关键词: 肺动脉高压, 氧化应激, 转化生长因子β, 炎症, 机器学习, 基因表达, 病理机制, 靶向治疗

Abstract: BACKGROUND: Recent studies have shown that there is a correlation between oxidative stress and pulmonary hypertension, but the key mechanism needs to be further explored.
OBJECTIVE: To investigate the mechanism of transforming growth factor β related to oxidative stress in pulmonary hypertension.
METHODS: Three data sets (GSE117261, GSE53408 and GSE113439) were obtained and integrated from the gene expression comprehensive database established and maintained by the National Center for Biotechnology Information (NCBI). The differentially expressed genes related to pulmonary hypertension were screened. The core genes related to oxidative stress of pulmonary hypertension were identified by combining the gene sets related to oxidative stress. Machine learning algorithms are used to further screen and determine the most important candidate genes. Subsequently, an experimental model of cells and animals was constructed to verify their functions: (1) Rat pulmonary mesenchymal stem cells were divided into control group, activation group, transforming growth factor β agonist group, and transforming growth factor β inhibitor group. Rat pulmonary mesenchymal stem cells were stimulated by monocrotaline, and the transforming growth factor β agonist and inhibitor were given for 48 hours respectively. (2) SD rats were divided into control group, model group, transforming growth factor β agonist group and transforming growth factor β inhibitor group. The rat model of pulmonary hypertension was established by intraperitoneal injection of monocrotaline. The agonist group and inhibitor group were injected intraperitoneally after modeling. The levels of transforming growth factor β, intercellular adhesion molecule -1, interleukin-1β and interleukin-6 in cell culture medium and lung tissue were detected by ELISA. DHE method was used to detect the level of reactive oxygen species in cells. The changes of mitochondrial membrane potential were detected by JC-1 method. The expression of key genes related to transforming growth factor β pathway was detected by RT-qPCR and western blot assay. Hematoxylin-eosin staining was used to observe the pathological changes of lung tissue in rats.
RESULTS AND CONCLUSION: (1) A total of 914 differentially expressed genes in pulmonary hypertension were identified by bioinformatics analysis, 39 of which were related to oxidative stress. Furthermore, through machine learning analysis, aquaporin, hemoglobin β chain, mitochondrial nicotinamide adenine dinucleotide dehydrogenase subunit 2 and Toll-like receptor 6 were identified as candidate genes related to oxidative stress in pulmonary hypertension. (2) At the cellular test, compared with the control group, the intracellular reactive oxygen species production in the activated group increased, the mitochondrial membrane potential decreased, the levels of transforming growth factor β, intercellular adhesion molecule -1, interleukin-1β and interleukin-6 increased significantly, and the mRNA and protein expressions of transforming growth factor β, hemoglobin β chain and Toll-like receptor 6 were up-regulated, while the expressions of aquaporin and mitochondrial nicotinamide adenine dinucleotide dehydrogenase subunit 2 mRNA and protein were down-regulated. Transforming growth factor β agonist further promoted the above changes, while transforming growth factor inhibitor significantly alleviated the above changes. (3) In the animal experiment, compared with the control group, the lung tissue of the model group showed obvious inflammatory infiltration, the level of inflammatory factors increased, with the expression of transforming growth factor β, hemoglobin β chain and Toll-like receptor 6 mRNA and protein up-regulated, while the expression of aquaporin and mitochondrial nicotinamide adenine dinucleotide dehydrogenase subunit 2 mRNA and protein down-regulated. The intervention of transforming growth factor β agonist further aggravated the pathological injury and inflammatory reaction of lung tissue, while transforming growth factor β inhibitor significantly reduced the pathological injury of lung tissue, inhibited the secretion of inflammatory factors, and reversed the change trend of related gene expression. The results exhibit that aquaporin, hemoglobin β chain, mitochondrial nicotinamide adenine dinucleotide dehydrogenase subunit 2 and Toll-like receptor 6 may be important genes that mediate oxidative stress injury of pulmonary hypertension, and transforming growth factor β signaling pathway plays an important role in regulating the expression of these genes and inflammatory injury. Targeted inhibition of transforming growth factor β signaling pathway may be a new way to improve the pathological process of pulmonary hypertension.

Key words: pulmonary hypertension, oxidative stress, transforming growth factor β, inflammation, machine learning, gene expression, pathological mechanism, targeted therapy