Table_2_An integrated co-expression network analysis reveals novel genetic biomarkers for immune cell infiltration in chronic kidney disease.xlsx

BackgroundChronic kidney disease (CKD) is characterized by persistent damage to kidney function or structure. Progression to end-stage leads to adverse effects on multiple systems. However, owing to its complex etiology and long-term cause, the molecular basis of CKD is not completely known.
MethodsTo dissect the potential important molecules during the progression, based on CKD databases from Gene Expression Omnibus, we used weighted gene co-expression network analysis (WGCNA) to identify the key genes in kidney tissues and peripheral blood mononuclear cells (PBMC). Correlation analysis of these genes with clinical relevance was evaluated based on Nephroseq. Combined with a validation cohort and receiver operating characteristic curve (ROC), we found the candidate biomarkers. The immune cell infiltration of these biomarkers was evaluated. The expression of these biomarkers was further detected in folic acid-induced nephropathy (FAN) murine model and immunohistochemical staining.
ResultsIn total, eight genes (CDCP1, CORO1C, DACH1, GSTA4, MAFB, TCF21, TGFBR3, and TGIF1) in kidney tissue and six genes (DDX17, KLF11, MAN1C1, POLR2K, ST14, and TRIM66) in PBMC were screened from co-expression network. Correlation analysis of these genes with serum creatinine levels and estimated glomerular filtration rate from Nephroseq showed a well clinical relevance. Validation cohort and ROC identified TCF21, DACH1 in kidney tissue and DDX17 in PBMC as biomarkers for the progression of CKD. Immune cell infiltration analysis revealed that DACH1 and TCF21 were correlated with eosinophil, activated CD8 T cell, activated CD4 T cell, while the DDX17 was correlated with neutrophil, type-2 T helper cell, type-1 T helper cell, mast cell, etc. FAN murine model and immunohistochemical staining confirmed that these three molecules can be used as genetic biomarkers to distinguish CKD patients from healthy people. Moreover, the increase of TCF21 in kidney tubules might play important role in the CKD progression.
DiscussionWe identified three promising genetic biomarkers which could play important roles in the progression of CKD.

Background: 慢性肾脏病（Chronic kidney disease, CKD）以肾脏功能或结构的持续性损伤为特征。病情进展至终末期时，会对全身多系统造成不良影响。然而，由于其病因复杂、病程漫长，目前慢性肾脏病的分子机制尚未完全阐明。
Methods: 为解析疾病进展过程中的潜在关键分子，本研究基于基因表达综合数据库（Gene Expression Omnibus, GEO）中的慢性肾脏病数据集，采用加权基因共表达网络分析（Weighted Gene Co-expression Network Analysis, WGCNA）分别鉴定肾脏组织与外周血单个核细胞（Peripheral Blood Mononuclear Cells, PBMC）中的核心基因。基于Nephroseq数据库对上述基因开展临床相关性分析，并结合验证队列与受试者工作特征曲线（Receiver Operating Characteristic curve, ROC）筛选候选生物标志物。此外，本研究还对这些生物标志物的免疫细胞浸润情况进行评估，并在叶酸诱导肾病（Folic acid-induced nephropathy, FAN）小鼠模型中验证其表达水平，同时通过免疫组化染色进一步确认。
Results: 本研究共从共表达网络中筛选得到肾脏组织相关基因8个（CDCP1、CORO1C、DACH1、GSTA4、MAFB、TCF21、TGFBR3、TGIF1）以及外周血单个核细胞相关基因6个（DDX17、KLF11、MAN1C1、POLR2K、ST14、TRIM66）。基于Nephroseq数据库的分析显示，上述基因与血清肌酐水平、估算肾小球滤过率均具有良好的临床相关性。验证队列与受试者工作特征曲线分析证实，肾脏组织中的TCF21、DACH1以及外周血单个核细胞中的DDX17可作为慢性肾脏病进展的生物标志物。免疫细胞浸润分析结果显示，DACH1与TCF21分别与嗜酸性粒细胞、活化CD8⁺T细胞、活化CD4⁺T细胞的浸润水平相关；而DDX17则与中性粒细胞、2型辅助性T细胞、1型辅助性T细胞、肥大细胞等的浸润水平相关。叶酸诱导肾病小鼠模型与免疫组化染色实验进一步验证，上述三种分子可作为区分慢性肾脏病患者与健康人群的遗传生物标志物。此外，肾小管中TCF21的表达上调可能在慢性肾脏病进展中发挥重要作用。
Discussion: 本研究筛选得到三种具有应用前景的遗传生物标志物，它们可能在慢性肾脏病的疾病进展过程中发挥关键作用。