Differentiation of human pulmonary type II cells in vitro by glucocorticoid plus cAMP
Differentiation of human pulmonary type II cells in vitro by glucocorticoid plus cAMP
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DOI：
10.1152/ajplung.00127.2002
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发表时间：
2002-11-01
影响因子：
4.9
通讯作者：
Ballard, PL
中科院分区：
文献类型：
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作者：
Gonzales, LW;Guttentag, SH;Ballard, PL
产生肺表面活性物质的成熟Ⅱ型肺泡细胞对于适应宫外生活以及预防婴儿呼吸窘迫综合征至关重要。我们开发了一种新的体外模型，以进一步研究Ⅱ型细胞分化的调控。从人胎肺分离的上皮细胞在塑料培养皿中用无血清培养基培养。用地塞米松 + 环磷腺苷酸类似物和异丁基甲基黄嘌呤处理4天的细胞，其磷脂酰胆碱合成增加，二饱和磷脂酰胆碱种类的含量增加，所有表面活性物质蛋白大量增加并加工为成熟形式，且板层小体丰富。DNA微阵列分析鉴定出约3100个表达基因，包括经激素处理诱导2 - >100倍（约2.5%）或抑制2 - 18倍（约1.2%）的基因子集。在高度调控的基因中，大多数由地塞米松和环磷腺苷酸试剂以相加或协同的方式共同调控。通过这种初步微阵列分析鉴定出的受调控基因中，约90%此前未被认为对激素有反应。一个新鉴定的激素诱导基因是Nkx2.1（甲状腺转录因子 - 1），它在表面活性物质蛋白基因表达中起关键作用。我们的研究结果表明，在这个体外系统中，糖皮质激素 + 环磷腺苷酸对于早熟诱导功能性Ⅱ型细胞是充分且必要的，并且这些激素主要联合作用以调控特定基因子集的表达。
Mature alveolar type II cells that produce pulmonary surfactant are essential for adaptation to extrauterine life and prevention of infant respiratory distress syndrome. We have developed a new in vitro model to further investigate regulation of type II cell differentiation. Epithelial cells isolated from human fetal lung were cultured in serum-free medium on plastic. Cells treated with dexamethasone + cAMP analog and isobutylmethylxanthine for 4 days exhibited increased phosphatidylcholine synthesis and content of disaturated phosphatidylcholine species, manyfold increases in all surfactant proteins with processing to mature forms, and abundant lamellar bodies. DNA microarray analysis identified similar to3,100 expressed genes, including subsets of genes induced 2- to >100-fold (similar to2.5%) or repressed 2- to 18-fold (similar to1.2%) by hormone treatment. Of the highly regulated genes, most were coregulated in an additive or synergistic manner by dexamethasone and cAMP agents. Approximately 90% of the regulated genes identified by this initial microarray analysis have not been previously recognized as hormone responsive. One newly identified hormone-induced gene is Nkx2.1 (thyroid transcription factor-1), which has a critical role in surfactant protein gene expression. Our findings indicate that glucocorticoid + cAMP is sufficient and necessary for precocious induction of functional type II cells in this in vitro system and that these hormones act primarily in combination to regulate expression of a subset of specific genes.