Adverse neurological outcome is a major cause of long-term morbidity in ex-preterm children. To investigate the effect of parturition and inflammation on the fetal brain, we utilized two in vivo mouse models of preterm birth. To mimic the most common human scenario of preterm birth, we used a mouse model of intrauterine inflammation by intrauterine infusion of lipopolysaccharide (LPS). To investigate the effect of parturition on the immature fetal brain, in the absence of inflammation, we used a non-infectious model of preterm birth by administering RU486. Pro-inflammatory cytokines (IL-10, IL-1β, IL-6 and TNF-α) in amniotic fluid and inflammatory biomarkers in maternal serum and amniotic fluid were compared between the two models using ELISA. Pro-inflammatory cytokine expression was evaluated in the whole fetal brains from the two models. Primary neuronal cultures from the fetal cortex were established from the different models and controls in order to compare the neuronal morphology. Only the intrauterine inflammation model resulted in an elevation of inflammatory biomarkers in the maternal serum and amniotic fluid. Exposure to inflammationinduced preterm birth, but not non-infectious preterm birth, also resulted in an increase in cytokine mRNA in whole fetal brain and in disrupted fetal neuronal morphology. In particular, Microtubuleassociated protein 2 (MAP2) staining was decreased and the number of dendrites was reduced (P < 0.001, ANOVA between groups). These results suggest that inflammation-induced preterm birth and not the process of preterm birth may result in neuroinflammation and alter fetal neuronal morphology. Keywords mouse model of preterm birth; neuroinflammation; neuronal injuryIn the United States, approximately 12% of all live births are delivered preterm (Green et al., 2005). Preterm birth (PTB) is the leading cause of neonatal mortality and morbidity in the NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptUnited States. Specifically, PTB is a risk factor for adverse neurological outcome for expreterm children (Anderson and Doyle, 2003;Hack et al., 2005).It has long been believed that cerebral palsy is the primary neurological outcome of clinical interest. However, it is now known that ex-preterm children also are at a significant risk for a spectrum of cognitive and neurobehavioral disorders Wood et al., 2005;Costeloe, 2006;Limperopoulos et al., 2007;Lindstrom et al., 2009) including autism spectrum disorders (Brimacombe et al., 2007;Limperopoulos et al., 2008;Schendel and Bhasin, 2008). Current understanding of the pathogenesis of fetal brain injury in a PTB focuses mainly on specific structural findings of white matter damage (WMD) (Cai et al., 2000;Paintlia et al., 2004;Rousset et al., 2006). This current paradigm may be insufficient to explain the increasing prevalence of adverse cognitive and neurobehavioral outcomes in ex-preterm infants.While adverse neurological outcomes are increasingly prevalent in ex-preterm children, it remains unknown whether th...

Objective-To elucidate possible mechanisms of fetal neuronal injury in inflammation-induced preterm birth.Study design-Utilizing mouse model of preterm birth, primary cultures were prepared from fetal brains: 1) control neurons (CN); 2) LPS-exposed neurons (LN); 3) control co-culture (CCC), consisting of neurons and glia; 4) LPS-exposed co-culture (LCC), consisting of LPS-exposed neurons and glia. CN and LN were treated with culture media from CN, LN, CCC and LCC after 24 hours in vitro. Immunocytochemistry was performed for culture characterization and neuronal morphology. Quantitative PCR was performed for neuronal differentiation marker, MAP2, and for cell death mediators, Caspases 1, 3 and 9.Results-LPS exposure in vivo did not influence neuronal or glial content in co-cultures but decreased expression of MAP2 in LN. Media from LN and LCC induced morphological changes in control neurons comparable to LN. The neuronal damage caused by in vivo exposure (LN) could not be reversed by media from control groups.Conclusion-LPS-induced preterm birth may be responsible for irreversible neuronal injury.

OBJECTIVE-To investigate whether magnesium sulfate (MgSO 4 ) prevents fetal brain injury in inflammation-associated preterm birth (PTB).STUDY DESIGN-Utilising a mouse model of PTB, LPS or normal saline (NS)-exposed mice via intrauterine injection, were randomized to intraperitoneal treatment with MgSO 4 or NS. From the 4 treatment groups, 1)NS+NS; 2)LPS+NS; 3)LPS+MgSO 4 ; and 4)NS+MgSO 4 , fetal brains were collected for QPCR studies and primary neuronal cultures. mRNA expression of cytokines, cell death, and markers of neuronal and glial differentiation were assessed.Immunocytochemistry and confocal microscopy were performed. RESULTS-There was no difference between LPS+NS and LPS+MgSO 4 groups in expression ofpro-inflammatory cytokines, cell death markers as well markers of pro-oligodendrocyte and astrocyte development (P>0.05 for all). Neuronal cultures from LPS+NS demonstrated morphological changes and this neuronal injury was prevented by MgSO 4 (P<0.001).CONCLUSION-Amelioration of neuronal injury in inflammation-associated PTB may be a key mechanism by which MgSO 4 prevents cerebral palsy.

Premature cervical ripening is believed to contribute to preterm birth (PTB). Preterm cervical ripening may be due to an aberrant regulation in timing of the same processes that occur at term, or may result from unique molecular mechanisms. Using mouse models of PTB, this study sought to investigate if the molecular mechanisms that govern cervical ripening were similar between preterm and term. Lipopolysaccharide (LPS) is infused into the uterine horn to create a mouse model of inflammation-induced PTB. For a noninfectious model of PTB, RU486 was administered. Both models result in delivery of pups in 8-24 h. Cervical tissues were collected from these models, as well as throughout gestation. Cervical tissues from E15 (preterm), E15 LPS (preterm inflammation), and E18.5 (term) were used for microarray analysis (n = 18). Additional experiments using gestational time course specimens were performed to confirm microarray results. Specific gene pathways were differentially expressed between the groups. Genes involved in immunity and inflammation were increased in the cervix in inflammation-induced PTB; term labor was not associated with differential expression of immune pathways. Cytokine expression was not increased in cervices during term labor, but was increased in the pospartum period. Epithelial cell differentiation pathway was significantly altered in term, but not preterm, labor. Activation of immune pathways may be sufficient for cervical ripening, but does not appear necessary. Differential expression of the epithelial cell differentiation pathway appears necessary in the process of cervical repair. Our results indicate that the molecular mechanisms governing preterm and term cervical ripening are distinctly different.

Inflammatory cytokines are important regulators of metabolism and food intake. Over production of inflammatory cytokines during bacterial and viral infections leads to anorexia and reduced food intake. However, it remains unclear whether any inflammatory cytokines are involved in the regulation of taste reception, the sensory mechanism governing food intake. Previously, we showed that tumor necrosis factor (TNF), a potent proinflammatory cytokine, is preferentially expressed in a subset of taste bud cells. The level of TNF in taste cells can be further induced by inflammatory stimuli. To investigate whether TNF plays a role in regulating taste responses, in this study, we performed taste behavioral tests and gustatory nerve recordings in TNF knockout mice. Behavioral tests showed that TNF-deficient mice are significantly less sensitive to the bitter compound quinine than wild-type mice, while their responses to sweet, umami, salty, and sour compounds are comparable to those of wild-type controls. Furthermore, nerve recording experiments showed that the chorda tympani nerve in TNF knockout mice is much less responsive to bitter compounds than that in wild-type mice. Chorda tympani nerve responses to sweet, umami, salty, and sour compounds are similar between TNF knockout and wild-type mice, consistent with the results from behavioral tests. We further showed that taste bud cells express the two known TNF receptors TNFR1 and TNFR2 and, therefore, are potential targets of TNF. Together, our results suggest that TNF signaling preferentially modulates bitter taste responses. This mechanism may contribute to taste dysfunction, particularly taste distortion, associated with infections and some chronic inflammatory diseases.