HuR/CEBPD调控PTX3表达机制及其在肺曲霉病中的免疫保护作用研究

The mortality of invasive pulmonary aspergillosis (IPA) is high. Aspergillus pattern recognition receptor (PRR) and macrophage function are associated with incidence of IPA. Our previous study showed that the genetic polymorphism of pentraxin-3 (PTX3), an aspergillus PRR, is closely related to the IPA susceptibility. Our preliminary data suggested that the expression of CCAAT enhancer binding protein (CEBPD) and PTX3 in macrophage increased upon the stimulation of aspergillus. After knockdown of CEBPD gene, the expression of PTX3 decreased and the macrophage phagocytosis of aspergillus is suppressed. Moreover, CEBPD can be modulated by RNA-binding protein HuR upon the stimulation of aspergillus, and directly bind to the PTX3 promoter. With these the findings and evidence, we propose that HuR upregulates the expression of CEBPD with the stimulation of aspergillus. Then, CEBPD binds to PTX3 promoter and increase the expression of PTX3. Finally, PTX3 enhances the phagocytosis of macrophage and protect the host from aspergillus infection. To verify our hypothesis, we will construct CEBPD-/+ and HuR -/+ macrophages and the model of aspergillosis in vitro. Then, we will confirm the effects and uncover the mechanism of HuR/CEBPD on the expression of PTX3 and phagocytosis of macrophage. We expect that this study will provide a new method to prevent and treat IPA.

侵袭性肺曲霉病（IPA）病死率高。曲霉模式识别受体（PRR）和巨噬细胞功能与IPA发病相关。我们前期研究发现，正五聚蛋白3（PTX3）作为一种曲霉PRR，其基因多态性与曲霉易感性密切相关。预实验结果显示，曲霉刺激巨噬细胞后，其转录增强子结合蛋白CEBPD和PTX3水平显著增高；沉默CEBPD基因后，PTX3减少，同时巨噬细胞对曲霉的吞噬能力减弱；而CEBPD受RNA结合蛋白HuR调节，可直接结合PTX3的启动子区域。因此，我们提出假说：曲霉感染时HuR可上调CEBPD表达，而CEBPD可与PTX3启动子结合，促进PTX3的转录表达，从而增强巨噬细胞对曲霉的识别吞噬。为此我们拟在巨噬细胞中沉默或高表达CEBPD和HuR，建立曲霉感染模型，揭示HuR/CEBPD调控PTX3表达和巨噬细胞吞噬曲霉孢子功能的机制，为防治IPA提供新策略。

侵袭性肺曲霉病（IPA）病死率高，治疗手段缺乏。正五聚蛋白3（PTX3）作为一种曲霉模式识别受体，能够识别曲霉孢子并促进巨噬细胞吞噬曲霉孢子。虽然PTX3参与了曲霉感染的免疫保护，但其表达调控机制研究甚少。在本研究中，我们发现肺曲霉病患者肺组织中CEBPD、PTX3的表达较正常肺组织明显增加，同样的体内侵袭性肺曲霉病小鼠模型和体外烟曲霉感染的巨噬细胞模型中CEBPD、PTX3的表达均高于正常对照组。接下来我们利用人单核细胞株THP-1诱导的巨噬细胞，通过siRNA和慢病毒构建CEBPD、HuR敲降/过表达细胞，再用烟曲霉孢子刺激细胞，观察PTX3以及巨噬细胞吞噬率的改变。在烟曲霉孢子刺激时，HuR由细胞核转移到细胞质，并稳定CEBPD mRNA，使得CEBPD表达增加，CEBPD能够与PTX3启动子区域结合进而导致PTX3表达增加，进而增强巨噬细胞对烟曲霉孢子的吞噬。由此我们得出课题研究结论：HuR-CEBPD通路在调控IPA重要模式识别受体PTX3的表达中起重要作用，从而影响巨噬细胞吞噬作用和IPA的发生发展。这一研究发现，为临床深入认知曲霉感染中的免疫保护机制提供了新的思路和帮助，为临床防治IPA提供新的证据和治疗靶点。