Department of Medicine, National Jewish Health, Denver, CO, USA.
Department of Medicine, National Jewish Health, Denver, CO, USA; Department of Medicine, University of Colorado Denver, Aurora, CO, USA.
Biochem Biophys Res Commun. 2014 Oct 3;452(4):1091-7. doi: 10.1016/j.bbrc.2014.09.060. Epub 2014 Sep 22.
Alveolar type II (ATII) cells remain differentiated and express surfactant proteins when cultured at an air-liquid (A/L) interface. When cultured under submerged conditions, ATII cells dedifferentiate and change their gene expression profile. We have previously shown that gene expression under submerged conditions is regulated by hypoxia inducible factor (HIF) signaling due to focal hypoxia resulting from ATII cell metabolism. Herein, we sought to further define gene expression changes in ATII cells cultured under submerged conditions. We performed a genome wide microarray on RNA extracted from rat ATII cells cultured under submerged conditions for 24-48h after switching from an A/L interface. We found significant alterations in gene expression, including upregulation of the HIF target genes stanniocalcin-1 (STC1), tyrosine hydroxylase (Th), enolase (Eno) 2, and matrix metalloproteinase (MMP) 13, and we verified upregulation of these genes by RT-PCR. Because STC1, a highly evolutionarily conserved glycoprotein with anti-inflammatory, anti-apoptotic, anti-oxidant, and wound healing properties, is widely expressed in the lung, we further explored the potential functions of STC1 in the alveolar epithelium. We found that STC1 was induced by hypoxia and HIF in rat ATII cells, and this induction occurred rapidly and reversibly. We also showed that recombinant human STC1 (rhSTC1) enhanced cell motility with extended lamellipodia formation in alveolar epithelial cell (AEC) monolayers but did not inhibit the oxidative damage induced by LPS. We also confirmed that STC1 was upregulated by hypoxia and HIF in human lung epithelial cells. In this study, we have found that several HIF target genes including STC1 are upregulated in AECs by a submerged condition, that STC1 is regulated by hypoxia and HIF, that this regulation is rapidly and reversibly, and that STC1 enhances wound healing moderately in AEC monolayers. However, STC1 did not inhibit oxidative damage in rat AECs stimulated by LPS in vitro. Therefore, alterations in gene expression by ATII cells under submerged conditions including STC1 were largely induced by hypoxia and HIF, which may be relevant to our understanding of the pathogenesis of various lung diseases in which the alveolar epithelium is exposed to relative hypoxia.
肺泡 II 型 (ATII) 细胞在气液 (A/L) 界面培养时保持分化并表达表面活性剂蛋白。当在浸没条件下培养时,ATII 细胞去分化并改变其基因表达谱。我们之前已经表明,由于 ATII 细胞代谢导致的局部缺氧,浸没条件下的基因表达受缺氧诱导因子 (HIF) 信号调节。在此,我们试图进一步定义在浸没条件下培养的 ATII 细胞的基因表达变化。我们对从切换到 A/L 界面后 24-48 小时在浸没条件下培养的大鼠 ATII 细胞提取的 RNA 进行了全基因组微阵列分析。我们发现基因表达发生了显著变化,包括 HIF 靶基因 STC1、酪氨酸羟化酶 (Th)、烯醇酶 (Eno)2 和基质金属蛋白酶 (MMP)13 的上调,并且我们通过 RT-PCR 验证了这些基因的上调。因为 STC1 是一种高度进化保守的糖蛋白,具有抗炎、抗凋亡、抗氧化和伤口愈合特性,广泛表达于肺部,因此我们进一步探讨了 STC1 在肺泡上皮中的潜在功能。我们发现 STC1 在大鼠 ATII 细胞中由缺氧和 HIF 诱导,这种诱导发生迅速且可逆。我们还表明,重组人 STC1 (rhSTC1) 在肺泡上皮细胞 (AEC) 单层中增强细胞迁移并形成延伸的伪足,但不抑制 LPS 诱导的氧化损伤。我们还证实 STC1 在人肺上皮细胞中由缺氧和 HIF 上调。在这项研究中,我们发现包括 STC1 在内的几种 HIF 靶基因在 AEC 中由浸没条件上调,STC1 受缺氧和 HIF 调节,这种调节是快速且可逆的,并且 STC1 适度增强 AEC 单层中的伤口愈合。然而,STC1 并未抑制 LPS 刺激的大鼠 AEC 中的氧化损伤。因此,包括 STC1 在内的 ATII 细胞在浸没条件下的基因表达改变主要由缺氧和 HIF 诱导,这可能有助于我们理解肺泡上皮暴露于相对缺氧时各种肺部疾病的发病机制。