed for the remaining outcomes. CONCLUSION: Football training involves numerous runs, accelerations and decelerations, which may be linked to marked increases in bone formation markers and preserved bone mass in middle-aged and elderly men with PCa undergoing ADT. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01711892.