This study was designed to investigate the role and mechanism of neutrophils in lung injury in mice with NEC. Combinational treatment with formula milk, hypoxia, and LPS was performed to establish NEC in 5-day-old mice. The pathological changes in intestinal and lung tissues were examined by HE staining. The terminal ileum was taken out for RNA-seq. Ly6G, NE, MPO staining, and flow cytometry were used for qualitative and quantitative analysis of neutrophils in lung tissues respectively. The mRNA relative expression levels of inflammatory factors in intestinal and lung tissues were detected by RT-qPCR; the ROS release level in neutrophils stimulated by fMLP was detected by an enzyme-labeled instrument. The role of ROS in NEC-induced lung injury was further validated by the NAC, a ROS-specific inhibitor. The expression levels of genes in the Keap1—Nrf2 pathway were determined by RT-qPCR. Results showed that compared with the Ctrl group, the intestinal and lung tissues of the mice in the NEC group were significantly damaged. The mRNA relative expression levels of inflammatory factors and anti-oxidant genes were increased, while the expression levels of anti-ROS genes were downregulated. Mouse intestinal RNA-seq results suggested a strong enrichment in the neutrophil degranulation pathway. The levels of NE, MPO, and ROS released by neutrophils were significantly higher than those in the Ctrl group. And the ROS released by neutrophils is involved in NEC-induced lung injury by regulating the Keap1-Nrf2 pathway as the inhibition of the release of ROS can alleviate the injury of intestinal and lung tissues. Thus our study demonstrates that NAC has therapeutic potential for NEC-induced lung injury.

Background: Prenatal exposure to adverse factors can cause congenital heart defects. Ketamine, a widely used anesthetic drug, produces several adverse reactions such as tachycardia, hypertension, and laryngospasm, especially in pediatric patients. This study aimed to detect the effects of ketamine exposure during pregnancy on the cardiogenesis of mouse offspring and the potential mechanisms.Methods: In this study, ketamine at an addictive dose (5 mg/kg) was administered to mice during early gestation to explore the epigenetic mechanism of its causing cardiac dysplasia. The cardiac morphology of the mouse offspring was observed through hematoxylin-eosin staining and transmission electron microscopy. The heart function of one-month-old neonates was detected by echocardiography. The expression of cardiomyogenesis-related genes was detected by western blot and RT-qPCR. The acetylation level of histone H3K9 at the Mlc2 promoter and its deacetylase level and activity were detected by CHIP-qPCR, RT-qPCR, and ELISA, respectively. Results: Our data revealed that ketamine exposure during pregnancy could cause cardiac enlargement, myocardial sarcomere disorganization, and decreased cardiac contractile function in mouse offspring. Moreover, ketamine reduced the expression of Myh6, Myh7, Mlc2, Mef2c, and cTnI. The histone H3K9 acetylation level at the Mlc2 promoter was down-regulated by increasing the histone deacetylase activity and HDAC3 level upon ketamine administration.Conclusions: Our work indicates that H3K9 acetylation is a vital player in cardiac dysplasia in offspring caused by prenatal ketamine exposure and HDAC3 is a key regulatory factor.