Differences in prevalence and associated factors between mild and severe vertebral fractures in Japanese men and women: the third survey of the ROAD study Abstract Vertebral fracture (VF) is a common osteoporotic fracture, while its epidemiology varies according to regions and ethnicities, little is known about it in Japan. Using whole-spine radiographs from a population-based cohort study, the Research on Osteoarthritis/Osteoporosis Against Disability study 3rd survey performed in 2012–2013, we estimated the sex- and age-specific prevalence of VF in the Japanese. Genant’s semiquantitative method (SQ) was used to define VF; SQ ≥ 1 as VF, SQ = 1 as mild VF, SQ≥ 2 as severe VF. We also revealed accurate site-specific prevalence, and associated factors with mild and severe VF. The participants were 506 men [mean age 66.3 years, standard deviation (SD):13.0] and 1038 women (mean age 65.3 years, SD: 12.6). The prevalence of VF in participants aged under 40, in their 40s, 50s, 60s, 70s, and ≥ 80 years was 17.4, 7.9, 18.5, 25.6, 26.3, and 41.5%, respectively, in men, and 2.9%, 2.4%, 7,3, 10.3, 27.1, and 53.0%, respectively, in women. Men had a significantly higher prevalence of mild VF (21.2%) than women (10.0%, p < 0.001); whereas, severe VF was significantly more prevalent in women (9.1%) than in men (4.7%, p = 0.003). VF was distributed with 2 peaks regarding site; one large peak at the thoracolumbar region, and another at the middle thoracic lesion. Low back pain and decreased walking ability were independently associated with severe VF, but not with mild VF, after adjustment for participant characteristics. Decreased walking ability was associated with multiple VFs in women, but not in men.

Incidence of osteonecrosis of the jaw in Japanese osteoporosis patients taking minodronic acid Abstract Osteonecrosis of the jaw (ONJ) associated with bisphosphonate therapy is a rare but severe side effect in osteoporosis patients. Recently, the number of osteoporosis patients with ONJ has dramatically increased in Japan. This has contributed to an increase in the number of patients avoiding extractions. However, there has been no prospective study providing definitive incidence data for ONJ in Japanese patients. The purpose of this study was to elucidate the true as well as suspected incidence of ONJ. A total of 3229 subjects (1612 subjects in the minodronic acid group and 1617 subjects in the raloxifene group) in the Japanese Osteoporosis Intervention Trial protocol number 4 participated in this study. ONJ was diagnosed by experienced dentists. Suspected Stage 0 and 1 (bone exposure of the jaw) ONJ was assessed by a structured questionnaire at baseline and at 6, 12, 18, and 24 months. No established ONJ cases were diagnosed during the study. The incidence of suspected Stage 0 and/or Stage 1 ONJ was 6.14 per 1000 patient-years in the minodronic acid group and 3.38 per 1000 patient-years in the raloxifene group [hazard ratio (95% confidence interval) = 1.82 (0.84–3.93), P = 0.13]. Approximately 50–60% of bone exposures that appeared during the study had disappeared at the next observation. Although the subjects in this study may have developed a greater interest in the health of the oral cavity, the incidence of ONJ after minodronic acid treatment would be lower than the expected incident rate.

Psoas cross-sectional area as a predictor of mortality and a diagnostic tool for sarcopenia in hip fracture patients Abstract This study aimed to determine: (1) the association between psoas cross-sectional area (PCA) and mortality in hip fracture patients, and (2) the usefulness of PCA as a diagnostic tool for sarcopenia. A total of 373 female and 121 male hip fracture patients aged 50 years or more who had surgical repair of their hip fracture between 2011 and 2017 were analyzed. PCA was measured at L4–L5 disc level using CT. PCA of gender-specific 20th percentile determined from this study cohort was used as a cutoff value. The effect of decreased PCA on mortality was analyzed. The association between PCA and appendicular lean mass (ALM) or/and grip strength was analyzed. In survival time of both genders, a significant difference was found between patients with the lowest quintile and upper 4 quintiles (p < 0.001 for females, p = 0.040 for males). The lowest quintile of PCA was associated with mortality, with hazard ratio (HR) of 2.33 (95% CI 1.44–3.47, p < 0.001) in females and 2.01 (95% CI 1.01–3.98, p = 0.046) in males. After adjustment of age, American Society of Anesthesiologists classification, body mass index, dementia, and a grip strength, the lowest quintile of PCA was significantly associated with mortality only in females, with HR of 1.76 (95% CI 1.05–2.70, p = 0.017). A moderate association between PCA and ALM was found in both genders (r = 0.358 for females, r = 0.455 for males). In conclusion, measurement of PCA has potential as a prognostic predictor and diagnostic tool for sarcopenia.

Predictive factors for the efficacy of denosumab in postmenopausal Japanese women with non-metastatic breast cancer receiving adjuvant aromatase inhibitors: a combined analysis of two prospective clinical trials Abstract Aromatase inhibitors (AIs) are the gold standard therapy for breast cancer in postmenopausal women. AI suppresses the conversion of androgens to estrogens; however, this results in osteopenia, osteoporosis, and bone fracture, thus reducing the patient’s quality of life. The use of adjuvant denosumab reduces the risk of clinical fractures in postmenopausal patients with breast cancer receiving AI. However, the efficacy of denosumab in the treatment of AI-associated bone loss has not been prospectively evaluated in Japan. In this study, we aimed to investigate the predictive factors for the efficacy of denosumab in postmenopausal patients with breast cancer treated with AI by analyzing the results of two prospective trials. The patients received 60 mg denosumab subcutaneously every 6 months. The primary endpoint was percentage change in lumbar spine bone mineral density (BMD) from baseline to month 12 in lumbar spine. Post hoc analysis and T tests were performed. A total of 205 patients were enrolled. At 12 and 24 months, the lumbar spine BMD increased by 5.6% [95% confidence interval (CI) 4.9–6.3] and 8.3% (95% CI 7.5–9.1), respectively. Subgroup analysis was conducted according to the time of AI therapy initiation, type of AI therapy, age, time since menopause, baseline body mass index, and BMD. The results showed that baseline lumbar and left femoral BMD was significantly associated with a percentage change in these sites, respectively. In addition, baseline left femoral BMD was also associated with a change in lumbar BMD. In conclusion, the baseline BMD in the lumbar spine was a predictive indicator for the efficacy of denosumab in this site and the baseline BMD in left femoral neck was a predictive indicator in lumbar spine and left femur.

A simple questionnaire for the prediction of vitamin D deficiency in Japanese adults (Vitaimn D Deficiency questionnaire for Japanese: VDDQ-J) Abstract Vitamin D deficiency (VDD) is associated with an increased risk of various diseases. Serum 25-hydroxyvitamin D [25(OH)D] concentration is the best marker for vitamin D status and its concentration < 20 ng/mL indicates VDD. However, its measurement is not easily applicable for the evaluation of vitamin D status in the general population because of its cost. Therefore, we aimed to develop a simple questionnaire for easily identifying the risk of VDD. From the total sample (649 healthy subjects aged 19–70 years), 434 and 215 subjects were randomly assigned to the derivation and the validation cohort, respectively. Prediction model for VDD was developed by backward logistic regression analysis. The regression β coefficients of the significant predictors were transformed into integral numbers and used for the individual score. These individual scores were summed to calculate the total risk score (VDD questionnaire for Japanese score: VDDQ-J score). VDD was present in 54.1% of the total subjects. The model for the prediction of VDD consisted of 7 predictors. Areas under the curve were 0.78 and 0.75 in the data set of internal validation and of the external validation, respectively. The cutoff value was determined to be 31 points (range 0–54) with the sensitivity/specificity and positive predictive value/negative predictive value of 61%/79%, and 81%/57%, respectively. Our VDDQ-J score is easy to answer by the wide range of subjects, and well predicts VDD. This risk score would be useful to identify subjects at risk for VDD both in clinical and epidemiological settings.

Local supplementation with plant-derived recombinant human FGF2 protein enhances bone formation in critical-sized calvarial defects Abstract Numerous studies have demonstrated the advantages of plant cell suspension culture systems in producing bioactive recombinant human growth factors. This study investigated the biological activity of recombinant basic human fibroblast growth factor (rhFGF2) protein produced by a plant culture system to enhance new bone formation in a bone defect mouse model. The human FGF2 cDNA gene was cloned into a plant expression vector driven by the rice α-amylase 3D promoter. The vector was introduced into rice calli (Oryza sativa L. cv. Dongjin), and the clone with the highest expression of rhFGF2 was selected. Maximum accumulation of rhFGF2 protein (approximately 28 mg/l) was reached at 13 day post-incubation. Male C57BL/6 mice underwent calvarial defect surgery and the defects were loaded with absorbable collagen sponge (ACS) only (ACS group) or ACS impregnated with 5 μg of plant-derived rhFGF2 (p-FGF2) protein or E. coli-derived rhFGF2 (e-FGF2) protein. Similar to the effects of e-FGF2, local delivery with p-FGF2 enhanced bone healing in the damaged region to higher levels than the ACS group. Exogenous addition of p-FGF2 or e-FGF2 exhibited similar effects on proliferation, mineralization, and osteogenic marker expression in MC3T3-E1 cells. Together, the current findings support the usefulness of this plant-based expression system for the production of biologically active rhFGF2.

Predicting radiological vertebral fractures with a combined physical function and body composition scoring system Abstract The objective of this study was to investigate the incidence of vertebral fractures (VFx) and the value of physical function (PF) and body composition (BC) for predicting VFx in a Japanese population. This study included 307 subjects (113 men, 194 women) at least 40 years of age who were assessed at community health check-ups in 2008 and 2016. PF was assessed by grip strength and by single-leg stance, timed up-and-go, and 30-s chair stand tests, each scored from 0 to 3 for a possible total of 12 points (higher scores reflect lower function). BC was scored on bioelectrical impedance measurements of trunk and appendage muscle volume, with 6 possible points. We diagnosed radiological VFx semiquantitatively on lateral views of the lumbar spine, and measured bone mineral status by quantitative ultrasound (QUS) of the calcaneus. We conducted logistic regression analysis with VFx as the dependent variable and age, sex, BMI, QUS, PF score, and BC score as independent variables. In 8 years, 36 participants (12%) sustained new VFx. After correcting for age, sex, BMI, and QUS, the odds of VFx increased with a PF score ≥ 8 (OR 5.6; 95% CI 1.21–25.90; P = 0.028) and increased further with a PF + BC score ≥ 9 (OR 8.1; 95% CI 1.80–36.00; P < 0.01). Both PF and BC are important for predicting fragility fractures. The scoring system used here may reflect small differences better than categorical (single cutoff) definitions of poor function.

Preventative effects of metformin on glucocorticoid-induced osteoporosis in rats Abstract This study evaluated the preventative effects of metformin (Met) on glucocorticoid (GC)-induced osteoporosis in a rat model, compared with alendronate (Aln). Twenty-eight 3-month-old female Sprague–Dawley rats were randomly assigned into four groups: normal control (Ctr), methylprednisolone (MP, 13 mg/kg/day, sc, 5 days per week), MP plus Aln orally (1 mg/kg/day), and MP plus Met orally (200 mg/kg/day). After 9 weeks, serum bone metabolic biochemistry, bone densitometry and histomorphometry were performed. The GC-induced osteoporosis model was characterized by decreased osteocalcin, increased tartrate-resistant acid phosphatase-5b (TRAP-5b), and decreased bone mineral density (BMD) in the femur and fifth lumbar vertebra (L5). Histomorphometrically, MP significantly decreased trabecular bone volume, decreased bone formation and increased bone resorption in proximal metaphysis, compared with the controls. Aln and Met increased the BMDs of femur (0.305 ± 0.011 vs. 0.280 ± 0.012, P < 0.05; 0.304 ± 0.019 vs. 0.280 ± 0.012, P < 0.05) and L5 (0.399 ± 0.029 vs. 0.358 ± 0.022, P < 0.05; 0.397 ± 0.022 vs. 0.358 ± 0.022, P < 0.05), compared with the model group. Met increased osteocalcin and decreased TRAP-5b, but Aln only decreased TRAP-5b, compared with model group. In histomorphometry of tibial proximal metaphysis, Aln and Met increased trabecular bone volume (39.21 ± 2.46 vs. 30.98 ± 5.83, P < 0.05; 38.97 ± 5.56 vs. 30.98 ± 5.83, P < 0.05), while Met increased the bone formation dynamic parameters and decreased bone resorption dynamic parameters, but Aln just decreased bone resorption dynamic parameters, compared with model group significantly. These findings suggest that metformin prevents GC-induced bone loss by suppressing bone resorption and stimulating bone formation in trabecular bone. The action mode of metformin was different from alendronate, which only suppressed bone resorption.

Pigment epithelium-derived factor (PEDF) reduced expression and synthesis of SOST/sclerostin in bone explant cultures: implication of PEDF-osteocyte gene regulation in vivo Abstract Mutations in Serpinf1 gene which encodes pigment epithelium-derived factor (PEDF) lead to osteogenesis imperfecta type VI whose hallmark is defective matrix mineralization. We reported previously that PEDF reduced expression and synthesis of Sost/Sclerostin as well as other osteocytes genes encoding proteins that regulate matrix mineralization [1]. To determine whether PEDF had an effect on osteocyte gene expression in bone, we used bone explant cultures. First, osteocytes were isolated from surgical waste of bone fragments obtained from patients undergoing elective foot surgeries under approved IRB protocol by Penn State College of Medicine IRB committee. Primary osteocytes treated with PEDF reduced expression and synthesis of Sost/Sclerostin and matrix phosphoglycoprotein (MEPE) as well as dentin matrix protein (DMP-1). On the whole, PEDF reduced osteocyte protein synthesis by 50% and by 75% on mRNA levels. For bone explants, following collagenase digestion, bone fragments were incubated in alpha-MEM supplemented with 250 ng/ml of PEDF or BSA. After 7 days of incubation in a medium supplemented with PEDF, analysis of mRNA by PCR and protein by western blotting of encoded osteocyte proteins showed reduced Sclerostin synthesis by 39% and MEPE by 27% when compared to fragments incubated in medium supplemented with BSA. mRNA expression levels of osteocytes in bone fragments treated with PEDF were reduced by 50% for both SOST and MEPE when compared to BSA-treated bone fragments. Taken together, the data indicate that PEDF has an effect on osteocyte gene expression in bone and encourage further studies to examine effect of PEDF on bone formation indices in animal models and its effect on osteocyte gene expression in vivo following PEDF administration.

ALPL mutations in adults with rheumatologic disorders and low serum alkaline phosphatase activity Abstract Tissue-nonspecific alkaline phosphatase (ALP), encoded by ALPL, is important for bone homeostasis and interacts with collagen type I. In the present study, we sequenced ALPL and a panel of collagen type I-related genes in 24 adults (age 22–80 years; 20 female) with persistently low serum ALP (< 40 U/L) and a range of rheumatologic symptoms. We found heterozygous pathogenic or likely pathogenic variants in ALPL in 14 (58%) of these individuals. In addition, 7 study participants had potentially damaging heterozygous variants of uncertain significance in genes related to collagen type I. Patients who were positive for ALPL variants had similar age and serum ALP levels to patients in whom no ALPL variants were detected, but had higher serum pyridoxal-5-phosphate concentrations (median 214 nmol/L vs. 64 nmol/L; p = 0.02; U test). In summary, heterozygous ALPL variants are frequent in individuals with rheumatologic symptoms and low ALP serum activity. It is possible that variants in genes that are involved in collagen type I production have a modifying effect on the clinical consequences of such ALPL variants.