Osteocyte dysfunction promotes osteoarthritis through MMP13-dependent suppression of subchondral bone homeostasis.

Osteoarthritis (OA), long considered a primary disorder of articular cartilage, is commonly associated with subchondral bone sclerosis. However, the cellular mechanisms responsible for changes to subchondral bone in OA, and the extent to which these changes are drivers of or a secondary reaction to cartilage degeneration, remain unclear. In knee joints from human patients with end-stage OA, we found evidence of profound defects in osteocyte function. Suppression of osteocyte perilacunar/canalicular remodeling (PLR) was most severe in the medial compartment of OA subchondral bone, with lower protease expression, diminished canalicular networks, and disorganized and hypermineralized extracellular matrix. As a step toward evaluating the causality of PLR suppression in OA, we ablated the PLR enzyme MMP13 in osteocytes while leaving chondrocytic MMP13 intact, using Cre recombinase driven by the 9.6-kb DMP1 promoter. Not only did osteocytic MMP13 deficiency suppress PLR in cortical and subchondral bone, but it also compromised cartilage. Even in the absence of injury, osteocytic MMP13 deficiency was sufficient to reduce cartilage proteoglycan content, change chondrocyte production of collagen II, aggrecan, and MMP13, and increase the incidence of cartilage lesions, consistent with early OA. Thus, in humans and mice, defects in PLR coincide with cartilage defects. Osteocyte-derived MMP13 emerges as a critical regulator of cartilage homeostasis, likely via its effects on PLR. Together, these findings implicate osteocytes in bone-cartilage crosstalk in the joint and suggest a causal role for suppressed perilacunar/canalicular remodeling in osteoarthritis

Yoga jam: remixing Kirtan in the Art of Living

Yoga Jam are a group of musicians in the United Kingdom who are active members of the Art of Living, a transnational Hindu-derived meditation group. Yoga Jam organize events—also referred to as yoga raves and yoga remixes—that combine Hindu devotional songs (bhajans) and chants (mantras) with modern Western popular musical genres, such as soul, rock, and particularly electronic dance music. This hybrid music is often played in a clublike setting, and dancing is interspersed with yoga and meditation. Yoga jams are creative fusions of what at first sight seem to be two incompatible phenomena—modern electronic dance music culture and ancient yogic traditions. However, yoga jams make sense if the Durkheimian distinction between the sacred and the profane is challenged, and if tradition and modernity are not understood as existing in a sort of inverse relationship. This paper argues that yoga raves are authenticated through the somatic experience of the modern popular cultural phenomenon of clubbing combined with therapeutic yoga practices and validated by identifying this experience with a reimagined Vedic tradition

Effects of Chondroitinase ABC-Mediated Proteoglycan Digestion on Decellularization and Recellularization of Articular Cartilage.

Articular cartilage has a limited capacity to heal itself and thus focal defects often result in the development of osteoarthritis. Current cartilage tissue engineering strategies seek to regenerate injured tissue by creating scaffolds that aim to mimic the unique structure and composition of native articular cartilage. Decellularization is a novel strategy that aims to preserve the bioactive factors and 3D biophysical environment of the native extracellular matrix while removing potentially immunogenic factors. The purpose of this study was to develop a procedure that can enable decellularization and recellularization of intact articular cartilage matrix. Full-thickness porcine articular cartilage plugs were decellularized with a series of freeze-thaw cycles and 0.1% (w/v) sodium dodecyl sulfate detergent cycles. Chondroitinase ABC (ChABC) was applied before the detergent cycles to digest glycosaminoglycans in order to enhance donor chondrocyte removal and seeded cell migration. Porcine synovium-derived mesenchymal stem cells were seeded onto the decellularized cartilage scaffolds and cultured for up to 28 days. The optimized decellularization protocol removed 94% of native DNA per sample wet weight, while collagen content and alignment were preserved. Glycosaminoglycan depletion prior to the detergent cycles increased removal of nuclear material. Seeded cells infiltrated up to 100 μm into the cartilage deep zone after 28 days in culture. ChABC treatment enhances decellularization of the relatively dense, impermeable articular cartilage by reducing glycosaminoglycan content. ChABC treatment did not appear to affect cell migration during recellularization under static, in vitro culture, highlighting the need for more dynamic seeding methods

Glucocorticoids cause mandibular bone fragility and suppress osteocyte perilacunar-canalicular remodeling

Osteocytes support dynamic, cell-intrinsic resorption and deposition of bone matrix through a process called perilacunar/canalicular remodeling (PLR). In long bones, PLR depends on MMP13 and is tightly regulated by PTH, sclerostin, TGFβ, and glucocorticoids. However, PLR is regulated differently in the cochlea, suggesting a mechanism that is anatomically distinct. Unlike long bones, the mandible derives from neural crest and exhibits unique susceptibility to medication and radiation induced osteonecrosis. Therefore, we sought to determine if PLR in the mandible is suppressed by glucocorticoids, as it is in long bone. Hemimandibles were collected from mice subcutaneously implanted with prednisolone or vehicle containing pellets for 7, 21, or 55 days (n = 8/group) for radiographic and histological analyses. Within 21 days, micro-computed tomography revealed a glucocorticoid-dependent reduction in bone volume/total volume and trabecular thickness and a significant decrease in bone mineral density after 55 days. Within 7 days, glucocorticoids strongly and persistently repressed osteocytic expression of the key PLR enzyme MMP13 in both trabecular and cortical bone of the mandible. Cathepsin K expression was significantly reduced only after 55 days of glucocorticoid treatment, at which point histological analysis revealed a glucocorticoid-dependent reduction in the lacunocanalicular surface area. In addition to reducing bone mass and suppressing PLR, glucocorticoids also reduced the stiffness of mandibular bone in flexural tests. Thus, osteocyte PLR in the neural crest-derived mandible is susceptible to glucocorticoids, just as it is in the mesodermally-derived femur, highlighting the need to further study PLR as a target of drugs, and radiation in mandibular osteonecrosis

Glucocorticoids cause mandibular bone fragility and suppress osteocyte perilacunar-canalicular remodeling

Osteocytes support dynamic, cell-intrinsic resorption and deposition of bone matrix through a process called perilacunar/canalicular remodeling (PLR). In long bones, PLR depends on MMP13 and is tightly regulated by PTH, sclerostin, TGFβ, and glucocorticoids. However, PLR is regulated differently in the cochlea, suggesting a mechanism that is anatomically distinct. Unlike long bones, the mandible derives from neural crest and exhibits unique susceptibility to medication and radiation induced osteonecrosis. Therefore, we sought to determine if PLR in the mandible is suppressed by glucocorticoids, as it is in long bone. Hemimandibles were collected from mice subcutaneously implanted with prednisolone or vehicle containing pellets for 7, 21, or 55 days (n = 8/group) for radiographic and histological analyses. Within 21 days, micro-computed tomography revealed a glucocorticoid-dependent reduction in bone volume/total volume and trabecular thickness and a significant decrease in bone mineral density after 55 days. Within 7 days, glucocorticoids strongly and persistently repressed osteocytic expression of the key PLR enzyme MMP13 in both trabecular and cortical bone of the mandible. Cathepsin K expression was significantly reduced only after 55 days of glucocorticoid treatment, at which point histological analysis revealed a glucocorticoid-dependent reduction in the lacunocanalicular surface area. In addition to reducing bone mass and suppressing PLR, glucocorticoids also reduced the stiffness of mandibular bone in flexural tests. Thus, osteocyte PLR in the neural crest-derived mandible is susceptible to glucocorticoids, just as it is in the mesodermally-derived femur, highlighting the need to further study PLR as a target of drugs, and radiation in mandibular osteonecrosis. Keywords: Osteocyte, Perilacunar/canalicular remodeling, Mandible, Glucocorticoid

H&E staining of longitudinal cross sections of decellularized cartilage.

<p>Compared to native cartilage (Ntv), both groups of decellularized cartilage contained less cellular material in the superficial, middle, and deep zones. The PBS group had slightly more residual nucleic material than the chABC group, especially in the deep zone (G-I). Scale bar = 100 μm and magnification = 100X.</p

Overview of optimized decellularization procedure used in study.

<p>Harvested cartilage plugs were frozen at -20°C then subjected to two dry FTCs followed by two wet FTCs with hypotonic buffer. Plugs were then incubated in multiwell plates on an orbital shaker with high frequency agitation (220 RPM) for the remaining steps. Samples were submerged in a 0.125 U/ml chABC solution to digest GAGs. This was followed by three wash cycles of alternating hypotonic buffer and 0.1% (w/v) SDS detergent to increase cell membrane permeability and promote cell lysis. Finally, DNase and RNase were added to degrade nuclear material.</p

Summary of Recellularization Results.

<p>(A-N) Fluorescent labeling of seeded SDSCs. Seeded SDSCs were labeled with red DiI and all cells (seeded and native) were labeled with blue DAPI; fluorescent images were superimposed onto differential interference contrast images to more easily view infiltration. (A-F) Within 6 days, SDSCs attached to all outer surfaces, with some infiltration into the deep zone. (G-L) After 28 days, the outer cell layers grew thicker and there were generally more cells in the deep zone. (M-N) SDSCs were found throughout the lengths of the channels (channel borders indicated by small white arrows). (O) dsDNA/WW after 28 days in culture, as quantified by the PicoGreen assay. The seeded groups had significantly more dsDNA/WW compared to their respective unseeded controls, with no significant difference between the seeded PBS and chABC groups. For histology, scale bar = 100 μm and magnification = 200X for superficial and middle zones and 100X for circumference and channels. For biochemistry, data is plotted as arithmetic mean ± SEM; asterisks denote significant differences between groups (<i>p</i><0.05; <i>n</i> = 6–10).</p

Safranin-O and Fast Green staining of longitudinal cross sections of decellularized cartilage.

<p>Compared to native cartilage (Ntv), both groups of decellularized cartilage had significantly less proteoglycan staining, with more proteoglycan removal in the chABC-treated scaffolds compared to the PBS controls. Scale bar = 100 μm and magnification = 100X.</p