Supplementary Material for: Uromodulin and α1-Antitrypsin Urinary Peptide Analysis to Differentiate Glomerular Kidney Diseases

Background/Aims: Glomerular kidney disease (GKD) is suspected in patients based on proteinuria, but its diagnosis relies primarily on renal biopsy. We used urine peptide profiling as a noninvasive means to link GKD-associated changes to each glomerular entity. Methods: Urinary peptide profiles of 60 biopsy-proven glomerular patients and 14 controls were analyzed by combining magnetic bead peptide enrichment, MALDI-TOF MS analysis, and ClinProTools v2.0 to select differential peptides. Tentative identification of the differential peptides was carried out by HPLC-MS/MS. Results: The HPLC-MS/MS results suggest that uromodulin (UMOD; m/z: 1682, 1898 and 1913) and α1-antitrypsin (A1AT; m/z: 1945, 2392 and 2505) are differentially expressed urinary peptides that distinguish between GKD patients and healthy subjects. Low UMOD and high A1AT peptide abundance was observed in 80–92% of patients with GKD. Proliferative forms of GKD were distinguished from nonproliferative forms, based on a combination of UMOD and A1AT peptides. Nonproliferative forms correlated with higher A1AT peptide levels – focal segmental glomerulosclerosis was linked more closely to high levels of the m/z 1945 peptide than minimal change disease. Conclusion: We describe a workflow – urinary peptide profiling coupled with histological findings – that can be used to distinguish GKD accurately and noninvasively, particularly its nonproliferative forms.背景与研究目的：肾小球肾病（Glomerular kidney disease, GKD）临床可依据蛋白尿疑似诊断，但确诊仍主要依赖肾活检。本研究采用尿液肽谱分析作为无创检测手段，将肾小球肾病相关的分子变化与各肾小球病变类型关联起来。 研究方法：本研究共纳入60例经肾活检确诊的肾小球肾病患者与14例健康对照者，通过磁珠肽富集、基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）分析及ClinProTools v2.0软件筛选差异肽段；随后采用高效液相色谱-串联质谱（HPLC-MS/MS）对差异肽段进行初步鉴定。 研究结果：HPLC-MS/MS分析结果显示，尿调蛋白（uromodulin, UMOD；质荷比m/z：1682、1898、1913）与α1-抗胰蛋白酶（α1-antitrypsin, A1AT；质荷比m/z：1945、2392、2505）为差异表达的尿液肽段，可有效区分肾小球肾病患者与健康受试者。在80%~92%的肾小球肾病患者中，可观察到尿调蛋白肽段丰度降低、α1-抗胰蛋白酶肽段丰度升高。基于尿调蛋白与α1-抗胰蛋白酶肽段的联合检测，可区分增殖性与非增殖性肾小球肾病亚型。非增殖性肾小球肾病亚型与α1-抗胰蛋白酶肽段高表达相关，其中局灶节段性肾小球硬化相较于微小病变肾病，与m/z 1945肽段的高表达关联更为紧密。 研究结论：本研究构建了一套结合尿液肽谱分析与组织病理学结果的检测流程，可实现肾小球肾病的精准无创鉴别，尤其针对其非增殖性亚型。