Effects of different concentrations of brusatol on silicosis fibrosis in mice
[Background]Silicosis is a diffuse fibrosis of the lungs caused by long-term inhalation of free silicon dioxide(SiO2).It has a complex pathogenesis and lacks effective treatment.Brusatol(Bru)has a variety of biological activities,and its role in silicosis fibrosis is unclear yet.[Objective]To investigate the effects of different concentrations of Bru on SiO2-induced silicosis fibrosis in mice.[Methods]Thirty male C57BL/6J mice were randomly divided into five groups:a control group,a silica group,and three Bru intervention groups with low,medium,and high doses(1,2,and 4 mg·kg-1),with 6 mice in each group.Except the control group,the remaining groups were establis-hed as SiO2-induced silicosis mouse models by using a single tracheal infusion of 50 μL 60 mg·mL-1 SiO2 suspension.The control group was dosed with equal amount of saline.The Bru intervention groups were injected intraperitoneally with Bru for 5 consecutive days and then injected every other day.After 28 d of exposure,the mice were executed and lung tissues were collected.The lung coefficient of the mice was measured,and the pathological changes of the lung tissues were observed after hematoxylin-eosin(HE)and Masson staining.The levels of apoptotic protein Cleaved-caspase 3,fibrosis-related protein α-smooth muscle actin(α-SMA),type I collagen(Col-I),autophagy-associated protein Beclin1,microtubule-associated protein 1 light chain 3(LC3),Sequestosome 1(p62/SQSTM1),Kelch like ECH-associated protein-1(Keap1),and nuclear factor erythroid 2 related factor 2(Nrf2)were detected by Western blot.The mRNA levels of Caspase 3,α-SMA,and Col-I were measured by realtime fluo-rescence-based quantitative PCR.[Results]Compared with the control group,the lung coefficient of mice in the silica group was significantly increased(P<0.01);the lung tissues of the silicosis mice showed damaged alveolar walls,along with infiltration of inflammatory cells,fibrous nodules,and collagen deposition;furthermore,the protein and mRNA levels of Cleaved-caspase 3,α-SMA,and Col-I were significantly increased(P<0.01);the expression levels of Beclin1,LC3-II/I,p62,and Nrf2 were increased,while that of Keap1 was decreased(P<0.05).The interventions with low and medium doses of Bru reduced lung coefficient(P<0.05)and protected against pathological damage and collagen deposition in the lung tissues of the silicosis mice;the protein and mRNA expression levels of Cleaved-caspase 3,α-SMA,and Col-I were significantly decreased in the low and medium dose groups(P<0.05,P<0.01),the expression levels of Beclin1,LC3-II/I,p62,and Nrf2 were also decreased(P<0.05,P<0.01),and the expression level of Keap1 was increased in the medium dose group(P<0.05).However,compared with the silica group,the differences in lung coefficient,pathological damage,and protein and mRNA expression levels of Cleaved-caspase 3,α-SMA,and Col-I in the Bru high dose group were not statistically significant(P>0.05).In addition,the high dose of Bru decreased Beclin1,LC3-II/I,and Nrf2 expression levels(P<0.01),did not change p62 protein expression level(P>0.05),while increased Keap1 protein level(P<0.01).[Conclusion]Low and medium doses of Bru might regulate autophagy through the Keap1-Nrf2 pathway,ameliorate autophagic degradation impairment,reduce pulmonary coefficient,attenuate apoptosis,and delay the progression of fibrosis in SiO2-induced silicosis mice.
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