Stuart-Tilley A K, Shmukler B E, Brown D, Alper S L
Molecular Medicine Unit, Beth Israel Deaconess Medical Center, Boston, Massachusetts 02215, USA.
J Am Soc Nephrol. 1998 Jun;9(6):946-59. doi: 10.1681/ASN.V96946.
In this study, an epitope-unmasking technique was used to immunolocalize AE2 anion exchanger polypeptide to basolateral plasma membranes of tubular epithelial cells in mouse kidney. Kidney AE2 immunostaining in mouse kidney was less prominent than in rat, consistent with the relative levels of AE2 mRNA and polypeptide in these two species. Glomeruli showed faint but consistent AE2 immunostaining, whereas proximal tubules were generally unstained. Macula densa epithelial cells displayed bright AE2 immunostaining, and cortical thick limbs were stained at a lower intensity. AE2 immunostaining was weak or absent in type B intercalated cells and principal cells of the cortical collecting duct, but increased in intensity in principal cells of the inner stripe of the outer medulla. AE2 staining in medullary thick limbs was also of greater intensity than in cortical thick limbs. AE2 staining was strong and uniform in the epithelial cells of the inner medullary collecting duct, and in epithelial cells of the papillary surface, the ureter, and the urinary bladder. Extratubular and epithelial cells of the inner medulla also showed punctate intracellular AE2 staining in a Golgi-like distribution that, in contrast to cell surface staining, was sodium dodecyl sulfate-sensitive. Golgi localization of AE2 epitope was confirmed by immunoperoxidase electron microscopy. Reverse transcription-PCR analysis of mouse kidney RNA detected AE2a, AE2b, and an AE2c2 transcript, but an AE2c1 transcript was absent. Unlike in rat, the mouse AE2c2 mRNA splice variant encoded a polypeptide with a novel predicted N-terminal amino acid sequence.
在本研究中，采用表位暴露技术将AE2阴离子交换蛋白多肽免疫定位到小鼠肾脏肾小管上皮细胞的基底外侧质膜上。小鼠肾脏中AE2的免疫染色不如大鼠明显，这与这两个物种中AE2 mRNA和多肽的相对水平一致。肾小球显示出微弱但一致的AE2免疫染色，而近端小管通常未染色。致密斑上皮细胞呈现明亮的AE2免疫染色，皮质厚升支的染色强度较低。B型闰细胞和皮质集合管主细胞中的AE2免疫染色较弱或缺失，但在外髓质内带主细胞中强度增加。髓质厚升支中的AE2染色强度也高于皮质厚升支。内髓质集合管的上皮细胞、乳头表面、输尿管和膀胱的上皮细胞中AE2染色强烈且均匀。内髓质的管外细胞和上皮细胞还显示出点状的细胞内AE2染色，呈高尔基体样分布，与细胞表面染色不同，这种染色对十二烷基硫酸钠敏感。通过免疫过氧化物酶电子显微镜证实了AE2表位的高尔基体定位。对小鼠肾脏RNA进行逆转录-聚合酶链反应分析检测到AE2a、AE2b和一个AE2c2转录本，但未检测到AE2c1转录本。与大鼠不同，小鼠AE2c2 mRNA剪接变体编码一种具有新预测N端氨基酸序列的多肽。
