收稿日期: 2023-04-14
网络出版日期: 2023-10-10
基金资助
国家自然科学基金地区基金(82160365);贵州医科大学附属医院 2021 年度院级临床研究课题(2021-GMHCT-003)
Regulatory effect of LPAR1 gene on tissue factor and plasminogen activator inhibitor-1 in LPSstimulated type II alveolar epithelial cells
Received date: 2023-04-14
Online published: 2023-10-10
目的 了解脂多糖（LPS）刺激下Ⅱ型肺泡上皮细胞（AECⅡ）中差异表达基因溶血磷脂酸受体1（LPAR1）的动态表达特点，以及该基因对LPS刺激下组织因子（TF）和纤溶酶原激活物抑制剂1（PAI-1）表达的调节作用。 方法 对LPS损伤后的RLE-6TN细胞进行RNA-seq测序，通过生物信息学分析，以NF-κB信号通路为核心，找到该信号通路富集到的的差异表达基因LPAR1。使用LPS刺激处于生长对数期的RLE-6TN细胞6、12、24、48及72 h，检测细胞中LPAR1基因的动态表达水平。在此基础上通过慢病毒转染敲低细胞中LPAR1基因的表达，观察该基因对细胞中TF和PAI-1表达的调节作用。 结果 RLE-6TN细胞中LPAR1的表达在刺激6 h后开始升高，24 h时候达到顶峰，之后逐渐降低；利用慢病毒转染技术，成功低敲了细胞中LPAR1基因水平。LPS刺激24 h后，RLE-6TN细胞中TF和PAI-1的mRNA和蛋白的表达水平与正常对照组比较显著增加，差异有统计学意义（P < 0.05）；低敲LPAR1基因后，TF和PAI-1的表达水平显著降低（P < 0.05），接近正常对照水平（P > 0.05）。 结论 LPAR1基因对LPS刺激下RLE-6TN细胞TF和PAI-1的表达具有调节作用。该基因有望成为纠正ARDS肺泡内纤维蛋白沉积的新靶标。
关键词： 溶血磷脂酸受体 1; Ⅱ型肺泡上皮细胞; 急性呼吸窘迫综合征; 组织因子; 纤溶酶原激活物抑制剂1
刘颖 , 陈先俊 , 肖川 , 袁佳 , 李清 , 李璐 , 杨金凤 , 沈锋 . LPAR1基因对脂多糖刺激下Ⅱ型肺泡上皮细胞中组织因子和纤溶酶原激活物抑制剂1表达的影响[J]. 实用医学杂志, 2023 , 39(18) : 2323 -2329 . DOI: 10.3969/j.issn.1006-5725.2023.18.007
Objective To investigate the dynamic expression characteristics of lysophosphatidic acid receptor 1 （LPAR1）， a differentially expressed gene， in type Ⅱ alveolar epithelial cells （AEC Ⅱ） stimulated by lipopolysaccharide （LPS）， and the role of the gene in the regulation of tissue factor （TF） and plasminogen activator inhibitor-1 （PAI-1）. Methods RNA-seq sequencing was performed on RLE-6TN cells after injured by LPS， and then using bioinformatics analysis， identify the differentially expressed gene LPAR1 in the NF-κB signaling pathway. LPAR1 gene was dynamically expressed in RLE-6TN cells at 6， 12， 24， 48 and 72 h after LPS stimulation. On this basis， the expression of LPAR1 gene was knocked down by lentiviral transfection to observe the effect of the gene on the expression of TF and PAI-1. Results The expression of LPAR1 in RLE-6TN cells started to increase after 6 h of stimulation， peaked at 24 h， and then gradually decreased. LPAR1 gene levels were successfully knocked down through lentiviral transfection. After 24 hours of LPS stimulation， the expression levels of TF and PAI-1 mRNA and protein in the cells increased significantly compared with normal controls， with statistical significance （P < 0.05）， and then after knocking down LPAR1 gene， the expression levels of TF and PAI-1 decreased significantly （P < 0.05） and approached normal control values（P > 0.05）. Conclusion The LPAR1 gene has a regulatory effect on TF and PAI-1 in LPS-stimulated RLE-6TN cells. The gene is expected to be a new target for correcting fibrin deposition in the alveoli of ARDS.
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