Microarchitectural Parameters Research Articles

7892 Longitudinal Changes in Bone Microarchitecture and Strength in Girls with Type 1 Diabetes

Abstract Disclosure: R. Dhaliwal: None. R.K. Saunders: None. M. Misra: None. M.L. Bouxsein: None. D.M. Mitchell: None. Introduction: Type 1 diabetes (T1D) confers an increased fracture risk, only partly explained by low areal bone mineral density (aBMD). We have previously shown in a cross-sectional study that bone microarchitecture is altered early in the course of T1D. The goal of this study is to evaluate how T1D influences longitudinal changes in volumetric BMD (vBMD) and microarchitecture. Methods: This is a 2-year prospective study of girls ages 10-16 years (62 with T1D and 59 controls). We used dual-energy x-ray absorptiometry (DXA) to measure aBMD, and high-resolution peripheral quantitative computed tomography (HR-pQCT) at the distal radius (7% site) and tibia (8% site) to measure vBMD and microarchitecture. We used linear mixed models to assess the effects of T1D on change in bone parameters over time, adjusting for bone age as an index of skeletal maturity, height, lean mass index (LMI), and fat mass index (FMI). Results: At baseline, mean duration of disease in T1D was 4.8 ± 3.2 years, and mean HbA1c was 8.6 ± 1.4%. The groups did not differ by age, bone age, and height, while T1D participants had higher LMI (p<0.001) and borderline higher FMI (p=0.107). Linear growth, weight gain, and changes in body composition over time did not differ between groups. Growth-mediated aBMD gains were observed in both groups at the subtotal body, total hip, femoral neck, and spine. While adjusted aBMD as well as trabecular bone score were lower in T1D, we did not observe between-group differences in changes in these endpoints over time. In both groups, at the distal radius, we observed significant increases over time in total and cortical vBMD, cortical thickness, trabecular (Tb) thickness, and estimated failure load, while cortical porosity decreased. While several microarchitectural parameters were altered in T1D participants, we did not observe significant between-group differences in changes in these measures over time. Of note, trabecular development was subtly altered, with T1D participants demonstrating a borderline significant decrease in Tb vBMD (-14.3 ± 9.0 mg HA/cm3, p=0.112) and Tb number (-0.14 ± 0.07 mm-[1], p=0.057), while control participants had no change in Tb vBMD or number, but a significant increase in Tb thickness (5.8 ± 2.5 µm, p=0.018). At the distal tibia, we observed similar overall detrimental effects of T1D at baseline. However, we did not observe between-group differences in microarchitectural change over time. Glycemic control as measured by HbA1c was not associated with changes in skeletal parameters among T1D participants. Conclusions: T1D leads to substantially altered areal and volumetric BMD as well as microarchitectural parameters in girls. While our data suggest that much of this deficit appears to occur early in the course of disease as prospective change is similar in T1D and controls, subtle changes in trabecular bone development at the radius may indicate ongoing impairment of bone accrual. Presentation: 6/3/2024

7793 Bone Mineral Density, Turnover And Bone Microarchitecture Alterations With Dapagliflozin In Post-menopausal Women With Type 2 Diabetes Mellitus: An Open-Label, Parallel-Arm, Randomized, Controlled Study

Abstract Disclosure: S.K. Bhadada: None. L. Das: None. M. Baruah: None. V. Singla: None. V. Dhiman: None. S. Ram: None. S. Sharma: None. N. Sachdeva: None. P. Dutta: None. Introduction: Evidence is limited and there is no consensus about the effects of SGLT2i on bone metabolism. The objective of the current study was to assess changes in bone mineral density (BMD), turnover markers (BTMs), and microarchitecture with the use of dapagliflozin in type 2 diabetes mellitus (T2DM). Methods: Post-menopausal women (<65 years) with T2DM (both menopause and T2DM duration > 5 years and HbA1c 7-11%) were randomly allocated (1:1) to receive either add-on dapagliflozin with standard-of-care or standard-of-care alone for T2DM management. Those with osteoporosis, on anti—osteoporotic medication, chronic glucocorticoids (>3 months), thiazolidinediones, phenytoin, tamoxifen, estrogen, eGFR<30ml/min/1.73m2, contraindication/past usage of SGLT2i, or with known endocrinopathy were excluded. Patients were made Vitamin D sufficient [25(OH)D > 30ng/ml] with oral cholecalciferol (60K daily for 1 week), if deficient. BMD (DXA, Hologic), BTMs (P1NP, CTX) and microarchitecture were assessed at baseline, 6 and 12 months. Results: A total of 131 patients were screened, of which 101 were randomized (50 dapagliflozin, 51 non-dapagliflozin). The mean age (± SD) in the dapagliflozin group was 58.1 ± 5.8 years, diabetes duration was 9.4 ± 5.2 years and HbA1c was 8.5 ± 1.7%. Baseline clinico-biochemical, areal BMD, volumetric BMD, trabecular structural (number, thickness, separation) and cortical structural (thickness and porosity) parameters were comparable between both groups. Only cortical pore diameter (Ct.Po.Dm) (±SD) at radius (0.199 ± 0.04 vs 0.180 ± 0.03, p =0.01) and tibia (0.256 ± 0.04 vs 0.226 ± 0.03, p =0.001) were higher in the dapagliflozin than the non-dapagliflozin group. At 6 months, BMD at lumbar spine (LS), hip, femoral neck (FN), and distal radius and BTMs were comparable between the dapagliflozin (n=38) and the non-dapagliflozin (n=39) groups. However, there was significantly higher trabecular thickness (Tb.Th) (0.230 ± 0.02 vs 0.218 ± 0.01, p=0.003), cortical thickness (Co.Th) (1.104 ± 0.2 vs 0.998 ± 0.2, p=0.04), intra-cortical porosity (Ct.Po) (0.008 ± 0.005 vs 0.005 ± 0.003, p= 0.020) and Ct.Po.Dm (0.192 ± 0.03 vs 0.169 ± 0.02, p=0.003), all at radius in the dapagliflozin group. At 12 months follow-up, BMD (g/cm2) at all 4 sites were again comparable between the dapagliflozin (n=36) and non-dapagliflozin (n=36) groups. Serum CTX (pg/ml) (388.8 ± 191.5 vs 327.4 ± 140.8, p = 0.06), and P1NP (ng/ml) (38.8 ± 16.3 vs 32.5 ± 16.3, p = 0.12) were non-significantly higher with dapagliflozin use. Among microarchitecture parameters, only Ct.Po.Dm at the radius (0.189 ± 0.03 vs 0.169 ± 0.03, p= 0.025) and tibia (0.251 ± 0.05 vs 0.228 ± 0.03, p= 0.030) remained significantly higher in the dapagliflozin group. Conclusion: Dapagliflozin has no long-term detrimental effects on BMD, BTMs or bone microarchitecture after 1 year of treatment in postmenopausal women with T2DM. Presentation: 6/1/2024

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

Microarchitectural analysis of the metacarpophalangeal joint using HR-pQCT in patients with rheumatoid arthritis: A comparison with healthy controls

ObjectiveTo investigate which joint microarchitectural parameters measured by high-resolution peripheral quantitative computed tomography (HR-pQCT) serve as imaging markers for rheumatoid arthritis (RA). MethodsThe second and third metacarpophalangeal (MCP) joints of 50 patients with RA and 50 healthy controls (HCs) (aged 50–79 years, all females) were scanned using a HR-pQCT. Joint space, trabecular bone microarchitecture, and erosion were measured and compared between RA patients and HCs. ResultsThere were no differences in joint space parameters between RA patients and HCs.For bone microarchitecture, RA patients had lower trabecular bone mineral density (127 vs. 167 mg/cm3), thinner trabecular thickness (0.20 vs. 0.21 mm), fewer trabecular number (1.49 vs. 1.55 /mm), more rod-like structure (1.68 vs. 1.23), and poorer trabecular connectivity (4.51 vs. 5.72 /mm3) than HCs.Regarding erosion, RA patients had a higher number of erosions per joint (36/100 vs. 18/100), larger volume (4.62 vs. 1.89 mm3), and longer width (2.40 vs. 1.82 mm) and longer length (2.34 vs. 1.64 mm) than HCs.Most of the erosions in HCs were <5 mm3 in volume (95 %) and located on the radial side (85 %). When erosions <5 mm3 were compared between RA patients and HCs, there were no differences in their location or morphology. ConclusionsDeterioration of bone microarchitecture and existences of erosions >5 mm3 in the MCP joints are sensitive imaging markers of RA. Erosions <5 mm3 in RA patients may include not only early pathological erosion but also physiological erosion because even HCs can have erosions <5 mm3.

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.

A Randomized Controlled Trial on the Effect of Luseogliflozin on Bone Microarchitecture Evaluated Using HR-pQCT in Elderly Type 2 Diabetes.

Bone fragility is a critical issue in the treatment of elderly people with type 2 diabetes (T2D). In the Canagliflozin Cardiovascular Assessment Study, the subjects with T2D who were treated with canagliflozin showed a significant increase in fracture events compared to a placebo group as early as 12weeks post-initiation. In addition, it has been unclear whether sodium-glucose co-transporter 2 (SGLT2) inhibitors promote bone fragility. We used high-resolution peripheral quantitative computed tomography (HR-pQCT) to prospectively evaluate the short-term effect of the SGLT2 inhibitor luseogliflozin on bone strength and microarchitecture in elderly people with T2D. This was a single-center, randomized, open-label, active-controlled pilot trial for ≥ 60-year-old Japanese individuals with T2D without osteoporosis. A total of 22 subjects (seven women and 15 men) were randomly assigned to a Lusefi group (added luseogliflozin 2.5mg) or a control group (added metformin 500mg) and treated for 48weeks. We used the second-generation HR-pQCT (Xtreme CT II®, Scanco Medical, Brüttisellen, Switzerland) before and 48weeks after the treatment to evaluate the subjects' bone microarchitecture and estimate their bone strength. Twenty subjects (Lusefi group, n = 9; control group, n = 11) completed the study, with no fracture events. As the primary outcome, the 48-week changes in the bone strength (stiffness and failure load) estimated by micro-finite element analysis were not significantly different between the groups. As the secondary outcome, the changes in all of the cortical/trabecular microarchitectural parameters at the radius and tibia from baseline to 48weeks were not significantly different between the groups. In the pilot trial, we observed no negative effect of 48-week luseogliflozin treatment on bone microarchitecture or bone strength in elderly people with T2D. UMIN-CTR no. 000036202 and jRCT 071180061.

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.

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.

Trabecular and cortical bone microarchitecture using high-resolution peripheral quantitative computed tomographic imaging in diabetic peripheral neuropathy

ContextType 2 Diabetes Mellitus (T2D) is associated with an increased risk of fragility fracture despite normal areal bone mineral density (BMD). The contribution of diabetic peripheral neuropathy (PN) to volumetric BMD (vBMD) and bone microarchitecture in T2D is not explored. ObjectiveTo assess vBMD and microarchitectural properties of bone using high-resolution peripheral quantitative computed tomography (HR-pQCT) in patients of T2D with or without PN. DesignThis is a cross-sectional study of patients of T2D divided into two groups [patients with T2D without PN (Group A) and T2D with PN (Group B)]. All patients underwent clinical examination, biochemical evaluation, dual-energy X-ray absorptiometry (DXA), and HR-pQCT of the radius and tibia. ResultsA total of 296 patients were included in the study [Group A (n = 98), Group B (n = 198)]. HR-pQCT demonstrated a significant difference in total vBMD[mg/cm3] at tibia (291.6 ± 61.8 vs. 268.2 ± 63.0; p-0.003); cortical vBMD[mg/cm3] at tibia [912.5 (863.3, 962.4) vs. 853.8 (795.3, 913.2) p-0.000], among groups A and B respectively. Among the microarchitecture parameters, there was a significant difference in cortical porosity at the tibia (2.5% ±1.7% vs. 3%±1.7%; p-0.004), trabecular number[mm-1] at the tibia [1.080 (0.896, 1.237) vs. 1.140 (0.983, 1.286), p-0.045] and trabecular thickness[mm] at the radius [0.228 (0.217, 0.247) Vs. 0.238 (0.224, 0.253); p-0.006], among groups A and B respectively. ConclusionDespite comparable areal BMD, T2D patients with PN have diminished vBMD and deteriorated skeletal microarchitecture, compared to those without PN.

Microstructural, Fluid Dynamic, and Mechanical Characterization of Zinc Oxide and Magnesium Chloride-Modified Hydrogel Scaffolds.

Scaffolds for the filling and regeneration of osteochondral defects are a current challenge in the biomaterials field, and solutions with greater functionality are still being sought. The novel approach of this work was to obtain scaffolds with biologically active additives possessing microstructural, permeability, and mechanical properties, mimicking the complexity of natural cartilage. Four types of scaffolds with a gelatin/alginate matrix modified with hydroxyapatite were obtained, and the relationship between the modifiers and substrate properties was evaluated. They differed in the type of second modifier used, which was hydrated MgCl2 in two proportions, ZnO, and nanohydroxyapatite. The samples were obtained by freeze-drying by using two-stage freezing. Based on microstructural observations combined with X-ray microanalysis, the microstructure of the samples and the elemental content were assessed. Permeability and mechanical tests were also performed. The scaffolds exhibited a network of interconnected pores and complex microarchitecture, with lower porosity at the surface (15 ± 7 to 29 ± 6%) and higher porosity at the center (67 ± 8 to 75 ± 8%). The additives had varying effects on the pore sizes and permeabilities of the samples. ZnO yielded the most permeable scaffolds (5.92 × 10-11 m2), whereas nanohydroxyapatite yielded the scaffold with the lowest permeability (1.18 × 10-11 m2), values within the range reported for trabecular bone. The magnesium content had no statistically significant effect on the permeability. The best mechanical parameters were obtained for ZnO samples and those containing hydrated MgCl2. The scaffold's properties meet the criteria for filling osteochondral defects. The developed scaffolds follow a biomimetic approach in terms of hierarchical microarchitecture and mechanical parameters as well as chemical composition. The obtained composite materials have the potential as biomimetic scaffolds for the regeneration of osteochondral defects.

High-resolution three-dimensional micro-computed tomography assessment of micro-architectural patterns in non-adults with cribra orbitalia: Correlation between macro- and micro-scale bone features.

Skeletal porous lesions such as cribra orbitalia (CO) have long been of interest to bioanthropologists worldwide, mainly due to their high prevalence in osteological material. Previous studies considered CO as an external morphological manifestation, and therefore, research has mainly focused on visible (macroscopic) CO patterns. However, the understanding of CO-induced micro-scale bone changes is still scarce. Therefore, we performed high-resolution micro-computed tomography imaging to investigate three-dimensional CO-induced micro-architectural patterns in non-adults, with a particular focus on the correlation between macroscopic and micro-architectural orbital features. Cortical and trabecular micro-architectural changes in the orbital roof were analyzed in non-adults younger than 15 years, using orbital roof samples with and without macroscopic traces of CO (n = 28). A widely accepted five-grade macroscopic CO scoring system was applied to analyze CO severity. Areas affected with CO (area 1) and areas without macroscopic CO traces (area 2) were analyzed separately. The conducted high-resolution analysis showed that cortical and trabecular micro-architecture varied with CO presence, lesion severity (CO grade), and the analyzed area. Inter-grade comparisons suggested that most of the analyzed micro-architectural parameters were not significantly different between adjacent CO grades. Based on the micro-architectural evaluation of areas 1 and 2, the porous lesions were much more extensive than revealed by gross examination. In addition, micro-architectural differences were particularly pronounced in younger non-adults. In summary, our pilot study suggests that the macroscopic examination of CO reflects only the tip of the iceberg, as the micro-architectural changes seem to be much larger than macroscopically identified. RESEARCH HIGHLIGHTS: Cribra orbitalia (CO) represents orbital porous lesions. A high-resolution microscopic assessment of CO-induced changes in non-adults was done by micro-computed tomography. The microarchitecture was affected by CO presence, CO grade, area, and age.

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.

Quantitative atlas of collagen hydrogels reveals mesenchymal cancer cell traction adaptation to the matrix nanoarchitecture

Collagen-based hydrogels are commonly used in mechanobiology to mimic the extracellular matrix. A quantitative analysis of the influence of collagen concentration and properties on the structure and mechanics of the hydrogels is essential for tailored design adjustments for specific in vitro conditions. We combined focused ion beam scanning electron microscopy and rheology to provide a detailed quantitative atlas of the mechanical and nanoscale three-dimensional structural alterations that occur when manipulating different hydrogel's physicochemistry. Moreover, we study the effects of such alterations on the phenotype of breast cancer cells and their mechanical interactions with the extracellular matrix. Regardless of the microenvironment's pore size, porosity or mechanical properties, cancer cells are able to reach a stable mesenchymal-like morphology. Additionally, employing 3D traction force microscopy, a positive correlation between cellular tractions and ECM mechanics is observed up to a critical threshold, beyond which tractions plateau. This suggests that cancer cells in a stable mesenchymal state calibrate their mechanical interactions with the ECM to keep their migration and invasiveness capacities unaltered. Statement of significanceThe paper presents a thorough study on the mechanical microenvironment in breast cancer cells during their interaction with collagen based hydrogels of different compositions. The hydrogels’ microstructure were obtained using state-of-the-art 3D microscopy, namely focused ion beam-scanning electron microscope (FIB-SEM). FIB-SEM was originally applied in this work to reconstruct complex fibered collagen microstructures within the nanometer range, to obtain key microarchitectural parameters. The mechanical microenvironment of cells was recovered using Traction Force Microscopy (TFM). The obtained results suggest that cells calibrate tractions such that they depend on mechanical, microstructural and physicochemical characteristics of the hydrogels, hence revealing a steric hindrance. We hypothesize that cancer cells studied in this paper tune their mechanical state to keep their migration and invasiveness capacities unaltered.

Sargassum polysaccharide attenuates osteoarthritis in rats and is associated with the up-regulation of the ITGβ1-PI3K-AKT signaling pathway

BackgroundOsteoarthritis (OA) presents a formidable challenge, characterized by as-yet-unclear mechanical intricacies within cartilage and the dysregulation of bone homeostasis. Our preliminary data revealed the encouraging potential of a Sargassum polysaccharide (SP), in promoting chondrogenesis. The aim of our study is to comprehensively assess the therapeutic effects of SP on OA models and further elucidate its potential mechanism. MethodsThe protective effects of SP were initially evaluated in an inflammation-induced human chondrocyte (C28) cell model. CCK-8 assays, Alcian blue staining, RT-qPCR and Western blotting were used to verify the chondrogenesis of SP in vitro. To assess the efficacy of SP in vivo, surgically induced medial meniscus destabilization (DMM) OA rats underwent an 8-week SP treatment. The therapeutic effects of SP in OA rats were comprehensively evaluated using X-ray imaging, micro-computed tomography (μ-CT), histopathological analysis, as well as immunohistochemical and immunofluorescent staining. Following these assessments, we delved into the potential signaling pathways of SP in inflammatory chondrocytes utilizing RNA-seq analysis. Validation of these findings was conducted through RT-qPCR and western blotting techniques. ResultsSP significantly enhance the viability of C28 chondrocytes, and increased the secretion of acidic glycoproteins. Moreover, SP stimulated the expression of chondrogenic genes (Aggrecan, Sox9, Col2a1) and facilitated the synthesis of Collagen II protein in C28 inflammatory chondrocytes. In vivo experiments revealed that SP markedly ameliorated knee joint stenosis, alleviated bone and cartilage injuries, and reduced the histopathological scores in the OA rats. μ-CT analysis confirmed that SP lessened bone impairments in the medial femoral condyle and the subchondral bone of the tibial plateau, significantly improving the microarchitectural parameters of the subchondral bone. Histopathological analyses indicated that SP notably enhanced cartilage quality on the surface of the tibial plateau, leading to increased cartilage thickness and area. Immunohistochemistry staining and immunofluorescence staining corroborated these findings by showing a significant promotion of Collagen II expression in OA joints treated with SP. RNA-seq analysis suggest that SP's effects were mediated through the regulation of the ITGβ1-PI3K-AKT signaling axis, thereby stimulating chondrogenesis. Verification through RT-qPCR and Western blot analyses confirmed that SP significantly upregulated the expression of ITGβ1, p110δ, AKT1, ACAN, and Col2a1. Notably, knock-down of ITGβ1 using siRNA in C28 chondrocytes inhibited the expression of ITGβ1, p110δ, AKT1, and ACAN. However, these inhibitory effects were not completely reversed by supplemental SP intervention. ConclusionsIn summary, our findings reveal that SP significantly enhances chondrogenesis both in vitro and in vivo, alleviating OA progression both in bone and cartilage. The observed beneficial effects are intricately linked to the activation of the ITGβ1-PI3K-AKT signaling axis. The translational potential of this articleOur research marks the first instance unveiling the advantageous effects and underlying mechanisms of SP in OA treatment. With its clinical prospects, SP presents compelling new evidence for the advancement of a next-generation polysaccharide drug for OA therapy.

Parathyroid hormone (1–34) retards the lumbar facet joint degeneration and activates Wnt/β-catenin signaling pathway in ovariectomized rats

PurposeFacet joint degeneration (FJD) is a major cause of low back pain. Parathyroid hormone (PTH) (1–34) is commonly used to treat osteoporosis. However, little is known about its effects on FJD induced by estrogen deficiency. This study aims to investigate the effects of PTH (1–34) on FJD induced by estrogen deficiency and the underlying pathogenesis of the disease.MethodsForty 3-month-old female Sprague-Dawley rats were randomly divided into four groups: 30 received bilateral ovariectomy (OVX) followed by 12 weeks of treatment with normal saline, PTH (1–34) or 17β-estradiol (E2), and 10 received sham surgery followed by administration of normal saline. Status and Wnt/β-catenin signaling activity in the cartilage and subchondral bone of the L4–L5 FJs and serum biomarkers were analyzed.ResultsAdministration of PTH (1–34) and E2 ameliorated cartilage lesions, and significantly decreased MMP-13 and caspase-3 levels and chondrocyte apoptosis. PTH (1–34) but not E2 significantly increased cartilage thickness, number of chondrocytes, and the expression of aggrecan. PTH (1–34) significantly improved microarchitecture parameters of subchondral bone, increased the expression of collagen I and osteocalcin, and decreased RANKL/OPG ratio. E2 treatment significantly increased the OPG level and decreased the RANKL/OPG ratio in the subchondral bone of ovariectomized rats, but it did not significantly improve the microarchitecture parameters of subchondral bone. Wnt3a and β-catenin expression was significantly reduced in the articular cartilage and subchondral bone in OVX rats, but PTH (1–34) could increase the expression of these proteins. E2 significantly increased the activity of Wnt/β-catenin pathway only in cartilage, but not in subchondral bone. The restoration of Wnt/β-catenin signaling had an obvious correlation with the improvement of some parameters associated with the FJs status.ConclusionWnt/β-catenin signaling may be a potential therapeutic target for FJD induced by estrogen deficiency. PTH (1–34) is effective in treating this disease with better efficacy than 17β-estradiol, and the efficacy may be attributed to its restoration of Wnt/β-catenin signaling.

Statistical investigation of interdependence of trabecular microarchitectural parameters from micro-computed tomography

Standard microarchitectural analysis of bone using micro-computed tomography produces a large number of parameters that quantify the structure of the trabecular network. Analyses that perform statistical tests on many parameters are at elevated risk of making Type I errors. However, when multiple testing correction procedures are applied, the risk of Type II errors is elevated if the parameters being tested are strongly correlated. In this article, we argue that four commonly used trabecular microarchitectural parameters (thickness, separation, number, and bone volume fraction) are interdependent and describe only two independent properties of the trabecular network. We first derive theoretical relationships between the parameters based on their geometric definitions. Then, we analyze these relationships with an aggregated in vivo dataset with 2987 images from 1434 participants and a synthetically generated dataset with 144 images using principal component analysis (PCA) and linear regression analysis. With PCA, when trabecular thickness, separation, number, and bone volume fraction are combined, we find that 92 % to 97 % of the total variance in the data is explained by the first two principal components. With linear regressions, we find high coefficients of determination (0.827–0.994) and fitted coefficients within expected ranges. These findings suggest that to maximize statistical power in future studies, only two of trabecular thickness, separation, number and bone volume fraction should be used for statistical testing.

Dietary supplementation with seeds of Sesamum indicum L. (pedaliaceae) mitigates the 7,12-dimethylbenz(a)anthracene-induced breast tumor burden in rats.

Sesamum indicum L. seeds; rich in zinc and lignans are endowed with antioxidant and immunomodulatory properties which attract research on their anticancer potential. Although many studies have reported the invitro antitumor potential of S.indicum and its phytoconstituents, much is yet to be known about its invivo effects. To fill this gap, the effects of dietary supplementation with seeds of S.indicum in 7,12-dimethylbenz(a)anthracene-exposed rats was assessed. 42 rats aged 30-35days were randomized into six groups (n=6) as follows: the normal (NOR) and negative (DMBA) control groups were fed with standard diet; the positive control group (DMBA+Zinc) was fed with standard diet supplemented with commercial zinc (0.01 %); the test groups were fed with standard diet supplemented with S.indicum seeds in different proportions (6.25 , 12.5 and 25 %). Breast cancer was induced by a single administration of DMBA (50 mg/kg BW, s.c.) diluted in corn oil. The experiment lasted 20weeks and afterward, tumor incidence; tumor burden, tumor volume, tumor micro-architecture andsome biochemical parameters were evaluated. As salient result, 100 % of rats in the DMBA group developed tumors, while rats feed with rat chow supplemented with S.indicum seeds (25 %) had a reduced incidence of tumors (33.3 %) and tumor volume (2.71 cm3 in sesame 25 % vs. 4.69 cm3 in the DMBA group, p˂0.01). The seeds (25 %) also slowed DMBA-induced neoplasm expansion in mammary ducts as compared to rats of DMBA group. In summary, supplementation with S.indicum seeds slowed breast tumorigenesis via its antioxidant capacity.

Pronounced cortical porosity and sex-specific patterns of increased bone and osteocyte lacunar mineralization characterize the human distal fibula with aging

The high occurrence of distal fibula fractures among older women suggests a potential link to impaired bone health. Here we used a multiscale imaging approach to investigate the microarchitecture, mineralization, and biomechanics of the human distal fibula in relation to age and sex. Micro-computed tomography was performed to analyze the local volumetric bone mineral density and various microarchitectural parameters of the trabecular and the cortical compartment. Bone mineral density distribution and osteocyte lacunar parameters were quantified using quantitative backscattered electron imaging in periosteal, endocortical, and trabecular regions. Additionally, cortical hardness and Young's modulus were assessed by nanoindentation. While cortical porosity strongly increased with age independent of sex, trabecular microarchitecture remained stable. Notably, nearly half of the specimens showed non-bony hypermineralized tissue located at the periosteum, similar to that previously detected in the femoral neck, with no consistent association with advanced age. Independent of this finding, cortical and trabecular mineralization, i.e., mean calcium content, as well as endocortical tissue hardness increased with age in males but not females. Importantly, we also observed mineralized osteocyte lacunae that increased with age specifically in females. In conclusion, our results indicate that skeletal aging of the distal fibula is signified not only by pronounced cortical porosity but also by an increase in mineralized osteocyte lacunae in females. These findings may provide an explanation for the increased occurrence of ankle fractures in older women.

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 cell response to additively manufactured 3D micro-architectures with controlled Poisson's ratio: Auxetic vs. non-auxetic meta-biomaterials

The Poisson's ratio and elastic modulus are two parameters determining the elastic behavior of biomaterials. While the effects of elastic modulus on the cell response is widely studied, very little is known regarding the effects of the Poisson's ratio. The micro-architecture of meta-biomaterials determines not only the Poisson's ratio but also several other parameters that also influence cell response, such as porosity, pore size, and effective elastic modulus. It is, therefore, very challenging to isolate the effects of the Poisson's ratio from those of other micro-architectural parameters. Here, we computationally design meta-biomaterials with controlled Poisson's ratios, ranging between -0.74 and +0.74, while maintaining consistent porosity, pore size, and effective elastic modulus. The 3D meta-biomaterials were additively manufactured at the micro-scale using two-photon polymerization (2PP), and were mechanically evaluated at the meso‑scale. The response of murine preosteoblasts to these meta-biomaterials was then studied using in vitro cell culture models. Meta-biomaterials with positive Poisson's ratios resulted in higher metabolic activity than those with negative values. The cells could attach and infiltrate all meta-biomaterials from the bottom to the top, fully covering the scaffolds after 17 days of culture. Interestingly, the meta-biomaterials exhibited different cell-induced deformations (e.g., shrinkage or local bending) as observed via scanning electron microscopy. The outcomes of osteogenic differentiation (i.e., Runx2 immunofluorescent staining) and matrix mineralization (i.e., Alizarin red staining) assays indicated the significant potential impact of these meta-biomaterials in the field of bone tissue engineering, paving the way for the development of advanced bone meta-implants. Statement of significanceWe studied the influence of Poisson's ratio on bone cell response in meta-biomaterials. While elastic modulus effects are well-studied, the impact of Poisson's ratio, especially negative values found in architected biomaterials, remains largely unexplored. The complexity arises from intertwined micro-architectural parameters, such as porosity and elastic modulus, making it challenging to isolate the Poisson's ratio. To overcome this limitation, this study employed rational computational design to create meta-biomaterials with controlled Poisson's ratios, alongside consistent effective elastic modulus, porosity, and pore size. The study reveals that two-photon polymerized 3D meta-biomaterials with positive Poisson's ratios displayed higher metabolic activity, while all the developed meta-biomaterials supported osteogenic differentiation of preosteoblasts as well as matrix mineralization. The outcomes pave the way for the development of advanced 3D bone tissue models and meta-implants.