Study On Relationship Of Caveolin-1 And TGF-β1 In Hyperoxia-Induced Lung Injury In Vivo And In Vitro

Objective Through the establishment of the model for hyperoxia-induced lung injury model in vivo and vitro, the dynamic changes of Caveolin-1 (Cav-1) and transforming growth factor batal(TGF-β1), and their relationship to the mechanism of lung fibrosis were investigated.Methods In vivo:Thirty premature Wistar rats at 21 days gestation were randomly and equally divided into room air group and experimental group. The lung tissues in two groups were obtained at 1,3 and 7 days after exposing to either room air or hyperoxia. The changes of pulmonary histopathology of different time points were observed by hematoxylin and eosin staining, the severity of pulmonary fibrosis was evaluated, and the expression of Caveolin-1, TGF-β1in the lung tissue was detected in immunohistochemistry. In vitro:The A549 cells were randomly divided into air group and experimental group. The cells in air group were cultured in the incubator with 5%CO2, and the incubator with 5%CO2-95%O2 mixture air was used to culture the experimental group cells. After exposure to oxygen or room air for 12,24 and 48 h, the changes of morphology were observed under inverted microscope; the relationship of expression between the Caveolin-1 and TGF-β1 was detected by double immunofluorescence staining technique, followed by laser scan with focus microscope.Results In vivo:(1) The pathological change of the lung tissue: the control group of 1,3,7day, the lung tissue gradually matured, which the alveolar structure developed to normalized gradually and the alveolar septum thinned. This group at different times had not manifest changes taken placed. On the first day of the experimental group, The lung showed no obvious pathologic change. On the third day there were a few inflammatory cells infiltration in interstitial and alveolar sac. On the seventh day, the damage was more serious:the framework was disordered, alveolar fusion with interstitial fibrosis could be found. (2) Expression of Caveolin-1 and TGF-β1 in the lung tissue:The expression of caveolin-1 remained at a higher level in the air group.Compared with control group, the expression of caveolin-1 in the hyperoxia exposure group began to decrease from 1 day,markedly at 3,7 days(31.21±4.04 vs 46.20±3.50; 19.94±5.58 vs 43.26±4.12, P<0.01)and the expression of Caveolin-1 was negatively correlated with score of interstitial fibrosis in lung (r=-0.88,-0.93, P<0.05). The expression of TGF-β1 remained at a lower level in the air group. Compared with control group, the expression of TGF-(31 in the hyperoxia exposure group began to increase significantly from 1 day, markedly at 3,7 days (20.45±2.75 vs 16.53±1.69; 35.78±2.28 vs 17.05±3.42, P<0.05), and a significant positive correlation was found between the expression of TGF-β1 and the score of interstitial fibrosis in lung (r=0.82,0.87, P<0.05). The expression of Caveolin-1 was highly negative relation to that of TGF-β1at the same time (r=-0.91,-0.86,-0.63,-0.82, P<0.05). In vitro:(1) Under inverted microscopy:A549 cells in control group were significantly increased, sticked to each other tightly and grew very quickly.Compared with control group, the changes in morphology were most serious and injured obvious in experimental group.Structure of the cells changed from typical multy-ang applan to off-normal round or ellpse. (2) The results of immunofluorescence double staining: Caveolin-1 positive cells showed red fluorescence, TGF-β1 positive cells showed green fluorescence, both of them showed yellow fluorescence. In the control group, the high expression of caveolin-1 associated with low expression of TGF-p1(44.25±3.23 vs 10.18±2.74); Compared with control group, with prolonged hyperoxia exposure,the content of Caveolin-1 reduced continuously, but TGF-β1 increased gradually (12h: 35.25±1.88 vs17.32±1.50; 24h:20.75±1.68vs26.51±1.82; 48h:9.86±2.80 vs41.94±4.47), and the expression of Caveolin-1 was highly negative relation to that of TGF-β1 at the same time (r=-0.91,-0.62,-0.53,-0.88, P<0.05). Conclusion The expression level of Caveolin-1 reduced significantly and that of TGF-β1 increased progressively, when exposed to hyperoxia,which means that Caveolin-1 maybe has the anti-pulmonary fibrosis function through decreasing TGF-β1 generation.