Characterization of mouse alveolar epithelial cell monolayers
Characterization of mouse alveolar epithelial cell monolayers
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DOI：
10.1152/ajplung.00021.2009
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发表时间：
2009-06-01
影响因子：
4.9
通讯作者：
Borok, Zea
中科院分区：
文献类型：
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作者：
DeMaio, Lucas;Tseng, Wanru;Borok, Zea
DeMaio L、Tseng W、Balverde Z、Alvarez JR、Kim K-J、Kelley DG、Senior RM、Crandall ED、Borok Z。小鼠肺泡上皮细胞单层的表征。 Am J Physiol Lung Cell Mol Physiol 296：L1051-L1058，2009。首次发表于 2009 年 3 月 27 日； doi: 10.1152/ajplung.00021.2009.-我们研究了细胞外基质对小鼠肺泡上皮细胞 (AEC) 单层 (MAECM) 运输特性的影响以及分离的小鼠 II 型肺泡上皮 (AT2) 细胞转分化为 I 型肺泡上皮 (AT1) 细胞样表型的影响。原代小鼠 AT2 细胞铺在层粘连蛋白 5 包被的聚碳酸酯滤膜上，形成单层细胞，培养 8 天后，跨上皮电阻 (R-T) 和等效短路电流 (I-EQ) 分别为 1.8 k Omega.cm(2) 和 5.3 mu A/cm(2)。阿米洛利 (10 μM)、哇巴因 (0.1 mM) 和匹莫齐特 (10 μM) 分别将 MAECM I-EQ 降低至其初始值的 40%、10% 和 65%。以任一顺序依次添加匹莫齐特和阿米洛利，都表明它们的抑制作用是相加的，这表明环核苷酸门控通道有助于阿米洛利不敏感的活性离子在 MAECM 中的转运。使用室测量短路条件下穿过 MAECM 的单向离子通量表明，从顶端到基底外侧方向的 Na+ 净吸收与根据观察到的短路电流计算的净离子通量相当：分别为 0.38 和 0.33 mu eq.cm(-2).h(-1)。在培养的第 1 天到第 9 天之间，AEC 表现出水通道蛋白 5 蛋白（一种 AT1 细胞标记）的表达增加，而前表面活性剂蛋白-C 蛋白（一种 AT2 细胞标记）的表达降低，这与向 AT1 细胞样表型的转变一致。这些结果表明，在层粘连蛋白 5 涂层的聚碳酸酯过滤器上生长的 AT1 细胞样 MAECM 表现出主动和被动运输特性，这可能反映了完整小鼠肺泡上皮的特性。该小鼠体外模型将加强对源自突变小鼠品系的 AEC 的研究，并有助于确定重要的结构-功能相关性。
DeMaio L, Tseng W, Balverde Z, Alvarez JR, Kim K-J, Kelley DG, Senior RM, Crandall ED, Borok Z. Characterization of mouse alveolar epithelial cell monolayers. Am J Physiol Lung Cell Mol Physiol 296: L1051-L1058, 2009. First published March 27, 2009; doi: 10.1152/ajplung.00021.2009.-We investigated the influence of extracellular matrix on transport properties of mouse alveolar epithelial cell (AEC) monolayers (MAECM) and transdifferentiation of isolated mouse alveolar epithelial type II (AT2) cells into an alveolar epithelial type I (AT1) cell-like phenotype. Primary mouse AT2 cells plated on laminin 5-coated polycarbonate filters formed monolayers with transepithelial resistance (R-T) and equivalent short-circuit current (I-EQ) of 1.8 k Omega.cm(2) and 5.3 mu A/cm(2), respectively, after 8 days in culture. Amiloride (10 mu M), ouabain (0.1 mM), and pimozide (10 mu M) decreased MAECM I-EQ to 40%, 10%, and 65% of its initial value, respectively. Sequential addition of pimozide and amiloride, in either order, revealed that their inhibitory effects are additive, suggesting that cyclic nucleotide-gated channels contribute to amiloride-insensitive active ion transport across MAECM. Ussing chamber measurements of unidirectional ion fluxes across MAECM under short-circuit conditions indicated that net absorption of Na+ in the apical-to-basolateral direction is comparable to net ion flux calculated from the observed short-circuit current: 0.38 and 0.33 mu eq.cm(-2).h(-1), respectively. Between days 1 and 9 in culture, AEC demonstrated increased expression of aquaporin-5 protein, an AT1 cell marker, and decreased expression of pro-surfactant protein-C protein, an AT2 cell marker, consistent with transition to an AT1 cell-like phenotype. These results demonstrate that AT1 cell-like MAECM grown on laminin 5-coated polycarbonate filters exhibit active and passive transport properties that likely reflect the properties of intact mouse alveolar epithelium. This mouse in vitro model will enhance the study of AEC derived from mutant strains of mice and help define important structure-function correlations.