salubrinal and Body-Weight

Osteoporosis is a major skeletal disease with low bone mineral density, which leads to an increased risk of bone fracture. Salubrinal is a synthetic chemical that inhibits dephosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α) in response to endoplasmic reticulum (ER) stress. To understand possible linkage of osteoporosis to ER stress, we employed an unloading mouse model and examined the effects of salubrinal in the pathogenesis of disuse osteoporosis. The results presented several lines of evidence that osteoclastogenesis in the development of osteoporosis was associated with ER stress, and salubrinal suppressed unloading-induced bone loss. Compared to the age-matched control, unloaded mice reduced the trabecular bone area/total area (B.Ar/T.Ar) as well as the number of osteoblasts, and they increased the osteoclasts number on the trabecular bone surface in a time-dependent way. Unloading-induced disuse osteoporosis significantly increased the expression of Bip, p-eIF2α and ATF4 in short-term within 6h of tail suspension, but time-dependent decreased in HU2d to HU14d. Furthermore, a significant correlation of ER stress with the differentiation of osteoblasts and osteoclasts was observed. Administration of salubrinal suppressed the unloading-induced decrease in bone mineral density, B.Ar/T.Ar and mature osteoclast formation. Salubrinal also increased the colony-forming unit-fibroblasts and colony-forming unit-osteoblasts. It reduced the formation of mature osteoclasts, suppressed their migration and adhesion, and increased the expression of Bip, p-eIF2α and ATF4. Electron microscopy showed that rough endoplasmic reticulum expansion and a decreased number of ribosomes on ER membrane were observed in osteoblast of unloading mice, and the abnormal ER expansion was significantly improved by salubrinal treatment. A TUNEL assay together with CCAAT/enhancer binding protein homologous protein (CHOP) expression indicated that ER stress-induced osteoblast apoptosis was rescued by salubrinal. Collectively, the results support the notion that ER stress plays a key role in the pathogenesis of disuse osteoporosis, and salubrinal attenuates unloading-induced bone loss by altering proliferation and differentiation of osteoblasts and osteoclasts via eIF2α signaling.

Topics: Animals; Apoptosis; Body Weight; Bone Resorption; Cell Count; Cell Differentiation; Cell Survival; Cinnamates; Colony-Forming Units Assay; Endoplasmic Reticulum Stress; Female; Femur; Fibroblasts; Hindlimb Suspension; Mice, Inbred C57BL; Muscular Disorders, Atrophic; NFATC Transcription Factors; Osteoblasts; Osteoclasts; Osteogenesis; Osteoporosis; Thiourea; X-Ray Microtomography

Phosphorylation of eukaryotic initiation factor 2α (eIF2α), transiently activated by various cellular stresses, is known to alleviate stress-induced cellular damage. Here, we addressed a question: does elevation of eIF2α phosphorylation by salubrinal (a pharmacological inhibitor of eIF2α dephosphorylation) enhance healing of bone wounds? We hypothesized that salubrinal would accelerate a closure of surgically generated bone holes by modifying expression of stress-sensitive genes. To examine this hypothesis, we employed a rat wound model. Surgical wounds were generated on anterior and posterior femoral cortexes, and salubrinal was locally administered on the anterior side. The results showed that, compared to a contralateral control, the size of surgical wounds was reduced by 10.8 % (day 10) and 18.0 % (day 20) on the anterior side (both p < 0.001), and 4.1 % (day 10; p < 0.05) and 11.1 % (day 20; p < 0.001) on the posterior side. In addition, salubrinal locally elevated cortical thickness and increased BMD and BMC. Pharmacokinetic analysis revealed that subcutaneous injection of salubrinal transiently increased its concentration in plasma followed by a rapid decrease within 24 h, and its half-life in plasma was 1.2 h. Salubrinal altered the phosphorylation level of eIF2α as well as the mRNA levels of ATF3, ATF4, and CHOP, and suppressed cell death induced by stress to the endoplasmic reticulum. In summary, the results herein demonstrate that subcutaneous administration of salubrinal accelerates healing of surgically generated bone holes through the modulation of eIF2α phosphorylation.

Topics: Animals; Apoptosis; Body Weight; Cells, Cultured; Cinnamates; Endoplasmic Reticulum; Eukaryotic Initiation Factor-2; Female; Femur; Half-Life; Phosphorylation; Rats; Rats, Sprague-Dawley; RNA, Messenger; Stress, Physiological; Thiourea; Wound Healing