Fejes-Tóth G, Rusvai E, Cleaveland E S, Náray-Fejes-Tóth A
Department of Physiology, Dartmouth Medical School, Lebanon, New Hampshire 03756, USA.
Am J Physiol. 1998 Mar;274(3):F596-601. doi: 10.1152/ajprenal.1998.274.3.F596.
AE2 mRNA and protein is expressed in several nephron segments, one of which is the cortical collecting duct (CCD). However, the distribution of AE2 among the different cell types of the CCD and the function of AE2 in the kidney are not known. The purpose of this study was to determine the distribution of AE2 mRNA among the three CCD cell types and to examine the effects of changes in acid/base balance on its expression. Following NH4Cl (acid) or NaHCO3 (base) loading of rabbits for approximately 18 h, CCD cells were isolated by immunodissection. AE2 mRNA levels were determined by RT-PCR and were normalized for beta-actin levels. We found that CCD cells express high levels of AE2 mRNA (approximately 500 copies/cell). AE2 mRNA levels were significantly higher in CCD cells originating from base-loaded than acid-loaded rabbits, with an average increase of 3.7 +/- 1.07-fold. The effect of pH on AE2 mRNA levels was also tested directly using primary cultures of CCD cells. CCD cells incubated in acidic media expressed significantly lower levels of AE2 mRNA than those in normal or alkaline media. Experiments with isolated principal cells, alpha-intercalated cells, and beta-intercalated cells (separated by fluorescence-activated cell sorting) demonstrated that AE2 mRNA levels are comparable in the three collecting duct cell subtypes and are similarly regulated by changes in acid/base balance. Based on these results, we conclude that adaptation to changes in extracellular H+ concentration is accompanied by opposite changes in AE2 mRNA expression. The observations that AE2 mRNA is not expressed in a cell-type-specific manner and that changes in acid/base balance have similar effects on each CCD cell subtype suggest that AE2 might serve a housekeeping function rather than being the apical anion exchanger of beta-intercalated cells.
AE2信使核糖核酸（mRNA）和蛋白在多个肾单位节段中表达，其中之一是皮质集合管（CCD）。然而，AE2在CCD不同细胞类型中的分布以及其在肾脏中的功能尚不清楚。本研究的目的是确定AE2 mRNA在CCD三种细胞类型中的分布，并研究酸碱平衡变化对其表达的影响。给兔子分别负荷氯化铵（酸）或碳酸氢钠（碱）约18小时后，通过免疫解剖分离出CCD细胞。通过逆转录聚合酶链反应（RT-PCR）测定AE2 mRNA水平，并根据β-肌动蛋白水平进行标准化。我们发现CCD细胞表达高水平的AE2 mRNA（约500拷贝/细胞）。来自碱负荷兔子的CCD细胞中AE2 mRNA水平显著高于酸负荷兔子，平均增加3.7±1.07倍。还直接使用CCD细胞原代培养物测试了pH对AE2 mRNA水平的影响。在酸性培养基中培养的CCD细胞表达的AE2 mRNA水平明显低于正常或碱性培养基中的细胞。对分离的主细胞、α-闰细胞和β-闰细胞（通过荧光激活细胞分选分离）进行的实验表明，AE2 mRNA水平在三种集合管细胞亚型中相当，并且受到酸碱平衡变化的类似调节。基于这些结果，我们得出结论，细胞外H⁺浓度变化的适应性伴随着AE2 mRNA表达的相反变化。AE2 mRNA不以细胞类型特异性方式表达以及酸碱平衡变化对每个CCD细胞亚型具有类似影响的观察结果表明，AE2可能发挥看家功能，而不是作为β-闰细胞的顶端阴离子交换体。
