Cultured human trabecular meshwork cells express aquaporin-1 water channels
Cultured human trabecular meshwork cells express aquaporin-1 water channels
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DOI：
10.3109/02713689508995815
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发表时间：
1995-12-01
影响因子：
2
通讯作者：
Regan, JW
中科院分区：
文献类型：
--
作者：
Stamer, WD;Seftor, REB;Regan, JW
水通道蛋白-1及其他相关蛋白的鉴定和表征为多种上皮组织通透性增强提供了分子解释。先前，我们记录了水通道蛋白-1在人眼中的分布，包括小梁网；房水的主要流出通道。本研究的目的是确定水通道蛋白-1是否可以在培养的人小梁网细胞中检测到。利用水通道蛋白-1的特异性引物，逆转录结合聚合酶链反应，从人小梁网细胞中制备出合适大小的总RNA产物。该产物的存在及其大小（298个碱基对）与这些细胞中水通道蛋白-1信息的存在一致。间接免疫荧光显微镜用亲和纯化的抗体对抗含有水通道蛋白-1的羧基尾的融合蛋白，显示出对质膜的特异性标记和免疫印迹，鉴定出M(r) 28000的条带，这与水通道蛋白-1的分子大小一致。水通道蛋白-1存在于人眼初级流出区主要的细胞类型——人小梁网细胞中，这表明水通道可能与水流体流出眼睛的运动有关。此外，水通道蛋白-1在人小梁网细胞培养物中的存在，为研究水通道蛋白-1的内源性表达及其在水流出调节中的可能作用提供了体外模型。
The identification and characterization of aquaporin-1 water channels and other related proteins has provided a molecular explanation for the enhanced permeability of a variety of epithelial tissues. Previously, we documented the distribution of aquaporin-1 in the human eye, which included the trabecular meshwork; the primary outflow channel for aqueous humor. The goal of this study was to determine if aquaporin-1 could be detected in cultures of human trabecular meshwork cells. Using primers specific for aquaporin-1, reverse transcription combined with polymerase chain reaction yielded a product of the appropriate size with total RNA prepared from the human trabecular meshwork cells. The presence of this product and its size (298 base pairs), is consistent with the presence of an aquaporin-1 message in these cells. Indirect immunofluorescence microscopy with affinity purified antibodies against a fusion protein containing the carboxy tail of aquaporin-1 showed specific labeling of the plasma membrane and immunoblotting, identified a band of M(r) 28,000 which agrees with the molecular size of aquaporin-1. The presence of aquaporin-1 in human trabecular meshwork cells, the predominant cell-type of the primary outflow region of the human eye, suggests that water channels may be involved with the movement of aqueous fluid out of the eye. In addition, the existence of aquaporin-1 on cultures of human trabecular meshwork cells provides an in vitro model to study the endogenous expression of aquaporin-1 and its possible role in the regulation of aqueous outflow.