Department of Medicine, and.
Department of Biomedical Informatics, Columbia University, New York, New York, USA.
JCI Insight. 2022 Mar 22;7(6):e146374. doi: 10.1172/jci.insight.146374.
The current strategy to detect acute injury of kidney tubular cells relies on changes in serum levels of creatinine. Yet serum creatinine (sCr) is a marker of both functional and pathological processes and does not adequately assay tubular injury. In addition, sCr may require days to reach diagnostic thresholds, yet tubular cells respond with programs of damage and repair within minutes or hours. To detect acute responses to clinically relevant stimuli, we created mice expressing Rosa26-floxed-stop uracil phosphoribosyltransferase (Uprt) and inoculated 4-thiouracil (4-TU) to tag nascent RNA at selected time points. Cre-driven 4-TU-tagged RNA was isolated from intact kidneys and demonstrated that volume depletion and ischemia induced different genetic programs in collecting ducts and intercalated cells. Even lineage-related cell types expressed different genes in response to the 2 stressors. TU tagging also demonstrated the transient nature of the responses. Because we placed Uprt in the ubiquitously active Rosa26 locus, nascent RNAs from many cell types can be tagged in vivo and their roles interrogated under various conditions. In short, 4-TU labeling identifies stimulus-specific, cell-specific, and time-dependent acute responses that are otherwise difficult to detect with other technologies and are entirely obscured when sCr is the sole metric of kidney damage.
目前检测肾小管细胞急性损伤的策略依赖于血清肌酐水平的变化。然而,血清肌酐(sCr)是功能和病理过程的标志物,不能充分检测肾小管损伤。此外,sCr 可能需要数天才能达到诊断阈值,但肾小管细胞在数分钟或数小时内就会对损伤和修复程序做出反应。为了检测与临床相关刺激的急性反应,我们创建了表达 Rosa26-floxed-stop 尿嘧啶磷酸核糖基转移酶(Uprt)的小鼠,并接种 4-硫代尿嘧啶(4-TU)以在选定时间点标记新生 RNA。来自完整肾脏的 Cre 驱动的 4-TU 标记 RNA 表明,体积减少和缺血在集合管和闰细胞中诱导了不同的基因程序。即使是谱系相关的细胞类型,对这两种应激源的反应也表达了不同的基因。TU 标记还表明反应具有瞬时性。因为我们将 Uprt 置于普遍活跃的 Rosa26 基因座中,许多细胞类型的新生 RNA 都可以在体内进行标记,并在各种条件下研究它们的作用。简而言之,4-TU 标记可识别刺激特异性、细胞特异性和时间依赖性的急性反应,而其他技术很难检测到这些反应,并且当 sCr 是唯一的肾脏损伤指标时,这些反应完全被掩盖。