Mechanism Investigation Of Hypoxia-inducible Factor In Neonatal Necrotizing Enterocolitis

Objective:To investigate the role of hypoxia-inducible factor（HIF-1α）in neonatal necrotizing enterocolitis（NEC）and its possible mechanism.Methods:The small intestine tissues of 8 children with NEC were collected as the disease group and 6 children with gastrointestinal abnormalities as the control group.Follow the random number method,forty7-day-old C57BL/6 newborn mice were divided into the NEC group（WT/NEC,n=20）and control group（WT/DF,n=20）;Forty 7-day-old Glutaredoxin1（Grx1）knockout mice(Grx1^-/-)on C57BL/6 background were divided into the NEC group(Grx1^-/-/NEC,n=20)and control group(Grx1^-/-/DF,n=20).NEC group mice were given formula milk and artificial feeding,hypoxia,cold stimulation,lipopolysaccharide gavage treatment;control group mice stay with dam in the same room,without any treatment,take the intestinal after 3 days of modeling.HE staining to detect histopathological changes in mouse small intestine.Western blot was used to detect HIF-1αin human and mouse intestinal tissues.It also detected Vascular Endothelial Growth Factor A（VEGFA）in mouse intestinal tissues.Mouse intestinal VEGFA m RNA was detected by RT-PCR.Immunofluorescence staining was used to detect HIF-1αin intestinal tissue of children and newborn mice;Platelet endothelial cell adhesion molecule（CD31）labeled small intestinal blood vessels,and immunofluorescence staining was used to detect the small intestinal blood vessel density of mice.Results:（1）The expression of HIF-1αin the small intestine of NEC children was significantly lower than the control group（P<0.05）;（2）The histopathology score of the small intestine of WT/NEC mice was significantly higher than the WT/DF group（P<0.05）,while the intestinal pathology score of the mice in the Grx1^-/-/NEC group was significantly lower than the WT/NEC group（P<0.05）;（3）The expression levels of HIF-1αand VEGFA in the small intestine of WT/NEC mice were significantly lower than the WT/DF group（P<0.05）;（4）The m RNA expression of VEGFA in WT/NEC mice intestine tissue was significantly lower than the WT/DF group（P<0.05）,and the expression of VEGFA in the Grx1^-/-/NEC group was significantly higher than WT/NEC group（P<0.05）;（5）HIF-1αwas mainly expressed in small intestinal epithelial cells.The immunofluorescence intensity of HIF-1αstaining in small intestinal tissues of NEC children and NEC mice was significantly lower than the control group,respectively.While Grx1 ablation significantly increased the intensity of HIF-1αimmunofluorescence staining in NEC mouse intestine tissue;（6）The small intestinal vessel density of WT/NEC mice was significantly lower than the WT/DF group（P<0.05）,while the small intestinal vessel density of mice in the Grx1^-/-/NEC group was significantly higher than the WT/NEC（P<0.05）.Conclusions:The results show that HIF-1αplays an important role in NEC children and NEC mice.Grx1 ablation can increase the expression of HIF-1αin the intestine of NEC mice,promote the expression of downstream target gene VEGFA m RNA,promote the development of small intestinal vessels in NEC mice,increase the density of small intestine vessels,and reduce the intestinal damage to protect the NEC mice.