去甲基化酶FTO调节IKKβm6A修饰在细颗粒物导致支气管哮喘气道屏障损伤机制研究

PM2.5与哮喘急性发作密切相关。前期在PM2.5暴露应激模型中，发现氧化应激增加，并发现m6A修饰酶FTO表达增加，同时RNA甲基化测序发现IKKβ3'-UTR序列 m6A修饰存在去甲基化。为进一步证实PM2.5是否通过氧化应激，影响FTO表达，从而导致IKKβm6A修饰去甲基化，导致气道屏障功能损伤。本项目拟以FTO调节IKKβm6A修饰为切入点在分子水平上，采用RNA免疫共沉淀、PCR和免疫印迹等方法分析PM2.5和氧化应激对FTO表达和IKKB m6A修饰影响，分析FTO过表达和低表达对IKKβm6A修饰的调节作用，以及IKKβ3'-UTR序列 m6A修饰对IKKβ表达影响，通过动物模型暴露PM2.5，分析PM2.5对IKKB m6A修饰的影响，研究其上下游调节机制，以阐明PM2.5对气道屏障功能损伤机制。由此发现潜在分子治疗靶点，并为相关靶点应用于临床转化奠定医学理论基础。

PM2.5 is closely related to acute asthma attack. In our study, we found the level of oxidative stress in cells was increased as well as the expression level of the demethylase FTO after PM2.5 exposure. Meanwhile, the level of m6A modification in 3'-UTR of IKKβ was decreased based on MeRIP-seq result. To further confirm whether PM2.5 destroyed airway barrier by reduction of IKKβm6A modification, which was regulated by FTO though oxidative stress, we will apply RNA immunoprecipitation, PCR, Westernblot to analyze the expression of FTO and IKKβm6A modification after expose to PM2.5, discover the regulation of OE-FTO or KO-FTO on IKKβm6A modification and the effect of m6A modification in 3'-UTR of IKKβ on its expression. Then analyzed the effects of PM2.5 on IKKB m6A modification and reveal the upstream and downstream regulation mechanism in animal model exposed to PM2.5 in order to elucidate the damage mechanism of PM2.5 on airway barrier. As a result, we can find potential molecular therapeutic targets and establish the theoretical evidence for the application of targets in clinical transformation.