Abstract：
OBJECTIVE: Autoantibodies targeting intracellular proteins are common in various autoimmune diseases. In the context of myositis, the pathologic significance of these autoantibodies has been questioned due to the assumption that autoantibodies cannot enter living muscle cells. This study aims to investigate the validity of this assumption.
METHODS: Confocal immunofluorescence microscopy was employed to localise antibodies and other proteins of interest in myositis muscle biopsies. Bulk RNA sequencing was used to examine the transcriptomic profiles of 669 samples, including those from patients with myositis, disease controls and healthy controls. Additionally, antibodies from myositis patients were introduced into cultured myoblasts through electroporation, and their transcriptomic profiles were analysed using RNA sequencing.
RESULTS: In patients with myositis autoantibodies, antibodies accumulated inside myofibres in the same subcellular compartment as the autoantigen. Bulk RNA sequencing revealed that muscle biopsies from patients with autoantibodies targeting transcriptional regulators exhibited transcriptomic patterns consistent with dysfunction of the autoantigen. For instance, in muscle biopsies from patients with anti-PM/Scl autoantibodies recognising components of the nuclear RNA exosome complex, an accumulation of divergent transcripts and long non-coding RNAs was observed; these RNA forms are typically degraded by the nuclear RNA exosome complex. Introducing patient antibodies into cultured muscle cells recapitulated the transcriptomic effects observed in human disease. Further supporting evidence suggested that myositis autoantibodies recognising other autoantigens may also disrupt the function of their targets.
CONCLUSIONS: This study demonstrates that, in myositis, autoantibodies are internalised into living cells, causing biological effects consistent with the disrupted function of their autoantigen.
METHODS: Confocal immunofluorescence microscopy was employed to localise antibodies and other proteins of interest in myositis muscle biopsies. Bulk RNA sequencing was used to examine the transcriptomic profiles of 669 samples, including those from patients with myositis, disease controls and healthy controls. Additionally, antibodies from myositis patients were introduced into cultured myoblasts through electroporation, and their transcriptomic profiles were analysed using RNA sequencing.
RESULTS: In patients with myositis autoantibodies, antibodies accumulated inside myofibres in the same subcellular compartment as the autoantigen. Bulk RNA sequencing revealed that muscle biopsies from patients with autoantibodies targeting transcriptional regulators exhibited transcriptomic patterns consistent with dysfunction of the autoantigen. For instance, in muscle biopsies from patients with anti-PM/Scl autoantibodies recognising components of the nuclear RNA exosome complex, an accumulation of divergent transcripts and long non-coding RNAs was observed; these RNA forms are typically degraded by the nuclear RNA exosome complex. Introducing patient antibodies into cultured muscle cells recapitulated the transcriptomic effects observed in human disease. Further supporting evidence suggested that myositis autoantibodies recognising other autoantigens may also disrupt the function of their targets.
CONCLUSIONS: This study demonstrates that, in myositis, autoantibodies are internalised into living cells, causing biological effects consistent with the disrupted function of their autoantigen.
摘要：
目的：靶向细胞内蛋白的自身抗体在各种自身免疫性疾病中很常见。在肌炎的背景下，由于假设自身抗体不能进入活肌细胞，因此这些自身抗体的病理意义受到质疑。本研究旨在探讨这一假设的有效性。
方法：采用共聚焦免疫荧光显微镜在肌炎肌肉活检中定位抗体和其他感兴趣的蛋白质。使用大量RNA测序来检查669个样本的转录组概况，包括肌炎患者，疾病控制和健康控制。此外,通过电穿孔将肌炎患者的抗体引入培养的成肌细胞中，并使用RNA测序分析了它们的转录组概况。
结果：在患有肌炎自身抗体的患者中，抗体在肌纤维内与自身抗原相同的亚细胞区室中积累。大量RNA测序显示，来自具有靶向转录调节因子的自身抗体的患者的肌肉活检显示出与自身抗原功能障碍一致的转录组模式。例如,在抗PM/Scl自身抗体识别核RNA外泌体复合物成分的患者的肌肉活检中，观察到不同转录物和长非编码RNA的积累;这些RNA形式通常被核RNA外泌体复合物降解。将患者抗体引入培养的肌肉细胞中，可以概括人类疾病中观察到的转录组效应。进一步的支持证据表明，识别其他自身抗原的肌炎自身抗体也可能破坏其靶标的功能。
结论：这项研究表明，在肌炎中，自身抗体被内化到活细胞中，引起与其自身抗原功能破坏一致的生物效应。
方法：采用共聚焦免疫荧光显微镜在肌炎肌肉活检中定位抗体和其他感兴趣的蛋白质。使用大量RNA测序来检查669个样本的转录组概况，包括肌炎患者，疾病控制和健康控制。此外,通过电穿孔将肌炎患者的抗体引入培养的成肌细胞中，并使用RNA测序分析了它们的转录组概况。
结果：在患有肌炎自身抗体的患者中，抗体在肌纤维内与自身抗原相同的亚细胞区室中积累。大量RNA测序显示，来自具有靶向转录调节因子的自身抗体的患者的肌肉活检显示出与自身抗原功能障碍一致的转录组模式。例如,在抗PM/Scl自身抗体识别核RNA外泌体复合物成分的患者的肌肉活检中，观察到不同转录物和长非编码RNA的积累;这些RNA形式通常被核RNA外泌体复合物降解。将患者抗体引入培养的肌肉细胞中，可以概括人类疾病中观察到的转录组效应。进一步的支持证据表明，识别其他自身抗原的肌炎自身抗体也可能破坏其靶标的功能。
结论：这项研究表明，在肌炎中，自身抗体被内化到活细胞中，引起与其自身抗原功能破坏一致的生物效应。
