Control of activity and expression of ion transporters in rat lung: Role of HIF and CREB in hypoxia and beta-adrenergic stimulation

ARDS、ALI、心血管疾病和肺泡缺氧引起肺水肿、肺泡缺氧，并通过降低转运蛋白的活性和表达损害肺泡液重吸收等保护机制。它还会引起炎症，从而进一步抑制重吸收。保护性机制如p2肾上腺素能（ß2AR）信号通路受损，通常通过增加离子转运体的表达和活性刺激肺泡重吸收。低氧对基因表达的影响是通过缺氧诱导因子（hypoxia-inducible factors， HIF）介导的，ß 2ar刺激通过CREB诱导基因表达。它们在离子转运体表达中的作用尚不清楚。由于这两个因素通过共激活蛋白p300和CBP相互作用，因此了解缺氧和p2ar刺激对肺泡重吸收和离子转运蛋白表达的影响之间的相互作用非常重要，这对治疗肺水肿具有重要的临床意义。在这个项目中，我们想测试假设，长时间的ß2AR刺激可以通过creb依赖性或非依赖性机制阻止缺氧诱导的肺泡离子转运蛋白活性和表达的抑制。将测试CREB和HIF的作用及其相互作用，以及长时间ß2AR刺激是否能阻止缺氧对肺泡运输的影响。我们将在体外对原代大鼠和人肺泡上皮细胞进行实验，并对长期服用ß 2ar激动剂特布他林后暴露于缺氧的大鼠进行体内实验，以特异性评估基因表达的作用。HIF和CREB的作用将通过基因沉默来研究。读出参数为离子转运体和ß 2ar信号的表达和活性。

ARDS, ALI, cardiovascular diseases, and alveolar hypoxia cause pulmonary edema, alveolar hypoxia, and impair protective mechanisms such as alveolar fluid reabsorption by decreasing activity and expression of transporters. It also induces inflammation, which further inhibits reabsorption. Protective mechanisms such as p2 adrenergic (ß2AR) signaling are impaired, which normally stimulates alveolar reabsorption by increasing expression and activity of ion transporters. Hypoxia-effects on gene expression are mediated by hypoxia-inducible factors (HIF), ß2AR-stimulation induces gene expression via CREB. Their role in the expression of ion transporters is unclear. Since both factors interact via co-activator proteins p300 and CBP, it is important to understand the interactions between effects of hypoxia and p2ARstimulation on alveolar reabsorption and expression of ion transporters, which is of great clinical relevance for treating pulmonary edema. In this project we want to test the hypotheses, that prolonged ß2AR stimulation can prevent the hypoxia-induced inhibition of activity and expression of alveolar ion transporters by CREB-dependent or -independent mechanisms. The roles of CREB and HIF and their interaction will be tested, and whether prolonged ß2AR stimulation prevents effects of hypoxia on alveolar transport. Experiments will be performed on primary rat and human alveolar epithelial cells invitro and on rats in-vivo exposed to hypoxia after treating with the ß2AR-agonist terbutaline for prolonged time to specifically evaluate the role of gene expression. The roles of HIF and CREB will be studied by gene silencing. Readout parameters are expression and activity of ion transporters and ß2AR-signaling.