Atypical fracture with long-term bisphosphonate therapy is associated with altered cortical composition and reduced fracture resistance.

Bisphosphonates are the most widely prescribed pharmacologic treatment for osteoporosis and reduce fracture risk in postmenopausal women by up to 50%. However, in the past decade these drugs have been associated with atypical femoral fractures (AFFs), rare fractures with a transverse, brittle morphology. The unusual fracture morphology suggests that bisphosphonate treatment may impair toughening mechanisms in cortical bone. The objective of this study was to compare the compositional and mechanical properties of bone biopsies from bisphosphonate-treated patients with AFFs to those from patients with typical osteoporotic fractures with and without bisphosphonate treatment. Biopsies of proximal femoral cortical bone adjacent to the fracture site were obtained from postmenopausal women during fracture repair surgery (fracture groups, n = 33) or total hip arthroplasty (nonfracture groups, n = 17). Patients were allocated to five groups based on fracture morphology and history of bisphosphonate treatment [+BIS Atypical: n = 12, BIS duration: 8.2 (3.0) y; +BIS Typical: n = 10, 7.7 (5.0) y; +BIS Nonfx: n = 5, 6.4 (3.5) y; -BIS Typical: n = 11; -BIS Nonfx: n = 12]. Vibrational spectroscopy and nanoindentation showed that tissue from bisphosphonate-treated women with atypical fractures was harder and more mineralized than that from bisphosphonate-treated women with typical osteoporotic fractures. In addition, fracture mechanics measurements showed that tissue from patients treated with bisphosphonates had deficits in fracture toughness, with lower crack-initiation toughness and less crack deflection at osteonal boundaries than that of bisphosphonate-naïve patients. Together, these results suggest a deficit in intrinsic and extrinsic toughening mechanisms, which contribute to AFFs in patients treated with long-term bisphosphonates.

Raman and Fourier Transform Infrared (FT-IR) Mineral to Matrix Ratios Correlate with Physical Chemical Properties of Model Compounds and Native Bone Tissue.

Raman and Fourier transform infrared (FT-IR) spectroscopic imaging techniques can be used to characterize bone composition. In this study, our objective was to validate the Raman mineral:matrix ratios (ν1 PO4:amide III, ν1 PO4:amide I, ν1 PO4:Proline + hydroxyproline, ν1 PO4:Phenylalanine, ν1 PO4:δ CH2 peak area ratios) by correlating them to ash fraction and the IR mineral:matrix ratio (ν3 PO4:amide I peak area ratio) in chemical standards and native bone tissue. Chemical standards consisting of varying ratios of synthetic hydroxyapatite (HA) and collagen, as well as bone tissue from humans, sheep, and mice, were characterized with confocal Raman spectroscopy and FT-IR spectroscopy and gravimetric analysis. Raman and IR mineral:matrix ratio values from chemical standards increased reciprocally with ash fraction (Raman ν1 PO4/Amide III: P < 0.01, R2 = 0.966; Raman ν1 PO4/Amide I: P < 0.01, R2 = 0.919; Raman ν1 PO4/Proline + Hydroxyproline: P < 0.01, R2 = 0.976; Raman ν1 PO4/Phenylalanine: P < 0.01, R2 = 0.911; Raman ν1 PO4/δ CH2: P < 0.01, R2 = 0.894; IR P < 0.01, R2 = 0.91). Fourier transform infrared mineral:matrix ratio values from native bone tissue were also similar to theoretical mineral:matrix ratio values for a given ash fraction. Raman and IR mineral:matrix ratio values were strongly correlated ( P < 0.01, R2 = 0.82). These results were confirmed by calculating the mineral:matrix ratio for theoretical IR spectra, developed by applying the Beer-Lambert law to calculate the relative extinction coefficients of HA and collagen over the same range of wavenumbers (800-1800 cm-1). The results confirm that the Raman mineral:matrix bone composition parameter correlates strongly to ash fraction and to its IR counterpart. Finally, the mineral:matrix ratio values of the native bone tissue are similar to those of both chemical standards and theoretical values, confirming the biological relevance of the chemical standards and the characterization techniques.

Altered distributions of bone tissue mineral and collagen properties in women with fragility fractures.

Heterogeneity of bone tissue properties is emerging as a potential indicator of altered bone quality in pathologic tissue. The objective of this study was to compare the distributions of tissue properties in women with and without histories of fragility fractures using Fourier transform infrared (FTIR) imaging. We extended a prior study that examined the relationship of the mean FTIR properties to fracture risk by analyzing in detail the widths and the tails of the distributions of FTIR properties in biopsies from fracture and non-fracture cohorts. The mineral and matrix properties of cortical and trabecular iliac crest tissue were compared in biopsies from women with a history of fragility fracture (+Fx; n=21, age: mean 54±SD 15y) and with no history of fragility fracture (-Fx; n=12, age: 57±5y). A subset of the patients included in the -Fx group were taking estrogen-plus-progestin hormone replacement therapy (HRT) (-Fx+HRT n=8, age: 58±5y) and were analyzed separately from patients with no history of HRT (-Fx-HRT n=4, age: 56±7y). When the FTIR parameter mean values were examined by treatment group, the trabecular tissue of -Fx-HRT patients had a lower mineral:matrix ratio (M:M) and collagen maturity (XLR) than that of -Fx+HRT patients (-22% M:M, -18% XLR) and +Fx patients (-17% M:M, -18% XLR). Across multiple FTIR parameters, tissue from the -Fx-HRT group had smaller low-tail (5th percentile) values than that from the -Fx+HRT or +Fx groups. In trabecular collagen maturity and crystallinity (XST), the -Fx-HRT group had smaller low-tail values than those in the -Fx+HRT group (-16% XLR, -5% XST) and the +Fx group (-17% XLR, -7% XST). The relatively low values of trabecular mineral:matrix ratio and collagen maturity and smaller low-tail values of collagen maturity and crystallinity observed in the -Fx-HRT group are characteristic of younger tissue. Taken together, our data suggest that the presence of newly formed tissue that includes small/imperfect crystals and immature crosslinks, as well as moderately mature tissue, is an important characteristic of healthy, fracture-resistant bone. Finally, the larger mean and low-tail values of mineral:matrix ratio and collagen maturity noted in our -Fx+HRT vs. -Fx-HRT biopsies are consistent with greater tissue age and greater BMD arising from decreased osteoclastic resorption in HRT-treated patients.

Multiscale contribution of bone tissue material property heterogeneity to trabecular bone mechanical behavior.

Heterogeneity of material properties is an important potential contributor to bone fracture resistance because of its putative contribution to toughness, but establishing the contribution of heterogeneity to fracture risk is still in an incipient stage. Experimental studies have demonstrated changes in distributions of compositional and nanomechanical properties with fragility fracture history, disease, and pharmacologic treatment. Computational studies have demonstrated that models with heterogeneous material properties predict apparent stiffness moderately better than homogeneous models and show greater energy dissipation. Collectively, these results suggest that microscale material heterogeneity affects not only microscale mechanics but also structural performance at larger length scales.