Cytokine Dysregulation in GM-CSF Autoimmunity

描述（由申请人提供）：特发性肺泡蛋白沉积症（PAP）是一种新发现的自身免疫性疾病，其特征是粒细胞-巨噬细胞集落刺激因子（GMCSF）的自身抗体中和和肺表面活性剂代谢严重失调，正常表面活性剂稳态中对GM-CSh的需求在GM-CSF缺陷小鼠的研究中得到证实，这些小鼠出现外源性GM-CSF可逆转的PAP样综合征。GM-CSF如何维持表面活性剂的稳态尚不清楚。人PAP肺泡巨噬细胞（AM）可以产生GM-CSF并对其产生反应，但其核转录因子过氧化物酶体增殖物激活受体qamma （PPARgamma）的表达水平较低。GM-CSF上调PPARgamma，提示GM-CSF-PPARgamma与人类PAP相关。有趣的是，在实验模型中，PPARgamma缺乏也与B细胞过度活跃和自身免疫有关。其他与B细胞功能障碍相关的细胞因子异常发生在PAP中。白细胞介素-10 （IL-10）是一种刺激抗体合成的多效细胞因子，在PAP肺和血液中升高，并阻断AM - GM-CSF的产生。激活素A，一种可能抑制B细胞循环的细胞因子，被GM-CSF上调，并且在PAP中也减少。因此，这些数据表明，PAP中存在GMCSF的中和性自身抗体会导致细胞因子失调，表面活性剂分解代谢严重受损和临床疾病。B淋巴细胞耗竭在自身免疫性疾病中显示出良好的临床效果。利妥昔单抗是一种针对B细胞特异性膜抗原CD20的嵌合鼠-人单克隆抗体，可选择性地消耗B细胞。根据我们的PAP数据，假设在利妥昔单抗临床试验的背景下减少PAP B细胞将降低抗GM-CSF抗体的水平，从而解决肺部疾病。在Aims 1-3中概述的机制研究将在基线和利妥昔单抗治疗后对肺和血液中的免疫细胞进行。目的1将研究：(a) IL-10、激活素a和ppγ对未经治疗的PAP B细胞增殖和抗GM-CSF合成的影响；(b) b细胞水平和抗gm - csf升数。目的2将研究PAP AM在局部b细胞调节中的作用，特别是：(a) GM-CSF对IL-10、激活素a和卵泡抑素（激活素a抑制剂）表达的影响；(b)激活素A对PPARgamma表达的影响。目的3将检测PAP T细胞：(a) PAP B细胞合成抗gm - csf的调节；(b)调节表型（CD4/CD25， PPARgamma表达），细胞因子（激活素A， IL-2, IL-10）和增殖。本应用程序的长期目标是描述免疫细胞，细胞因子和抗gm - csf自身抗体在PAP病理生理中的关系。利妥昔单抗试验提供了一个独特的机会来探索B细胞耗竭对PAP肺AM功能的影响，同时研究可能改善PAP肺部疾病的新疗法。

DESCRIPTION (provided by applicant): Idiopathic pulmonary alveolar proteinosis (PAP) is a newly recognized autoimmune disorder characterized by neutralizing autoantibodies to granulocyte-macrophage colony stimulating factor (GMCSF) and severely dysregulated lung surfactant metabolism, the GM-CSh requirement in normal surfactant homeostasis is evidenced by studies of GM-CSF-deficient mice which develop a PAP-like syndrome reversible with exogenous GM-CSF. How GM-CSF maintains surfactant homeostasis is unknown. Human PAP alveolar macrophages (AM) can produce and respond to GM-CSF but have low expression of the nuclear transcription factor, peroxisome proliferator-activated receptor qamma (PPARgamma). GM-CSF upregulates PPARgamma, suggesting a GM-CSF-PPARgamma link in human PAP. Interestingly, PPARgamma deficiency is also linked to B cell hyperactivity and autoimmunity in experimental models. Other cytokine anomalies associated with B cell dysfunction occur in PAP. lnterleukm-10 (IL-10), a pleiotropic cytokine that stimulates antibody synthesis, is elevated in PAP lung and blood and blocks AM GM-CSF production. Activin A, a cytokine that may restrain B cell cycling, is upregulated by GM-CSF, and is also reduced in PAP. Thus, these data suggest that the presence of neutralizing auto-antibodies to GMCSF in PAP results in cytokine dysregulation, severely impaired surfactant catabolism and clinical disease. Depletion of B lymphocytes has shown promising clinical results in autoimmune disease. Rituximab, a chimeric murine-human monoclonal antibody directed against the B cell-specific membrane antigen CD20, selectively depletes B cells. Based upon our PAP data, it is hypothesized that reducing PAP B Cells in the context of a Rituximab clinical trial will diminish levels of anti- GM-CSF antibodies, and thus resolve pulmonary disease. Mechanism studies outlined in Aims 1-3 will be conducted on immune cells from lung and blood at baseline and post Rituximab therapy. Aim 1 will investigate: (a) effects of IL-10, activin A, and PPARgamma on untreated PAP B cell proliferation and anti- GM-CSF synthesis; and (b) B cell levels and anti-GM-CSF liters. Aim 2 will examine the role of PAP AM in localized B-cell regulation, specifically: (a) GM-CSF effect on expression of IL-10, activin A and follistatin, (activin A inhibitor); and (b) activin A effect on PPARgamma expression. Aim 3 will examine PAP T cells for: (a) regulation of PAP B cell synthesis of anti-GM-CSF; and (b) regulatory phenotype (CD4/CD25, PPARgamma expression), cytokines (activin A, IL-2, IL-10), and proliferation. The long-term objective of this application is to delineate relationships among immune cells, cytokines and anti-GM-CSF autoantibody in PAP pathophysiology. The rituximab trial offers a unique opportunity to explore effects of B cell depletion on PAP pulmonary AM function and at the same time investigate new therapy that may improve the lung disease in PAP.