Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, and Center for Research in Complex Systems Science, Mahidol University, Bangkok, Thailand.
Medical Proteomics Unit, Office for Research and Development, Faculty of Medicine Siriraj Hospital, and Center for Research in Complex Systems Science, Mahidol University, Bangkok, Thailand.
Metabolism. 2017 Sep;74:47-61. doi: 10.1016/j.metabol.2016.12.014. Epub 2017 Jan 4.
Chronic potassium (K) deficiency can cause renal damage namely hypokalemic nephropathy with unclear pathogenic mechanisms. In the present study, we investigated expression and functional alterations in renal tubular cells induced by prolonged K deficiency.
MDCK cells were maintained in normal-K (CNK) (K=5.3mmol/L), low-K (CLK) (K=2.5mmol/L), or K-depleted (CKD) (K=0mmol/L) medium for 10days (n=5 independent cultures/condition). Differentially expressed proteins were identified by a proteomics approach followed by various functional assays.
Proteomic analysis revealed 46 proteins whose levels significantly differed among groups. The proteomic data were confirmed by Western blotting. Gene Ontology (GO) classification and protein network analysis revealed that majority of the altered proteins participated in metabolic process, whereas the rest involved in cellular component organization/biogenesis, cellular process (e.g., cell cycle, regulation of cell death), response to stress, and signal transduction. Interestingly, ATP measurement revealed that intracellular ATP production was increased in CLK and maximum in CKD. Flow cytometry showed cell cycle arrest at S-phase and G2/M-phase in CLK and CKD, respectively, consistent with cell proliferation and growth assays, which showed modest and marked degrees of delayed growth and prolonged doubling time in CLK and CKD, respectively. Cell death quantification also revealed modest and marked degrees of increased cell death in CLK and CKD, respectively.
In conclusion, prolonged K deficiency increased intracellular ATP, cell cycle arrest and cell death in renal tubular cells, which might be responsible for mechanisms underlying the development of hypokalemic nephropathy.
慢性钾（K）缺乏可导致肾脏损伤，即低钾血症性肾病，其发病机制尚不清楚。在本研究中，我们研究了长期 K 缺乏诱导的肾小管细胞表达和功能改变。
MDCK 细胞在正常 K（CNK）（K=5.3mmol/L）、低 K（CLK）（K=2.5mmol/L）或 K 耗竭（CKD）（K=0mmol/L）培养基中维持 10 天（每组 5 个独立培养物）。通过蛋白质组学方法鉴定差异表达蛋白，然后进行各种功能测定。
蛋白质组学分析显示 46 种蛋白的水平在各组间有显著差异。蛋白质组学数据通过 Western blot 得到验证。GO 分类和蛋白质网络分析表明，大多数改变的蛋白参与代谢过程，而其余的则参与细胞成分组织/生物发生、细胞过程（如细胞周期、细胞死亡调控）、应激反应和信号转导。有趣的是，ATP 测量显示 CLK 和 CKD 中细胞内 ATP 生成增加，CKD 中最高。流式细胞术显示 CLK 和 CKD 分别在 S 期和 G2/M 期出现细胞周期阻滞，与细胞增殖和生长测定一致，分别显示 CLK 和 CKD 中细胞生长延迟和倍增时间延长的轻度和显著程度。细胞死亡定量也显示 CLK 和 CKD 中细胞死亡程度增加。
总之，长期 K 缺乏增加了肾小管细胞内的 ATP、细胞周期阻滞和细胞死亡，这可能是低钾血症性肾病发展的机制。
