Aged Male Rats Regenerate Cortical Bone with Reduced Osteocyte Density and Reduced Secretion of Nitric Oxide After Mechanical Stimulation

Abstract

Mechanical loading is integral to the repair of bone damage. Osteocytes are mechanosensors in bone and participate in signaling through gap junction channels, which are primarily comprised of connexin 43 (Cx43). Nitric oxide (NO) and prostaglandin E2 (PGE2) have anabolic and catabolic effects on bone, and the secretion of these molecules occurs after mechanical stimulation. The effect of age on the repair of bone tissue after damage and on the ability of regenerated bone to transduce mechanical stimulation into a cellular response is unexplored. The goal of this study was to examine (1) osteocytes and their mineralized matrix within regenerated bone from aged and mature animals and (2) the ability of regenerated bone explants from aged and mature animals to transduce cyclic mechanical loading into a cellular response through NO and PGE2 secretion. Bilateral cortical defects were created in the diaphysis of aged (21-month-old) or mature (6-month-old) male rats, and new bone tissue was allowed to grow into a custom implant of controlled geometry. Mineralization and mineral-to-matrix ratio were significantly higher in regenerated bone from aged animals, while lacunar and osteocyte density and phosphorylated (pCx43) and total Cx43 protein were significantly lower, relative to mature animals. Regenerated bone from mature rats had increased pCx43 protein and PGE2 secretion with loading and greater NO secretion relative to aged animals. Reduced osteocyte density and Cx43 in regenerated bone in aged animals could limit the establishment of gap junctions as well as NO and PGE2 secretion after loading, thereby altering bone formation and resorption in vivo.

Osteocyte apoptosis: the roles and key molecular mechanisms in resorption-related bone diseases

“…Trigger the generation of pro-inflammatory and pro-osteoclastic factors via positive feedback loop [109][110][111][112]114 FADD Trigger a caspase cascade; induce GCs-induced apoptosis 6,118 Sclerostin Promote osteocyte cells death upon unloading; inhibit bone formation 53,107,108 BNIP3 Promote cell death during hypoxia 60,62 CCN2 Promote osteocyte apoptosis upon excess mechanical stress 33,69 TNF-α Stimulate osteocyte apoptosis upon inflammation and cancer 92,115,116 Caspase-3 Regulate osteocyte apoptosis via physical interactions in mechanistic stimulus 27,127 CTSK Breakdown the bone matrix adjacent to the osteocyte; increase the size of the osteocyte lacunae and mineralization decrease vitality of osteocytes 23 DMP-1 Regulate osteocyte formation and phosphate homeostasis; involve in osteocytic apoptosis 29,84,154 Pyk2 Promote GCs-induced osteocytic apoptosis via focal adhesion 82 Panx-1 Promote fatigue-induced osteocytic apoptosis 160 Anti-apoptotic function SOD2 Suppress aging and loss of bone mass; decrease degeneration of the osteocyte LCN 24,61 AMPK Protects against Hcy-induced osteocyte apoptosis 96,101,102 NO Maintain osteocytic vitality by pulsatile fluid flow 44,54,100…”

“…Of note, the osteocyte, one of the "permanent" cells of bone, regulates cell-to-cell communication of signals via Cx43 gap junctions to maintain cell survival, whereas, Table 2 Key pathological factors that are involved in the osteocyte apoptosis and their potential role. production of phosphorylated Cx43, prostaglandin E2 (PGE2), and nitric oxide (NO) would decrease with aging [41][42][43][44] (Table 1). It should be emphasized that miR21 deletion in miR21 fl/fl bones increased apoptosis-related gene expression, such as phosphatase and tensin homolog (PTEN), in contrast, a miR21 analog prevented apoptosis in Cx43 def osteocytes, indicating that disconnection of the Cx43/miR21 pathway leads to aging-induced osteocyte apoptosis 41 (Fig.…”

Senescent and apoptotic osteocytes and aging: Exercise to the rescue?

“…Aging and physical inactivity are each associated with a decrease in lacunae that contain osteocytes, loss of directional orientation of lacunae, and the number and length of dendrites that connect through canniculi; all of which would interfere with the detection of strain or fluid-flow shear stress [25][26][27][28]. Connexin43, a gap junction protein involved in mechanotransduction, protects from osteocyte apoptosis, but also decreases with aging [29][30][31][32]. Alternatively, unloading could induce senescence indirectly through the effect of inactivity on reductions in myokines, adverse changes in whole body nutrient trafficking, and the resulting lipid accumulation in both osteoblasts and osteocytes.…”

Role of connexins and pannexins during ontogeny, regeneration, and pathologies of bone

“…The effect of age on bone repair after damage and mechanical stimulation also involves gap junctional intercellular communication (GJIC) and Cx43 activity. For example, reduced osteocyte density and Cx43 levels were observed in regenerated bone in aged animals, limiting the establishment of GJIC, altering bone formation and bone resorption, as well NO and PGE 2 secretion [ 31 ].…”