High bone mass in mice can be linked to lower osteoclast formation, resorptive capacity, and restricted in vitro sensitivity to inhibition by stable sulforaphane.

Mouse strains can have divergent basal bone mass, yet this phenotype is seldom reflected in the design of studies seeking to identify new modulators of bone resorption by osteoclasts. Sulforaphane exerts inhibitory effects on in vitro osteoclastogenesis in cells from C57BL/6 mice. Here, we explore whether a divergent basal bone mass in different mouse strains is linked both to in vitro osteoclastogenic potential and to SFX-01 sensitivity. Accordingly, osteoclasts isolated from the bone marrow (BM) of C57BL/6, STR/Ort and CBA mice with low, high, and intermediate bone mass, respectively, were cultured under conditions to promote osteoclast differentiation and resorption; they were also treated with chemically stabilised sulforaphane (SFX-01) and respective sensitivity to inhibition evaluated by counting osteoclast number/resorption activity on dentine discs. We observed that osteoclastogenesis exhibited different macrophage colony-stimulating factor/receptor activator of nuclear factor kappa-Β ligand sensitivity in these mouse strains, with cells from C57BL/6 and CBA generating higher osteoclast numbers than STR/Ort; the latter formed only half as many mature osteoclasts. We found that 100 nM SFX-01 exerted a potent and significant reduction in osteoclast number and resorptive activity in cells derived from C57BL/6 mice. In contrast, 10-fold higher SFX-01 concentrations were required for similar inhibition in CBA-derived cells and, strikingly, a further 2.5-fold greater concentration was required for significant restriction of osteoclast formation/function in STR/Ort. These data are consistent with the notion that the BM osteoclast precursor population contributes to the relative differences in mouse bone mass and that mice with higher bone mass exhibit lower in vitro osteoclastogenic potential as well as reduced sensitivity to inhibition by SFX-01.

Stable Sulforaphane Targets the Early Stages of Osteoclast Formation to Engender a Lasting Functional Blockade of Osteoclastogenesis.

Sulforaphane, the native but unstable form of SFX-01, is an antioxidant that activates the NRF2 and inhibits the NF-KB pathways to achieve its actions. Resolving the mechanism(s) by which SFX-01 serves to control the various osteoclastogenic stages may expose pathways that could be explored for therapeutic use. Here we seek to identify the stage of osteoclastogenesis targeted by SFX-01 and explore whether, like SFN, it exerts its actions via the NRF2 and NF-KB pathways. Osteoclasts generated from the bone marrow (BM) of mice were cultured with SFX-01 at different timepoints to examine each phase of osteoclastogenesis separately. This showed that SFX-01 exerted actions throughout the process of osteoclastogenesis, but had its largest effects in the early osteoclast precursor differentiation stage. Thus, treatment with SFX-01 for the duration of culture, for the initial 3 days differentiation or for as little as the first 24 h was sufficient for effective inhibition. This aligned with data suggesting that SFX-01 reduced DC-STAMP levels, osteoclast nuclear number and modified cytoskeletal architecture. Pharmacological regulation of the NRF2 pathways, via selective inhibitors/activators, supported the anti-osteoclastogenic roles of an SFX-01-mediated by NRF2 activation, as well as the need for tight NF-KB pathway regulation in osteoclast formation/function.

Reduced Field-of-View Diffusion-Weighted Imaging of the Lumbosacral Enlargement: A Pilot In Vivo Study of the Healthy Spinal Cord at 3T.

Diffusion tensor imaging (DTI) has recently started to be adopted into clinical investigations of spinal cord (SC) diseases. However, DTI applications to the lower SC are limited due to a number of technical challenges, related mainly to the even smaller size of the SC structure at this level, its position relative to the receiver coil elements and the effects of motion during data acquisition. Developing methods to overcome these problems would offer new means to gain further insights into microstructural changes of neurological conditions involving the lower SC, and in turn could help explain symptoms such as bladder and sexual dysfunction. In this work, the feasibility of obtaining grey and white matter (GM/WM) DTI indices such as axial/radial/mean diffusivity (AD/RD/MD) and fractional anisotropy (FA) within the lumbosacral enlargement (LSE) was investigated using a reduced field-of-view (rFOV) single-shot echo-planar imaging (ss-EPI) acquisition in 14 healthy participants using a clinical 3T MR system. The scan-rescan reproducibility of the measurements was assessed by calculating the percentage coefficient of variation (%COV). Mean FA was higher in WM compared to GM (0.58 and 0.4 in WM and GM respectively), AD and MD were higher in WM compared to GM (1.66 µm2ms-1 and 0.94 µm2ms-1 in WM and 1.2 µm2ms-1 and 0.82 µm2ms-1 in GM for AD and MD respectively) and RD was lower in WM compared to GM (0.58 µm2ms-1 and 0.63 µm2ms-1 respectively). The scan-rescan %COV was lower than 10% in all cases with the highest values observed for FA and the lowest for MD. This pilot study demonstrates that it is possible to obtain reliable tissue-specific estimation of DTI indices within the LSE using a rFOV ss-EPI acquisition. The DTI acquisition and analysis protocol presented here is clinically feasible and may be used in future investigations of neurological conditions implicating the lower SC.