Expression Of CD40 In Brain White Matter Damage Of Preterm Rats Induced By Intrauterine Infection Infection

Objective: 1.To preliminary understand the expression of CD40 and CD40 L in the brain white matter injury in of premature rats caused by intrauterine infection.2.To investigate the effects and potential mechanism of CD40 and CD40 L on white matter brain injury induced by intrauterine infection in premature rats.Methods: 1.Establishment and grouping of animal models: rats on the 17 th day of pregnancy were randomly selected for intraperitoneal injection of lipopolysaccharide(LPS)for 2 consecutive days,and premature rats with a gestational age of less than 22 days were selected as the brain white matter injury group(experimental group).The rats pregnant on the day of 17 th were randomly selected to administer the equivalent amounts of mifepristone intraperitoneally for 2 consecutive days,and the neonatal preterm rats with a gestational age of less than 22 days were selected as the preterm infant group(control group).2.Materials: 16 premature rats were randomly selected from the two groups on the postnatal day 1(P1),P3,P7 and P14,8rats out of 16 were randomly selected to be decapitated from which the periventricular white matter tissue was immediately stored in refrigerator at-80℃ for RT-PCR,western blot and ELISA;The remaining rats were decapitated after cardiac perfusion for hematoxylin-eosin(HE)staining and immunofluorescence staining.In addition,8premature rats in two groups were randomly selected and fed to the 30 th day after birth for water maze experiments.3.Detection methods: HE staining was used to observe the pathological changes of placenta in two groups of pregnant rats and the pathological changes of brain tissue in two groups of premature rats at different time points;RT-PCR was used to detect the expression of CD40,CD40 L and myelin basic protein(MBP)in periventricular white matter tissues of the two groups of premature rats at different timepoints;Western blot was used to detect the protein expression of CD40 and CD40 L in the periventricular white matter tissues of the two groups at different time points;Immunofluorescence was used to detect the morphological changes of microglia in the periventricular white matter areas of the two groups at different time points;ELISA was used to determine concentrations of interieukin(IL)-6 and tumor necrosis factor(TNF)-αin the periventricular white matter tissue of the two groups at different time points;Water maze experiment was used to detect learning and cognitive ability of two groups of preterm rats.Results: 1.Successful establishment of animal models: HE staining of placental tissue in brain injury group(experimental group)showed tissue congestion,edema,with massive neutrophil infiltration;There were no obvious pathological damages in the placenta of the preterm infant group(control group).In the experimental group,pathological manifestations such as loose structure,softening focus and fiber disorder were observed in the periventricular white matter area of premature rats at P7 and P14.There were no obvious pathological changes in the control group.2.Expression of CD40 and CD40 L in the periventricular white matter tissue of premature rats in two groups: The expression level of CD40 m RNA and the expression levels of protein of CD40 and CD40 L in the experimental group were significantly higher than those in the control group at P3 and P7(P<0.05).3.Morphological changes of microglia in the periventricular white matter areas of premature rats in two groups: Microglia in the experimental group showed amoeboid morphology at P1,P3 and P7,while microglia in the control group showed branch-like morphology at P3 and P7.4.Expression of TNF-αand IL-6 in the periventricular white matter tissue of two premature rats: Compared with the control group,TNF-α and IL-6 in the periventricular of the experimental rats were significantly increased on the 3rd and 7th day after birth(P <0.05).5.Expression of MBP m RNA in periventricular white matter tissues of premature rats in two groups:The expression of MBP m RNA in periventricular white matter tissue of premature rats in the experimental group was significantly lower than that in the control group at P14(P<0.05).6.Water maze experiment: The escape latency period of premature rats in the experimental group was significantly longer than that in the control group(P<0.05),and the number of crossing platforms times and the time spent in the target quadrant were significantly less than those in the control group(P<0.05).Conclusion: 1.The model of white matter injury in premature rats induced by intrauterine infection can be successfully established by intraperitoneal injection of LPS in pregnant rats.2.The Up-regulation of CD40 expression may be involved in the pathogenesis of brain white matter injury in premature rats by activating microglia to release inflammatory cytokines.