Musculoskeletal Research Laboratory, SH Ho Scoliosis Research Laboratory, Department of Orthopaedics and Traumatology, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong.
Center for Neuromusculoskeletal Restorative Medicine, CUHK InnoHK Centres, Hong Kong Science Park, Hong Kong.
J Cachexia Sarcopenia Muscle. 2023 Jun;14(3):1349-1364. doi: 10.1002/jcsm.13228. Epub 2023 Apr 19.
The progressive deterioration of tissue-tissue crosstalk with aging causes a striking impairment of tissue homeostasis and functionality, particularly in the musculoskeletal system. Rejuvenation of the systemic and local milieu via interventions such as heterochronic parabiosis and exercise has been reported to improve musculoskeletal homeostasis in aged organisms. We have shown that Ginkgolide B (GB), a small molecule from Ginkgo biloba, improves bone homeostasis in aged mice by restoring local and systemic communication, implying a potential for maintaining skeletal muscle homeostasis and enhancing regeneration. In this study, we investigated the therapeutic efficacy of GB on skeletal muscle regeneration in aged mice.
Muscle injury models were established by barium chloride induction into the hind limb of 20-month-old mice (aged mice) and into C2C12-derived myotubes. Therapeutic efficacy of daily administrated GB (12 mg/kg body weight) and osteocalcin (50 μg/kg body weight) on muscle regeneration was assessed by histochemical staining, gene expression, flow cytometry, ex vivo muscle function test and rotarod test. RNA sequencing was used to explore the mechanism of GB on muscle regeneration, with subsequent in vitro and in vivo experiments validating these findings.
GB administration in aged mice improved muscle regeneration (muscle mass, P = 0.0374; myofiber number/field, P = 0.0001; centre nucleus, embryonic myosin heavy chain-positive myofiber area, P = 0.0144), facilitated the recovery of muscle contractile properties (tetanic force, P = 0.0002; twitch force, P = 0.0005) and exercise performance (rotarod performance, P = 0.002), and reduced muscular fibrosis (collagen deposition, P < 0.0001) and inflammation (macrophage infiltration, P = 0.03). GB reversed the aging-related decrease in the expression of osteocalcin (P < 0.0001), an osteoblast-specific hormone, to promote muscle regeneration. Exogenous osteocalcin supplementation was sufficient to improve muscle regeneration (muscle mass, P = 0.0029; myofiber number/field, P < 0.0001), functional recovery (tetanic force, P = 0.0059; twitch force, P = 0.07; rotarod performance, P < 0.0001) and fibrosis (collagen deposition, P = 0.0316) in aged mice, without an increased risk of heterotopic ossification.
GB treatment restored the bone-to-muscle endocrine axis to reverse aging-related declines in muscle regeneration and thus represents an innovative and practicable approach to managing muscle injuries. Our results revealed the critical and novel role of osteocalcin-GPRC6A-mediated bone-to-muscle communication in muscle regeneration, which provides a promising therapeutic avenue in functional muscle regeneration.
随着年龄的增长，组织间相互作用逐渐恶化，导致组织内稳态和功能显著受损，尤其是在肌肉骨骼系统中。通过诸如异体共生和运动等干预措施来恢复系统和局部环境，据报道可以改善老年生物体的肌肉骨骼内稳态。我们已经表明，来自银杏的小分子银杏内酯 B（GB）通过恢复局部和全身通讯来改善衰老小鼠的骨内稳态，这意味着维持骨骼肌内稳态和增强再生的潜力。在这项研究中，我们研究了 GB 对老年小鼠骨骼肌再生的治疗功效。
通过氯化钡诱导 20 月龄（老年）小鼠的后肢和 C2C12 衍生的肌管建立肌肉损伤模型。通过组织化学染色、基因表达、流式细胞术、离体肌肉功能测试和旋转棒测试评估每日给予 GB（12mg/kg 体重）和骨钙素（50μg/kg 体重）对肌肉再生的治疗效果。使用 RNA 测序来探索 GB 对肌肉再生的作用机制，并随后进行体外和体内实验验证这些发现。
GB 给药可改善老年小鼠的肌肉再生（肌肉质量，P=0.0374；肌纤维数/视野，P=0.0001；中心核，胚胎肌球蛋白重链阳性肌纤维面积，P=0.0144），促进肌肉收缩特性的恢复（强直力，P=0.0002；抽搐力，P=0.0005）和运动表现（旋转棒性能，P=0.002），并减少肌肉纤维化（胶原沉积，P<0.0001）和炎症（巨噬细胞浸润，P=0.03）。GB 逆转了与年龄相关的骨钙素（一种成骨细胞特异性激素）表达的下降，从而促进肌肉再生。外源性骨钙素补充足以改善肌肉再生（肌肉质量，P=0.0029；肌纤维数/视野，P<0.0001）、功能恢复（强直力，P=0.0059；抽搐力，P=0.07；旋转棒性能，P<0.0001）和纤维化（胶原沉积，P=0.0316）老年小鼠，而不会增加异位骨化的风险。
GB 治疗恢复了骨-肌肉内分泌轴，逆转了与年龄相关的肌肉再生下降，因此代表了一种管理肌肉损伤的创新和可行方法。我们的结果揭示了骨钙素-GPRC6A 介导的骨-肌肉通讯在肌肉再生中的关键和新颖作用，为功能性肌肉再生提供了有前途的治疗途径。
