自噬缺陷与小鼠模型关节老化过程中软骨损伤的关系。
The relationship of autophagy defects to cartilage damage during joint aging in a mouse model.
机构信息
The Scripps Research Institute La Jolla California and Instituto de Investigación Biomédica de A Coruña Complexo Hospitalario Universitario de A Coruña SERGAS and Universidade da Coruña, A Coruña, Spain.
The Scripps Research Institute, La Jolla, California.
出版信息
Arthritis Rheumatol. 2015 Jun;67(6):1568-76. doi: 10.1002/art.39073.
OBJECTIVE
Aging is a main risk factor for osteo arthritis (OA), the most prevalent musculoskeletal disorder. Defects in autophagy, an essential cellular homeostasis mechanism, have recently been observed in OA articular cartilage. The objectives of this study were to establish the constitutive level of autophagy activation in normal cartilage and to monitor the temporal relationship between changes in autophagy and aging-related degradation of cartilage in a mouse model.
METHODS
In GFP-LC3-transgenic mice, green fluorescent protein (GFP)-light chain 3 (LC3) is ubiquitously expressed, and the accumulation of GFP puncta, representing autophagosomes, was quantified by confocal microscopy as a measure of autophagy activation. Expression of the autophagy proteins autophagy-related protein 5 (ATG-5) and microtubule-associated protein 1 light chain 3 (LC3) was analyzed by immunohistochemistry. Cartilage cellularity, apoptotic cell death, and cartilage structural damage and changes in synovium and bone were examined by histology and immunohistochemistry.
RESULTS
Basal autophagy activation was detected in liver and knee articular cartilage from young (6-month-old) mice, with higher levels in cartilage than in liver in the same animals. In 28-month-old mice, there was a statistically significant reduction in the total number of autophagic vesicles per cell (P < 0.01) and in the total area of vesicles per cell (P < 0.01) in the articular cartilage as compared to that from young 6-month-old mice. With increasing age, the expression of ATG-5 and LC3 decreased, and this was followed by a reduction in cartilage cellularity and an increase in the apoptosis marker poly(ADP-ribose) polymerase p85. Cartilage structural damage progressed in an age-dependent manner subsequent to the autophagy changes.
CONCLUSION
Autophagy is constitutively activated in normal cartilage. This is compromised with aging and precedes cartilage cell death and structural damage.
目的
衰老是骨关节炎（OA）的主要危险因素，OA 是最常见的肌肉骨骼疾病。最近在 OA 关节软骨中观察到自噬这一重要细胞内稳态机制的缺陷。本研究的目的是确定正常软骨中自噬激活的组成性水平，并在小鼠模型中监测自噬与软骨衰老降解之间的时相关系。
方法
在 GFP-LC3 转基因小鼠中，绿色荧光蛋白（GFP）-轻链 3（LC3）广泛表达，通过共聚焦显微镜定量 GFP 斑点的积累，代表自噬体，作为自噬激活的衡量标准。通过免疫组织化学分析自噬蛋白自噬相关蛋白 5（ATG-5）和微管相关蛋白 1 轻链 3（LC3）的表达。通过组织学和免疫组织化学检查软骨细胞密度、凋亡细胞死亡以及软骨结构损伤和滑膜及骨的变化。
结果
在年轻（6 个月大）小鼠的肝脏和膝关节关节软骨中检测到基础自噬激活，同一动物的软骨中的自噬水平高于肝脏。与年轻的 6 个月大的小鼠相比，28 个月大的小鼠每个细胞的自噬小体总数（P<0.01）和每个细胞的小体总面积（P<0.01）均有统计学显著减少。随着年龄的增长，ATG-5 和 LC3 的表达减少，随后软骨细胞密度降低，凋亡标志物多聚（ADP-核糖）聚合酶 p85 增加。软骨结构损伤在自噬变化后呈年龄依赖性进展。
结论
自噬在正常软骨中持续激活。随着年龄的增长，自噬会受到影响，并且会先于软骨细胞死亡和结构损伤。
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