Rheumatic Disease Sera: Probes of Disease Mechanism

描述（由申请人提供）：本申请的广泛、长期目标是确定人类自身免疫性肌病的致病机制，以期确定这类疾病的治疗相关途径。最近的几项研究已经确定细胞毒性淋巴细胞颗粒途径是自身免疫性肌炎的潜在重要途径。在多发性肌炎中，细胞毒性淋巴细胞颗粒经常向肌肉细胞极化，表明这些淋巴细胞处于活跃的脱颗粒过程中。此外，所有针对多发性肌炎和皮肌炎的自身抗原都是统一的，因为它们容易被颗粒酶B （GrB）有效地切割，产生在其他形式的细胞死亡中不会产生的独特片段。我们假设细胞毒性淋巴细胞颗粒通路在诱导肌肉细胞死亡和产生独特形式的自身抗原中起双重作用，从而驱动自身免疫性肌炎的自身免疫反应。该提案的具体目的是：(1)确定细胞毒性淋巴细胞在体外诱导肌肉细胞功能障碍和死亡的机制。这将通过阐明体外肌肉细胞中GrB下游的效应途径，并通过定义在成肌细胞中观察到的显著抑制caspases的机制来实现；(2)明确自身免疫性肌炎患者体内肌细胞损伤的途径。在Aim 1中被证明与功能相关的GrB下游通路的活性将使用一组新的试剂直接询问，这些试剂专门报告GrB、半胱天蛋白酶、钙蛋白酶、nNOS和肌肉结构和调节蛋白的状态和活性；(3)在动物模型中阐明grb介导的裂解和肌肉细胞分化状态对肌肉抗原免疫原性的影响。这些研究将探讨GrB切割在产生针对肌炎特异性自身抗原EF-1alpha的特异性免疫应答中的作用，并确定成肌细胞是否是针对肌肉特异性自身抗原的自身抗体应答的优先启动者。综上所述，这些研究将增强我们对参与自身免疫性肌炎发病机制的效应机制的理解。

DESCRIPTION (provided by applicant): The broad, long-term objectives of this proposal are to define pathogenic mechanisms in human autoimmune myopathies, with a view to defining pathways of therapeutic relevance in this group of diseases. Several recent studies have identified the cytotoxic lymphocyte granule pathway as a pathway of potential importance in autoimmune myositis. In polymyositis, cytotoxic lymphocyte granules are frequently polarized towards muscle cells, indicating that these lymphocytes are in the active process of degranulation. Furthermore, all autoantigens targeted in polymyositis and dermatomyositis are unified by their susceptibility to efficient cleavage by granzyme B (GrB), generating unique fragments not generated during other forms of cell death. We hypothesize that the cytotoxic lymphocyte granule pathway plays a dual role in inducing muscle cell death and in generating the unique forms of autoantigens, which drive the autoimmune response in autoimmune myositis. The specific aims of the proposal are to (1) Define the mechanisms whereby cytotoxic lymphocytes induce muscle cell dysfunction and death in muscle cells in vitro. This will be accomplished by elucidating the effector pathways downstream of GrB in muscle cells in vitro, and by defining the mechanisms responsible for the prominent inhibition of caspases observed in myoblasts; (2) Define the pathways of muscle cell damage in vivo in affected tissues from patients with autoimmune myositis. The activity of the pathways downstream of GrB demonstrated to be of functional relevance in Aim 1 will be directly interrogated using a set of novel reagents that specifically report on the state and activity of GrB, caspases, calpains, nNOS, and muscle structural and regulatory proteins; (3) Elucidate the role of GrB-mediated cleavage and muscle cell differentiation state on the immunogenicity of muscle antigens in an animal model. These studies will address the role of cleavage by GrB in generating the specific immune response to the myositis-specific autoantigen EF-1alpha and define whether the myoblast is the preferential initiator of autoantibody responses to muscle-specific autoantigens. Taken together, these studies will enhance our understanding of the effector mechanisms that participate in the pathogenesis of autoimmune myositis.