Erythrocytes as Time Capsules of Disease Activity in Systemic Lupus Erythematosus

描述(申请人提供)：系统性红斑狼疮(SLE)是一种影响多个器官的典型自身免疫性疾病。利用最近开发的一种流式细胞仪分析，我们发现SLE患者红细胞表面有大量补体C4衍生的激活产物沉积。从初步研究中出现的其他有趣的发现包括：1)在SLE患者中检测到与红细胞结合的C4衍生激活产物(E-C4)水平升高和波动，2)同一患者中同时存在不同E-C4水平的不同红细胞亚群。这些发现导致了这一假设的中心假设，即红细胞和网织红细胞可能分别作为体内炎症状态(从而疾病活动)的“时间胶囊”和“即时信使”。一方面，在疾病暴发期间循环的红细胞可能会在其表面获得更多的C4衍生的激活产物，而从疾病爆发过程中产生的红细胞 缓解期间的骨髓可能会承受C4衍生产物水平的下降。另一方面，网织红细胞(红细胞的最年轻形式)，当在疾病活跃状态下从骨髓中出现时，可能会立即接触到并获得高水平的C4衍生产品。因此，测定血液循环中的红细胞和网织红细胞表面C4衍生的激活产物的水平应能揭示在过去120天(红细胞的寿命)内的疾病活动，并可能为正在进行或即将发生的疾病激活提供线索。本申请中提出的研究旨在验证这一假说(特异性目标1和2)，研究C4多态与C4衍生产物在红细胞/网织红细胞上沉积的关系(特异性目标3)，并阐明C4衍生产物沉积在红细胞/网织红细胞上的生化基础(特异性目标4)。建议的研究将通过i)年龄分级红细胞和网织红细胞的流式细胞术分析，ii)E-C4水平、网织红细胞结合C4水平与SLE疾病活动之间的相关性的统计分析，iii)SLE患者和健康对照的C4基因分型和表型，以及iv)红细胞和C4之间相互作用的生化研究来完成。这些拟议的研究将是在SLE疾病活动的背景下，旨在调查补体和红细胞/网织红细胞之间相互作用的生化和临床相关性的第一次努力。最终，从拟议的研究中获得的信息可能有助于开发新的实验室测试，以便更早和更准确地检测SLE耀斑，从而促进制定和评估SLE的新治疗方法。

DESCRIPTION (provided by applicant): Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease affecting multiple organs. Using a recently developed flow cytometric assay, we discovered the deposition of significant amounts of complement C4-derived activation product on the surface of erythrocytes of SLE patients. Additional interesting findings emerging from the preliminary studies include: 1) elevated and fluctuating levels of erythrocyte-bound C4-derived activation products (E-C4) were detected among SLE patients, and 2) distinct subpopulations of erythrocytes with different E-C4 levels were concurrently present in the same patient. These findings have led to the central hypothesis of this proposal that erythrocytes and reticulocytes may serve as, respectively, "time capsules" and "instant messenger" of the inflammatory condition in vivo (and thus disease activity) in SLE patients. On one hand, erythrocytes circulating during a disease flare may acquire an increased amount of C4-derived activation products on their surface, whereas erythrocytes emerging from the bone marrow during remission may bear decreased levels of C4-derived products. On the other hand, reticulocytes (the youngest form of erythrocytes), when emerging from the bone marrow during an active disease state, may immediately be exposed to and acquire high levels of C4-derived products. Therefore, determination of the levels of C4-derived activation products on the surface of erythrocytes and reticulocytes circulating at any given time should reveal disease activity during the preceding 120 days (the life span of erythrocytes) and may provide clues to ongoing/forthcoming disease activation. The research proposed in this application is aimed to verify this hypothesis (Specifics Aim 1 and 2), to investigate the relationship between C4 polymorphism and the deposition of C4-derived products on erythrocytes/reticulocytes (Specific Aim 3), and to elucidate the biochemical basis of the deposition of C4-derived products on erythrocytes/reticulocytes (Specific Aim 4). The proposed studies will be accomplished by i) flow cytometric analysis of age-fractionated erythrocytes and reticulocytes, ii) statistical analysis of the correlation between E-C4 levels, reticulocyte-bound C4 levels, and SLE disease activity, iii) genotyping and phenotyping of C4 in SLE patients and healthy controls, and iv) biochemical studies of the interaction between erythrocytes and C4. These proposed studies will constitute the first endeavor aimed to investigate the biochemical and clinical relevance of the interaction between complement and erythrocytes/reticulocytes, in the context of SLE disease activity. Ultimately, information derived from the proposed studies may aid in the development of new laboratory tests for earlier and more accurate detection of SLE flares, and thereby facilitating the formulation and assessment of new therapeutic approaches for SLE.