Reduced local mechanical stimuli in spaceflight diminishes osteocyte lacunar morphometry and spatial heterogeneity in mouse cortical bone

Three-dimensional (3D) imaging of osteocyte lacunae has recently substantiated the connection between lacunar shape and size, and osteocyte age, viability, and mechanotransduction. Yet it remains unclear why individual osteocytes reshape their lacunae and how networks of osteocytes change in response to local alterations in mechanical loads. We evaluated the effects of local mechanical stimuli on osteocyte lacunar morphometrics in tibial cortical bone from young female mice flown on the Space Shuttle for ~13 days. We optimized scan parameters, using a laboratory-based submicrometer-resolution X-Ray Microscope, to achieve large ~ 0.3 mm3 fields of view with sufficient resolution (≥ 0.3 μm) to visualize and measure thousands of lacunae per scan. Our novel approach avoids large measurement errors that are inherent in 2D and enables a facile 3D solution as compared to the lower resolution from benchtop micro-computed tomography (CT) systems or the cost and inaccessibility of synchrotron-based CT. Osteocyte lacunae were altered following microgravity exposure in a region-specific manner: more elongated (+7.0% Stretch) in predominately tensile-loaded bone as compared to those in compressively-loaded regions. In compressively-loaded bone, lacunae formed in microgravity were significantly larger (+6.9% Volume) than in the same region formed on Earth. We also evaluated lacunar heterogeneity (i.e., spatial autocorrelation of lacunar morphometric parameters) via kriging models. These statistical models demonstrated that heterogeneity varied with underlying spatial contributors, i.e. the local mechanical and biological environment. Yet in the absence of gravitational loading, osteocyte lacunae in newly formed bone were larger and were collectively more homogenous than in bone formed on Earth. Overall, this study shows that osteocyte reshape their lacunae in response to changes, or absence, in local mechanical stimuli and different biological environments. Additionally, spatial relationships among osteocytes are complex and necessitate evaluation in carefully selected regions of interest and of large cell populations.

Large-scale osteocyte lacunar morphological analysis of transiliac bone in normal and osteoporotic premenopausal women

Bone's adaptation ability is governed by the network of embedded osteocytes that inhabit individual crevasses called lacunae. The morphology of these lacunae and their resident osteocytes are known to change with age and diseases such as postmenopausal osteoporosis. However, it is unclear whether alterations in lacunar morphology are present in younger populations with osteoporosis. To investigate this, we implemented a previously validated methodology to image and quantify the three-dimensional morphometries of lacunae on a large scale (26.2 million cells) with ultra-high-resolution micro-computed tomography (microCT) in transiliac bone biopsies from three groups of premenopausal women: control n=39; idiopathic osteoporosis (IOP) n=45; idiopathic low BMD (ILBMD) n=19. Important lacunar morphometric parameters were measured in both trabecular and cortical bone: lacunar density (Lc.N/BV), lacunar porosity (Lc.TV/BV), lacunar number (Lc.N), lacunar volume (Lc.V), lacunar surface area (Lc.S), lacunar alignment (Lc.θ), lacunar stretch (Lc.St), lacunar oblateness (Lc.Ob), lacunar equancy (Lc.Eq), and lacunar sphericity (Lc.Sr). These were then compared against each other and also with previously measured tissue morphometries including: bone volume density (BV/TV), trabecular separation (Tb.Sp), trabecular number (Tb.N), and trabecular thickness (Tb.Th), structure model index (SMI), cortical porosity (Ct.Po) and cortical pore spacing (Ct.Sp). We detected no differences in lacunar morphology between the IOP, ILBMD and healthy premenopausal women. In contrast, we did find significant differences between lacunar morphologies in cortical and trabecular regions within all three groups, which was consistent with our previous findings on a subgroup of the healthy group. Furthermore, we discovered strong correlations between Lc.Sr from both trabecular and cortical regions with the measured BV/TV. The findings and comprehensive lacunar dataset we present here will be a crucial foundation for future investigations of the relationship between osteocyte lacunar morphology and disease.

Phenomenon of osteocyte lacunar mineralization: indicator of former osteocyte death and a novel marker of impaired bone quality?

An increasing number of patients worldwide suffer from bone fractures that occur after low intensity trauma. Such fragility fractures are usually associated with advanced age and osteoporosis but also with long-term immobilization, corticosteroid therapy, diabetes mellitus, and other endocrine disorders. It is important to understand the skeletal origins of increased bone fragility in these conditions for preventive and therapeutic strategies to combat one of the most common health problems of the aged population. This review summarizes current knowledge pertaining to the phenomenon of micropetrosis (osteocyte lacunar mineralization). As an indicator of former osteocyte death, micropetrosis is more common in aged bone and osteoporotic bone. Considering that the number of mineralized osteocyte lacunae per bone area can distinguish healthy, untreated osteoporotic and bisphosphonate-treated osteoporotic patients, it could be regarded as a novel structural marker of impaired bone quality. Further research is needed to clarify the mechanism of lacunar mineralization and to explore whether it could be an additional target for preventing or treating bone fragility related to aging and various endocrine diseases.

Hyperglycemia compromises Rat Cortical Bone by Increasing Osteocyte Lacunar Density and Decreasing Vascular Canal Volume

Uncontrolled diabetes is associated with increased risk of bony fractures. However, the mechanisms have yet to be understood. Using high-resolution synchrotron micro-CT, we calculated the changes in the microstructure of femoral cortices of streptozotocin-induced hyperglycemic (STZ) Wistar Albino rats and tested the mechanical properties of the mineralized matrix by nanoindentation. Total lacunar volume of femoral cortices increased in STZ group due to a 9% increase in lacunar density. However, total vascular canal volume decreased in STZ group due to a remarkable decrease in vascular canal diameter (7 ± 0.3 vs. 8.5 ± 0.4 µm). Osteocytic territorial matrix volume was less in the STZ group (14,908 ± 689 µm3) compared with healthy controls (16,367 ± 391 µm3). In conclusion, hyperglycemia increased cellularity and lacunar density, decreased osteocyte territorial matrix, and reduced vascular girth, in addition to decreasing matrix mechanical properties in the STZ group when compared with euglycemic controls.