Yurinskaya Valentina E, Vereninov Igor A, Vereninov Alexey A
Laboratory of Cell Physiology, Institute of Cytology, Russian Academy of Sciences, St-Petersburg, Russia.
Peter the Great St-Petersburg Polytechnic University, St-Petersburg, Russia.
Front Cell Dev Biol. 2020 Nov 6;8:591872. doi: 10.3389/fcell.2020.591872. eCollection 2020.
Fluxes of monovalent ions through the multiple pathways of the plasma membrane are highly interdependent, and their assessment by direct measurement is difficult or even impossible. Computation of the entire flux balance helps to identify partial flows and study the functional expression of individual transporters. Our previous computation of unidirectional fluxes in real cells ignored the ubiquitous cotransporters NKCC and KCC. Here, we present an analysis of the entire balance of unidirectional Na, K, and Cl fluxes through the plasma membrane in human lymphoid U937 cells, taking into account not only the Na/K pump and electroconductive channels but all major types of cotransporters NC, NKCC, and KCC. Our calculations use flux equations based on the fundamental principles of macroscopic electroneutrality of the system, water balance, and the generally accepted thermodynamic dependence of ion fluxes on the driving force, and they do not depend on hypotheses about the molecular structure of the channel and transporters. A complete list of the major inward and outward Na, K, and Cl fluxes is obtained for human lymphoid U937 cells at rest and during changes in the ion and water balance for the first 4 h of staurosporine-induced apoptosis. It is shown how the problem of the inevitable multiplicity of solutions to the flux equations, which arises with an increase in the number of ion pathways, can be solved in real cases by analyzing the ratio of ouabain-sensitive and ouabain-resistant parts of K (Rb) influx (OSOR) and using additional experimental data on the effects of specific inhibitors. It is found that dynamics of changes in the membrane channels and transporters underlying apoptotic changes in the content of ions and water in cells, calculated without taking into account the KCC and NKCC cotransporters, differs only in details from that calculated for cells with KCC and NKCC. The developed approach to the assessment of unidirectional fluxes may be useful for understanding functional expression of ion channels and transporters in other cells under various conditions. Attached software allows reproduction of all calculated data under presented conditions and to study the effects of the condition variation.
单价离子通过质膜多种途径的通量高度相互依赖，通过直接测量对其进行评估困难甚至不可能。计算整个通量平衡有助于识别部分流并研究单个转运蛋白的功能表达。我们之前对真实细胞中单向通量的计算忽略了普遍存在的协同转运蛋白NKCC和KCC。在此，我们对人类淋巴U937细胞中通过质膜的单向钠、钾和氯通量的整体平衡进行了分析，不仅考虑了钠/钾泵和导电通道，还考虑了所有主要类型的协同转运蛋白NC、NKCC和KCC。我们的计算使用基于系统宏观电中性、水平衡以及离子通量对驱动力的普遍接受的热力学依赖性的基本原理的通量方程，并且不依赖于关于通道和转运蛋白分子结构的假设。获得了人类淋巴U937细胞在静息状态下以及在星形孢菌素诱导凋亡的前4小时离子和水平衡变化期间主要内向和外向钠、钾和氯通量的完整列表。展示了随着离子途径数量增加而出现的通量方程必然存在多种解的问题如何在实际情况中通过分析钾（铷）流入的哇巴因敏感和哇巴因抗性部分的比率（OSOR）并使用关于特定抑制剂作用的额外实验数据来解决。发现未考虑KCC和NKCC协同转运蛋白计算的细胞中离子和水含量凋亡变化背后的膜通道和转运蛋白的变化动态与考虑KCC和NKCC的细胞计算的变化动态仅在细节上有所不同。所开发的评估单向通量的方法可能有助于理解各种条件下其他细胞中离子通道和转运蛋白的功能表达。随附的软件允许在给定条件下重现所有计算数据并研究条件变化的影响。
