| Background and ObjectivePerinatal infection play an important role in inducing both preterm labor andbrain injury in the premature infant. White matter injury (WMI) is one of the mostcommon and predominant pattern of brain injury in premature infants, it canseriously affect the health and life quality of preterm infants. Studies have shownthat postnatal infection is one of the most important risk factors for adverseneurodevelopmental outcomes in preterm newborns. Many studies have appliedbacterial lipopolysaccharide (LPS) to establish animal model of infection inducedneonatal brain injury, the most common model of neonatal brain damage are causedby intrauterine infection or postnatal infection. There are several pathway for LPSusing, which including intraperitoneal injection,intravenous or intracerebral injection.Currently, safe and effective postnatal interventions are urgently needed to minimizeneurological deficits of preterm infants. Erythropoietin (EPO) is a new neurotrophicand protective factors of the central nervous system, it has been demonstrated thatexogenous EPO can through the blood-brain barrier into the brain tissue and shows aprotective effect to neonatal brain injury. However, it is still unclear what the rational time of administration of EPO for brain damage in premature neonate caused byinfections should be. Base on the previous model of neonatal rat brain damageinduced by intraperitoneal injection of LPS, this experimental study aims toinvestigate the best EPO administer opportunity for white matter injury caused bypostnatal infection in premature infants and the related mechanism, and to provides atheoretical basis of clinical application of EPO to prevent the brain damage.Materials and MethodsPostnatal day2(P2) newborn SD rats were randomly divided into4groups:control group(A),LPS group(B),the early EPO group (C)and the late EPO group (D).Pups in A,B and C groups were injected intraperitoneally with different drugs(A forsaline, B for0.6mg/kg of LPS, and C for0.6mg/kg of LPS and5000U/kg ofEPO)once a day for consecutive5days(P2-P6).Pups in D group were injectedintraperitoneally with0.6mg/kg of LPS once a day for consecutive5days(P2-P6),and then with5000U/kg of EPO once a day for consecutive5days(P7-P11).10newborn rats in A and B groups were randomly selected on P2(at6h afterintraperitoneal injection of drugs for the first time),P7and P12, the brains weredissected into left and right hemispheres marked by sagittal suture, using ELISAmethod to evaluate the Erythropoietin Receptor (EPOR) protein level with the rightcerebral hemisphere and RT-PCR method to investigate EPOR mRNA level with theleft cerebral hemisphere. Immunohistochemistry was adopted to evaluate theexpression of myelin basic protein (MBP), glial fibrillary acidic protein (GFAP) andEPOR at specified time point. HE dyeing was used to observe the pathologicalchanges of brain damage in different groups.Results1. Pathological changes of newborn rats brain tissue in different groupsHE staining of the control group presented normal structure in brainhippocampal.Reduced numbers of hippocampal pyramidal cells and unclearedstructure were found in LPS infected group. It was better in EPO group than LPS group,and the early EPO group showed better outcome than the later EPO group.Normal lateral ventricles,normal structure of periventricular white matter,clearlygray and white matter boundaries and the neatly arranged neurons were found in thecontrol group.Expansion of the lateral ventricles and periventricular leukomalaciawere found in LPS infected group. No leukomalacia was found in EPO group,andlateral ventricles expansion of EPO groups was alleviated and it was more obviousin early EPO group than later EPO group.2. The MBP expression of neonatal rats brain in difference groupsThe MBP expression of LPS infection group was significantly reducedcompared with control group(P<0.05), and the MBP expression of both early andlate EPO groups increased in contrast to the LPS infection group(P<0.05), moreover,the early EPO group increased more obviously than the later EPO group(P<0.05).3. The GFAP expression of neonatal rat brain in different groupsThe GFAP expression of LPS infection group at P7increased significantly thanthe control group(P<0.05),and there is no statistically significant difference betweenEPO group and LPS infection group (P>0.05).The GFAP expression of LPSinfection group at P12increased significantly than the control group (P<0.05),andthat of EPO groups were lower than LPS infection group at P12(P<0.05), butcompared the early EPO group with the later EPO group, no statistically significantdifference was found(P>0.05).4. Effects of LPS on EPOR protein and EPOR mRNA expression in neonatalrat brain tissueImmunohistochemistry showed that the EPOR expression was significantlyincreased in infection group than control group at P7,the average integral opticaldensity (IOD) of the two groups had statistically significant difference (P <0.05).The expression of EPOR in infection group increased more than the control group atP12, but there was no statistically significant differences between the two groups(P>0.05).ELISA and RT-PCR indicated that the level of EPOR protein and EPORmRNA in infection group was up-regulated compared with the control group, therewas statistically significant differences between the two groups both at P2and P7(P<0.05), but not at P12(P>0.05).The EPOR protein and mRNA level had a decline tendency with the increase of age, the EPOR level of infection group and controlgroup at P2was higher than that at P7and P12(P <0.05),the expression of the twogroups at P7was higher than P12,but there was no statistically significantdifferences (P>0.05).Conclusions1.EPO shows protective effects to neonatal rat brain injury induced bypostpartum infection, manifests by improving pathological changes, promoting whitematter myelin development, reducing excessive proliferation of astrocytes, it issuperior to be administered at early time than at later time.2.Infection can up-regulate the EPOR expression in rat brain and the EPORexpression level has a decline tendency with the increase of age. |
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