| Objective Hyperoxia may impair lung growth and is a major contributing factor to the development of bronchopulmonary dysplasia (BPD). We have previously demonstrated that 95% hyperoxia in the newborn rat could temporarily up-regulate the gene expression of p53 and p21WAF/CIP1, which could lead to of cell cycle arrest. Recent clinical study showed that elevated apoptotic ratio was evident in stages II, III, and IV of BPD among alveolar and bronchiolar cells. Animal experiments also proved that hyperoxia may induce apoptosis in newborn murine lung, but the molecular mechanism remains unclear. The aim of this study is to investigate the effect of Bax and Bcl-XL expression in newborn rat with moderate hyperoxic exposure.Methods Hyperoxic lung injury model was established by exposing to 60% O2 in the neonatal period of Spraque-Dawley rats. Rats exposed to air were used as control groups, with 8 animals in each group on repeated experiments. The pathology of lung was detected by HE stain. Mean alveolar area and alveolar number perμm2 were applied for estimating the pathological effects of prolonged hyperoxia in neonatal rats. The levels of Bax and Bcl-XL protein in lung were detected by immunohistochemistry. Under light microscope, five areas of lung parenchyma were systematically and randomly photographed from each slide. With HMIAS-2000 medical color image documentation and analysis system the absorbance (A) values of areas were obtained to semi-quantify the levels of the proteins. The levels of Bax and Bcl-XL mRNA in lungs were detected by reverse transcription-polymerase chain reaction (RT-PCR). PCR products were electrophoretically separated with 2.5% agarose gels. The A values of the DNA bands were quantified by complete gel documentation and analysis system. The A ratios of target gene/β-actin denoted the relative content of target gene mRNA. All numeric results were represented as mean±standard deviation. Results Alveolar dysplasia appeared 4 days after hyperoxia. Mean alveolar area increased in hyperoxia groups. Alveolar number perμm2 decreased from the 4th day after hyperoxia. Bax and Bcl-XL proteins were mainly expressed on bronchiolar epithelial cells and vascular endothelial cells. Compared with control group, the expression of Bax increased from the 1st day after hyperoxia, the level of its mRNA decreased from the 11th day (q=8.4802, P<0.05) and the level of its protein decreased until the 14th day (q=6.8364, P<0.05). The expression of Bcl-XL mRNA augmented from the 1st to 14th day, while the expression of its protein diminished on the 1st day (q=10.4299, P<0.05) and then raised. As hyperoxic exposure prolonged, the levels of Bax mRNA, its protein and Bcl-XL mRNA degraded, while the level of Bcl-XL protein upgraded. In air groups, the level of Bax mRNA increased on the 11th day after exposure and the level of its protein increased on the 14th day after exposure. The levels of Bcl-XL mRNA and protein were the highest among air groups on the 1st day.Conclusion Regulation of moderate hyperoxia induced apoptosis in lungs of newborn rat may be related to the abnormal expression of Bax and Bcl-XL. At the early stage after hyperoxia, expression of Bax increases, which promotes apoptosis in lung. Apoptosis of lung epithelial cells and endothelial cells impaired normal lung development. Expression of Bax decreases while the level of Bcl-XL increases along with the oxygen exposure. This change leads to lung repair process and newborn rats become resistant to hyperoxia. The relationship between apoptosis and time of oxygen exposure is also indicated in the present study, which is that apoptosis effect on newborn lung is significant at the early stage of oxygen exposure and attenuates later. |
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