CYP450表氧化酶途径作为治疗哮喘气道重塑新靶向的研究

There are no effective therapeutic drugs for airway remodeling which characteristic is airway epithelial cells and smooth muscle cells proliferation leading to airway luminal stenosis. Our previous studies found that active metabolites of cytochrome P-450 epoxygenase (CYP450), epoxyeicosatrienoic acids (EETs) can inhibit cigarette-induced expression of CYP2C and CYP2J in the lung epithelial cells and endothelial cells, and reduce the release of inflammatory and growth factors. Oral soluble epoxide hydrolase (sEH) inhibitors AUDA or ZDHXB-101(a novel EET derivative was synthesized by our lab) can inhibit ovalbumine-induced the expression of airway remodeling biomarkers as well as reduce bleomycin-induced pulmonary fibrosis in mice. But whether the lung tissues of asthmatic models in mice abnormally express CYP450 epoxygenase and sEH, and the pathways of CYP450 epoxygenase metabolites EETs and sEH inhibitors as the therapeutic targets of airway remodeling have not been reported. In the project, we will collect the lung tissue samples from asthmatic models in mice, and detect and analysis the differences of the airway remodeling biomarkers, CYP450 epoxygenase genes (CYP2C, CYP2J) and sEH after exposure of the antigen challenge. We will study the roles and the signaling pathways of CYP450 epoxygenase metabolites EETs, AUDA and ZDHXB-101 on remodeling markers CYP2C, CYP2J and sEH in the antigen exposed or growth factors TGF-β1 to the primary epithelial cells and smooth muscle cells. Furthermore, we will further research that the effects and mechanism of EETs, AUDA and ZDHXB-101 in the airway remodeling of animal models. These results will provide evidences which the CYP450 epoxygenase pathway as a novel target for therapy of asthmatic airway remodeling.

哮喘气道重塑以气道上皮和平滑肌等细胞增生导致气道管腔狭窄为特征，目前尚无有效治疗药物。我们的前期研究发现细胞色素P450（CYP）表氧化酶代谢物二十碳三烯酸类（EETs）能抑制香烟诱导肺上皮细胞和氧化低密度脂蛋白诱导的内皮细胞CYP2J和CYP2C表达，减少炎性和纤维化因子的释放；预试验发现新合成的能升高EETs水平的小分子化合物ZDHXB-101和可溶性环氧水解酶(sEH)抑制剂AUDA能抑制博来霉素诱导的肺纤维化和减少卵白蛋白诱导的小鼠哮喘模型气道重塑标志物的表达从而减轻气道重塑。至今尚无哮喘动物模型是否异常表达CYP450表氧化酶和sEH的报道，本项目将测定动物模型肺组织EETs和CYP2J、CYP2C表达；采用分离和培养原代上皮细胞和平滑肌细胞以及建立动物模型，研究ZDHXB-101和sEH抑制剂抗气道重塑的作用和机制，以确立CYP450表氧化酶途径能否作为治疗哮喘气道重塑新靶向。

哮喘气道重塑以气道上皮和平滑肌等细胞增生导致气道管腔狭窄为特征，目前尚无有效治疗药物。我们的前期研究发现细胞色素P450（CYP）表氧化酶代谢物二十碳三烯酸类（EETs）能抑制香烟诱导肺上皮细胞和氧化低密度脂蛋白诱导的内皮细胞CYP2J和CYP2C表达，减少炎性和纤维化因子的释放；预试验发现新合成的能升高EETs水平的小分子化合物ZDHXB-101和可溶性环氧水解酶(sEH)抑制剂AUDA能抑制博来霉素诱导的肺纤维化和减少卵白蛋白诱导的小鼠哮喘模型气道重塑标志物的表达从而减轻气道重塑。而至今尚无哮喘动物模型是否异常表达CYP450表氧化酶和sEH的报道。本项目通过sEH抑制剂AUDA以及EETs的衍生物ZDHXB-101对抗原暴露12周的小鼠哮喘模型气道重塑和气道高反应性进行研究，结果表明，1）ZDHXB-101和AUDA能显著抑制小鼠肺组织sEH的表达，增加CYP2J2和14,15-EET的表达水平，2）抑制乙酰甲胆碱诱导的气道高反应性，明显改善小鼠肺功能，3）显著抑制肺组织炎症浸润，减少炎症因子白介素（IL）-13，IL-17和金属蛋白MMP-9 mRNA和蛋白的表达，4）减轻重塑模型小鼠肺纤维化病变，减少胶原沉积和杯状细胞增生，5）明显减少肺纤维化相关因子α肌动蛋白（α-SMA），S100A4，N-cadherin，Twist和MMP-9的表达。另外，在细胞方面，ZDHXB-101能有效抑制TGF-β1诱导的支气管上皮细胞（16HBE）炎症因子（TNF-α，IL-17）和纤维化因子（MMP-9，MMP-2，Vimentin和COL1A1）的表达升高，而这些指标的抑制依赖于EPHX-2表达的降低。ZDHXB-101和AUDA抗气道重塑可能通过Erk1/2，JNK MAPKs和STAT3信号通路起作用。上述研究结果表明CYP450表氧化酶途径可能是治疗哮喘气道重塑的潜在靶点。