Institute of Neuroscience - Padova Section, Consiglio Nazionale delle Ricerche, viale G, Colombo 3, 35121 Padova, Italy.
Arthritis Res Ther. 2010;12(2):R52. doi: 10.1186/ar2963. Epub 2010 Mar 24.
The endoplasmic reticulum (ER) stress-response, evoked in mice by the overexpression of class I major histocompatibility complex antigen (MHC-I), was proposed as a major mechanism responsible for skeletal muscle damage and dysfunction in autoimmune myositis. The present study was undertaken to characterize in more detail the ER stress-response occurring in myofibers of patients with inflammatory myopathies, focusing on the expression and distribution of Grp94, calreticulin and Grp75, three ER chaperones involved in immunomodulation.
Muscle biopsies were obtained from seven healthy subjects and 29 myositis patients, who were subdivided into groups based on the morphological evidence of inflammation and/or sarcolemmal immunoreactivity for MHC-I. Biopsies were analyzed by means of immunohistochemistry and western blot using anti-Grp94, anti-calreticulin and anti-Grp75 specific antibodies. Parallel analyses on these ER chaperones were conducted in rabbit and/or murine skeletal muscle after experimental induction of regeneration or systemic inflammation.
Upregulation of Grp94 characterized regenerating myofibers of myositis patients (P = 0.03, compared with values detected in biopsies without signs of muscle regeneration) and developing and regenerating myofibers of mouse muscles. Conversely, levels of calreticulin and Grp75 increased about fourfold and twofold, respectively, in patient biopsies positive for sarcolemmal MHC-I immunoreactivity, compared with healthy subjects and patients negative for both inflammation and MHC-I labeling (P < 0.005). Differently from calreticulin, the Grp75 level increased significantly also in patient biopsies that displayed occasional sarcolemmal MHC-I immunoreactivity (P = 0.002), suggesting the interference of other mechanisms. Experimental systemic inflammation achieved in mice and rabbits by a single injection of bacterial lipopolysaccharide significantly increased Grp75 and calreticulin but not MHC-I expression in muscles.
These results indicate that, in myositis patients, muscle regeneration and inflammation, in addition to MHC-I upregulation, do evoke an ER stress-response characterized by the increased expression of Grp94 and Grp75, respectively. The increase in the muscle Grp75 level in patients showing occasional immunoreactivity for sarcolemmal MHC-I might be considered further as a broader indicator of idiopathic inflammatory myopathy.
内质网（ER）应激反应，在小鼠中由 I 类主要组织相容性复合体抗原（MHC-I）的过表达引起，被认为是导致自身免疫性肌炎中骨骼肌损伤和功能障碍的主要机制。本研究旨在更详细地描述发生在炎症性肌病患者肌纤维中的 ER 应激反应，重点研究参与免疫调节的三种 ER 伴侣蛋白 Grp94、钙网蛋白和 Grp75 的表达和分布。
从 7 名健康受试者和 29 名肌炎患者中获取肌肉活检组织，根据炎症的形态学证据和/或肌细胞膜免疫反应性 MHC-I 将患者分为组。使用抗 Grp94、抗钙网蛋白和抗 Grp75 特异性抗体通过免疫组织化学和 Western blot 分析活检组织。在兔和/或鼠骨骼肌中进行这些 ER 伴侣蛋白的平行分析，以实验诱导再生或全身炎症。
Grp94 的上调特征在于肌炎患者再生肌纤维（P = 0.03，与无肌肉再生迹象的活检组织中检测到的值相比）和发育和再生的鼠肌纤维。相反，在肌细胞膜免疫反应性 MHC-I 呈阳性的患者活检组织中，钙网蛋白和 Grp75 的水平分别增加了约四倍和两倍，与健康受试者和既无炎症又无 MHC-I 标记的患者相比（P < 0.005）。与钙网蛋白不同，Grp75 水平在仅偶尔出现肌细胞膜 MHC-I 免疫反应性的患者活检组织中也显著增加（P = 0.002），这表明其他机制的干扰。通过单次注射细菌脂多糖在小鼠和兔中实现的实验性全身炎症显著增加了肌肉中的 Grp75 和钙网蛋白表达，但不增加 MHC-I 表达。
这些结果表明，在肌炎患者中，肌肉再生和炎症，除了 MHC-I 的上调外，还分别引起 ER 应激反应，其特征是 Grp94 和 Grp75 的表达增加。在仅偶尔出现肌细胞膜 MHC-I 免疫反应性的患者中，肌肉 Grp75 水平的增加可进一步被视为特发性炎症性肌病的更广泛指标。
