The interventional effect of Xuebijing injection on expression of mitochondrial fusion protein 2 and the ultrastructure changes in lung tissues in rats with paraquat poisoning

Abstract

To investigate the mechanism of pulmonary fibrosis induced by paraquat (PQ), and the effect of Xuebijing injection in treatment of PQ poisoning. Seventy-two male Wistar rats were randomly divided into control group, PQ poisoning group, and Xuebijing intervention group, with 24 rats in each group. Pulmonary fibrosis was induced by single gavage at the dosage of 50 mg/kg of PQ, while 1 mL of distilled water was given by gavage in control group. Xuebijing injection at the dosage of 4 mL/kg were given intraperitoneally at 30 minutes after exposure to PQ in Xuebijing group, and it was repeated every 12 hours; same amount of physiological saline was given intraperitoneally in PQ group and control group. The experiment lasted for 14 days. Six rats in each group were sacrificed on 1, 3, 7, 14 days, respectively, after insult, and 30 minutes after the last intervention. The lung tissues were harvested, the changes in pathology in lung tissue and the degree of pulmonary fibrosis were observed with optical microscope with hematoxylin-eosin (HE) staining and Masson stain. The ultrastructure changes in lung tissues were observed with transmission electron microscopic, and the content of hydroxyproline (HYP) in the lung tissue was determined by alkaline hydrolysis. The expression of mitochondrial fusion protein 2 (Mfn2) was determined by Western Blot. (1) HE staining: in PQ group, inflammation was most marked on the 3rd day. On the 7th day, exudates in the alveoli started to be organized, and hypertrophic fibroblasts were seen to secrete slim collagen fibers, and fibrosis could be seen in alveoli. On the 14th day, intensive hyperplasia of fibroblasts could be observed, and the alveolar structure was destroyed and collapsed, with deposition of collagen deposited with formation of pulmonary fibrosis. At the same time, pathologic changes were milder in Xuebijing group than those in PQ group. (2) Masson staining: the degree of inflammation in alveoli and pulmonary fibrosis were less marked in Xuebijing group than those of PQ group on the 14th day. (3) Under the transmission electron microscopy, it was found that the mitochondria of lung tissue cells was relatively less in number on the 14th day in PQ group, and the majority of them underwent degeneration, swelling and damage. Basement membrane became folded, alveoli were collapsed, and fibrosis was obvious. These changes were less serious in Xuebijing group. (4) Content of HYP: contents of HYP in lung tissues on the 3rd day in PQ group and Xuebijing group were significantly higher than those in control group (743.3±50.2 μg/g, 718.1±34.0 μg/g vs. 665.8±6.6 μg/g, both P<0.05), it then increased gradually, but the contents of HYP in Xuebijing group were significantly lower on the 7th day and 14th day than those in PQ group (790.5±23.8 μg/g vs. 876.7±42.0 μg/g, 812.9±72.3 μg/g vs. 931.3±33.0 μg/g, both P<0.05). (5) Expression of Mfn2: the expression of Mfn2 in control group was relatively lower. The expression of Mfn2 in PQ group was increased gradually under stress, but its rate was low. The expression of Mfn2 (A value) in Xuebijing group was significantly higher than that in PQ group on the 1st day (0.731±0.035 vs. 0.618±0.029, P<0.05), and it was elevated steadily, reaching the peak on the 7th day (0.732±0.037 vs. 0.669±0.034, P<0.05), but it was lower than that of PQ group on the 14th day (0.708±0.034 vs. 0.765±0.041, P<0.05). Xuebijing reduces lung inflammatory reaction and pulmonary fibrosis as a result of PQ poisoning. The mechanism is that Xuebijing regulates and increases expression of Mfn2 in lung tissue.