Bone Microarchitecture Parameters Research Articles

In vivo impact on rabbit subchondral bone of viscosupplementation with a hyaluronic acid antioxidant conjugate

ObjectiveThis study aimed to assess the effects of an antioxidant-conjugated Hyaluronic Acid (HA), specifically HA-4-aminoresorcinol (HA4AR), on articular cartilage and subchondral bone in osteoarthritis (OA). We conducted a comparative analysis between HA4AR and a commercially available high molecular weight HA formulation in a rabbit model of OA.Materials and methodsEighteen rabbits underwent unilateral anterior cruciate ligament transection (ACLT) and were divided into three groups of six: Saline-group, HA-group, and HA4AR-group, based on the type of intra-articular injection received. Additionally, eight non-operated contralateral knees served as reference points (Contralateral-group). Six weeks post-surgery, iodine-enhanced micro-computed tomography imaging was used to evaluate articular cartilage volume and thickness, and to assess subchondral bone microarchitecture and mineral density.ResultsCartilage thickness in both the HA and HA4AR groups was comparable to that of the Contralateral group. Notably, there was a significant reduction in subchondral bone plate tissue mineral density in the HA-group when compared to both the HA4AR and Saline groups (p < 0.05). However, no significant differences in trabecular subchondral bone microarchitectural parameters and mineral densities were observed between the HA4AR-group and the Saline-group. When compared to the Contralateral, Saline, and HA4AR groups, the HA-group exhibited a marked decrease in subchondral bone plate tissue mineral density (p < 0.05). Additionally, a significant reduction in trabecular bone volume fraction was noted in the HA-group compared to the Contralateral-group (p < 0.05).ConclusionsThe HA-4AR hydrogel demonstrated significant preservation of subchondral bone plate tissue mineral density compared to HA alone, while other bone microarchitectural parameters remained largely unchanged. These findings indicate that HA-4AR may provide enhanced osteoprotective benefits in the treatment of osteoarthritis.

Effects of Semaglutide and Tirzepatide on Bone Metabolism in Type 2 Diabetic Mice.

Background/Objectives: Type 2 diabetes and weight loss are associated with detrimental skeletal health. Incretin-based therapies (GLP-1 receptor agonists, and dual GIP/GLP-1 receptor agonists) are used clinically to treat diabetes and obesity. The potential effects of semaglutide and tirzepatide on bone metabolism in type 2 diabetic mice remain uncertain. Methods: Combined streptozotocin and high fat feeding were employed in female C57BL/6J mice to promote hyperglycemia. Mice were administered for 4 weeks with a saline vehicle (sc., once-daily), semaglutide (40 μg/kg/d, sc., every three days), or tirzepatide (10 nmol/kg, sc., once-daily). Bone strength was assessed by three-point bending. Femur microarchitecture was determined by micro-CT, and bone formation and resorption parameters were measured by histomorphometric analysis. Serum was collected to measure bone resorption (C-telopeptide fragments of type I collagen, CTX) and formation (procollagen type 1 N-terminal propeptide, P1NP) biomarkers, respectively. The expression of bone metabolism-related genes was evaluated in the bone using RT-PCR. Results: Glucose levels significantly reduced after 4 weeks of semaglutide and tirzepatide treatment (both p < 0.05) compared with vehicle treatment. Tirzepatide led to more weight loss than semaglutide. Compared to saline-treated diabetic mice, the mean femur length was shorter in the tirzepatide group. After treatment with tirzepatide or semaglutide, cortical bone and trabecular bone parameters did not change significantly compared to saline-treated diabetic mice, except that cortical thickness was lower in the semaglutide group compared to the saline group (p = 0.032). Though CTX and P1NP levels decreased, however, the change in CTX and P1NP levels did not differ among the four groups during the 4 weeks of treatment (all p > 0.05). Semaglutide affected RANKL and OPG mRNA expression and increased the ratio of OPG/RANKL. No significant difference was found in the quantity of Col1a1, RANKL, OPG, and RUNX2 between tirzepatide- and saline-treated diabetic mice. Conclusions: The 4-week treatment with semaglutide and tirzepatide had a neutral effect on bone mass compared with the controls, and most of the bone microarchitecture parameters were also comparable between groups in diabetic mice. A better understanding of incretin-based therapies on bone metabolism in patients with diabetes requires further evaluation in large clinical trials.

8693 Dexamethasone Increases Bone Mass In Mice

Abstract Disclosure: F. Sultana: None. M.L. Temple: None. S. Kim: None. V. Ryu: None. F. Korkmaz: None. A. Gumerova: None. P. Georgii: None. U. Cheliadinova: None. D. Vasilyeva: None. V. Laurencin: None. R. Witztum: None. O. Barak: None. L. Cullen: None. A.R. Pallapati: None. S. Rojekar: None. J. Gimenez Roig: None. D. Lizneva: None. W. Zhou: None. T. Yuen: None. M. Zaidi: None. Severe osteoporosis and bone fractures are strongly associated with long-term or recurring glucocorticoid use in people. Steroids have traditionally been used as an anti-inflammatory and immunosuppressive agent, and while their use has been widely implicated in causing bone loss by suppressing osteoblastic bone formation, recent data have consistently shown mixed results in mice. It is also important to stress that dexamethasone (DEX) in particular has been used for over three decades in cell cultures to stimulate osteoblast formation. We therefore posit that mechanisms other than direct steroid effects on osteoblasts contribute to bone loss, and that, at least physiologically, steroids stimulate (rather than inhibit) bone formation. Here, we have aimed to clarify the effects of DEX on bone metabolism. We first studied the effects of low and high doses of DEX (1 mg/kg and 10 mg/kg), given daily as intraperitoneal injections to 8-week-old C57BL/6 mice. Micro-CT showed that DEX markedly increased bone microarchitecture, including the bone volume fraction, connectivity density, and trabecular number, while decreasing trabecular spacing in femoral epiphyses compared with vehicle-injected controls. Serum levels of adrenocorticotropic hormone (ACTH) and corticosterone did not change following DEX. These findings suggest that both low and high DEX doses are anabolic to the skeleton. Reaffirming these data, we found that adrenalectomy (with salt replacement) caused a significant decline in bone mineral density. Likewise, Tpit-/- mice, in which ACTH and corticosterone are both reduced, display reduced bone microarchitectural parameters, together with low mineralizing surface and bone formation rates on dynamic histomorphometry, impaired osteoblast formation in Cfu-f cultures, and lowered expression of the osteoblastogenesis genes Bmp2, Ibsp, Runx2 and Sp7. Finally, deleting the glucocorticoid receptor (Gr) specifically in osteoclasts in Acp5-Cre;Grfl/fl mice had no effect on microarchitectural parameters at the spine or proximal tibial metaphysis. However, and importantly, absent GR in the osteoclast, DEX increased bone mass significantly, suggesting that its notable anabolic action was mediated by the osteoblast. Our findings challenge the longstanding premise that steroids directly impact the osteoblast to cause bone loss, and that other mechanisms, such as reduced ACTH levels and/or central effects of DEX, may play a key role. Presentation: 6/1/2024

Quantification of bone microarchitecture using photon-counting CT at different radiation doses: A comparison with µCT

PurposeAccurate measurements of trabecular bone microarchitecture are required for a proper assessment of bone fragility. Photon-counting detector CT (PCD-CT) has different technical properties than conventional CT, resulting in higher resolution and thereby potentially enabling in-vivo measurement of trabecular microarchitecture. The purpose of this study was to quantify trabecular bone microarchitectural parameters with PCD-CT at varying radiation doses and compare this to µCT as gold standard. MethodBoth distal radii, distal tibiae, femoral heads, and two vertebrae were dissected from one human. All specimens were scanned ex-vivo on a PCD-CT system (slice increment 0.1 mm; pixel size 0.1042–0.127 mm) and a µCT system (isotropic voxel size 49–68.4 µm). The radiation doses of the PCD-CT scans were varied from 2.5 to 120 mGy based on the volume CT dose index (CTDIvol32). For the PCD-CT scans, contrast-to-noise ratio and trabecular sharpness were calculated and compared between radiation doses. µCT and PCD-CT scans were registered. The trabecular bone was then segmented from all PCD-CT and µCT scans and split into cubes with 6-mm edge length. For each cube, bone volume over total volume, trabecular thickness, trabecular number, and trabecular heterogeneity were calculated and compared between corresponding PCD-CT and µCT cubes. ResultsWith increasing dose, contrast-to-noise ratio and trabecular sharpness values increased for the PCD-CT images. Already at the lowest dose, high correlations between the trabecular microarchitectural parameters between µCT and PCD-CT were found (R2 = 0.55–0.95), which improved with increasing radiation dose (R2 = 0.76–0.96 at 20 mGy). ConclusionsPCD-CT can be used to quantify trabecular bone microarchitecture, with accuracy comparable to µCT and at clinically relevant radiation doses.

Trabecular bone microstructure parameters as predictors for chronological age: a systematic review.

Estimating chronological age is crucial in forensic identification. The increased application of medical imaging in age analysis has facilitated the development of new quantitative methods for the macroscopic evaluation of bones. This study aimed to determine the association of age-related changes in the trabecular microstructure with chronological age for age estimation in forensic science through different non-invasive imaging techniques. This systematic review was reported according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. An electronic search was performed with PubMed/MEDLINE, Scopus, and Cochrane databases as well as with a Google Scholar search. Qualitative synthesis was performed using the Anatomical Quality Assessment tool. A detailed literature search yielded 3467 articles. A total of 14 articles were ultimately included in the study. A narrative approach was employed to synthesize the data. Microcomputed tomography, high-resolution peripheral quantitative computed tomography, and cone beam computed tomography have been used for the quantitative estimation of age. These imaging techniques aid in identifying the trabecular bone microarchitectural parameters for chronological age estimation. Age-related changes in trabecular bone included a decrease in the bone volume fraction, trabecular number, and connectivity density and an increase in trabecular separation. This study also revealed that morphometric indices vary with age and anatomical site. This study is registered with the International Prospective Register of Systematic Reviews (PROSPERO) with the registration number CDRD42023391873.

Freezing does not influence the microarchitectural parameters of the microstructure of the freshly harvested femoral head bone.

The femoral head is one of the most commonly used bones for allografts and biomechanical studies. However, there are few reports on the trabecular bone microarchitectural parameters of freshly harvested trabecular bones. To our knowledge, this is the first study to characterize the microstructure of femoral heads tested immediately after surgery and compare it with the microstructure obtained with conventional freezing. This study aims to investigate whether freezing at -80°C for 6weeks affects the trabecular microstructure of freshly harvested bone tissue. This study was divided into two groups: one with freshly harvested human femoral heads and the other withthe same human femoral heads frozen at -80°C for 6weeks. Each femoral head was scanned using an X-ray microcomputed tomography scanner (µCT) to obtain the microarchitectural parameters, including the bone volume fraction (BV/TV), the mean trabecular thickness (Tb.th), the trabecular separation (Tb.sp), the degree of anisotropy (DA), and the connectivity density (Conn.D). There was no statistically significant difference between the fresh and the frozen groups for any of the parameters measured. This study shows that freezing at -80°C for 6weeks does not alter bone microstructure compared with freshly harvested femoral heads tested immediately after surgery.

Bone microarchitecture and strength assessed by HRpQCT in individuals with type 2 diabetes and prediabetes: the Maastricht study.

Type 2 diabetes (T2D) is a prevalent disease and has been associated with an increased fracture risk despite normal or even higher areal BMD. The aim of this study was to estimate the association between glucose metabolism status (GMS) and measurements of glycemic control with HRpQCT parameters of bone microarchitecture and strength. Participants of the Maastricht study who underwent an HRpQCT scan at the distal radius and tibia were included. GMS was determined by use of an oral glucose tolerance test and grouped into a normal glucose metabolism (NGM), prediabetes, or T2D. Linear regression models were used, stratified by sex with multiple adjustments. This study incorporated cross-sectional data from 1400 (796 [56.9%] NGM, 228 [16.3%] prediabetes, and 376 [26.9%] T2D) men and 1415 (1014 [71.7%] NGM, 211 [14.9%] prediabetes, and 190 [13.4%] T2D) women. The mean age was 59.8 ± 8.6 and 57.6 ± 9.0 yr for men and women, respectively. After adjustment, T2D was associated with a higher total BMD measured by HRpQCT and cortical thickness, and a smaller total and trabecular area in men and women compared with NGM. In women, T2D was additionally associated with a higher stiffness and failure load at the radius. Results were more pronounced at the distal radius than at the distal tibia. To conclude, these findings suggest that in this cohort of Maastricht study participants, total and trabecular bone area are smaller, but bone microarchitecture, density, and bone strength assessed by HRpQCT are not impaired in individuals with T2D.

PPI use is not associated with bone microarchitecture and strength assessed with HR-pQCT after three-years follow-up in patients visiting the Fracture Liaison Service

BackgroundThe use of proton pump inhibitors (PPIs) has been associated with an increased fracture risk in observational studies. However, the reported association between PPI use and bone mineral density (BMD), bone microarchitecture, and bone strength is inconsistent. This study aims to assess the association between PPI use and bone microarchitecture and strength using high-resolution peripheral quantitative CT (HR-pQCT) in a three-year follow-up study in patients with a recent fracture visiting the Fracture Liaison Service (FLS). MethodsThis three-year prospective cohort study included FLS patients aged ≥ 50 years with a recent fracture (median age 62 [IQR 56–69] years, 68.7 % females) and without anti-osteoporosis treatment indication. HR-pQCT scans (distal radius and tibia) were obtained at baseline (T0) and three-year follow-up (T3). Volumetric bone mineral density and bone area, microarchitecture, and strength (micro-finite element analysis) were determined. The association between three-year continuous PPI use and the percentage change in HR-pQCT parameters between T0 and T3 was assessed using sex-stratified multivariate linear regression analyses. Covariates included age, BMI, vitamin-D deficiency (< 50 nmol/l), glucocorticoid use, and cardiovascular co-morbidity (males and females) fracture type (major/hip vs. all others, only males) and probable sarcopenia (only females). ResultsIn total, 282 participants had available medication data throughout follow-up, of whom 20.6 % were continuous PPI users. In both males and females with complete HR-pQCT follow-up data (males: N = 69 radius, N = 84 tibia; females: N = 147 radius, N = 168 tibia), PPI use was not associated with the percentage change of any of the bone microarchitecture or strength parameters between T0 and T3 at the radius and tibia as compared to non-use. ConclusionCompared to non-use, PPI use was not associated with the change of bone microarchitecture and strength in FLS patients at three years of follow-up. These results do not support that an altered bone microarchitecture or strength may contribute to the increased fracture risk associated with PPI use, as reported in observational studies.

Bone microarchitectural alterations associated with spinal cord injury: Relation to sex hormones, metabolic factors, and loading

ContextPeople living with spinal cord injury (SCI) are at high risk for bone fractures. Neural, hormonal and metabolic contributors to bone microarchitectural alterations are incompletely understood. ObjectiveTo determine the relationship of physical, metabolic and endocrine characteristics with bone microarchitecture, characterized using high-resolution peripheral quantitative computed tomography (HRpQCT) in SCI. DesignCross-sectional analyses of bone properties in people with SCI. ParticipantsTwenty adults with SCI and paraplegia (12) or motor incomplete quadriplegia (8). Outcome measuresDistal tibia and radius HRpQCT parameters, including density, microstructure and strength by microfinite element anaysis (μFEA); sex hormones; metabolic and inflammatory markers. ResultsThe mean age of the participants with SCI was 41.5 ± 10.3 years, BMI 25.7 ± 6.2 kg/m2, time since injury 10.4 ± 9.0 years. Participants with SCI had significantly lower median total (Z score − 3.3), trabecular (−2.93), and cortical vBMD (−1.87), and Failure Load by μFEA (−2.48) at the tibia than controls. However, radius vBMD, aBMD and microarchitecture were similar in participants with SCI and un-injured controls. Unexpectedly, C-Reactive Protein (CRP) was positively associated with tibial trabecular vBMD (β = 0.77, p = 0.02), thickness (β = 0.52, p = 0.04) and number (β = 0.92, p = 0.02). At the radius, estradiol level was positively associated with total vBMD (β = 0.59, p = 0.01), trabecular thickness (β = 0.43, p = 0.04), cortical thickness (β = 0.63, p = 0.01) and cortical porosity (β = 0.74 p = 0.04). ConclusionsRadius vBMD and microarchitecture is preserved but tibial total, cortical and trabecular vBMD, and estimated bone strength are markedly lower and bone microarchitectural parameters substantially degraded in people with SCI. The alterations in bone microarchitecture in people with SCI are likely multifactorial, however marked degradation of bone microarchitecture in tibia but not radius suggests that unloading is an important contributor of site-specific alterations of bone microarchitecture after SCI. Fracture prevention in SCI should focus on strategies to safely increase bone loading.ClinicalTrials.gov registration #: (NCT03576001).

Menopause-Associated Changes in Mandibular Bone Microarchitecture Are Site-Specific

BackgroundThere are conflicting reports on the effects of decreased estrogen levels on mandibular bone microarchitecture, and whether these effects are consistent throughout the mandible is unclear and may have important implications for treatment planning. PurposeThe goal of this study was to evaluate trabecular and cortical bone microstructure in the mandibular condyle and the mandibular basal bone and compare these sites between premenopausal and postmenopausal women. Study Design, Setting, SampleParticipants were recruited for a cross-sectional cohort study at Columbia University Irving Medical Center. Each participant had CBCT taken of their mandibular condyles and of the basal bone. Exclusion criteria for the population included a) current chemo- or immunotherapy; b) history of bisphosphonate or other osteoporosis therapy; and c) currently pregnant, nursing, or on hormonal birth control. Independent VariableThe predictor variables are menopausal status (before or after menopause) and mandibular region of interest (condyle/basal bone). Outcome VariableParameters of interest included the following indicators of bone quality: trabecular bone volume fraction (BVF), trabecular thickness, trabecular number, trabecular separation, cortical BVF, cortical thickness, and cortical porosity. CovariatesCovariates included demographic variables such as age, estrogen levels, and ethnicity. AnalysesQuantitative microstructure analyses were conducted on CBCT images, and differences between groups for continuous measures (including age) were assessed with unpaired t-test, and demographic variables were assessed by Chi-square. Statistical significance was recorded at p<0.05. ResultsThe premenopausal and postmenopausal groups each had 31 participants, with the following average age: premenopausal= 43.9±6.9 vs. postmenopausal = 57.5±7.6 years old; p<0.001, and estrogen levels: premenopausal =91.77±80.13 pg/ml vs. postmenopausal= 41.44±61.62 pg/ml; p <0.01). Postmenopausal women had significantly greater condylar trabecular separation (0.61±0.18 vs 0.47±0.11 mm; p<0.001) and lower trabecular number (1.03±0.18 vs 1.21±0.19 1/mm; p<0.001) compared to premenopausal women. There were no significant differences in the basal bone microarchitectural parameters between the menopausal groups. Conclusion and RelevanceMenopause is associated with mandibular condylar trabecular bone loss but has minimal effects on the mandibular basal bone. This may have important ramifications for treatment planning in advanced age individuals.

Targeting FGFR3 signaling and drug repurposing for the treatment of SLC26A2-related chondrodysplasia in mouse model

BackgroundMutations in Slc26a2 cause a spectrum of autosomal-recessive chondrodysplasia with a significant and negligible influence on the quality of life. It has been reported that Slc26a2 deficiency triggers the ATF6 branch of the UPR, which may, in turn, activate the negative regulator of the FGFR3 signaling pathway. However, the correlation between the deletion of Slc26a2 and the augmentation of downstream phosphorylation of FGFR3 has not been investigated in vivo. MethodsFirst, we constructed Slc26a2 and Fgfr3 double knockout mouse lines and observed gross views of the born mice and histological staining of the tibial growth plates. The second approach was to construct tamoxifen-inducible Cre-ERT2 mouse models to replicate SLC26A2-related non-lethal dysplastic conditions. Pharmacological intervention was performed by administering the FGFR3 inhibitor NVP-BGJ398. The effect of NVP-BGJ398 on chondrocytes was assessed by Alcian blue staining, proliferation, apoptosis, and chondrocyte-specific markers and then verified by western blotting for variations in the downstream markers of FGFR3. The growth process was detected using X-rays, micro-CT examination, histomorphometry staining of growth plates, and immunofluorescence. ResultsGenetic ablation of Fgfr3 in embryonic Slc26a2-deficient chondrocytes slightly attenuated chondrodysplasia. Subsequently, in the constructed mild dysplasia model, we found that postnatal intervention with Fgfr3 gene in Slc26a2-deficient chondrocytes partially alleviated chondrodysplasia. In chondrocyte assays, NVP-BGJ398 suppressed the defective phenotype of Slc26a2-deficient chondrocytes and restored the phosphorylation downstream of FGFR3 in a concentration-dependent manner. In addition, in vivo experiments showed significant alleviation of impaired chondrocyte differentiation, and micro-CT analysis showed a clear improvement in trabecular bone microarchitectural parameters. ConclusionOur results suggested that inhibition of FGFR3 signaling pathway overactivation and NVP-BGJ398 has promising therapeutic implications for the development of SLC26A2-related skeletal diseases in humans. The translational potential of this articleOur data provide genetic and pharmacological evidence that targeting FGFR3 signaling via NVP-BGJ398 could be a route for the treatment of SLC26A2-associated skeletal disorders, which promisingly advances translational applications and therapeutic development.

Exploring the potential mechanism of Heng-Gu-Gu-Shang-Yu-He-Ji therapy for osteoporosis based on network pharmacology and transcriptomics

Ethnopharmacological relevanceHeng-Gu-Gu-Shang-Yu-He-Ji (Osteoking, OK) is a well-known formula for fracture therapy. In clinic, OK is effective in treating fractures while alleviating osteoporosis (OP) symptoms. However, active components of OK and the associated molecular mechanisms remain not fully elucidated. Aim of the studyThis study aims to systematically evaluate the anti-osteoporosis efficacy of OK and for the first time combine network pharmacology with high-throughput whole gene transcriptome sequencing to study its underlying mechanism. Materials and methodsIn this study, the osteoporosis model was established by the castration of both ovaries. The level of serum bone turnover factor was detected by enzyme-linked immunosorbent assay. Micro-CT and HE staining were used to observe the changes of bone histopathology, and nano-indentation technique was used to detect the biomechanical properties of rat bone. The main active Chemical components of OK were identified using UPLC-DAD. Efficacy verification and mechanism exploration were conducted by network pharmacology, molecular docking, whole gene transcriptomics and in vivo experiments. ResultsIn our study, OK significantly improved bone microarchitecture and bone biomechanical parameters in OVX rats, reduced osteoclast indexes such as C-telopeptide of type I collage (CTX-I) and increased Osteoprotegerin (OPG)/Receptor activator of NF-κB ligand (RANKL) levels. Mechanistically, PI3K/AKT pathway was a common pathway for genome enrichment analysis (KEGG) of both network pharmacology and RNA-seq studies. G protein-β-like protein (GβL), Ribosomal-protein S6 kinase homolog 2 (S6K2), and Phosphoinositide 3-kinase (PI3K) appeared differentially expression in the PI3K-AKT signaling pathway. These results were also confirmed by qRT-PCR and immunohistochemistry. ConclusionsOK may be used to treat osteoporosis, at least partly by activating PI3K/AKT/mTORC1 signaling pathway.

The effect of zoledronic acid on hip geometry in renal transplant recipients: a double-blind placebo-controlled randomized study

BackgroundIn renal transplant patients, bisphosphonates may prevent bone loss, but little is known about their effects on bone microarchitecture and geometrical hip parameters, as the key factors of bone stability. This study aimed to analyze the effect of zoledronic acid on the mentioned parameters in kidney transplant patients.MethodsIn this double-blind, randomized trial, 33 patients were followed for six months after administering either 4mg of zoledronic acid or a placebo. Bone mineral density (BMD) measurement of the spine, hip, radius, and whole body was obtained, and trabecular bone score (TBS) was evaluated using the software. Geometric assessment at the proximal femur was performed by the HSA program.ResultsEighteen patients in the intervention group and 15 in the control group completed the study. The mean percentages of the changes in the BMD at the lumbar spine and whole body were significantly different between the placebo and intervention groups (-0.23% vs. 4.91% and -2.03% vs. 1.23%) (P < 0.05). Zoledronic acid appeared to enhance the subperiosteal diameter, endocortical diameter, and cross-sectional moment of inertia (CSMI) at the narrow neck in comparison with placebo (P < 0.05); however, no difference in TBS was observed between both groups (P > 0.05).ConclusionsWe concluded that a single administration of zoledronic acid might ameliorate bone loss at the lumbar spine and the whole body and maintain the subperiosteal diameter, endocortical diameter, and CSMI as parameters of bone strength at the narrow neck of the proximal femur after six months in renal-transplant recipients.Trial registrationThis study was registered in IRCT (ID: IRCT20181202041821N1) on 04–05-2019.

THU419 Comprehensive Evaluation Of Osteoporosis And Impaired Bone Micro Architecture In Patients With Transfusion Dependent Beta Thalassemia

Abstract Disclosure: P. Dutta: None. comprehensive evaluation of osteoporosis and impaired bone micro architecture in patients with transfusion dependent beta thalassemia Authors: Pinaki Dutta, Liza Das, Alka Khadwal, Pankaj Malhotra, Sanjay Kumar Bhadada, Vandana Dhiman, Vivek Sharma, Raman Introduction: Thalassemia-related bone disease is an entity that includes low bone mass, fractures and impaired bone microarchitecture. The current study was designed to evaluate the prevalence and determinants of low bone mass, osteoporosis and microarchitectural parameters using high-resolution peripheral quantitative computed tomography (HR-pQCT) in patients with transfusion-dependent thalassemia (TDT). Patients and Methods TDT patients (n=42) and age- and BMI-matched controls (n=40) were recruited. Demographic, biochemical and hormonal parameters were assessed. Bone mineral density (BMD), T- and Z- scores were measured by DEXA using the HOLOGIC Discovery A Viewer 6 machine. Bone microarchitecture was estimated by HR-pQCT (XtremeCT II, Scanco Medical AG, Switzerland). Results: The mean age of thalassemia patients at recruitment was 26.9 ± 6.5 years and that of controls was 28.9 ± 7.9 years. The mean age of transfusion initiation was 7 months (IQR 3-18) and the median number of transfusions required was 24 (15-25) annually. Serum calcium in cases (8.87 ± 0.67mg/dl) was significantly lower than the controls (9.40 ± 0.45mg/dl), with 9 patients having hypocalcemia and 7 of these having hypoparathyroidism. Hypogonadism was present in 40.4% patients. Bone turnover markers (P1NP, β-CTX) were not significantly different between cases and controls, except alkaline phosphatase (ALP) which was higher in the cases (102.6 ± 60.7 U/l) as opposed to controls (78.2 ± 32.1 U/l). There was a significant correlation between ALP and liver dysfunction (alanine transaminase exceeding the upper normal limit) (r = 0.36, p=0.01) and 63.6% of those with elevated ALP had liver dysfunction. Low bone mass (Z ≤−2) was present in significantly greater proportion of patients versus controls at femoral neck (29.3% vs 2.4%) and at lumbar spine (52.4% vs 14.7%). Total, cortical and trabecular volumetric bone mineral density were significantly lower in cases as compared to controls(p&amp;lt;0.05). The trabecular number was significantly lower (p&amp;lt;0.0001) and trabecular separation significantly higher at radius and tibia (p&amp;lt;0.001). Cortical thickness at radius and tibia were also significantly lower in cases than controls (p&amp;lt;0.0001). Conclusion: Patients with transfusion-dependent thalassemia have significantly lower areal and volumetric BMD (both cortical and trabecular). The bone microarchitecture is also significantly impaired in terms of lower number and wider spacing of trabeculae as well as lower cortical thickness at both radius and tibia. Presentation: Thursday, June 15, 2023

Arctiin attenuates iron overload‑induced osteoporosis by regulating the PI3K/Akt pathway.

Iron overload is a prevalent pathological factor observed among elderly individuals and those with specific hematological disorders, and is frequently associated with an elevated incidence of osteoporosis. Although arctiin (ARC) has been shown to possess antioxidant properties and the ability to mitigate bone degeneration, its mechanism of action in the treatment of iron overload‑induced osteoporosis (IOOP) remains incompletely understood. To explore the potential molecular mechanisms underlying the effects of ARC, the MC3T3‑E1 cell osteoblast cell line was used. Cell Counting Kit was used to assess MC3T3‑E1 cell viability. Alkaline phosphatase staining and alizarin red staining were assessed for osteogenic differentiation. Calcein AM assay was used to assess intracellular iron concentration. In addition, intracellular levels of reactive oxygen species (ROS), lipid peroxides, mitochondrial ROS, apoptosis rate and mitochondrial membrane potential changes in MC3T3‑E1 cells were examined using flow cytometry and corresponding fluorescent dyes. The relationship between ARC and the PI3K/Akt pathway was then explored by western blotting and immunofluorescence. In addition, the effects of ARC on IOOP was verified using an iron overload mouse model. Immunohistochemistry was performed to evaluate expression of osteogenesis‑related proteins. Micro-CT and H&E were used to analyze bone microstructural parameters and histomorphometric indices in the bone tissue. Notably, ARC treatment reversed the decreased viability and increased apoptosis in MC3T3‑E1 cells originally induced by ferric ammonium citrate, whilst promoting the formation of mineralized bone nodules in MC3T3‑E1 cells. Furthermore, iron overload induced a decrease in the mitochondrial membrane potential, augmented lipid peroxidation and increased the accumulation of ROS in MC3T3‑E1 cells. ARC not only positively regulated the anti‑apoptotic and osteogenic capabilities of these cells via modulation of the PI3K/Akt pathway, but also exhibited antioxidant properties by reducing oxidative stress. Invivo experiments confirmed that ARC improved bone microarchitecture and biochemical parameters in a mouse model of iron overload. In conclusion, ARC exhibits potential as a therapeutic agent for IOOP by modulating the PI3K/Akt pathway, and via its anti‑apoptotic, antioxidant and osteogenic properties.

Osteogenic Effects of KY-054, a Novel Coumarin Derivative on Femur Cortical Bone in Ovariectomized Female Rats.

Osteoporosis is treated with oral and parenteral bone resorption inhibitors such as bisphosphonates, and parenteral osteogenic drugs including parathyroid hormone (PTH) analogues and anti-sclerostin antibodies. In the present study, we synthesized KY-054, a 4,6-substituted coumarin derivative, and found that it potently promoted osteoblast differentiation with an increase in alkaline phosphatase (ALP) activity at 0.01-1 µM in mouse-derived mesenchymal stem cells (ST2 cells) and rat bone marrow-derived mesenchymal stem cells (BMSCs). In the ovariectomized (OVX) rats, KY-054 (10 mg/kg/d, 8 weeks) increased plasma bone-type ALP activity, suggesting in vivo promoting effects on osteoblast differentiation and/or activation. In dual-energy X-ray absorption (DEXA) scanning, KY-054 significantly increased the distal and diaphyseal femurs areal bone mineral density (aBMD) that was decreased by ovariectomy, indicating its beneficial effects on bone mineral contents (BMC) and/or bone volume (BV). In micro-computed tomography (micro-CT) scanning, KY-054 had no effect on metaphysis trabecular bone loss and microarchitecture parameters weakened by ovariectomy, but instead increased metaphysis and diaphysis cortical bone volume (Ct.BV) and cortical BMC (Ct.BMC) without reducing medullary volume (Med.V), resulting in increased bone strength parameters. It is concluded that KY-054 preferentially promotes metaphysis and diaphysis cortical bone osteogenesis with little effect on metaphysis trabecular bone resorption, and is a potential orally active osteogenic anti-osteoporosis drug candidate.

Bone parameters in T1D and T2D assessed by DXA and HR-pQCT – A cross-sectional study: The DIAFALL study

People with type 1 diabetes (T1D) and type 2 diabetes (T2D) have an increased risk of fractures due to skeletal fragility. We aimed to compare areal bone mineral density (aBMD), volumetric BMD (vBMD), cortical and trabecular measures, and bone strength parameters in participants with diabetes vs. controls. In a cross-sectional study, we included adult participants with T1D (n=111, MA=52.9years), T2D (n=106, MA=62.1years) and controls (n=328, MA=57.7years). The study comprised of DXA scans and HR-pQCT scans, biochemistry, handgrip strength (HGS), Timed Up and GO (TUG), vibration perception threshold (VPT), questionnaires, medical histories, alcohol use, and previous fractures. Group comparisons were performed after adjustment for sex, age, BMI, diabetes duration, HbA1c, alcohol, smoking, previous fractures, postmenopausal, HGS, TUG, and VPT. We found decreased aBMD in participants with T1D at the femoral neck (p=0.028), whereas T2D had significantly higher aBMD at peripheral sites (legs, arms, p<0.01) vs. controls. In T1D we found higher vBMD (p<0.001), cortical vBMD (p<0.001), cortical area (p=0.002) and thickness (p<0.001), lower cortical porosity(p=0.008), higher stiffness (p=0.002) and failure load (p=0.003) at radius and higher vBMD (p=0.003), cortical vBMD(p<0.001), bone stiffness (p=0.023) and failure load(p=0.044) at the tibia than controls. In T2D we found higher vBMD (p<0.001), cortical vBMD (p<0.001), trabecular vBMD (p<0.001), cortical area (p<0.001) and thickness (p<0.001), trabecular number (p=0.024), lower separation (p=0.010), higher stiffness (p<0.001) and failure load (p<0.001) at the radius and higher total vBMD (p<0.001), cortical vBMD (p<0.011), trabecular vBMD (p=0.001), cortical area (p=0.002) and thickness (p=0.021), lower trabecular separation (p=0.039), higher stiffness (p<0.001) and failure load (p=0.034) at tibia compared with controls. aBMD measures were as expected lower in T1D and higher in T2D than controls. Favorable bone microarchitecture and strength parameters were seen at the tibia and radius for T1D and T2D.

High-resolution peripheral quantitative computed tomography - Towards clinical use for osteoporosis

Dual X-ray absorptiometry (DXA) and the FRAX score lack sensitivity to identify postmenopausal women at high risk of fracture. To overcome this shortcoming, evaluation of bone microarchitecture using high-resolution peripheral quantitative computed tomography (HR-pQCT) has been suggested to improve fracture risk prediction. In several prospective studies, bone microarchitectural parameters, evaluated using the finite element analysis (FEA) method, have provided better prediction of fracture risk than BMD alone, measured using DXA, or FRAX score. Most cohorts with prospective data have been combined in the Bone Microarchitecture International Consortium analysis, which confirmed on a large scale the improvement of fracture risk prediction, especially with FEA at the radius, but the magnitude of the improvement was not substantial. A recent study has shown that analyzing the microarchitecture to identify women to treat was cost-effective when using zoledronate. A deep learning model using only the images of the distal forearm, including both the bone and soft tissues, has also improved fracture risk prediction substantially. The adoption of deep learning to analyze bone microarchitecture is likely to simplify and speed up the process of fracture risk evaluation. This will allow for adequate preventive therapy of a large proportion of postmenopausal women at high risk who are currently left untreated.

A vitamin D deficient diet increases weight gain and compromises bone biomechanical properties without a reduction in BMD in adult female mice

Vitamin D contributes to the development and maintenance of bone. Evidence suggests vitamin D status can also alter energy balance and gut health. In young animals, vitamin D deficiency (VDD) negatively affects bone mineral density (BMD) and bone microarchitecture, and these effects may also occur due to chronic ethanol intake. However, evidence is limited in mature models, and addressing this was a goal of the current study. Seven-month-old female C57BL/6 mice (n = 40) were weight-matched and randomized to one of four ad libitum diets: control, alcohol (Alc), vitamin D deficient (0 IU/d), or Alc+VDD for 8 weeks. A purified (AIN-93) diet was provided with water or alcohol (10 %) ad libitum. Body weight and food intake were recorded weekly, and feces were collected at 0, 4, and 8 weeks. At the age of 9 months, intestinal permeability was assessed by oral gavage of fluorescein isothiocyanate–dextran. Thereafter, bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry. The microarchitecture of the distal femur was assessed by micro-computed tomography and biomechanical properties were evaluated by cyclic reference point indentation. VDD did not affect BMD or most bone microarchitecture parameters, however, the polar moment of inertia (p < 0.05) was higher in the VDD groups compared to vitamin D sufficient groups. VDD mice also had lower whole bone water content (p < 0.05) and a greater average unloading slope (p < 0.01), and energy dissipated (p < 0.01), indicating the femur displayed a brittle phenotype. In addition, VDD caused a greater increase in energy intake (p < 0.05), weight gain (p < 0.05), and a trend for higher intestinal permeability (p = 0.08). The gut microbiota of the VDD group had a reduction in alpha diversity (p < 0.05) and a lower abundance of ASVs from Rikenellaceae, Clostridia_UCG-014, Oscillospiraceae, and Lachnospiraceae (p < 0.01). There was little to no effect of alcohol supplementation on outcomes. Overall, these findings suggest that vitamin D deficiency causes excess weight gain and reduces the biomechanical strength of the femur as indicated by the higher average unloading slope and energy dissipated without an effect on BMD in a mature murine model.

The impact of motion induced artifacts in the evaluation of HR-pQCT scans of the scaphoid bone: an assessment of inter- and intraobserver variability and quantitative parameters.

In-vivo high-resolution peripheral quantitative computed tomography (HR-pQCT) has high potential in scaphoid bone pathologies' scientific and clinical fields. The manufacturer's visual grading scale (VGS) classifies motion artifacts and divides scans into five quality grades ranging from grade 1 (good quality) to grade 5 (poor quality). This prospective study aimed to investigate the feasibility of the VGS and the influence of image quality on bone density and microarchitecture parameters for the scaphoid bone. Within one year, twenty-two patients with scaphoid fractures received up to six scans of their fractured and contralateral wrist (each consisting of three stacks) using second-generation HR-pQCT (total 256 scans). Three experienced observers graded each stack following the visual grading system, and inter- and intraobserver variability were assessed. The contralateral uninjured scaphoids were then compared pairwise within each patient to high-quality grade 1 scans to determine the influence of image quality on density and microarchitecture parameters. Inter- and intraobserver variability among the three observers significantly revealed fair to moderate agreement, P<0.001 and P<0.05, respectively. Bone volume (BV) fraction tended to increase with poorer image quality but did not exceed four percent. Trabecular bone mineral density (Tb.BMD) decreased with poorer image quality but did not exceed five percent. Trabecular number and trabecular thickness significantly increased by 15.5% and 6.8% at grade five (P<0.001), respectively, and trabecular separation significantly decreased by 13.7% at grade five (P<0.001). This study revealed a considerable influence of motion on bone morphometry parameters of the scaphoid. Therefore, high image quality must be a central point in studies focusing on the histomorphometry of small objects. The high inter- and intraobserver variability limit the VGS. Future research may focus on other grading systems or automated techniques leading to more consistent and reproducible results. Currently, the use of microarchitectural analysis should be limited to cases without motion artefacts or, at most low graded motion artefacts.