Roberto Güerri

Microindentation for In Vivo Measurement of Bone Tissue Mechanical Properties in Humans

Bone tissue mechanical properties are deemed a key component of bone strength, but their assessment requires invasive procedures. Here we validate a new instrument, a reference point indentation (RPI) instrument, for measuring these tissue properties in vivo. The RPI instrument performs bone microindentation testing (BMT) by inserting a probe assembly through the skin covering the tibia and, after displacing periosteum, applying 20 indentation cycles at 2 Hz each with a maximum force of 11 N. We assessed 27 women with osteoporosis‐related fractures and 8 controls of comparable ages. Measured total indentation distance (46.0 ± 14 versus 31.7 ± 3.3 µm, p = .008) and indentation distance increase (18.1 ± 5.6 versus 12.3 ± 2.9 µm, p = .008) were significantly greater in fracture patients than in controls. Areas under the receiver operating characteristic (ROC) curve for the two measurements were 93.1% (95% confidence interval [CI] 83.1–100) and 90.3% (95% CI 73.2–100), respectively. Interobserver coefficient of variation ranged from 8.7% to 15.5%, and the procedure was well tolerated. In a separate study of cadaveric human bone samples (n = 5), crack growth toughness and indentation distance increase correlated (r = –0.9036, p = .018), and scanning electron microscope images of cracks induced by indentation and by experimental fractures were similar. We conclude that BMT, by inducing microscopic fractures, directly measures bone mechanical properties at the tissue level. The technique is feasible for use in clinics with good reproducibility. It discriminates precisely between patients with and without fragility fracture and may provide clinicians and researchers with a direct in vivo measurement of bone tissue resistance to fracture.

Are the High Hip Fracture Rates Among Norwegian Women Explained by Impaired Bone Material Properties

Hip fracture rates in Norway rank among the highest in the world, more than double that of Spanish women. Previous studies were unable to demonstrate significant differences between the two populations with respect to bone mass or calcium metabolism. In order to test whether the difference in fracture propensity between both populations could be explained by differences in bone material quality we assessed bone material strength using microindentation in 42 Norwegian and 46 Spanish women with normal BMD values, without clinical or morphometric vertebral fractures, no clinical or laboratory signs of secondary osteoporosis, and without use of drugs with known influence on bone metabolism. Bone material properties were assessed by microindentation of the thick cortex of the mid tibia following local anesthesia of the area using the Osteoprobe device (Active Life Scientific, Santa Barbara, CA, USA). Indentation distance was standardized against a calibration phantom of methylmethacrylate and results, as percentage of this reference value, expressed as bone material strength index units (BMSi). We found that the bone material properties reflected in the BMSi value of Norwegian women was significantly inferior when compared to Spanish women (77 ± 7.1 versus 80.7 ± 7.8, p < 0.001). Total hip BMD was significantly higher in Norwegian women (1.218 g/cm2 versus 0.938 g/cm2, p < 0.001) but regression analysis revealed that indentation values did not vary with BMD r2 = 0.03 or age r2 = 0.04. In conclusion Norwegian women show impaired bone material properties, higher bone mass, and were taller than Spanish women. The increased height will increase the impact on bone after falls, and impaired bone material properties may further enhance the risk fracture after such falls. These ethnic differences in bone material properties may partly explain the higher propensity for fracture in Norwegian women.

Bone Density, Microarchitecture, and Tissue Quality Long-term After Kidney Transplant.

Background Bone mineral density (BMD) measured by dual-energy x-ray absorptiometry is used to assess bone health in kidney transplant recipients (KTR). Trabecular bone score and in vivo microindentation are novel techniques that directly measure trabecular microarchitecture and mechanical properties of bone at a tissue level and independently predict fracture risk. We tested the bone status of long-term KTR using all 3 techniques. Methods Cross-sectional study including 40 KTR with more than 10 years of follow-up and 94 healthy nontransplanted subjects as controls. Bone mineral density was measured at lumbar spine and the hip. Trabecular bone score was measured by specific software on the dual-energy x-ray absorptiometry scans of lumbar spine in 39 KTR and 77 controls. Microindentation was performed at the anterior tibial face with a reference-point indenter device. Bone measurements were standardized as percentage of a reference value, expressed as bone material strength index (BMSi) units. Multivariable (age, sex, and body mass index-adjusted) linear regression models were fitted to study the association between KTR and BMD/BMSi/trabecular bone score. Results Bone mineral density was lower at lumbar spine (0.925 ± 0.15 vs 0.982 ± 0.14; P = 0.025), total hip (0.792 ± 0.14 vs 0.902 ± 0.13; P < 0.001), and femoral neck (0.667 ± 0.13 vs 0.775 ± 0.12; P < 0.001) in KTR than in controls. BMSi was also lower in KTR (79.1 ± 7.7 vs 82.9 ± 7.8; P = 0.012) although this difference disappeared after adjusted model (P = 0.145). Trabecular bone score was borderline lower (1.21 ± 0.14 vs 1.3 ± 0.15; adjusted P = 0.072) in KTR. Conclusions Despite persistent decrease in BMD, trabecular microarchitecture and tissue quality remain normal in long-term KTR, suggesting important recovery of bone health.

COL1A1 haplotypes and hip fracture

Fragility fractures resulting from low‐trauma events such as a fall from standing height are associated with osteoporosis and are very common in older people, especially women. Three single nucleotide polymorphisms (SNPs) at the COL1A1 gene (rs1107946, rs11327935, and rs1800012) have been widely studied and previously associated with bone mineral density (BMD) and fracture. A rare haplotype (T‐delT‐T) of these three SNPs was found to be greatly overrepresented in fractured individuals compared with nonfractured controls, thus becoming a good candidate for predicting increased fracture risk. The aim of our study was to assess the association of this haplotype with fracture risk in Spanish individuals. We recruited two independent groups of ∼100 patients with hip fracture (a total of 203 individuals) and compared the genotype and haplotype distributions of the three SNPs in the fractured patients with those of 397 control individuals from the BARCOS Spanish cohort. We found no association with risk of fracture at the genotype level for any of the SNPs, and no differences in the SNP frequencies between the two groups. At the haplotype level, we found no association between the T‐delT‐T haplotype and fracture. However, we observed a small but significant (p = 0.03) association with another rare haplotype, G‐insT‐T, which was slightly overrepresented in the patient group.

Genetic Analysis of High Bone Mass Cases from the BARCOS Cohort of Spanish Postmenopausal Women

The aims of the study were to establish the prevalence of high bone mass (HBM) in a cohort of Spanish postmenopausal women (BARCOS) and to assess the contribution of LRP5 and DKK1 mutations and of common bone mineral density (BMD) variants to a HBM phenotype. Furthermore, we describe the expression of several osteoblast-specific and Wnt-pathway genes in primary osteoblasts from two HBM cases. A 0.6% of individuals (10/1600) displayed Z-scores in the HBM range (sum Z-score >4). While no mutation in the relevant exons of LRP5 was detected, a rare missense change in DKK1 was found (p.Y74F), which cosegregated with the phenotype in a small pedigree. Fifty-five BMD SNPs from Estrada et al. [NatGenet 44:491-501,2012] were genotyped in the HBM cases to obtain risk scores for each individual. In this small group of samples, Z-scores were found inversely related to risk scores, suggestive of a polygenic etiology. There was a single exception, which may be explained by a rare penetrant genetic variant, counterbalancing the additive effect of the risk alleles. The expression analysis in primary osteoblasts from two HBM cases and five controls suggested that IL6R, DLX3, TWIST1 and PPARG are negatively related to Z-score. One HBM case presented with high levels of RUNX2, while the other displayed very low SOX6. In conclusion, we provide evidence of lack of LRP5 mutations and of a putative HBM-causing mutation in DKK1. Additionally, we present SNP genotyping and expression results that suggest additive effects of several genes for HBM.

Maintenance low dose systemic glucocorticoids have limited impact on bone strength and mineral density among incident renal allograft recipients: A pilot prospective cohort study

Soon after kidney transplant (KT), a decrease in parathormone and bone mineral density (BMD) occur, but little is known on the impact of KT on novel bone quality parameters including trabecular bone score (TBS) and bone material strength index (BMSi). We aimed to study BMD, TBS and BMSi in the first year after KT, in patients not treated with any bone therapy. A cohort including 36 patients underwent KT on a low-glucocorticoid-dose protocol (5 mg daily-prednisone from post-operative-day 42 onwards) and was observed for 12 months prospectively. At 3 months, phosphorus and parathormone decreased, while calcium increased. We also observed at 3 months a transient mild 2.9% bone loss at femoral neck (BMD change 0.752 ± 0.15 vs 0.730 ± 0.15; p = 0.004), but no change at either spine or total hip. Both TBS and BMSi remained stable. At 12 months, lumbar (but not total hip or femoral neck) BMD slightly decreased by 2.1% vs baseline (0.950 ± 0.15 vs 0.930 ± 0.5; p = 0.046), while TBS and BMSi remained unmodified. In KT patients on low-dose glucocorticoids and no bone therapy, there were small BMD decreases at femoral neck (at 3 months) and lumbar spine (at 12 months), but no change in either TBS or BMSi. Low-dose post-KT glucocorticoid treatment shows limited impact on bone, supporting steroid-restrictive protocols.