Accurate profiles of IgA1 O-glycan heterogeneity in IgA nephropathy

IgA 肾病中 IgA1 O-聚糖异质性的准确分析

• 批准号: 7670445
• 负责人: Matthew B Renfrow
• 金额: $ 18.13万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-08-15 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7670445
• 关键词: Acetylgalactosamine; Address; Antibodies; Antigen-Antibody Complex; Binding; Biological Assay; Biopsy; Classification; Complex; Deposition; Diagnosis; Digestion; Disaccharides; Disease; Disease Marker; Dissociation; Electrons; End stage renal failure; Enzyme-Linked Immunosorbent Assay; Fourier transform ion cyclotron resonance; Future; Galactose; Glomerulonephritis; Glycopeptides; Heterogeneity; IgA1; Immunoglobulin A; Individual; Inpatients; Investigation; Kidney; Kidney Diseases; Laboratories; Lead; Lectin; Link; Lupus Nephritis; Mass Spectrum Analysis; Methodology; Methods; Molecular; Myeloma Proteins; N-terminal; Pathogenesis; Pathology; Patients; Pattern; Peptide Hydrolases; Polysaccharides; Population; Protein Glycosylation; Protocols documentation; Recruitment Activity; Renal Circulation; Reporting; Research; Role; Sampling; Series; Serum; Site; Techniques; Testing; Time; Trypsin; design; disorder control; glycoprotein structure; glycosylation; insight; mesangial cell; novel; public health relevance; research study; tandem mass spectrometry

DESCRIPTION (provided by applicant): IgA nephropathy (IgAN) is the most common form of glomerulonephritis in the world and is a leading cause of end-stage renal disease. Although the mechanisms of its pathogenesis remain unclear, reports from several laboratories indicate a key role for an aberrantly glycosylated IgA1 present in the circulation and renal deposits. This IgA1 is galactose (Gal)-deficient in some of its 3-5 O-linked glycans in the hinge region (HR) of the heavy chain. Thus, in patients with IgAN, the IgA1 O-linked glycans are truncated, with terminal N acetylgalactosamine (GalNAc) or sialylated GalNAc. A variety of methods (glycan-specific lectin binding assays, MALDI-TOF MS methods, and chromatographic analysis of glycan composition) have detected clear differences between normal healthy controls' and IgAN patients' O-glycan populations in IgA1. However, the sites of attachment of normal and Gal-deficient glycans remain to be defined. Thus, a method is needed that can provide detailed structural information about the population of O-glycopeptides within each sample. Fourier transform-ion cyclotron resonance mass spectrometry (FT-ICR MS) provides unmatched mass accuracy in the identification of biomolecules, including the population of IgA1 O-glycopeptides enzymatically released from serum IgA1. We have recently localized (for the first time by a direct method) the multiple O-glycan chains in three different IgA1 HR glycopeptides by electron capture dissociation (ECD) FT-ICR tandem MS. This methodology enables us to test our hypothesis that the populations of serum IgA1 O-glycans differ in composition and sites of attachment in patients with IgAN when compared to normal healthy and disease controls and, furthermore, that these differences can serve as markers of the disease and define the pathological features. To establish accurate profiles of IgA1 glycoforms in IgAN patients we propose the following: 1. Provide an FT-ICR accurate mass profile of serum IgA1 glycoforms found inpatients with IgAN as well as a baseline profile from normal healthy and disease controls. and 2. Localize sites of O-glycan attachment in individual serum IgA1 glycoforms from patients with IgAN, normal healthy controls, and patients with other forms of glomerulonephritis by ECD FT-ICR tandem mass spectrometry. We perform this analysis on a total of 115 samples (45 IgAN, 25 lupus nephritis, and 45 healthy controls). These studies will provide detailed structural information about the aberrant glycosylation patterns found in IgAN, give insights into the pathogenesis of this disease, and may provide a novel noninvasive method for diagnosis of IgAN. PUBLIC HEALTH RELEVANCE: This two year project seeks to use novel mass spectrometry techniques to define the sites of aberrant O-glycosylation of serum IgA1 in patients with IgA nephropathy (IgAN). This can only be done in context of understanding the sites of IgA1 O-glycosylation in normal healthy controls and disease controls. The results will provide insights into the pathology of the IgAN and may provide a non-invasive method of diagnosing th3disease.

描述（由申请人提供）：IGA肾病（IGAN）是世界上肾小球肾炎的最常见形式，是终末期肾脏疾病的主要原因。尽管其发病机理的机制尚不清楚，但几个实验室的报告表明，循环和肾脏沉积物中存在异常糖基化的IgA1的关键作用。该IgA1在重链的铰链区域（HR）中的某些3-5 o连接的聚糖中缺乏半乳糖（GAL）。因此，在患有IGAN的患者中，Iga1 O连接的聚糖被截短，末端N乙酰乳糖胺（GalNAC）或乙二醇酸化的GALNAC。多种方法（聚糖特异性凝集素结合测定，MALDI-TOF MS方法和聚糖组成的色谱分析）在IGA1中发现了正常健康对照组和IGAN患者的O-聚糖种群之间的明显差异。然而，正常和gal的聚糖的附着位置仍有待定义。因此，需要一种可以提供有关每个样品中O-糖肽群的详细结构信息的方法。傅里叶转化离子回旋共振质谱法（FT-ICR MS）在鉴定生物分子（包括从血清IGA1中释放的IgA1 O-糖肽种群）提供了无与伦比的质量精度。我们最近（首次通过直接方法将多个O-聚糖链首次定位在三种不同的IgA1 HR糖肽中，通过电子捕获解离（ECD）FT-ICR串联MS。这种方法使我们能够检验我们的假设，即与正常的健康和疾病控制相比，Igan患者的血清IGA1 O-聚糖种群在组成和附着位置有所不同，此外，这些差异可以作为疾病的标志物，并定义病理特征。为了建立IgA1糖型在IGAN患者中的准确曲线，我们提出以下建议：1。提供FT-ICR精确的血清IGA1糖型的精确质量特征，发现患有IGAN的患者患者以及正常健康和疾病对照的基线谱。 2。来自IGAN，正常健康对照患者和其他形式的肾小球肾炎患者的O-Glycan附着的位置，通过ECD FT-ICR串联质谱法，来自IGAN，正常健康对照的患者和患有其他形式的肾小球肾炎患者的位置。我们对总共115个样本（45个Igan，25卢普斯肾炎和45个健康对照组）进行了此分析。这些研究将提供有关IGAN中发现异常的糖基化模式的详细结构信息，可深入了解该疾病的发病机理，并可能为诊断Igan提供新颖的无创方法。公共卫生相关性：这个为期两年的项目旨在使用新型的质谱技术来定义IgA肾病（IGAN）患者血清IGA1异常O-糖基化的部位。这只能在理解正常健康对照和疾病对照中IgA1 O-糖基化的位点的背景下完成。结果将提供有关IGAN病理学的见解，并可能提供一种诊断th3disease的非侵入性方法。