Effect Of Intrauterine Inflammation On Clinical Outcome Of Preterm Infants And Characteristic Genes Expression Profiles Of TLR Pathway In Immune Cells From Cord Blood

Fetus and neonate are probably the most susceptible to infection for life and have high mortality related with infection. More than half of neonatal mortality is related with intrauterine and extrauterine infections. Twenty seven percent of neonatal death is due to preterm delivery, for which intrauterine infection is the important responsible risk factor, and twenty six percent of neonatal death is caused by sepsis and pneumonia. Intrauterine infection/inflammation is not only related with preterm delivery but also with the clinical outcome of newborns. The susceptibility of fetus and neonates is attributed to the immature immune responses to microbial infection. In this paper, we explored the relationship between intrauterine inflammation and neonatal outcome by a large cohort. To probe into the molecular mechanisms underlying the increased susceptibility of newborns to infectious diseases, we detected the gene expression profiles of TLR signaling pathway in immune cells from cord blood and compared with that from adult. These results hopefully will enrich our knowledge about fetal and neonatal infection, which is very important challenge for health care providers.Part I:Effect of intrauterine inflammation response on clinical outcome of preterm infantsObjective The prospective cohort study was designed to explore the effects of intrauterine inflammation on neonatal outcome of premature birth.Method216singleton newborns at a gestational age34weeks or less were prospectively recruited into this study, all neonates were born in the international peace maternity&children hospital of shanghai Jiao Tong University between Jan2008and Oct2010. Antenatal, perinatal, and neonatal data were prospectively collected and stored in a database. Neonates who were transferred to another hospital were followed up to complete the data. According to histological evidence of placental infection, intrauterine inflammation was divided into maternal inflammatory response (MIR) and fetal inflammatory response (FIR). Uterine cavity microbial culture was obtained from76maternity patients. Serial head ultrasounds were performed in95premature infants and cranial MRI was obtained at1-3months of corrected age from36premature infants. Brainstem auditory evoked potential and Bayley Scales of Infant Development were obtained at6months corrected age and12months of corrected age in25cases, respectively.Result (1) The Placenta histopathology results showed that intrauterine inflammation response was present in104of216cases (48.1%), and of104cases,53cases (51.0%) the inflammation involved of only maternal side and51cases (49.0%) the inflammation involved both maternal and fetal side. There is no case of only fetal inflammation response. According to the pathological findings, these premature infants were divided into three groups, both maternal and fetal inflammation response negative group (MIR-FIR-), both maternal and fetal inflammation response positive group (MIR+FIR+) and maternal inflammation response positive but fetal inflammation response negative group (MIR+FIR-).(2) To analyze the relationship between preterm and intrauterine inflammation response,216preterm were categorized as27-29weeks,30-32weeks and>33weeks according to gestational age. The incidence of MIR+(whatever FIR status) is significantly higher in27-29weeks (50.0%) and30-32weeks (54.8%) than in>33weeks (35.4%), p<0.05. Furthermore, The incidence of MIR+FIR+is significantly higher in27-29weeks (27.8%) and30-32weeks (28.7%) than in>33weeks (12.1%), p<0.05.(3)17(22.4%) mycoplasma and14(18.4%) microbial species were identified from the microbiological examinations of the76uterine cavity samples, in and samples. There is no significant difference of culture positive rate among MIR-FIR-group, MIR+FIR-group and MIR+FIR+group.(4)95premature infants with serial head ultrasound examination were included into the analysis of the relationship between intrauterine inflammation and neonatal brain injury. Although the overall incidence of intraventricular hemorrhage (IVH) was not significantly different among three groups, the incidence of＜2grade IVH in the MIR+FIR+group (42.3%) was significantly higher than that in the MIR+FIR-group (13.0%) and the MIR-FIR-group(15.2%), p<0.05. Logistic regression analysis showed that MIR+FIR+was associated with increased>2grade neonatal IVH (odds ratio,4.08;95%confidence interval,1.259-13.24; P<.05).(5) The incidence of neonatal respiratory distress syndrome (RDS) in the MIR+FIR-group (5.7%) and the MIR+FIR+(2.0%) group were significantly lower than that in the MIR-FIR-group (19.6%), p<0,05. Logistic regression analysis showed that MIR+FIR+was associated with decreased neonatal RDS (odds ratio,0.08;95%confidence interval,0.010-0.661; P<.05).(6) The incidence of neonatal early onset sepsis in MIR-FIR-group, MIR+FIR-group, and MIR+FIR+group were15.7%,11.3%and7.1%, respectively. Although the trend of increasing neonatal early onset sepsis was occured in the intrauterine inflammation exposure newborns, there was no statistically significant diffidence.(7) The results of follow up for part of study cohort did not show that intrauterir inflammation exposure was associated with brainstem auditory evoked potential and Bayley Scales of Infant Development.Conclusion (1) The Placenta histopathology is the important evidence for judging intrauterine inflammation response. Although it is difficult to distinguish FIE as separate processes from MIR, both are typically different based upon the location of placental histologic findings.(2) The intrauterine inflammation response is the important cause for preterm delivery. Especially under circumstance of preterm premature rupture of membrane, the incidence of intrauterine inflammation response was inversely related to gestational age.(3) The intrauterine inflammation response definitely affect neonatal outcome of premature newborns. MRI+FIR+increased neonatal>2grade IVH and deceased th risk of neonatal RDS and needs for assisted ventilation. Intrauterine inflammation showed the trend of increasing neonatal onset sepsis. Part Ⅱ:Characteristic Gene Expression Profiles of TLR signaling pathway in immune cells from cord bloodObjectives To explore the molecular mechanisms underlying the increased susceptibility of newborns to infectious diseases, we compared the genes expression profiles of Toll-like receptor (TLR) signaling pathway in immune cells from cord blood with that from adult peripheral blood in response to lipopolysaccharide (LPS) stimulation in vitro.Methods Cord Blood samples of10health term newborns and peripheral blood from and10health adult donors were obtained. Whole blood cells were cultured in vitro and LPS was added. Immune cells were isolated at0,2,4,6,8and24hours after LPS induction and total RNA was extracted. Commercially available TLR signaling microarray (Oligo GEArray(?), SuperArray, USA) was used to detect the gene expression profiles of TLR pathway, and differentially expressed genes were confirmed by PCR Array.Results The microarray results showed that the number of differentially expressed genes (i.e., genes that were differentially expressed by^2folds in newborns compared to adults) varied according to time. The maximum differential expression genes occurred at8hours, there were40genes up-regulated and39genes down-regulated. Time-series clustering pattern showed that up-regulated gene expression peaked at2hours after stimulation for the newborn but at6hours for the adult. Some genes were up-regulated or down-regulated consistently in newborns compared with the adults at all or most time point. These up-regulated genes included RP105, Tollip, GPC1, HRAS, HSPA4, FADD, IKKα, NFKBIL1, SAPK3, JNK2and JNK1, and the down-regulated genes included TLR1, TLR2, CD14, MyD88, HSPA1A, MAL, IRAK3and RELB. The expressions of some important negative regulative genes were higher but most proinflammatory factor genes were lower in the newborn than in the adult.Conclusion (1) There were obvious differences in gene expression patterns and dynamical changes in TLR signaling mediators between newborns and adults after LPS stimulation.(2) It appeared that negative regulators might predominate over positive regulators of TLR signaling pathway in response to LPS stimulation in the newborn, hence contributing to neonatal inadequate immunity against infection.