甘油激酶5对气道炎症和上皮损伤修复的调控效应及其机制

Currently, there is still a lack of efficient treatments for Chronic Obstructive Pulmonary Disease (COPD). Reducing airway inflammation and.maintaining epithelial barrier function are critical in the prevention and treatment of COPD. Glycerol kinase deficiency-induced elevation of free fat acid plays important roles in chronic inflammation and metabolic disorders. Our previous studies found that,glycerol kinase 5 (GK5) is highly expressed on airway epithelial cells; cigarette smoke extract compensatorily upregulated the expression of GK5 and aquaglyceroporin 3 (AQP3) and AQP9 48hours after epithelial cell injury; GK5 siRNA significantly decreased the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), AQP3, and AQP9 on airway epithelial cells, and reduced epithelial cell migration. Therefore, we hypothesize that GK5 plays an important role in reducing airway inflammation and maintaining airway epithelial barrier function. We propose to study the effects of GK5 on the airway inflammation and the repair of epithelial cells after injury using both in vivo and in vitro models; the effects and mechanism of GK5 knockdown and overexpression on the gene and protein expression of aquaporins and aquaglyceroporins; the effects and mechanism of AQP9 on wound healing of airway epithelial cells. Our studies will provide potential pharmacological targets for the treatment of COPD.

目前对慢阻肺仍然缺乏针对呼吸道病理改变有效的治疗措施。有效降低气道炎症及维持上皮细胞屏障稳定性对于防治慢阻肺至关重要。甘油激酶参与游离脂肪酸代谢，在慢性炎症和代谢疾病中具有重要作用，但其在慢阻肺病变进展中的作用未明。我们前期研究发现，甘油激酶5（GK5）在气道上皮细胞表达；动物及细胞实验表明，吸烟代偿性上调GK5及水甘油通道AQP3和AQP9表达；GK5基因敲除显著降低上皮细胞内GAPDH、AQP3和AQP9表达，并伴随细胞迁移能力下降。因此，我们推测GK5在气道炎症及上皮损伤修复中具有重要作用。本项目拟采用吸烟大鼠慢阻肺及烟熏细胞损伤模型，明确GK5对气道炎症和上皮损伤修复的影响；GK5对气道上皮水和甘油通道蛋白表达的影响，水甘油通道AQP9对气道上皮损伤修复的影响及机制，为慢阻肺的治疗提供新的思路。

目前对慢阻肺仍然缺乏针对呼吸道病理改变有效的治疗措施。有效降低气道炎症及维持上皮细胞屏障稳定性对于防治慢阻肺至关重要。甘油激酶参与游离脂肪酸代谢，在慢性炎症和代谢疾病中具有重要作用，但其在慢阻肺病变进展中的作用未明。本项目通过Gk5基因敲除小鼠和上皮细胞GK5基因敲减/过表达，阐明了Gk5通过水甘油通道和脂质代谢途径，加重烟熏引起的气道损伤和炎症反应。. 临床研究：COPD稳定期患者血浆外泌体中GK5 mRNA表达显著高于健康吸烟人群。体内研究：烟熏上调Gk5在气道上皮表达；通过构建Gk5基因敲除小鼠，发现连续烟熏30天后，Gk5-/-烟熏组小鼠状态变差，出现死亡现象，Gk5-/-小鼠气道损伤和炎症反应加重，肺组织中凋亡相关蛋白表达增加。体外研究：烟雾提取物上调GK5在气道上皮表达；在气道上皮细胞中GK5基因敲减降低细胞增殖和迁移；降低水甘油通道AQP3和AQP9基因及蛋白表达；降低脂滴合成相关信号分子SREBP1和SCD1基因及蛋白表达；GK5基因过表达诱导脂滴，显著增加细胞内SREBP1、ACSL1、和PLIN3等蛋白表达。. GK5基因敲减显著降低AQP3基因及蛋白表达，AQP3在烟熏小鼠肺组织中表达上调，小鼠AQP3基因敲除加重烟熏引起的气道上皮损伤和炎症反应；降低肺上皮细胞Ki67和Scd1表达。烟雾提取物上调AQP3在气道上皮表达；上皮细胞AQP3基因敲减显著降低细胞内SCD1、survivin、和PLIN2的基因表达。. GK5和AQP3基因敲减都显著降低SCD1基因及蛋白表达，提示SCD1是GK5和AQP3的下游信号分子。烟熏小鼠肺内Scd1表达水平升高，抑制Scd1加重烟熏引起的小鼠体重和肝脏系数下降、气道损伤、和炎症反应等。烟雾提取物上调SCD1在气道上皮表达；上皮细胞内SCD1基因敲减显著降低细胞增殖，降低细胞内survivin和PLIN2的基因表达。. 慢阻肺患者肺癌发病率高，肺癌靶向耐药患者血浆外泌体中GK5 mRNA表达显著高于吉非替尼敏感患者，在吉非替尼耐药细胞中GK5表达上调，GK5基因敲减通过SREBP1/SCD1信号通路诱导线粒体损伤、半胱天冬酶活化、细胞周期停滞、和细胞凋亡，表明GK5通过SREBP1/SCD1信号传导途径介导吉非替尼耐药。

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