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Paget's disease of bone (PDB) is a common metabolic bone disorder with a significant genetic component. To date, only one gene associated with PDB has been identified, the p62-Sequestosome1 gene (SQSTM1), and more than 20 mutations of this gene have been reported in PDB, the most common being the P392L substitution. In order to search for differentially expressed genes in PDB, we investigated the relative gene expression profile of candidate genes in osteoclast (OCL) cultures from 12 PDB patients and six unmatched healthy controls with known genetic status regarding p62, including healthy carriers of the P392L mutation. We selected 48 OCL-expressed candidate genes that may be involved in relevant pathways of PDB pathogenesis, such as OCL signaling, survival, bone resorption activity, or adhesion. In OCL cultures derived from peripheral blood mononuclear cells, total RNA extraction was performed, followed by real-time PCR experiments. Relative quantification analysis utilized the qBase method where relative expression levels were normalized with respect to a set of reference primer pairs for three housekeeping genes. When compared to non-mutated healthy controls, OCL cultures from PDB patients displayed a significant down-regulation in genes involved in apoptosis (CASP3 and TNFRSF10A), in cell signaling (TNFRSF11A), in the OCL bone resorbing function (ACP5 and CTSK) and in the gene coding for Tau protein (MAPT) (all comparisons, p

Atypical femur fractures (AFF) arise in the subtrochanteric and diaphyseal regions. Because of this unique distribution, we hypothesized that patients with AFF demonstrate specific geometrical variations of their lower limb whereby baseline tensile forces applied to the lateral cortex are higher and might favor the appearance of these rare stress fractures, when exposed to bisphosphonates. Using the low irradiation 2D-3D X-ray scanner EOS™ imaging technology we aimed to characterize and compare femur geometric parameters between women who sustained bisphosphonate-associated AFF and those who had experienced similar duration of exposure to bisphosphonates but did not sustain fractures. Conditional logistic regression models were constructed to estimate the association between selected geometric parameters and the occurrence of AFF. We identified 16 Caucasian women with AFF and recruited 16 ethnicity-, sex-, age-, height- and cumulative bisphosphonate exposure-matched controls from local osteoporosis clinics. Compared to controls, those with AFF had more lateral femur bowing (-3.2° SD [3.4] versus -0.8° SD [1.9] p=0.02). In regression analysis, lateral femur bowing was associated with the risk of AFF (aOR 1.54; 95% CI 1.04-2.28, p=0.03). Women who sustained a subtrochanteric AFF demonstrated a lesser femoral neck shaft angle (varus geometry) than those with a fracture at a diaphyseal site (121.9 [3.6]° versus 127.6 [7.2]°, p=0.07), whereas femur bowing was more prominent in those with a diaphyseal fracture compared to those with a subtrochanteric fracture (-4.3 [3.2]° versus -0.9 [2.7]°, p=0.07). Our analyses support that subjects with AFF exhibit femoral geometry parameters that result in higher tensile mechanical load on the lateral femur. This may play a critical role in the pathogenesis of AFF and requires further evaluation in a larger size population.

More than 20 mutations of the Sequestosome 1 (SQSTM1) gene have been reported in patients of European descent affected by Paget's disease of bone (PDB). In this investigation, a systematic screening for SQSTM1 mutations was conducted in consecutively evaluated unrelated patients with phenotypical PDB living in the New York City area (NY, United States). Seventy unrelated PDB patients with a multiethnic background, mostly of Jewish, Italian American, and Western European ancestries, were recruited. Sequencing of exons 7 and 8 was performed on DNA samples isolated from peripheral blood. Seven patients (10%) had SQSTM1 mutations, of which three had a family history of PDB. Four patients carried the C1215T (P392L) mutation, and three patients carried novel SQSTM1 missense mutations: T1085A (S349T), C1209T (A390V), and T1290A (L417Q) mutations. All PDB patients with SQSTM1 mutations had polyostotic involvement, and the mean number of affected bones was significantly higher in pagetic patient carriers of a SQSTM1 mutation when compared to non-mutated PDB patients (4.0 vs. 2.0, respectively; P = 0.003). Haplotype analysis in patient carriers of the P392L mutation revealed that all P392L mutations were carried by haplotype 2. The SQSTM1 mutation rate in unrelated American patients described in the present study was similar to that reported in European populations.