Tartrate-resistant Acid Phosphatase 5b Research Articles

Effects of I-125 seeds combined with anlotinib on tumor growth and bone metabolism in A549 tumor-bearing mice

Purpose This study aimed to investigate the therapeutic potential of tumor suppression and mechanism for different implantation modes of iodine-125 (I-125) seeds irradiation in a mice xenograft model, and its skeletal complications. Materials and methods A total of 24 mice carrying A549 lung tumor-derived xenografts were randomly assigned to four groups, including non-radioactive (sham) seeds implantation, I-125 seeds fractional implantation, I-125 seeds single implantation and I-125 seeds single implantation combined with anlotinib. Ki67 immunohistochemistry, TUNEL immunofluorescence and CD31 morphometric analysis were used to determine the proliferation index, rate of apoptotic cells and microvessel density, respectively. Additionally, the side effects on the skeletal system in mice treated with I-125 seeds implantation were evaluated by histomorphometric staining with tartrate-resistant acid phosphate (TRAP) and alkaline phosphatase (ALP) expression in femur, tartrate-resistant acid phosphatase 5b (TRACP-5b) and procollagen type I N-terminal propeptide (PINP) levels in serum were evaluated by enzyme-linked immunosorbent assay (ELISA). Results The I-125 seeds single and fractionated implantation had similar therapeutic effects and complications when the total number of I-125 seeds was the same. A single implantation of I-125 seeds with or without anlotinib could analogously inhibit the tumor growth in xenografts mice, while the single implantation combined with anlotinib had more effective in tumor inhibition. The results of Ki67, TUNEL and CD31 staining confirmed an evident reduction in tumor cell proliferation and angiogenesis, as well as an increase in apoptosis. A relatively integrated bone metabolism was indicated after I-125 seeds single implantation with or without anlotinib, and the results were similar in I-125 seeds fractional implantation, including a reduction in the number of TRAP-positive cells and an increase in ALP expression level. Additionally, the serum TRACP-5b activity was decreased and the serum PINP concentration was increased following I-125 seeds implantation. Conclusions Single and fractionated implantation pattern of I-125 radioactive seeds had similar therapeutic efficacy against tumor growth, while brachytherapy with I-125 seeds implantation may be an effective and safe treatment strategy for its potential protection against cancer treatment-induced bone loss.

Gujiansan Ameliorates Avascular Necrosis of the Femoral Head by Regulating Autophagy via the HIF-1α/BNIP3 Pathway.

Background Clinically, the traditional Chinese medicine compound Gujiansan has been widely used in the treatment of steroid-induced avascular necrosis of the femoral head (SANFH). The present study aimed to investigate the mechanisms underlying the therapeutic effect of Gujiansan. Methods A rat model of SANFH was established by the injection of dexamethasone (DEX) at a high dosage of 25 mg/kg/d. Then, Gujiansan was intragastrically administered for 2 weeks, 4 weeks, and 8 weeks, and histological examination of the femoral head was performed. The expression levels of related mRNAs and proteins were analyzed by qRT-PCR, Western blotting, and immunohistochemistry, and the levels of bone biochemical markers and cytokines were detected with ELISA kits. Results Gujiansan administration ameliorated SANFH and induced the expression of hypoxia-inducible factor-1α (HIF-1α), Bcl-2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3), LC3, and Beclin-1 in the rat model in a dose- and time-dependent manner, and Gujiansan promoted osteocalcin secretion at the femoral head. In addition, Gujiansan increased the levels of bone formation- and bone resorption-specific markers (osteocalcin (OC), bone-specific alkaline phosphatase (BAP), tartrate resistant acid phosphatase-5b (TRACP-5b), N-terminal telopeptides of type I collagen (NTX-1), and C-terminal telopeptide of type I collagen (CTX-1)) and decreased the levels of proinflammatory cytokines (TNF-α, IL-6, and CRP) in a dose- and time-dependent manner. Conclusions Gujiansan accelerates the formation of a new bone, promotes the absorption of the damaged bone, inhibits the inflammatory response, induces autophagy of the femoral head via the HIF-1α/BNIP3 pathway, and ultimately ameliorates SANFH.

Interrelationships Between Sclerostin, Secondary Hyperparathyroidism, and Bone Metabolism in Patients on Hemodialysis.

Sclerostin is an osteocyte-derived inhibitor of bone formation and is increased in kidney failure, but its role in the pathogenesis of renal bone disease remains unknown. We aimed to explore the association of serum sclerostin with bone metabolism in patients undergoing hemodialysis, with a particular focus on parathyroid hormone (PTH)-dependent and PTH-independent pathways. This cross-sectional and prospective cohort study included 654 patients undergoing hemodialysis at 10 facilities in Japan. We employed multivariable linear regression to explore whether sclerostin levels were associated with metacarpal bone mineral density (BMD), intact PTH, bone alkaline phosphatase (BAP), and tartrate-resistant acid phosphatase-5b (TRACP-5b). We employed mediation analyses to explore whether and to what extent the association of PTH with bone turnover markers is mediated by sclerostin. We also compared sclerostin levels between patients with and without previous or incident fractures. The median sclerostin level in hemodialysis patients was 3- to 4-fold higher than that in healthy individuals. Higher sclerostin levels were associated with higher metacarpal BMD and lower levels of intact PTH, BAP, and TRACP-5b. However, the relationships of sclerostin with bone turnover markers were substantially attenuated after adjustment for PTH. Mediation analysis suggested that the effects of PTH on bone turnover markers were mainly direct rather than mediated by sclerostin. Sclerostin levels were not associated with previous or incident fractures. These findings suggest that in patients undergoing dialysis, sclerostin has only a limited role in bone metabolism and may not mediate the effect of PTH on bone turnover.

Efficacy of Evocalcet in Previously Cinacalcet-Treated Secondary Hyperparathyroidism Patients

IntroductionEvocalcet is a recently approved calcimimetic agent for secondary hyperparathyroidism (SHPT). In this study, the efficacy and safety of once-daily oral evocalcet were evaluated in patients without prior cinacalcet use (nonusers) and previously treated patients (users).MethodsThis post hoc analysis of a previous phase III head-to-head comparison study included SHPT patients treated with evocalcet with or without prior cinacalcet use. Endpoints included trends in the median intact and whole parathyroid hormone (PTH), mean corrected calcium, phosphate, and bone metabolic markers, and whole-to-intact PTH ratios throughout the 30-week study period; proportions of patients achieving target intact PTH, corrected calcium, and phosphate at weeks 28 to 30; and adverse drug reactions (ADRs).ResultsThis study included 127 nonusers and 190 users with significant differences in age; duration of dialysis; use of intravenous vitamin D receptor activators; levels of intact PTH, corrected calcium, tartrate-resistant acid phosphatase 5b, procollagen type 1 N-terminal-propeptide; and largest parathyroid gland volume (P < 0.05 for all characteristics) between 2 groups at baseline. Users required higher evocalcet dosages than nonusers. Similar efficacy results were found in the 2 groups except for a significantly higher proportion of nonusers achieving the intact PTH target (81.6% vs 67.1%, difference [95% confidence interval], −14.5% [−24.59, −3.34]), and a significant reduction in largest parathyroid gland volume from week 0 to week 30 (−120.6 [567.2] mm3, P = 0.043). No difference was found in ADRs between the 2 groups.ConclusionTreatment with evocalcet is effective and safe irrespective of prior cinacalcet treatment in SHPT patients.

Effect of TNF inhibitors with bisphosphonates vs bisphosphonates alone on bone mineral density and bone and cartilage biomarkers at 1 year in patients with rheumatoid arthritis: A prospective study.

The present study aimed prospectively to investigate the effect of a combination of tumour necrosis factor inhibitors and bisphosphonates (TNFi with BP) on bone mineral density (BMD) and bone and cartilage biomarkers compared to that of BP alone at 1 year in patients with rheumatoid arthritis (RA). Two groups of patients with RA and osteoporosis were enrolled. One group (37 patients) had already received BP, while the other group (37 patients) had already received TNFi with BP. The serum bone resorption and formation markers, cartilage markers, BMD in the lumbar spine, femoral neck, and distal radius were prospectively investigated at the beginning of the study and at 6 and 12 months. The percentages of change recorded for the various assessment categories were as follows in the TNFi with BP group: (1) tartrate-resistant acid phosphatase-5b had significantly decreased and osteocalcin had increased; (2) matrix metalloproteinase-3 and cartilage oligomeric matrix protein had significantly decreased; and (3) each BMD did not differ significantly between the groups. Our data suggested that TNFi with BP therapy not only suppressed cartilage degradation and bone resorption but also increased bone formation; however, this treatment did not affect the BMD at 1 year.

Bone responsiveness to parathyroid hormone is negatively associated with parathyroid hormone-lowering drug use in patients undergoing hemodialysis: a cross-sectional study

BackgroundParathyroid hormone (PTH) acts on bone to indirectly increase the number and activity of osteoclasts. Thus, PTH has a stimulatory effect on bone resorption and upregulates bone turnover. However, the responsiveness of bone to PTH varies widely among patients receiving dialysis. In fact, relative to the serum PTH level, the level of serum tartrate-resistant acid phosphatase-5b (TRACP-5b), a bone resorption marker derived from osteoclasts, varies as well. This study aimed to examine factors related to bone responsiveness to PTH in patients undergoing chronic hemodialysis (HD).MethodsThis study included patients receiving chronic HD in Kawasaki Municipal Tama Hospital (Kanagawa, Japan) and Yonaha Medical Clinic (Okinawa, Japan) and excluded patients who received HD for less than 6 months, those who received a combination of HD and peritoneal dialysis, and those who had cancer bone metastases or myeloma. The TRACP-5b/intact PTH (iPTH) ratio was created as an index of bone responsiveness to PTH, categorized into tertiles (low, medium, and high), and a cross-sectional study was conducted. P < 0.05 indicated statistically significant differences.ResultsOne hundred and six patients were analyzed. Age (P = 0.010), body mass index (BMI) (P = 0.003), use of calcium-sensing receptor (CaSR) agonists (P = 0.008), use of vitamin D receptor activators (VDRAs) (P = 0.012), plasma iPTH level (P < 0.001), serum 1,25(OH)2D level (P = 0.003), and serum TRACP-5b level (P < 0.001) were significantly different among the three categories. In the single linear regression analysis, age (P = 0.016), corrected serum calcium level (P = 0.007), and ln [1,25(OH)2D] (P = 0.044) showed a significant positive correlation with ln [TRACP-5b/iPTH], whereas BMI (P = 0.026), use of CaSR agonists (P = 0.001), use of VDRAs (P = 0.009), and serum phosphorus level (P = 0.018) showed a significant negative correlation. Upon conducting multiple linear regression analysis incorporating significant variables in the single linear regression analysis, a significant negative correlation was observed between the TRACP-5b/iPTH ratio and intravenous administration of a CaSR agonist (etelcalcetide) and/or a VDRA (calcitriol or maxacalcitol) in all the adjusted models.ConclusionsBone responsiveness to PTH is negatively correlated with the intravenous administration of a CaSR agonist and/or a VDRA in patients undergoing chronic HD.

Osteoclastic activity in chronic otitis media with cholesteatoma-related bone destruction.

Cholesteatoma-related bone destruction is the cause of many complications due to chronic otitis media. This study aimed to evaluate osteoclastic activity in cholesteatoma-related bone destruction using tartrate-resistant acid phosphatase 5b, an enzyme specific to osteoclastic activity. Seventy-two patients diagnosed with chronic otitis media were included in this study and were divided into two groups: with and without bone destruction. The blood serum and tissue tartrate-resistant acid phosphatase 5b levels from both groups were compared. There were no significant differences in the level of serum enzymes between both groups. However, in tissue samples, tartrate-resistant acid phosphatase 5b levels were significantly lower in the bone destruction group than the group without bone destruction. This study determined that the level of tartrate-resistant acid phosphatase 5b, a specific enzyme for osteoclastic activity in cholesteatoma-related bone destruction, is locally decreased. This data suggests that osteoclastic activity may decrease in cholesteatoma-related bone destruction. However, further experimental and clinical studies are required to clarify this highly complex mechanism.

Uncoupling of Bone Turnover may Compromise Skeletal Health of Young Patients With Haemophilia A

There is evidence that bone mass is decreased and bone metabolism is dysregulated in children with haemophilia (CWH). The objective of this study was to investigate the impact of haemophilia on skeletal health in children, with regards to bone mineral density (BMD) and metabolic bone profile. This study included 51 male CWH A. Dual-energy X-ray absorptiometry (DXA) was performed to assess BMD in lumbar spine (LS) and total body less head (TBLH) and Z-scores were calculated (low BMD Z-score<-2, low-normal BMD Z-score between -1 and -2). Serum levels of osteocalcin (OC), procollagen type I C-terminal propeptide (PICP), bone alkaline phosphatase (bALP), bone tartrate-resistant acid phosphatase 5b (TRAP5b), vitamin D, parathormone (PTH), urinary calcium/creatinine (uCa/uCr) and urine deoxypyridinoline/creatinine (uDPD/uCr) were measured. Mean BMD Z-scores were lower than predicted at both sites of measurement. More specifically, 10% of CWH A had low and 20% low-normal BMD Z-scores in LS, whereas 9.1% had low-normal TBLH BMD Z-scores and there were no patients with low BMD Z-scores at this site of measurement. 36.7% of CWH had low vitamin D levels and 19.6% had a history of fracture. Also, patients with haemophilia had lower OC and higher uDPD/uCr levels while OC positively correlated to BMD Z-scores and uDPD/uCr negatively correlated to BMD Z-scores at both sites. No statistically significant differences were observed with regards to mode of treatment, number of haemorrhages and the presence of target-joints. CWH A had decreased BMD Z-scores at both sites with an uncoupling of bone turnover LS BMD seemed to be more affected than TBLH BMD.

Hemodiafiltration Is Associated With Reduced Inflammation and Increased Bone Formation Compared With Conventional Hemodialysis in Children: The HDF, Hearts and Heights (3H) Study

BackgroundPatients on dialysis have a high burden of bone-related comorbidities, including fractures. We report a post hoc analysis of the prospective cohort study HDF, Hearts and Heights (3H) to determine the prevalence and risk factors for chronic kidney disease-related bone disease in children on hemodiafiltration (HDF) and conventional hemodialysis (HD).MethodsThe baseline cross-sectional analysis included 144 children, of which 103 (61 HD, 42 HDF) completed 12-month follow-up. Circulating biomarkers of bone formation and resorption, inflammatory markers, fibroblast growth factor-23, and klotho were measured.ResultsInflammatory markers interleukin-6, tumor necrosis factor-α, and high-sensitivity C-reactive protein were lower in HDF than in HD cohorts at baseline and at 12 months (P < .001). Concentrations of bone formation (bone-specific alkaline phosphatase) and resorption (tartrate-resistant acid phosphatase 5b) markers were comparable between cohorts at baseline, but after 12-months the bone-specific alkaline phosphatase/tartrate-resistant acid phosphatase 5b ratio increased in HDF (P = .004) and was unchanged in HD (P = .44). On adjusted analysis, the bone-specific alkaline phosphatase/tartrate-resistant acid phosphatase 5b ratio was 2.66-fold lower (95% confidence interval, −3.91 to −1.41; P < .0001) in HD compared with HDF. Fibroblast growth factor-23 was comparable between groups at baseline (P = .52) but increased in HD (P < .0001) and remained unchanged in HDF (P = .34) at 12 months. Klotho levels were similar between groups and unchanged during follow-up. The fibroblast growth factor-23/klotho ratio was 3.86-fold higher (95% confidence interval, 2.15–6.93; P < .0001) after 12 months of HD compared with HDF.ConclusionChildren on HDF have an attenuated inflammatory profile, increased bone formation, and lower fibroblast growth factor-23/klotho ratios compared with those on HD. Long-term studies are required to determine the effects of an improved bone biomarker profile on fracture risk and cardiovascular health.

Association between Musculoskeletal Pain and Bone Turnover Markers in Long-Term Pb-Exposed Workers.

Background: On chronic exposure, Lead (Pb) deposits in the skeletal system, replaces calcium ions, and alters the normal physiological processes, which in turn, lead to stunting, delayed fracture healing, and high resorption of collagen molecules. The present study aimed to assess the association of musculoskeletal pain and discomfort with bone turnover markers (BTMs) among long-term Pb-exposed workers. Study design: A cross-sectional study. Methods: The study recruited 176 male Pb-exposed workers and 80 control subjects who were matched for age, gender, and socio-economic status. Blood lead levels (BLLs), bone growth markers, such as serum osteocalcin (OC), alkaline phosphatase (ALP), bone alkaline phosphatase (BAP), and bone resorption markers: serum pyridinoline (Pry), deoxypyridinoline (DPry), tartrate-resistant acid phosphatase-5b(TRACP-5b), and hydroxyproline in urine (HyP-U) of participants were investigated. Pain and discomfort in the musculoskeletal system were assessed using Nordic Musculoskeletal Questionnaire. Results: Pb-exposure was significantly associated with musculoskeletal discomfort of the lower back (P<0.001), upper back (P<0.001), and ankle/foot (P=0.011). Among bone formation markers, serum OC was significantly lower in musculoskeletal discomfort of elbows (P=0.033) and ankle/foot (P=0.042). Among bone resorption markers, serum DPry was significantly lower in musculoskeletal discomfort of the neck (P=0.049) and shoulders (P=0.023). HyP-U was significantly higher in musculoskeletal discomfort of shoulders (P=0.035) and lower back (P=0.036). Conclusion: As evidenced by the obtained results, Pb-exposure was associated with musculoskeletal discomfort of the lower back, upper back, and ankle/foot. Lower bone formation (serum OC) marker was noted with musculoskeletal discomfort of elbows and ankle/foot. Furthermore, bone resorption markers were associated with musculoskeletal discomfort of the neck, shoulders, and lower back. The findings of the present study suggested that long-term Pb-exposure and BTMs were associated with musculoskeletal discomfort.

Bone turnover and mineral metabolism in adult patients with hypophosphatasia treated with asfotase alfa

SummaryThere is limited understanding of how asfotase alfa affects mineral metabolism and bone turnover in adults with pediatric-onset hypophosphatasia. This study showed that adults with hypophosphatasia treated with asfotase alfa experienced significant changes in biochemical markers of bone and mineral metabolism, possibly reflecting enhanced bone remodeling of previously osteomalacic bone.IntroductionHypophosphatasia (HPP), due to a tissue nonspecific alkaline phosphatase (TNSALP) deficiency, can cause impaired bone mineralization and turnover. Although HPP may be treated with asfotase alfa, an enzyme replacement therapy, limited data are available on how treatment with asfotase alfa affects mineral metabolism and bone turnover in adults with HPP.MethodsALP substrates, bone turnover and mineral metabolism markers, and bone mineral density (BMD) data from EmPATHY, a single-center, observational study of adults (≥ 18 years) with pediatric-onset HPP treated with asfotase alfa (NCT03418389), were collected during routine clinical care and analyzed from baseline through 24 months of treatment.ResultsData from 21 patients showed significantly increased ALP activity and reduced urine phosphoethanolamine (PEA)/creatinine (Cr) ratios after baseline through 24 months of asfotase alfa treatment. There were significant transient increases in parathyroid hormone 1-84 (PTH), osteocalcin, and procollagen type 1 N-propeptide (P1NP) levels at 3 and 6 months and in tartrate-resistant acid phosphatase 5b (TRAP5b) levels at 3 months, with a significant decrease in N-terminal telopeptide of type 1 collagen (NTX) levels at 24 months. Lumbar spine BMD T scores continuously increased during treatment.ConclusionSignificant changes in bone turnover and mineral metabolism markers after asfotase alfa treatment suggest that treatment-mediated mineralization may enable remodeling and bone turnover on previously unmineralized surfaces. Urine PEA/Cr ratios may be a useful parameter in monitoring treatment during routine care.

Bone mass protective potential mediated by bovine milk basic protein requires normal calcium homeostasis in mice

ObjectivesMilk provide protective effects against bone loss caused by an impaired calcium balance. Although the effects of some elements have previously been confirmed, the involvement of milk basic protein (MBP) in bone mineral metabolism remains poorly characterized. Moreover, the importance of mineral nutrition sufficiency to establish the effect of MBP must be evaluated. MethodsFirst, to evaluate the physiological conditions required for MBP activity, we examined the bone and mineral phenotypes of mice that suffer from insufficient calcium absorption due to a lack of intestinal vitamin D signaling. Second, to determine whether vitamin D signaling affects the effect of MBP on bone resorption, in vitro osteoclastogenesis were assessed using bone marrow cells. ResultsIn mice with systemic vitamin D receptor (Vdr) inactivation, dietary MBP supplementation was unable to normalize hypercalcemia and hyperparathyroidism and failed to rescue bone mineralization impairments. In contrast, calcium and bone homeostasis responded to MBP supplementation when Vdr inactivation was restricted to the intestines. Hyperparathyroidism in intestine-specific Vdr knockout mice was also improved by MBP supplementation, along with a decrease in bone resorption in response to the level of serum tartrate-resistant acid phosphatase 5b. These results corresponded with a reduction in tartrate-resistant acid phosphatase-stained osteoclast numbers and the eroded surface on the tibia. MBP treatment dose-dependently suppressed osteoclastogenesis in cultured bone marrow macrophages regardless of vitamin D activity. These effects of MBP were blunted when parathyroid hormone was added to the culture medium, which is in line with the in vivo phenotype observed with systemic Vdr inactivation and suggests that severe hyperparathyroidism limits MBP activity in the bone. ConclusionsTherefore, adaptive calcium homeostasis is an essential requirement when MBP exerts protective effects through the inhibition of bone resorption.

Adjuvant endocrine therapy effects on bone mineral density and microstructure in women with breast cancer

Although aromatase inhibitors (AIs) are typical drugs for cancer treatment-induced bone loss, their effects on the bone microstructure remain unclear. In this study, we evaluated changes in the bone mineral density (BMD) and bone microstructure associated with AI treatment using high-resolution peripheral quantitative computed tomography (HR-pQCT) in patients with early breast cancer. This prospective, single-arm, observational study included non-osteoporotic, postmenopausal women with hormone receptor-positive breast cancer. Patients underwent dual-energy X-ray absorptiometry (DXA), HR-pQCT, and tartrate-resistant acid phosphatase-5b (TRACP-5b) or procollagen type-I N-terminal propeptide measurements at baseline and 6 and 12months after AI therapy. The primary endpoint was changes in the total volumetric BMD (Tt.vBMD), trabecular vBMD (Tb.vBMD), and cortical vBMD (Ct.vBMD) longitudinally at the distal radius and tibia. Twenty women were included (median age 57.5years; range 55-72years). At 12months, HR-pQCT indicated a significant decrease in the Tt.vBMD (median distal radius -5.3%, p < 0.01; distal tibia -3.2%, p < 0.01), Tb.vBMD (-3.2%, p < 0.01; -1.0%, p < 0.05, respectively), and Ct.vBMD (-3.2%, p < 0.01; -2.7%, p < 0.01, respectively). Estimated bone strength was also significantly decreased. The DXA BMD value in the total hip (p < 0.01) and femoral neck (p = 0.03), but not in the lumbar spine, was significantly decreased. The TRACP-5b levels was significantly negatively associated with changes in the Tt.vBMD in both the distal radius and tibia (r = -0.53, r = -0.47, respectively) CONCLUSION: Postmenopausal women who received AIs for early breast cancer experienced significant trabecular and cortical bone deterioration and a decrease in estimated bone strength within only 1year.

Phase 2 Study of Carfilzomib and Bone Metabolism in Patients with Relapsed Multiple Myeloma

Background: Carfilzomib is an epoxyketone proteasome inhibitor (PI) that binds selectively and irreversibly to the constitutive proteasome and immunoproteasome. In addition to its direct cytotoxicity, PIs possess bone anabolic effect, by facilitating osteoblast differentiation, which in turn, inhibits multiple myeloma (MM) growth. However, the relative contribution of carfilzomib on direct tumor killing and bone formation has been difficult to discern when it is used in combination with other agents such as steroids, chemotherapeutics, or immunomodulatory agents (iMiDs); most of which increase bone loss.Objective: We explored the effect of single agent carfilzomib on bone metabolism and disease response in patients with relapsed MM.Methods: We conducted a phase II study of single-agent carfilzomib at 56 mg/m2 day 1, 2, 8, 9, 15, 16 of a 28 day cycle in patients with relapsed MM. Serum markers of bone resorption including carboxyterminal cross-linked telopeptide of type I collagen (ICTP) and tartrate resistant acid phosphatase 5b (TRACP 5b) and bone formation markers including N-terminal type 1 procollagen (P1NP) and osteocalcin (OC) were analyzed at baseline and 6 months after therapy, and compared using paired t-tests. Imaging using 18F-NaF PET-CT scans to monitor bone response and 8F-FDG PET-CT to measure localized tumor burden were done at baseline and 6 months post treatment. For each patient, the three sites with the highest baseline SUVs and an iliac crest (representing an uninvolved site) were selected for quantitative comparisons using paired t-test.Results: Ten patient were accrued. Average age was 70.1 +/- 6.79 (mean +/- SD) years. Median prior lines of therapy was 2. 9 were evaluable for response. Overall response rate was 77.8%: VGPR (1/9: 11%), partial response (6/9: 67%), stable disease (2/9: 22%). Median PFS was 11.5 months. Overall survival was 21.9 months. Common treatment adverse events (AE) were comparable to previous reports including diarrhea (40%), fever (40%), dyspnea (30%) and fatigue (30%). Grade 3/4 AEs, occurring in 10% each included neutropenia, thrombocytopenia, dyspnea, fatigue, vomiting, ataxia, heart failure and pulmonary edema.Serum ICTP and TRACP were elevated at baseline in 80% of subjects; means (+/- SD) of serum ICTP (14.4 +/- 3.1 ug/L) and TRACP (8.2 +/- 4.2 U/L), respectively. Both markers declined significantly after therapy (means +/- SD of serum ICTP 8.4 +/- 3.1 ug/L and TRACP (5.2 +/- 4.2 U/L); p<0.01), suggesting osteoclast suppression. Serum PINP and OC varied among subjects; 5 patients with low levels and 4 normal levels and 1 with elevated level: mean (+/- SD) of PINP 25 +/- 23 ug/L and OC (18 +/- 12 ug/L). Neither post treatment mean PINP nor OC differed significantly from baseline overall (p=0.16). However, all subjects who achieve PR or better had a more than 25% increase in PINP and OC, suggesting increased new bone formation.8F-FDG and 18F-NaF were positive in all subjects. Number of active sites and SUVs vary largely among subjects. 18F-NaF images mirrored the 8F-FDG signals, appearing as rings around the tumor sites, suggesting a bone reaction to the localized tumors. 6 of 9 subjects had post treatment PET/CTs for paired comparison. Post treatment 8F-FDG SUV of representative sites decreased significantly, while 18F-NaF SUVs varied; 58% of lesions had a decline in SUV, while 32% were unchanged and 10% were brighter, suggesting heterogeneous bone response. 18F-NaF SUVs of iliac sites decreased post treatment, suggesting a lower bone turnover overall. CT scans showed increased sclerosis at the rim of lytic lesions without significant change in lesion size, at this early time point.Conclusion: Carfilzomib at 56 mg/m2 dosing strongly inhibits bone resorption and demonstrates bone anabolic effects, in addition to cytotoxicity, in relapsed MM. Single agent use allows bone response analysis without the confounding effects of other anti-MM agents used in combination therapy. Response rates and side effects are favorable to prior reports. Serum markers of bone remodeling are useful for an assessment of overall bone metabolism while the 18F-NaF PET/CT allows assessment of both local and systemic changes but lacks specificity. Studies of osteoblast differentiation from subject's bone marrow stromal cells pre and post treatment are ongoing. Whether carfilzomib can heal bone lesions requires longer follow up. DisclosuresSuvannasankha:Celgene: Research Funding; Bristol-Myers Squibb: Research Funding; Takeda: Research Funding; Janssen Pharmaceuticals: Research Funding. Abonour:Celgene: Other: Steering committee, Research Funding; Prothena: Research Funding; Takeda: Other: Steering committee, Research Funding. Roodman:PharmaMar: Research Funding.

Depletion of Mature B Cells By Anti CD20-Directed Antibody Induces Trabecular Bone Loss in Young Mice: An Alert for Clinicians?

Targeted B cell therapy by anti-CD20 antibodies has revolutionized the treatment of B cell malignancies, immune cytopenias as well as connective tissue diseases. A growing body of evidence links B cell homeostasis and bone metabolism, including reports demonstrating severe bone loss in transgenic mice lacking IgM+ B cells. Here, we addressed the hypothesis that pharmacological depletion of mature B cells induces bone loss in an age-dependent manner.Ten-week and one-year old C57BL/6J female mice were treated with 150µg and 250µg, respectively, of mouse anti-mouse CD20 (hereafter αCD20), administered in two equal doses, two weeks apart. Four weeks after the first injection, distal femurs and L3 vertebrae (old mice only) were subjected to bone morphometric analysis using µCT. The efficiency of B cell depletion as well as immunophenotyping of the bone marrow (BM) and spleen were assessed by flow cytometry. We used ELISA to measure serum markers of bone formation (N-terminal propeptide of procollagen 1, P1NP)and bone resorption (tartrate-resistant acid phosphatase 5b, TRAcP 5b). The expression of receptor activator of nuclear factor κ-B ligand (RANKL), osteoprotegerin (OPG) and interleukin 7 (IL-7) in both BM containing bone fragment (BM-BF) and spleen was analyzed by means of RT-qPCR. To investigate the osteoclastogenic potential of various BM B cell subsets, total BM cells from wild-type untreated mice were flow-sorted and subjected to an osteoclastogenesis assay in RANKL- and macrophage colony stimulating factor (M-CSF)-containing medium.In young mice with >90% splenic B cell depletion (5 out of 7), αCD20 treatment (compared to diluent controls) induced a significant trabecular bone loss, reflected by 25% decrease in bone mineral density (30.1±2.5 vs 40±3.7 mgHA/cm3), 44% decrease in connectivity density (10.3±1.5 vs 18.9±2.8 mm-3), 13% reduction in trabecular number (2.8±0.13 vs 3.24±0.13 mm-1), and corresponding 14% increase in trabecular separation (p<0.05 for all parameters, n=5-7). However, no effect of αCD20 on bone mass was noted in the old mice (neither in femurs nor in L3 vertebrae, n=6 in each group).αCD20 treatment induced a compensatory increase in BM pro-B cells in young animals (2.8±0.5% vs 1.8±0.1%, p<0.05) which was even more pronounced in old ones. However, only the former exhibited a 2.5-fold increase in M-CSF receptor (CD115) positivity of this population (p<0.05). In that respect, our osteoclastogenic assays showed that among various BM B cell populations, only CD115+ pro-B cells could give rise to functional osteoclasts (18±6.5% OC area vs none). Moreover, serum P1NP levels were significantly lower in the young but not in the old αCD20-treated animals as compared to their corresponding diluent controls (48.2±8.7 ng/mL vs 74.5±4.6 and 27.9±8.2 ng/mL vs 30.3±10.8, respectively), indicating decreased bone formation in the former. αCD20 treatment did not affect the levels of TRAcP 5b, a surrogate marker of the total osteoclast number. Following B cell depletion, the expression of IL-7, known to induce bone loss, was upregulated in both BM-BF and spleen by the magnitude of 37% and 50%, respectively (p<0.05). Yet, the expression of RANKL and OPG in BM-BF and in spleen, were similar in control and in the αCD20 treatment groups.The current study demonstrates for the first time the adverse skeletal outcomes of pharmacological depletion of mature B cells, conceivably fueled by B cell-derived osteoclastogenesis in the presence of decreased bone formation. Since many patients treated with clinically available anti-CD20 antibodies already have other risk factors for bone loss, e.g. female gender and glucocorticoid therapy, these data also have immediate clinical implications. DisclosuresNo relevant conflicts of interest to declare.

FC 133HEMODIAFILTRATION IS ASSOCIATED WITH REDUCED INFLAMMATION AND INCREASED BONE TURNOVER COMPARED TO CONVENTIONAL HEMODIALYSIS IN CHILDREN - THE HDF, HEART AND HEIGHT (3H) STUDY

Abstract Background and Aims Children on dialysis have a high burden of bone related comorbidities and fractures. We report a post-hoc analysis of the HDF-Hearts-Height study to determine the prevalence and risk factors for mineral bone disease in children on hemodiafiltration (HDF) and conventional hemodialysis (HD). Method 144 children were included in baseline cross-sectional analysis, of which 103 (61 HD, 42 HDF) completed 12-month follow-up. Biomarkers of bone formation and resorption, inflammatory markers, fibroblast growth factor-23 (FGF23) and klotho were measured. Results Inflammatory markers interleukin-6 [IL-6], tumor necrosis factor-alfa [TNF-α], and high-sensitivity CRP [hsCRP] were lower in the HDF compared to HD cohorts at baseline and 12 months (p&amp;lt;0.001). Concentrations of bone formation (bone-specific alkaline phosphatase, BAP) and resorption (tartrate-resistant acid phosphatase 5b [TRAP5b]) markers were comparable between cohorts at baseline, but after 12-months the BAP/TRAP5b ratio increased in HDF (p=0.004) and was unchanged in HD (p=0.44). On adjusted analysis the BAP/TRAP5b ratio was 2.66-fold lower (95%CI -3.91, -1.41; p&amp;lt;0.0001) in HD compared to HDF. FGF23 was comparable between groups at baseline (p=0.52) but increased in HD (p&amp;lt;0.0001) and remained static in HDF (p=0.34) at 12-months. Klotho levels were similar between groups and unchanged during follow-up. The FGF23/klotho ratio was 3.86-fold higher (95% CI 2.15, 6.93; p&amp;lt;0.0001) in HD compared to HDF. Conclusion We conclude that children on HDF have increased bone turnover, an attenuated inflammatory profile and lower FGF23/klotho ratios compared to those on HD. Long-term studies are required to determine the effect, if any, of an improved bone biomarker profile on fracture risk and growth.

Metabolomics analysis of the therapeutic effects of Qiwei Tongbi oral liquid on rheumatoid arthritis in rats

Qiwei Tongbi oral liquid (QWTB), a classical traditional Chinese medicine (TCM) formula, has a good therapeutic effect on rheumatoid arthritis (RA) and is widely used in China. To comprehensively elucidate the therapeutic mechanism of QWTB in the treatment of RA, the effects of QWTB on biomarkers and metabolic pathways in a rat model of kidney deficiency arthritis were investigated in this study. The effects of QWTB on pharmacodynamic indicators, including paw swelling, arthritis score; interleukin-1β, interleukin-6, interleukin-17 F, tumor necrosis factor-α, tartrate-resistant acid phosphatase 5b, bone alkaline phosphatase, bone-specific alkaline phosphatase, bone glaprotein, urea, and creatinine levels; and histopathology, suggested that QWTB significantly improved renal function, inhibited the inflammatory response, and reduced bone loss. In total, 39 differential metabolites were screened by comparing the endogenous components between blank and model rat plasma, among which 16 metabolites were altered by QWTB. The metabolism pathway analysis revealed that α-linolenic acid metabolism, phenylalanine metabolism, sphingolipid metabolism, histidine metabolism and glycerophospholipid metabolism were greatly disturbed. Thus, the biomarkers investigated included (1) α-linolenic acid, (2) hippuric acid, (3) phosphatidylethanolamine (15:0/22:2(13Z,16Z)), (4) phenylpyruvic acid, (5) sphinganine, and (6) urocanic acid. QWTB affected three abnormal biomarkers: (3), (4), and (6). Phenylphruvic acid, sphinganine and urocanic acid were significantly associated with pharmacodynamic indicators, as shown by Pearson correlation analysis. These results indicated that RA-related biomarkers had certain reliability and biological significance. In summary, QWTB regulated the metabolic disorders in rats with RA. Its therapeutic mechanism may involve the regulation of phenylalanine metabolism, histidine metabolism, and glycerophospholipid metabolism. The results of this study are useful for understanding the therapeutic mechanisms of TCM.

The role of Serpina3n in the reversal effect of ATRA on dexamethasone-inhibited osteogenic differentiation in mesenchymal stem cells

BackgroundGlucocorticoid-induced osteoporosis (GIOP) is the most common secondary osteoporosis. Patients with GIOP are susceptible to fractures and the subsequent delayed bone union or nonunion. Thus, effective drugs and targets need to be explored. In this regard, the present study aims to reveal the possible mechanism of the anti-GIOP effect of all-trans retinoic acid (ATRA).MethodsBone morphogenetic protein 9 (BMP9)-transfected mesenchymal stem cells (MSCs) were used as an in vitro osteogenic model to deduce the relationship between ATRA and dexamethasone (DEX). The osteogenic markers runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), and osteopontin were detected using real-time quantitative polymerase chain reaction, Western blot, and immunofluorescent staining assay. ALP activities and matrix mineralization were evaluated using ALP staining and Alizarin Red S staining assay, respectively. The novel genes associated with ATRA and DEX were detected using RNA sequencing (RNA-seq). The binding of the protein–DNA complex was validated using chromatin immunoprecipitation (ChIP) assay. Rat GIOP models were constructed using intraperitoneal injection of dexamethasone at a dose of 1 mg/kg, while ATRA intragastric administration was applied to prevent and treat GIOP. These effects were evaluated based on the serum detection of the osteogenic markers osteocalcin and tartrate-resistant acid phosphatase 5b, histological staining, and micro-computed tomography analysis.ResultsATRA enhanced BMP9-induced ALP, RUNX2 expressions, ALP activities, and matrix mineralization in mouse embryonic fibroblasts as well as C3H10T1/2 and C2C12 cells, while a high concentration of DEX attenuated these markers. When DEX was combined with ATRA, the latter reversed DEX-inhibited ALP activities and osteogenic markers. In vivo analysis showed that ATRA reversed DEX-inhibited bone volume, bone trabecular number, and thickness. During the reversal process of ATRA, the expression of retinoic acid receptor beta (RARβ) was elevated. RARβ inhibitor Le135 partly blocked the reversal effect of ATRA. Meanwhile, RNA-seq demonstrated that serine protease inhibitor, clade A, member 3N (Serpina3n) was remarkably upregulated by DEX but downregulated when combined with ATRA. Overexpression of Serpina3n attenuated ATRA-promoted osteogenic differentiation, whereas knockdown of Serpina3n blocked DEX-inhibited osteogenic differentiation. Furthermore, ChIP assay revealed that RARβ can regulate the expression of Serpina3n.ConclusionATRA can reverse DEX-inhibited osteogenic differentiation both in vitro and in vivo, which may be closely related to the downregulation of DEX-promoted Serpina3n. Hence, ATRA may be viewed as a novel therapeutic agent, and Serpina3n may act as a new target for GIOP.

Voluntary running of defined distances alters bone microstructure in C57BL/6 mice fed a high-fat diet.

Obesity increases the risk for pathological conditions such as bone loss. On the other hand, physical exercise reduces body adiposity. To test the hypothesis that physical activity improves bone quality, we evaluated voluntary running of defined distances on trabecular and cortical microstructure in mice fed a high-fat diet (HFD). Sedentary mice were fed the standard AIN93G diet or the HFD. Mice fed the HFD remained sedentary or were assigned to unrestricted running or 75%, 50%, and 25% of unrestricted running with an average running activity at 8.3, 6.3, 4.2, and 2.1km per day, respectively. The bone structural differences found in sedentary mice were that HFD, compared with the AIN93G diet, resulted in a lower bone volume fraction (BV/TV) and a higher structure model index (SMI) in vertebrae. Running had a greater effect on trabecular microstructure in femurs than in vertebrae; the decrease in SMI and an increase in trabecular thickness (Tb.Th) were in dose-dependent manners. Running was positively correlated with BV/TV and Tb.Th and inversely correlated with SMI in femurs. The HFD increased plasma concentrations of tartrate-resistant acid phosphatase 5b, a marker of bone resorption, in sedentary mice, while running decreased it in a dose-dependent manner. The findings show that voluntary running improves bone quality in young adult mice fed an HFD. Novelty: The high-fat diet alters bone microstructure by increasing bone resorption. Quantitative voluntary running improves bone microstructure through its attenuation of bone resorption in mice fed a high-fat diet.

Correlation of Osteoporosis in Patients With Newly Diagnosed Type 2 Diabetes: A Retrospective Study in Chinese Population.

This study aimed to explore the risk factors attributed to osteoporosis in newly type 2 diabetes mellitus (T2DM) patients. This study aimed to recruit 244 T2DM patients and 218 non-diabetic controls. We collected demographic characteristics, medical history, bone mineral density and biomarkers including bone specific alkaline phosphatase (BALP), osteocalcin, N-terminal peptide of type I procollagen (P1NP), tartrate-resistant acid phosphatase 5b (TRCAP-5b), β-Cross Laps of type I collagen-containing cross-linked C-telopeptide (β-CTX), 25-hydroxyvitamin D, parathyroid hormone were recorded or detected. Bone mineral density (BMD) was our primary outcome. Based on the result of BMD, we divided both the control group and T2DM group into three subgroups: normal bone mass, osteopenia and osteoporosis. In control group, we found age, sex, menopausal status, BMI, P1NP, BALP, TRACP-5b, osteocalcin, and corrected serum calcium are differential among three subgroups. In T2DM group, we found age, sex, menopausal status, drinking status, BMI, HbA1c, TRACP-5b and OC were differential among three subgroups. In T2DM and control groups, age, female, postmenopausal status, BALP, TRACP-5b and osteocalcin were positively correlated while BMI was negatively correlated with osteoporosis. In control group, β-CTX was positively correlated with osteoporosis. In T2DM group, HbA1c and corrected serum calcium concentration were positively correlated with osteoporosis. After further adjustment of age, BMI in male, TRACP-5b was positively correlated with the risk of osteoporosis in newly diagnosed T2DM. After adjusted of age, BMI and menopausal status in female, OC was positively correlated with the risk of osteoporosis in newly diagnosed T2DM and controls. In female T2DM, BALP and P1NP were positively correlated with the risk of osteoporosis. In conclusion, age, BMI and menopausal status are common risk factors for osteoporosis in diabetic and non-diabetic patients, however TRACP-5b, BALP and osteocalcin are special risk factors for osteoporosis in newly diagnosed T2DM patients but not non-diabetic patients, which may be applied to identify osteoporosis risk in T2DM patients, but this result needs to be proven with fracture data.