Transcriptome-based network analysis reveals renal cell type-specific dysregulation of hypoxia-associated transcripts [AB81]

Accumulating evidence suggests that dysregulation of hypoxia-regulated transcriptional mechanisms is involved in development of chronic kidney diseases (CKD). However, it remains unclear how hypoxia-induced transcription factors (HIFs) and subsequent biological processes contribute to CKD development and progression. In our study, genome-wide expression profiles of more than 200 renal biopsies from patients with different CKD stages revealed significant correlation of HIF-target genes with eGFR in glomeruli and tubulointerstitium. These correlations were positive and negative and in part compartment-specific. Microarrays of proximal tubular cells and podocytes with stable HIF1α and/or HIF2α suppression displayed cell type-specific HIF1/HIF2-dependencies as well as dysregulation of several pathways. WGCNA analysis identified gene sets that were highly coregulated within modules. Characterization of the modules revealed common as well as cell group- and condition-specific pathways, GO-Terms and transcription factors. Gene expression analysis of the hypoxia-interconnected pathways in patients with different CKD stages revealed an increased dysregulation with loss of renal function. In conclusion, our data clearly point to a compartment- and cell type-specific dysregulation of hypoxia-associated gene transcripts and might help to improve the understanding of hypoxia, HIF dysregulation, and transcriptional program response in CKD. Immortalized human podocytes (AB81) and human proximal tubular epithelial cells (HK2) with stable HIF1 and/or HIF2 suppression were subjected to normoxia (N, 20 % O2) or hypoxia (H, 1% O2) for 24h and gene expression profiles were generated. 3 replicates per group.

越来越多的研究证据表明，缺氧调控的转录机制失调参与了慢性肾脏病（chronic kidney disease, CKD）的发生发展。然而，目前尚不清楚缺氧诱导转录因子（hypoxia-induced transcription factors, HIFs）及其后续生物学过程如何推动CKD的发生与进展。本研究中，我们对200余例不同分期CKD患者的肾脏活检标本开展全基因组表达谱分析，结果显示HIF靶基因的表达与肾小球及肾小管间质的估算肾小球滤过率（estimated glomerular filtration rate, eGFR）存在显著相关性，此类相关性可呈正向或负向，且部分具有组织分区特异性。对稳定抑制HIF1α和/或HIF2α的近端肾小管上皮细胞及足细胞进行基因芯片分析，结果显示其存在细胞类型特异性的HIF1/HIF2依赖性调控模式，同时多条通路出现表达失调。加权基因共表达网络分析（weighted gene co-expression network analysis, WGCNA）结果显示，模块内存在高度共调控的基因集；对这些模块的特征解析表明，其既包含共有通路、基因本体术语（Gene Ontology terms, GO-Terms）及转录因子，也存在细胞群和实验条件特异性的通路与调控元件。对不同分期CKD患者体内缺氧关联通路的基因表达分析显示，随着肾功能减退，此类通路的失调程度逐渐加重。综上，本研究数据明确证实，缺氧相关基因转录本的失调具有组织分区和细胞类型特异性，该发现或有助于加深对CKD中缺氧、HIF失调及转录程序应答的理解。本研究将稳定抑制HIF1和/或HIF2的永生化人足细胞（AB81）及人近端肾小管上皮细胞（HK2）分别置于常氧（N，20% O₂）或缺氧（H，1% O₂）环境中培养24小时，随后获取其基因表达谱；每组设置3次生物学重复。