强心苷对LLC-PK1和MDCK细胞中钠泵表达及功能的影响。

Effects of cardiac glycosides on sodium pump expression and function in LLC-PK1 and MDCK cells.

作者信息

Liu Jiang, Periyasamy Sankaridrug M, Gunning William, Fedorova Olga V, Bagrov Alexei Y, Malhotra Deepak, Xie Zijian, Shapiro Joseph I

机构信息

Department of Medicine, Medical College of Ohio, Toledo, Ohio 43614, USA.

出版信息

Kidney Int. 2002 Dec;62(6):2118-25. doi: 10.1046/j.1523-1755.2002.00672.x.

DOI:10.1046/j.1523-1755.2002.00672.x

PMID:12427136

Abstract

BACKGROUND

The decreases in proximal tubule sodium reabsorption seen with chronic renal failure and volume expansion have been ascribed to circulating digitalis-like substances (DLS). However, the circulating concentrations of DLS do not acutely inhibit the sodium pump to a degree consistent with the observed changes in proximal tubule sodium reabsorption.

METHODS

We examined how cell lines that simulated proximal (LLC-PK1) and distal tubule (MDCK) cells responded to acute (30 min) and long-term (up to 12 hours) Na+,K+-ATPase inhibition with DLS.

RESULTS

In LLC-PK1, but not MDCK cells, low concentrations of ouabain decreased 86Rb uptake profoundly in a time and dose dependent manner. In LLC-PK1 cells grown to confluence, transcellular 22Na flux was markedly reduced in concert with the decreases in 86Rb uptake. Similar findings were observed with marinobufagenin (MBG) and deproteinated extract of serum derived from patients with chronic renal failure. However, inhibition of the Na+,K+-ATPase with low extracellular potassium concentrations did not produce any of these effects. Western and Northern blots detected no change in alpha1 Na+,K+-ATPase protein and message RNA, respectively, in LLC-PK1 cells treated with ouabain for 12 hours. However, the decrease in enzymatic activity of Na+,K+-ATPase of these cells was comparable to observed decreases in 86Rb uptake. Differential centrifugation as well as biotinylation experiments demonstrated a shift of the Na+,K+-ATPase from the plasmalemma with prolonged ouabain treatment.

CONCLUSIONS

The results show that binding of cardiac glycosides by proximal (but not distal) tubular cells results in internalization of Na+,K+-ATPase with the net effect to amplify inhibition of the Na+,K+-ATPase. As the circulating concentrations of DLS increase with chronic renal failure and volume expansion, we suggest that this phenomenon explains some of the decreased sodium reabsorption by the proximal tubule seen in these conditions.

摘要

背景

慢性肾衰竭和容量扩张时近端小管钠重吸收的减少被归因于循环中的类洋地黄物质（DLS）。然而，DLS的循环浓度并不能急性抑制钠泵至与近端小管钠重吸收所观察到的变化相一致的程度。

方法

我们研究了模拟近端（LLC-PK1）和远端小管（MDCK）细胞的细胞系对DLS急性（30分钟）和长期（长达12小时）抑制Na⁺,K⁺-ATP酶的反应。

结果

在LLC-PK1细胞而非MDCK细胞中，低浓度哇巴因以时间和剂量依赖的方式显著降低⁸⁶Rb摄取。在生长至汇合的LLC-PK1细胞中，跨细胞²²Na通量与⁸⁶Rb摄取的减少一致地显著降低。用海蟾蜍毒配基（MBG）和慢性肾衰竭患者血清的脱蛋白提取物也观察到类似结果。然而，用低细胞外钾浓度抑制Na⁺,K⁺-ATP酶并未产生这些效应中的任何一种。蛋白质免疫印迹法和Northern印迹法分别检测到用哇巴因处理12小时的LLC-PK1细胞中α1 Na⁺,K⁺-ATP酶蛋白和信使RNA无变化。然而，这些细胞的Na⁺,K⁺-ATP酶活性的降低与观察到的⁸⁶Rb摄取的减少相当。差速离心以及生物素化实验表明，随着哇巴因处理时间延长，Na⁺,K⁺-ATP酶从质膜发生移位。