Passive siRNA transfection method for gene knockdown in air-liquid interface airway epithelial cell cultures
Passive siRNA transfection method for gene knockdown in air-liquid interface airway epithelial cell cultures
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DOI：
10.1152/ajplung.00122.2021
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发表时间：
2021-07-01
影响因子：
4.9
通讯作者：
Chiarella, Sergio E.
中科院分区：
文献类型：
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作者：
Bartman, Colleen M.;Stelzig, Kimberly E.;Chiarella, Sergio E.
人支气管上皮细胞（HBE）在空气-液体界面（ALI）培养物中的分化概括了体内环境的器官型建模。尽管ALI培养物对于研究呼吸道上皮屏障是非常宝贵的，但是功能丧失研究受到经典转染方法中的潜在细胞毒性试剂、分化方案的长度和原代上皮细胞传代次数的限制。在这里，我们提出了一种简单的方法，小干扰RNA（siRNA）转染正常HBE（NHBE）在ALI文化，不需要潜在的细胞毒性转染试剂，并不改变生理学或形态NHBE在分化过程中的功效和使用。该转染方案为HBE ALI培养物的功能丧失研究引入了一种可重复且有效的方法，该培养物可用于模拟呼吸系统和气道疾病。
Differentiation of human bronchial epithelial cells (HBEs) in air-liquid interface (ALI) cultures recapitulates organotypic modeling of the in vivo environment. Although ALI cultures are invaluable for studying the respiratory epithelial barrier, loss-of-function studies are limited by potentially cytotoxic reagents in classical transfection methods, the length of the differentiation protocol, and the number of primary epithelial cell passages. Here, we present the efficacy and use of a simple method for small interfering RNA (siRNA) transfection of normal HBEs (NHBEs) in ALI cultures that does not require potentially cytotoxic transfection reagents and does not detrimentally alter the physiology or morphology of NHBEs during the differentiation process. This transfection protocol introduces a reproducible and efficient method for loss-of-function studies in HBE ALI cultures that can be lever-aged for modeling the respiratory system and airway diseases.