Increased Heterogeneity of Bone Matrix Mineralization in Pediatric Patients Prone to Fractures: A Biopsy Study

Idiopathic osteoporosis (IOP) in children is characterized by fragility fractures and/or low bone mineral density in otherwise healthy individuals. The aim of the present work was to measure bone mineralization density distribution (BMDD) based on quantitative backscattered electron imaging (qBEI) in children with suspected IOP. Entire cross‐sectional areas of transiliac bone biopsy samples from children (n = 24, 17 boys; aged 6.7–16.6 years) with a history of fractures (n = 14 with at least one vertebral fracture) were analyzed for cancellous (Cn) and cortical (Ct) BMDD. Outcomes were compared with normal reference BMDD data and correlated with the patients' clinical characteristics and bone histomorphometry findings. The subjects had similar average degree but significantly higher heterogeneity of mineralization in both Cn and Ct bone (Cn.CaWidth +23%, Ct.CaWidth +15%, p < 0.001 and p = 0.002, respectively), together with higher percentages of low mineralized cancellous (Cn.CaLow +35%, p < 0.001) and highly mineralized cortical bone areas (Ct.CaHigh +82%, p = 0.032). Ct.CaWidth and Ct.CaLow were positively correlated with mineralizing surface per bone surface (MS/BS; a primary histomorphometric determinant of bone formation) and with serum bone turnover markers (all p < 0.05). The correlations of the mineralization heterogeneity with histomorphometric and serum bone turnover indices suggest that an enhanced variation in bone turnover/formation contributes to the increased heterogeneity of mineralization. However, it remains unclear whether the latter is cause for, or the response to the increased bone fragility in these children with suspected IOP. © 2014 American Society for Bone and Mineral Research.     

Annual intravenous zoledronic acid for three years increased cancellous bone matrix mineralization beyond normal values in the HORIZON biopsy cohort

The efficacy of 3 years of annual intravenous administration of zoledronic acid (ZOL) in reducing vertebral and nonvertebral fractures in postmenopausal osteoporosis has been shown by the HORIZON pivotal fracture trial. Histomorphometric analysis of transiliac bone biopsies from the HORIZON participants revealed significantly improved trabecular architecture and reduced bone remodeling for the ZOL‐treated versus placebo‐treated patients. The aim of our study was to evaluate the cancellous and cortical bone mineralization density distribution (BMDD) in these biopsies by quantitative backscattered electron imaging (qBEI). The study cohort comprised 82 patients on active treatment (ZOL, yearly doses of 5 mg) and 70 treated with placebo, and all received adequate Ca and VitD supplementation. Comparison of ZOL‐treated versus placebo‐treated cancellous (Cn.) and cortical (Ct.) BMDD‐derived variables resulted in significantly higher average (Cn.CaMean + 3.2%, Ct.CaMean + 2.7%) and mode calcium concentrations (Cn.CaPeak + 2.1%, Ct.CaPeak + 1.5%), increased percentages of highly mineralized bone areas (Cn.CaHigh + 64%, Ct.CaHigh + 31%), lower heterogeneity of mineralization (Cn.CaWidth ?14%, Ct.CaWidth ?13%), and decreased percentages of low mineralized bone areas (Cn.CaLow ?22%, Ct.CaLow ?26%) versus placebo (all p < 0.001). Cn. BMDD from the patients on active treatment also revealed a statistically significant shift to higher Ca concentrations when compared to a historical normal reference BMDD. These differences in BMDD from ZOL patients compared to the other groups were in line with the correlation of BMDD variables with previously determined cancellous mineralizing surface per bone surface (Cn. MS/BS, a primary histomorphometric index for bone turnover), showing that those with lower Cn. MS/BS had a higher degree of bone matrix mineralization. However, the differences in BMDD variables between the study groups remained when adjusted for Cn. MS/BS, suggesting that other factors in addition to reduced bone turnover might contribute to the higher bone matrix mineralization after ZOL treatment. © 2013 American Society for Bone and Mineral Research.     

PTH(1‐84) Administration in Hypoparathyroidism Transiently Reduces Bone Matrix Mineralization

Patients with hypoparathyroidism have low circulating parathyroid (PTH) levels and higher cancellous bone volume and trabecular thickness. Treatment with PTH(1‐84) was shown to increase abnormally low bone remodeling dynamics. In this work, we studied the effect of 1‐year or 2‐year PTH(1‐84) treatment on cancellous and cortical bone mineralization density distribution (Cn.BMDD and Ct.BMDD) based on quantitative backscattered electron imaging (qBEI) in paired transiliac bone biopsy samples. The study cohort comprised 30 adult hypoparathyroid patients (14 treated for 1 year; 16 treated for 2 years). At baseline, Cn.BMDD was shifted to higher mineralization densities in both treatment groups (average degree of mineralization Cn.Ca_Mean +3.9% and +2.7%, p < 0.001) compared to reference BMDD. After 1‐year PTH(1‐84), Cn.Ca_Mean was significantly lower than that at baseline (–6.3%, p < 0.001), whereas in the 2‐year PTH(1‐84) group Cn.Ca_Mean did not differ from baseline. Significant changes of Ct.BMDD were observed in the 1‐year treatment group only. The change in histomorphometric bone formation (mineralizing surface) was predictive for Cn.BMDD outcomes in the 1‐year PTH(1‐84) group, but not in the 2‐year PTH(1‐84) group. Our findings suggest higher baseline bone matrix mineralization consistent with the decreased bone turnover in hypoparathyroidism. PTH(1‐84) treatment caused differential effects dependent on treatment duration that were consistent with the histomorphometric bone formation outcomes. The greater increase in bone formation during the first year of treatment was associated with a decrease in bone matrix mineralization, suggesting that PTH(1‐84) exposure to the hypoparathyroid skeleton has the greatest effects on BMDD early in treatment. © 2015 American Society for Bone and Mineral Research.     

Intravenous Treatment With Ibandronate Normalizes Bone Matrix Mineralization and Reduces Cortical Porosity After Two Years in Male Osteoporosis: A Paired Biopsy Study

The spectrum of therapeutic options and the amount of clinical trials for male osteoporosis (mOP) is lower than those for postmenopausal osteoporosis. Therefore, we examined the effects of 24 months of ibandronate (IBN) treatment (3 mg/3 mL intravenously every 3 months) on bone material quality in 19 subjects with mOP within an open‐label, single‐center, prospective phase III study (Eudract number 2006‐006692‐20). Patients (median age [25th, 75th percentiles] 53.0 [44.5; 57.0] years) were included if they had low bone mineral density (BMD) and/or at least one low trauma fracture and no secondary cause of osteoporosis. The primary endpoint was to evaluate IBN effects on cancellous (Cn.) and cortical (Ct.) bone mineralization density distribution (BMDD) based on quantitative backscattered electron imaging (qBEI) of paired transiliacal bone biopsies (baseline, 24 months). Secondary endpoints included changes in areal bone mineral density (BMD by dual‐energy X‐ray absorptiometry [DXA]) and serum markers of bone turnover including type I collagen peptides CrossLaps (CTX), procollagen type 1 amino‐terminal propeptide (P1NP), and osteocalcin (OC). At baseline, cancellous bone matrix mineralization from mOP was lower than published reference data (mean degree of mineralization Cn.CaMean ?1.8%, p < 0.01). IBN treatment increased calcium concentrations versus baseline (Cn.CaMean +2.4%, Ct.CaMean, +3.0% both p < 0.01), and reduced heterogeneity of mineralization (Cn.CaWidth ?14%, p = 0.044; Ct.CaWidth, ?16%, p = 0.001), leading to cancellous BMDD within normal range. IBN treatment was associated with a decrease in porosity of mineralized cortical tissue (?25%, p = 0.01); increases in BMD at the lumbar spine, the femoral neck, and the total hip (+3.3%, +1.9%, and +5.6%, respectively, p ≤ 0.01); and reductions in CTX (?37.5%), P1NP (?44.4%), and OC (?36.3%, all p < 0.01). Our BMDD findings are in line with the reduction of bone turnover markers and the increase in BMD by IBN in our patients and suggest that the latter mainly reflects the increase in matrix mineralization and the reduction of cortical porosity in this cohort with mOP. © 2014 American Society for Bone and Mineral Research.     

Relationship of Bone Mineralization Density Distribution (BMDD) in Cortical and Cancellous Bone Within the Iliac Crest of Healthy Premenopausal Women

Bone mineralization density distribution (BMDD) is an important determinant of bone mechanical properties. The most available skeletal site for access to the BMDD is the iliac crest. Compared to cancellous bone much less information on BMDD is available for cortical bone. Hence, we analyzed complete transiliac crest bone biopsy samples from premenopausal women (n = 73) aged 25–48 years, clinically classified as healthy, by quantitative backscattered electron imaging for cortical (Ct.) and cancellous (Cn.) BMDD. The Ct.BMDD was characterized by the arithmetic mean of the BMDD of the cortical plates. We found correlations between Ct. and Cn. BMDD variables with correlation coefficients r between 0.42 and 0.73 (all p < 0.001). Additionally to this synchronous behavior of cortical and cancellous compartments, we found that the heterogeneity of mineralization densities (Ct.Ca_Width), as well as the cortical porosity (Ct.Po) was larger for a lower average degree of mineralization (Ct.Ca_Mean). Moreover, Ct.Po correlated negatively with the percentage of highly mineralized bone areas (Ct.Ca_High) and positively with the percentage of lowly mineralized bone areas (Ct.Ca_Low). In conclusion, the correlation of cortical with cancellous BMDD in the iliac crest of the study cohort suggests coordinated regulation of bone turnover between both bone compartments. Only in a few cases, there was a difference in the degree of mineralization of >1wt % between both cortices suggesting a possible modeling situation. This normative dataset of healthy premenopausal women will provide a reference standard by which disease- and treatment-specific effects can be assessed at the level of cortical bone BMDD.     

Bone Matrix Quality After Sclerostin Antibody Treatment

Sclerostin antibody (Scl‐Ab) is a novel bone‐forming agent that is currently undergoing preclinical and clinical testing. Scl‐Ab treatment is known to dramatically increase bone mass, but little is known about the quality of the bone formed during treatment. In the current study, global mineralization of bone matrix in rats and nonhuman primates treated with vehicle or Scl‐Ab was assayed by backscattered scanning electron microscopy (bSEM) to quantify the bone mineral density distribution (BMDD). Additionally, fluorochrome labeling allowed tissue age–specific measurements to be made in the primate model with Fourier‐transform infrared microspectroscopy to determine the kinetics of mineralization, carbonate substitution, crystallinity, and collagen cross‐linking. Despite up to 54% increases in the bone volume after Scl‐Ab treatment, the mean global mineralization of trabecular and cortical bone was unaffected in both animal models investigated. However, there were two subtle changes in the BMDD after Scl‐Ab treatment in the primate trabecular bone, including an increase in the number of pixels with a low mineralization value (Z₅) and a decrease in the standard deviation of the distribution. Tissue age–specific measurements in the primate model showed that Scl‐Ab treatment did not affect the mineral‐to‐matrix ratio, crystallinity, or collagen cross‐linking in the endocortical, intracortical, or trabecular compartments. Scl‐Ab treatment was associated with a nonsignificant trend toward accelerated mineralization intracortically and a nearly 10% increase in carbonate substitution for tissue older than 2 weeks in the trabecular compartment (p < 0.001). These findings suggest that Scl‐Ab treatment does not negatively impact bone matrix quality. © 2014 American Society for Bone and Mineral Research.     

Histomorphometry and Bone Matrix Mineralization Before and After Bisphosphonate Treatment in Boys With Duchenne Muscular Dystrophy: A Paired Transiliac Biopsy Study

Duchenne muscular dystrophy (DMD) is a genetic disorder causing progressive muscle weakness. To prolong independent ambulation, DMD patients are treated with glucocorticoids, which, in turn, can increase bone fragility. In a cohort with vertebral fractures, intravenous bisphosphonate (iv BP) therapy stabilized vertebrae and reduced back pain. To characterize the effects of glucocorticoid therapy and bisphosphonate treatment on bone tissue and material properties, paired transiliac biopsy samples (before and after on average 2.4 years of iv BP) from 9 boys with DMD were studied for histomorphometry and bone mineralization density distribution (BMDD) and compared to reference values. Before iv BP, the boys had low cancellous bone volume (BV/TV) and cortical thickness (Ct.Wi) (both on average 56% of the healthy average, p < 0.001 versus reference), and mineralizing surface (MS/BS) in the lower normal range (on average 74% of the healthy average). The average degree of mineralization of cancellous (Cn.CaMean) and cortical compartments (Ct.CaMean) was 21.48 (20.70, 21.90) wt% and 20.42 (19.32, 21.64) wt%, respectively (median [25th, 75th percentiles]), which was not different from reference. After iv BP, BV/TV and Ct.Wi were, on average, unchanged. However, at the individual patient level, BV/TV Z‐scores increased in 2, remained unchanged in 4, and declined in 3 patients. Additionally, on average, MS/BS decreased (–85%, p < 0.001), Cn.CaMean (+2.7%) increased, whereas the heterogeneity of cancellous (Cn.CaWidth –19%) and cortical bone mineralization (Ct.CaWidth –8%, all p < 0.05) decreased versus baseline. The changes in bone mineralization are consistent with the antiresorptive action of iv BP. At the same time, our observations point to the need for novel therapies with less or absent bone turnover suppression, including the fact that bone turnover was low even before bisphosphonate therapy, that bone turnover declined further (as expected) with treatment, and that declines in trabecular bone volume were observed in some boys despite bisphosphonate therapy. © 2015 American Society for Bone and Mineral Research     

Examining the Relationships Between Bone Tissue Composition,Compositional Heterogeneity,and Fragility Fracture: A Matched Case‐Controlled FTIRI Study

Fourier transform infrared imaging (FTIRI) provides information on spatial distribution of the chemical composition of thin tissue specimens at ～7 µm spatial resolution. This study of 120 age‐ and bone mineral density (BMD)‐matched patients was designed to investigate the association of FTIRI variables, measured in iliac crest biopsies, with fragility fractures at any site. An earlier study of 54 women found hip BMD to be a significant explanatory variable of fracture risk for cortical bone but not for cancellous bone. In the current study, where age and BMD were controlled through matching, no such association was observed, validating the pairing scheme. Our first study of unmatched iliac crest biopsies found increases in collagen maturity (cancellous and cortical bone) and mineral crystal size (cortical bone only) to be a significant explanatory variable of fracture when combined with other covariates. The ratio for collagen maturity has been correlated to the amount of enzymatic collagen cross‐links. To assess the impact of other FTIRI variables (acid phosphate substitution, carbonate‐to‐phosphate ratio, and the pixel distribution [heterogeneity] of all relevant FTIRI variables), we examined biopsies from a matched case‐controlled study, in which 60 women with fractures were each paired with an age‐ and BMD‐matched female control. With the matched data set of 120 women, conditional logistic regression analyses revealed that significant explanatory variables of fracture were decreased carbonate‐to‐phosphate ratio in both cancellous (odds ratio [OR] = 0.580, 95% confidence interval [CI] 0.37–0.909, p = 0.0176) and cortical bone (OR = 0.519, 95% CI 0.325–0.829, p = 0.0061), and increased heterogeneity (broadened pixel distribution) of collagen maturity for cancellous bone (OR = 1.549, 95% CI 1.002–2.396, p = 0.0491). The observation that collagen maturity was no longer linked to fracture in age‐ and BMD‐matched samples suggests that age‐dependent variation in collagen maturity may be a more important contributory factor to fragility fractures than previously thought. © 2015 American Society for Bone and Mineral Research.     

Subtle changes in bone mineralization density distribution in most severely affected patients with chronic obstructive pulmonary disease

Chronic obstructive pulmonary disease (COPD) is associated with low aBMD as measured by DXA and altered microstructure as assessed by bone histomorphometry and microcomputed tomography. Knowledge of bone matrix mineralization is lacking in COPD. Using quantitative backscatter electron imaging (qBEI), we assessed cancellous (Cn.) and cortical (Ct.) bone mineralization density distribution (BMDD) in 19 postmenopausal women (62.1 ± 7.3 years of age) with COPD. Eight had sustained fragility fractures, and 13 had received treatment with inhaled glucocorticoids. The BMDD outcomes from the patients were compared with healthy reference data and were correlated with previous clinical and histomorphometric findings.In general, the BMDD outcomes for the patients were not significantly different from the reference data. Neither the subgroups of with or without fragility fractures or of who did or did not receive inhaled glucocorticoid treatment, showed differences in BMDD. However, subgroup comparison according to severity revealed 10% decreased cancellous mineralization heterogeneity (Cn.CaWidth) for the most severely affected compared with less affected patients (p = 0.042) and compared with healthy premenopausal controls (p = 0.021). BMDD parameters were highly correlated with histomorphometric cancellous bone volume (BV/TV) and formation indices: mean degree of mineralization (Cn.CaMean) versus BV/TV (r = 0.58, p = 0.009), and Cn.CaMean and Ct.CaMean versus bone formation rate (BFR/BS) (r = -0.71, p < 0.001). In particular, those with lower BV/TV (< 50th percentile) had significantly lower Cn.CaMean (p = 0.037) and higher Cn.CaLow (p = 0.020) compared with those with higher (> 50th percentile) BV/TV.The normality in most of the BMDD parameters and bone formation rates as well as the significant correlations between them suggests unaffected mineralization processes in COPD. Our findings also indicate no significant negative effect of treatment with inhaled glucocorticoids on the bone mineralization pattern. However, the observed concomitant occurrence of relatively lower bone volumes with lower bone matrix mineralization will both contribute to the reduced aBMD in some patients with COPD.     

Abnormally High and Heterogeneous Bone Matrix Mineralization After Childhood Solid Organ Transplantation: A Complex Pathology of Low Bone Turnover and Local Defects in Mineralization

Chronic renal, liver, and heart failure in children associates with multiple skeletal complications. Increased fracture incidence often persists after transplantation and could be related to alterations in bone material properties. In the present cohort study we evaluated bone mineralization density distribution (BMDD) by quantitative backscattered electron imaging (qBEI) in 23 pediatric solid organ allograft recipients with suspected osteoporosis. We measured BMDD in the entire cross‐sectional area of transiliac bone biopsies obtained from kidney (n = 9), liver (n = 9), and heart (n = 5) transplant recipients (aged 7.6 to 19.7 years; 6.0 ± 5.6 years posttransplantation, patients with a history of clinical fractures: n = 14). The BMDD findings were compared with age‐appropriate references and with a previously presented cohort of children with chronic kidney disease on dialysis (CKD5D, n = 18). Furthermore, we related the BMDD parameters with patients’ clinical and bone histomorphometric outcomes. Compared to healthy children, qBEI results for cancellous and cortical bone in transplant recipients revealed an increase in the most frequently occurring calcium concentration (+2.9%, p = 0.001; +3.5%, p = 0.014), in the portion of fully mineralized bone (fivefold; 10‐fold, both p < 0.0001) and in heterogeneity of mineralization (+26,5% and +27.8%, both p < 0.0001), respectively. Moreover, the BMDD parameters were nonsignificantly distinct from CKD5D cohort except that the heterogeneity in mineralization was higher posttransplantation. There was a strong inverse correlation between the average calcium content of the bone matrix and patients’ biochemical ALP levels, histomorphometric indices of bone formation and resorption. The abnormally high bone matrix mineralization in transplant recipients, consistent with serum and histomorphometric outcomes, suggests a history of low bone turnover with accumulation of fully mineralized bone packets. Additionally, the increased heterogeneity of mineralization suggests local alterations in mineralization kinetics, which may be linked to dysfunctional osteocytes that were recently shown to accumulate within the bone matrix during organ failure and concomitant glucocorticoid and immunosuppressive medication. © 2017 American Society for Bone and Mineral Research.     

Duration-Dependent Increase of Human Bone Matrix Mineralization in Long-Term Bisphosphonate Users with Atypical Femur Fracture

Bisphosphonates (BPs) are the most widely used drugs for the treatment of osteoporosis but prolonged use of BPs might increase the risk of atypical femur fracture (AFF). There are only a few studies that address the bone material quality in patients on long-term BP treatment with or without AFFs. We analyzed 52 trans-iliac bone biopsies from patients on long-term BP therapy with (n = 26) and without (n = 26) AFF. At the microscopic level, the degree of mineralization of bone (DMB) was assessed on whole bone by X-ray digitized microradiography while microhardness by Vickers microindentation, and bone matrix characteristics by Fourier transform infrared microspectroscopy (FTIRM) (mineral/organic ratio, mineral maturity and crystallinity, and collagen maturity) were measured at random focal areas. The AFF patients were treated longer than non-AFF patients (9.7 ± 3.3 years versus 7.9 ± 2.7 years). As expected, bone remodeling was low in both groups, without difference between them. The AFF group had significantly higher DMB in cortical bone (+2.9%, p = .001), which remained so after adjusting for treatment duration (p = .007), and showed a trend in cancellous bone (+1.6%, p = .05). Consistent with higher DMB, heterogeneity index (HI) was lower in the AFF than in the non-AFF group, illustrating lower heterogeneity of mineralization in the AFF group. A significant positive correlation between the duration of treatment and DMB in cortical bone was found in AFF, and not in the non-AFF group. Microhardness and bone matrix characteristics were similar between groups. We conclude that the AFF group had a duration-dependent increase in DMB leading to a significantly higher DMB than the non-AFF. Because BPs have high affinity to bone mineral and lining the walls of the osteocyte lacunae, the accumulation of matrix-bound BPs in AFF could lead to inhibition of the osteocyte cytoskeleton blunting their response to mechanical strains, a hypothesis to be further investigated. © 2021 American Society for Bone and Mineral Research (ASBMR).     

Altered bone composition in children with vertebral fracture

Primary osteoporosis in children often leads to vertebral fractures, but it remains unknown whether these fractures associate with changes in bone composition. This study aimed to determine the differences in bone composition in fracture‐prone children with and without vertebral fractures, as assessed by Fourier transform infrared spectroscopic imaging (FTIRI) and bone histomorphometry. Iliac crest bone biopsies (n = 24) were obtained from children who were suspected of primary osteoporosis based on evidence from the fracture history and/or low bone mineral density (BMD) by dual‐energy X‐ray absorptiometry. Vertebral morphology was determined by radiography. Bone biopsies were analyzed using histomorphometry and FTIRI. Phosphate‐to‐amide I, carbonate‐to‐phosphate, carbonate‐to‐amide I, and cross‐link ratio (collagen maturity) were calculated. Children with (n = 14) and without (n = 10) vertebral fracture were compared. Low cancellous bone volume (BV/TV) was detected by histomorphometry in 36% of the children with vertebral fracture, and bone turnover rate was abnormal in 64% of them. Children with vertebral fractures had lower carbonate‐to‐phosphate ratios (p < .05) and higher collagen maturity (p < .05) than children without vertebral fracture. The children with low BV/TV in biopsy showed lower carbonate‐to‐amide I ratios (p < .05) than the children with normal bone volume. This study showed changes in bone composition among fracture‐prone children who had sustained a vertebral fracture. The observed changes in bone composition in these children may contribute to their greater propensity to sustain vertebral fractures. © 2011 American Society for Bone and Mineral Research     

Women With Pregnancy and Lactation–Associated Osteoporosis (PLO) Have Low Bone Remodeling Rates at the Tissue Level

Pregnancy and lactation–associated osteoporosis (PLO) is a rare, severe, early form of osteoporosis in which young women present with fractures, usually multiple vertebral fractures, during late pregnancy or lactation. In studies of idiopathic osteoporosis (IOP) in premenopausal women, we enrolled 78 women with low-trauma fractures and 40 healthy controls, all with normal menses and no secondary cause of bone loss. In 15 of the affected women, the PLO subgroup, fractures had occurred during late pregnancy or lactation. We hypothesized that clinical, bone structural, and metabolic characteristics would differ between women with PLO and those with (non-PLO) IOP and controls. All were evaluated > 12 months postpartum, when structural and remodeling characteristics would be expected to reflect baseline premenopausal status rather than transient postpartum changes. As previously reported, affected subjects (PLO and IOP) had BMD and microarchitectural deficiencies compared to controls. Women with PLO did not differ from those with IOP in terms of age, BMI, body fat, menarcheal age, parity, or age at first pregnancy. However, women with PLO had a more severe clinical presentation than those with IOP: more fractures (5.5 ± 3.3 versus 2.6 ± 2.1; p = 0.005); more vertebral fractures (80% versus 17%; p < 0.001); and higher prevalence of multiple fractures. BMD deficits were more profound and cortical width tended to be lower in PLO. PLO subjects also had significantly lower tissue-level mineral apposition rate and bone formation rates (0.005 ± 0.005 versus 0.011 ± 0.010 mm²/mm/year; p = 0.006), as well as lower serum P1NP (33 ± 12 versus 44 ± 18 µg/L; p = 0.02) and CTX (257 ± 102 versus 355 ± 193 pg/mL; p = 0.01) than IOP. The finding that women with PLO have a low bone remodeling state assessed more than a year postpartum increases our understanding of the pathogenic mechanism of PLO. We conclude that women with PLO may have underlying osteoblast functional deficits which could affect their therapeutic response to osteoanabolic medications. © 2019 American Society for Bone and Mineral Research.     

Fragility Fractures in Men with Idiopathic Osteoporosis Are Associated with Undermineralization of the Bone Matrix without Evidence of Increased Bone Turnover

The pathogenesis of primary osteoporosis in younger individuals is still elusive. An important determinant of the biomechanical competence of bone is its material quality. In this retrospective study we evaluated bone material quality based on quantitative backscattered electron imaging to assess bone mineralization density distribution (BMDD) in bone biopsies of 25 male patients (aged 18–61 years) who sustained fragility fractures but were otherwise healthy. BMDD of cancellous bone was compared with previously established adult reference data. Complementary information was obtained by bone histomorphometry. The histomorphometric results showed a paucity of osteoblasts and osteoclasts on the bone surface in the majority of patients. BMDD revealed a significant shift to lower mineralization densities for cancellous bone values: CaMean (weighted mean Ca content, –5.9%), CaPeak (mode of the BMDD, −5.6%), and CaHigh (portion of fully mineralized bone, −76.8%) were decreased compared to normative reference; CaWidth (heterogeneity in mineralization, +18.5%) and CaLow (portion of low mineralized bone, +68.8; all P < 0.001) were significantly increased. The shift toward lower mineral content in the bone matrix in combination with reduced indices of bone formation and bone resorption suggests an inherent mineralization defect leading to undermineralized bone matrix, which might contribute to the susceptibility to fragility fractures of the patients. The alteration in bone material might be related to osteoblastic dysfunction and seems fundamentally different from that in high bone turnover osteoporosis with a negative bone balance.     

Bone matrix mineralization is preserved during early perimenopausal stage in healthy women: a paired biopsy study

Summary

Bone matrix mineralization based on quantitative backscatter electron imaging remained unchanged during the first year of menopause in paired transiliac biopsy samples from healthy women. This suggests that the reported early perimenopausal reductions in bone mineral density are caused by factors other than decreases in the degree of mineralization.

Introduction

It is unknown whether perimenopausal loss of bone mass is associated with a drop in bone matrix mineralization.

Methods

For this purpose, we measured the bone mineralization density distribution (BMDD) by quantitative backscatter electron imaging (qBEI) in n?=?17 paired transiliac bone biopsy samples at premenopausal baseline and 12 months after last menses (obtained at average ages of 49?±?2 and 55?±?2 years, respectively) in healthy women. For interpretation of BMDD outcomes, previously measured bone mineral density (BMD) and biochemical and histomorphometric markers of bone turnover were revisited for the present biopsy cohort.

Results

Menopause significantly decreased BMD at the lumbar spine (?4.5 %) and femoral neck (?3.8 %), increased the fasting urinary hydroxyproline/creatinine ratio (+60 %, all p?<?0.01) and histomorphometric bone formation rate (+25 %, p?<?0.05), but affected neither cancellous nor cortical BMDD variables (paired comparison p?>?0.05). Mean calcium concentrations of cancellous (Cn.CaMean) and cortical bone (Ct.CaMean) were within normal range (p?>?0.05 compared to established reference data). Ct.CaMean was significantly correlated with Cn.CaMean before (R?=?0.81, p?<?0.001) and after menopause (R?=?0.80, p?<?0.001) and to cortical porosity of mineralized tissue (Ct.Po.) after menopause (R?=??0.57, p?=?0.02).

Conclusions

Surprisingly, the BMDD was found not affected by the changes in bone turnover rates in this cohort. This suggests that the substantial increase in bone formation rates took place shortly before the second biopsy, and the bone mineralization changes lag behind. We conclude that during the first year after the last menses, the degree of bone matrix mineralization is preserved and does not contribute to the observed reductions in BMD.

     

Alendronate increases degree and uniformity of mineralization in cancellous bone and decreases the porosity in cortical bone of osteoporotic women

The strength of bone is correlated with bone mass but is also influenced significantly by other factors such as structural properties of the matrix (e.g., collagen mutations) and the mineral. Changes at all levels of this organization could contribute to fracture risk. We investigated the effects of alendronate (Aln) treatment on the density of mineralization and the ultrastructure of the mineral/collagen composite, size and habitus of mineral particles in iliac cancellous bone, as well as on the porosity of iliac cortical bone from postmenopausal osteoporotic women. Twenty-four transiliac bone biopsies from Phase III Aln (10 mg/day) trials (placebo and Aln after 2 and 3 years of treatment, n = 6 per group) were studied. The mineral structure was investigated by quantitative backscattered electron imaging (qBEI) and by scanning small-angle X-ray scattering (scanning-SAXS). qBEI histograms reflect the bone mineralization density distribution (BMDD), whereas SAXS patterns characterize the size and arrangement of the mineral particles in bone. We found that: (i) the relative calcium content of osteoporotic bone was significantly lower than that of data-base controls; (ii) mineralization was significantly higher and more uniform after Aln treatment; (iii) size and habitus of the mineral particles was not different between placebo and Aln-treated groups; and (iv) the porosity of cortical bone was reduced significantly by Aln treatment. We conclude that Aln treatment increases the degree and uniformity of bone matrix mineralization without affecting the size and habitus of the mineral crystals. It also decreases the porosity of the corticalis. Together these effects may contribute to the observed reduction in fractures.     

IGF‐1 Receptor Expression on Circulating Osteoblast Progenitor Cells Predicts Tissue‐Based Bone Formation Rate and Response to Teriparatide in Premenopausal Women With Idiopathic Osteoporosis

We have previously reported that premenopausal women with idiopathic osteoporosis (IOP) have profound microarchitectural deficiencies and heterogeneous bone remodeling. Those with the lowest bone formation rate have higher baseline serum insulin‐like growth factor‐1 (IGF‐1) levels and less robust response to teriparatide. Because IGF‐1 stimulates bone formation and is critical for teriparatide action on osteoblasts, these findings suggest a state of IGF‐1 resistance in some IOP women. To further investigate the hypothesis that osteoblast and IGF‐1‐related mechanisms mediate differential responsiveness to teriparatide in IOP, we studied circulating osteoblast progenitor (COP) cells and their IGF‐1 receptor (IGF‐1R) expression. In premenopausal women with IOP, peripheral blood mononuclear cells (PBMCs) were obtained at baseline (n = 25) and over 24 months of teriparatide treatment (n = 11). Flow cytometry was used to identify and quantify COPs (non‐hematopoetic lineage cells expressing osteocalcin and RUNX2) and to quantify IGF‐1R expression levels. At baseline, both the percent of PBMCs that were COPs (%COP) and COP cell‐surface IGF‐1R expression correlated directly with several histomorphometric indices of bone formation in tetracycline‐labeled transiliac biopsies. In treated subjects, both %COP and IGF‐1R expression increased promptly after teriparatide, returning toward baseline by 18 months. Although neither baseline %COP nor increase in %COP after 3 months predicted the bone mineral density (BMD) response to teriparatide, the percent increase in IGF‐1R expression on COPs at 3 months correlated directly with the BMD response to teriparatide. Additionally, lower IGF‐1R expression after teriparatide was associated with higher body fat, suggesting links between teriparatide resistance, body composition, and the GH/IGF‐1 axis. In conclusion, these assays may be useful to characterize bone remodeling noninvasively and may serve to predict early response to teriparatide and possibly other bone formation–stimulating medications. These new tools may also have utility in the mechanistic investigation of teriparatide resistance in premenopausal IOP and perhaps in other populations. © 2017 American Society for Bone and Mineral Research.     

Bone Mineral and Organic Properties in Postmenopausal Women Treated With Denosumab for Up to 10 years

In postmenopausal women with osteoporosis, denosumab (DMAb) therapy through 10 years resulted in significantly higher degree of mineralization of bone, with a subsequent increase from years 2–3 to year 5 and no further difference between years 5 and 10. Our aim was to assess the variables reflecting the quality of bone mineral and organic matrix (Fourier transform infrared microspectroscopy), and the microhardness of bone (Vickers microindentation). Cross-sectional assessments were performed in blinded fashion on iliac bone biopsies from osteoporotic women (72 from FREEDOM trial, 49 from FREEDOM Extension trial), separately in cortical and cancellous compartments. After 2–3 years of DMAb, mineral/matrix ratio and microhardness of cortical bone were significantly higher compared with placebo, whereas mineral maturity, mineral crystallinity, mineral carbonation, and collagen maturity were not different in both bone compartments. Through 5 years of DMAb, mineral carbonation was significantly lower and mineral/matrix ratio, mineral maturity, and crystallinity were significantly higher versus 2–3 years and were not different between 5 and 10 years, with the exception of mineral maturity in cancellous bone. These data support a transition of mineral to more mature crystals (within physiological range) and the completeness of secondary mineralization within 5 years of DMAb treatment. Microhardness in cortical and cancellous compartments was significantly lower at 5 years of DMAb versus 2–3 years and was not different from years 5 to 10. The lower microhardness at years 5 and 10 is likely the result of maturation of the organic matrix in a persistently low state of bone remodeling over 5 and 10 years. © 2022 American Society for Bone and Mineral Research (ASBMR).     

Bone micromechanical properties are compromised during long-term alendronate therapy independently of mineralization

In the treatment of postmenopausal osteoporosis (PMOP), the use of alendronate (ALN) leads to a decrease in the risk of vertebral and nonvertebral fractures. To explore the possible adverse effects of prolonged ALN therapy, we studied the effects of 8 ± 2 years (6–10 years) of ALN treatment on the iliac cortical bone mineral and collagen quality and micromechanical properties; by design, our study examined these parameters, independent of the degree of mineralization. From six ALN‐treated and five age‐matched untreated PMOP women, 153 bone structural units have been chosen according their degree of mineralization to obtain the same distribution in each group. In those bone structural units, Fourier transform infrared spectroscopy, quantitative microradiography, and nanoindentation were used to assess bone quality. Irrespective of the degree of mineralization, ALN treatment was associated with higher collagen maturity (+7%, p < 0.001, c.v. = 13% and 16% in treated and untreated women, respectively) and lower mineral crystallinity than that observed in the untreated PMOP group (?2%, p < 0.0001, c.v. = 3% in both groups). Bone matrix from ALN‐treated women also had lower elastic modulus (?12%, p < 0.0001, c.v. = 14% in both groups) and, contact hardness (?6%, p < 0.05, c.v. = 14% in both groups) than that of untreated women. Crystallinity (which reflects the size and perfection of crystals) was associated with both elastic modulus and contact hardness in treated women exclusively (r = 0.43 and r = 0.54, p < 0.0001, respectively), even after adjustment for the amount of mineral. We infer that long‐term ALN treatment compromises micromechanical properties of the bone matrix as assessed ex vivo. The strength deficits are in part related to difference in crystallinity, irrespective of the mineral amount and mineral maturity. These novel findings at local levels of bone structure will have to be taken into account in the study of the pathophysiology of bone fragilities associated with prolonged ALN treatment. © 2012 American Society for Bone and Mineral Research.     

Hypermineralization and High Osteocyte Lacunar Density in Osteogenesis Imperfecta Type V Bone Indicate Exuberant Primary Bone Formation

In contrast to “classical” forms of osteogenesis imperfecta (OI) types I to IV, caused by a mutation in COL1A1/A2, OI type V is due to a gain‐of‐function mutation in the IFITM5 gene, encoding the interferon‐induced transmembrane protein 5, or bone‐restricted interferon‐inducible transmembrane (IFITM)‐like protein (BRIL). Its phenotype distinctly differs from OI types I to IV by absence of blue sclerae and dentinogenesis imperfecta, by the occurrence of ossification disorders such as hyperplastic callus and forearm interosseous membrane ossification. Little is known about the impact of the mutation on bone tissue/material level in untreated and bisphosphonate‐treated patients. Therefore, investigations of transiliac bone biopsy samples from a cohort of OI type V children (n = 15, 8.7 ± 4 years old) untreated at baseline and a subset (n = 8) after pamidronate treatment (2.6 years in average) were performed. Quantitative backscattered electron imaging (qBEI) was used to determine bone mineralization density distribution (BMDD) as well as osteocyte lacunar density. The BMDD of type V OI bone was distinctly shifted toward a higher degree of mineralization. The most frequently occurring calcium concentration (CaPeak) in cortical (Ct) and cancellous (Cn) bone was markedly increased (+11.5%, +10.4%, respectively, p < 0.0001) compared to healthy reference values. Treatment with pamidronate resulted in only a slight enhancement of mineralization. The osteocyte lacunar density derived from sectioned bone area was elevated in OI type V Ct and Cn bone (+171%, p < 0.0001; +183.3%, p < 0.01; respectively) versus controls. The high osteocyte density was associated with an overall immature primary bone structure (“mesh‐like”) as visualized by polarized light microscopy. In summary, the bone material from OI type V patients is hypermineralized, similar to other forms of OI. The elevated osteocyte lacunar density in connection with lack of regular bone lamellation points to an exuberant primary bone formation and an alteration of the bone remodeling process in OI type V. © 2017 American Society for Bone and Mineral Research.