HO-1通过BV-BVR-TJ途径修复皮肤屏障在特应性皮炎中的作用研究

Oxidative stress is an important factor in skin barrier dysfunction and immune abnormalities in atopic dermatitis (AD). Heme oxygenase 1 (HO-1) and its product bilverdin (BV) are important endogenous antioxidants in the body, with strong antioxidant and anti-inflammatory cytokines. The study found that the application of HO-1 can improve the AD lesions, but the mechanism is unknown. Our previous results showed that exogenous BV inhibits UV-induced inflammation and repairs the skin barrier by activating BV reductase (BVR). Whether HO-1 and its product BV have a protective effect in AD by regulating the expression of skin barrier-associated proteins or by inhibiting the immune inflammatory reaction, or both, is an issue to be elucidated in this study. This study intends to analyze the metabolic levels of HO-1 and BV in AD patients and their relationship with the severity of AD ；To determine the protective effect of exogenous HO-1 and BV on AD through in vivo experiments； And by using biological techniques including gene knockout through cell experiments, to elucidate the molecular mechanisms underlying the HO-1-BV-BVR pathway affecting the expression of TJ protein and inflammatory cytokines through the regulation of key pathways TLR4/NF-KB and PI3K/Art. This study first explores the impact of HO-1-BV-BVR on AD, and lays a theoretical foundation for exploring new therapeutic targets.

氧化应激是引起特应性皮炎（AD）皮肤屏障功能障碍及免疫异常的重要因素。HO-1及其酶促产物胆绿素（BV）具有强大的抗氧化及抗炎等细胞保护作用。应用外源性HO-1能减轻AD皮损但机制不清。我们的前期结果表明外源性BV通过激活BV还原酶（BVR）抑制紫外线引起的炎症并修复皮肤屏障。HO-1通过BV-BVR途径是调控TJ蛋白表达，修复皮肤屏障，还是抑制炎症反应而在AD中具有保护作用，抑或兼而有之是本研究亟待阐明的问题。本研究拟首先分析内源性HO-1及BV在AD患者中的代谢水平及它们与AD严重程度的关系；其次通过动物实验，利用基因敲除等技术，明确外源性HO-1及BV对AD的保护作用；通过细胞实验，阐明HO-1-BV-BVR途径通过调控TLR4/NF-KB及PI3K/Art通路而影响TJ蛋白及炎症因子表达的分子机制。本研究首次探讨HO-1-BV-BVR对AD的影响，为探索新的治疗靶点奠定理论基础。

目的：皮肤屏障功能障碍和Th2型炎症是特应性皮炎（AD）的关键特征。研究表明NRf2/HO-1通路的激活是有效治疗AD的主要机制之一。然而，血红素加氧酶-1（HO-1）在AD中的作用机制尚不清楚。本研究旨在探讨AD患者中HO-1、氧化应激和炎症之间的关系，并进一步探讨AD的风险基因。同时明确HO-1及其代谢产物BV对AD小鼠和角质形成细胞氧化应激模型的治疗作用及潜在机制。..方法：通过ELISA及RT-PCR检测AD患者和对照组外周血中HO-1、氧化应激及炎症因子的水平。应用氧化应激PCR-array检测AD患者和对照组中的差异表达基因。应用孟德尔随机化（MR）方法分析AD的风险基因。建立H2O2诱导的人角质形成细胞氧化损伤模型及MC903诱导的AD小鼠模型，通过RT-PCR，Western Blotting及免疫荧光等方法，探讨HO-1及BV对紧密连接蛋白Claudin-1和ZO-1及炎症因子的表达及调控机制。..结果：AD患者的HO-1、氧化应激及炎症水平明显高于对照组，AD患者抗氧化基因、ROS代谢基因及氧转运体的表达水平明显降低，并且氧化应激水平与炎症呈正相关。另外，AD患者omega-3脂肪酸水平可能与AD风险降低相关。在细胞水平上，HO-1显著增加SOD活性，降低ROS和MDA水平，并显著抑制H2O2诱导的HaCaT细胞中HMGB1、IL-4和NF-B等炎症因子的表达。在动物水平上，HO-1及BV显著减轻MC903诱导的AD小鼠的临床症状，HO-1通过上调Claudin-1和ZO-1的表达来修复皮肤屏障功能，并通过减少炎症细胞的浸润和炎症细胞因子的表达改善AD皮肤炎症。体内外研究进一步证实了低剂量的HO-1通过抑制磷酸化STAT1/3信号通路发挥治疗作用。..结论：氧化损伤和抗氧化防御能力的降低可能参与了AD的病理生理过程。HO-1治疗对小鼠AD样皮损和H2O2诱导的人角质形成细胞氧化应激损伤具有显著的保护作用，提示HO-1有望成为一种新的AD治疗候选药物。另外，补充omega-3脂肪酸或丰富饮食可能有助于预防AD的发生。