Skeletal Muscle Contraction Reduces Effects of Unloading on Bone Independently from the Central Nervous System: Studies Using Functional Electrical Stimulation after Spinal Cord Transection

Spinal cord injury (SCI) causes severe bone loss and disrupts connections between higher centers in the central nervous system (CNS) and bone. Muscle contraction elicited by functional electrical stimulation (FES) partially protects against loss of bone but cellular and molecular events by which this occurs are unknown. Here, using a rat model, we characterized effects of 7 days of contraction-induced loading of tibia and fibula due to FES when begun 16 weeks after SCI. SCI reduced tibial and femoral BMD by 12-17% and promoted bone resorption, as indicated by increased serum CTX; SCI-related changes in CTX were reversed by FES. In cultures of bone marrow cell-derived cells, SCI increased the number of osteoclasts and mRNA levels of the several osteoclast differentiation markers; these changes were significantly reversed by FES. The number of osteoblasts was also reduced by SCI as was the ratio of OPG/RANKL mRNAs therein; the unfavorable change in OPG/RANKL ratio was partially reversed by FES. cDNA microarray analysis revealed that alterations in genes involved in signaling through Wnt, FSH/LH, PTH and calcineurin/NFAT pathways may be linked to the favorable action of FES on SCI-induced bone resorption. In particular, SCI increased levels of the Wnt inhibitors DKK1, sFRP2 and SOST in osteoblasts, These effects were completely or partially reversed by FES. Our results demonstrate an anti-bone resorptive activity of acute FES in bone loss after SCI and suggest potential underlying mechanisms, among them involving increased Wnt signaling to cause more favorable ratios of OPG and RANKL for the inhibition of osteoclastogenesis. The present study indicates that the effects of bone reloading on SCI- related bone remodeling occurred independently of the effects of higher CNS centers on bone.脊髓损伤（Spinal cord injury, SCI）可引发严重骨丢失，并破坏中枢神经系统（central nervous system, CNS）高级中枢与骨骼之间的信号连接。功能性电刺激（functional electrical stimulation, FES）诱发的肌肉收缩可部分缓解骨丢失，但介导该保护作用的细胞与分子机制尚未阐明。本研究以大鼠为模型，针对脊髓损伤造模16周后，通过FES诱导胫骨与腓骨产生收缩负载、持续7天的干预效果进行了系统表征。 SCI可使胫骨与股骨的骨密度（bone mineral density, BMD）降低12%~17%，并促进骨吸收，该现象可通过血清CTX水平升高得到印证；FES可逆转SCI导致的血清CTX水平异常变化。 在骨髓细胞衍生的细胞培养体系中，SCI可增加破骨细胞（osteoclast）数量以及多种破骨细胞分化标志物的mRNA表达水平，上述变化均可被FES显著逆转。SCI同时降低了成骨细胞（osteoblast）数量及其内骨保护素/核因子κB受体活化因子配体（OPG/RANKL）的mRNA比值，而FES可部分逆转OPG/RANKL比值的不利改变。 cDNA微阵列（cDNA microarray）分析显示，Wnt、卵泡刺激素/黄体生成素（FSH/LH）、甲状旁腺激素（PTH）以及钙调神经磷酸酶/NFAT（calcineurin/NFAT）等通路相关基因的表达异常，可能与FES对SCI诱导骨吸收的保护作用相关。具体而言，SCI可上调成骨细胞内Wnt抑制因子DKK1、sFRP2及SOST的表达水平，而FES可完全或部分逆转上述效应。 本研究结果证实，急性FES可对脊髓损伤后骨丢失发挥抗骨吸收作用，并揭示了其潜在的作用机制，其中包括增强Wnt信号通路活性，以优化OPG与RANKL的比值，从而更有效地抑制破骨细胞生成。此外，本研究表明，骨骼负载恢复对SCI相关骨重塑的调控作用，并不依赖于中枢神经系统高级中枢对骨骼的直接调控。