Protective Effects Of Resveratrol On Hyperoxia-induced Lung Injury In Neonatal Rats By Alleviating Apoptosis

objective:Bronchopulmonary dysplasia（BPD）is one of the most common long-term lung complications of prematurely born infants caused by prolonged injury and repair during immature lung development.Resveratrol has a variety of pharmacological activities,including anti-inflammatory,antiapoptotic,antioxidant,antifibrotic,anticancer and immune-regulating effects.It has been reported that resveratrol can reduce lung injury induced by hyperoxia in neonatal rats,but its protective mechanism is not clear.This study aimed to investigate the possible protective mechanism of resveratrol in BPD.Methods:Neonate rats were delivered spontaneously and randomized divided into four groups on postnatal day（PN）0.5:room air（21%O₂）+dimethyl sulfoxide（DMSO）,room air+resveratrol（Res）,hyperoxia（80%）+DMSO,hyperoxia+resveratrol.The rat pups in hyperoxia+DMSO group and hyperoxia+Res group were placed inside a high oxygen culture chamber at a flow rate of 2 L/min to maintain 80%O₂ concentration.Neonate rats in air+DMSO group and air+Res group were placed inside a high oxygen culture chamber the same as hyperoxia groups,but they were exposed to room air.Room air+Res group and hyperoxia+Res group were daily given intraperitoneal injection（i.p.）of resveratrol 60mg/kg（Res was dissolved in 35%DMSO）from PN0.5 to the end of experiment.Room air+DMSO group and hyperoxia+DMSO group received 35%DMSO（i.p.）per day at the same dose and time with Res groups.The dams in the room air groups and hyperoxia groups were exchanged every 24h to avoid oxygen toxicity.Lung tissues were collected on PN1,PN7 and PN14.Protective effects of resveratrol on hyperoxia-induced lung injury were evaluated by HE staining,TUNEL staining,ROS detection,qRT-PCR and western blotting.Results:1.Lung histopathological changes:with the extension of hyperoxia exposure time,alveolar wall thickening and pulmonary hemorrhage,alveolarization disorders appeared in hyperoxia+DMSO group on PN7 and PN14.Resveratrol alleviatedhyperoxia-inducedlunghistopathologicalchanges.Alveolarization of lung tissues was evaluated by MLI and RAC,resveratrol reduced hyperoxia-induced increase of MLI and decrease of RAC.2.TUNEL staining:in the hyperoxia+DMSO group,the apoptosis index significantly increased compared with other groups on PN7 and PN14.The hyperoxia+Res group displayed lower apoptosis index than the hyperoxia+DMSO group.3.ROS detection:there was an increase in ROS level in hyperoxia+DMSO group compared with other groups.In neonatal rats pre-treated with resveratrol and exposed to hyperoxia,the level of ROS was significantly lower than hyperoxia+DMSO group.4.qRT-PCR:on PN7 and PN14,the SIRT1 mRNA level in the hyperoxia+DMSO group was significantly decreased compared with that in other groups.Resveratrol treatment raised the decreased SIRT1 mRNA level induced by hyperoxia.Although the relative p53 mRNA level in hyperoxia+DMSO group was higher than other groups on PN14,this result was not statistically significant.5.Western blotting:the protein level of SIRT1,p53 and Acetyl-p53 were found no statistical difference on PN1;The lung protein expression of SIRT1 were significantly decreased while p53 and Acetyl-p53 were increased in the hyperoxia+DMSO group compared with that in the other groups,and resveratrol treatment attenuated the decreases of SIRT1 and the increase of p53 and Acetyl-p53 induced by hyperoxia on PN7 and PN14.Conclusion:Hyperoxia-induced alveolar simplification and apoptosis were alleviated by resveratrol;Resveratrol reduced ROS production,up-regulated SIRT1,decreased the expressing of p53 and Acetyl-p53 in the lung of hyperoxia-exposed neonatal rats.This study showed that resveratrol alleviated hyperoxia-induced apoptosis in neonate rats lung tissue via reducing ROS and p53.Resveratrol induced SIRT1upregulation and Acetyl-p53 reduction may also be involved in lung protection.