Whole-genome expression profiling revealed Escherichia coli MG1655 genes induced by growth on mucus, conditions designed to mimic nutrient availability in the mammalian intestine. Most were nutritional genes corresponding to catabolic pathways for nutrients found in mucus. We knocked out several pathways and tested the relative fitness of the mutants for colonization of the mouse intestine in competition with their wild-type parent. We found that only mutations in sugar pathways affected colonization, not phospholipid and amino acid catabolism, not gluconeogenesis, not the tricarboxylic acid cycle, and not the pentose phosphate pathway. Gluconate appeared to be a major carbon source used by E. coli MG1655 to colonize, having an impact on both the initiation and maintenance stages. N-acetylglucosamine and N-acetylneuraminic acid appeared to be involved in initiation, but not maintenance. Glucuronate, mannose, fucose, and ribose appeared to be involved in maintenance, but not initiation. The in vitro order of preference for these seven sugars paralleled the relative impact of the corresponding metabolic lesions on colonization: gluconate > N-acetylglucosamine > N-acetylneuraminic acid ‫؍‬ glucuronate > mannose > fucose > ribose. The results of this systematic analysis of nutrients used by E. coli MG1655 to colonize the mouse intestine are intriguing in light of the nutrientniche hypothesis, which states that the ecological niches within the intestine are defined by nutrient availability. Because humans are presumably colonized with different commensal strains, differences in nutrient availability may provide an open niche for infecting E. coli pathogens in some individuals and a barrier to infection in others.

Infect. Immun. 58: [2438][2439][2440][2441][2442][2443][2444][2445] 1990), but what nutrients and metabolic pathways are employed during colonization has not been determined. In this study, when the wild-type EDL933 strain was fed to mice along with an EDL933 ⌬ppsA ⌬pckA mutant, which is unable to utilize tricarboxylic acid cycle intermediates and gluconeogenic substrates for growth, both strains colonized the mouse intestine equally well. Therefore, EDL933 utilizes a glycolytic substrate(s) for both initial growth and maintenance when it is the only E. coli strain fed to the mice. However, in the presence of large numbers of MG1655, a K-12 strain, it is shown that EDL933 utilizes a glycolytic substrate(s) for initial growth in the mouse intestine but appears to utilize both glycolytic and gluconeogenic substrates in an attempt to maintain colonization. It is further shown that MG1655 predominantly utilizes glycolytic substrates for growth in the mouse intestine whether growing in the presence or absence of large numbers of EDL933. Data are presented showing that although small numbers of EDL933 grow to large numbers in the intestine in the presence of large numbers of MG1655 when both strains are fed to mice simultaneously, precolonization with MG1655 affords protection against subsequent colonization by EDL933. Moreover, in mice that are precolonized with EDL933, small numbers of MG1655 are able to grow rapidly in the intestine and EDL933 is eliminated. In situ hybridization experiments using E. coli-specific rRNA probes showed that while MG1655 is found only in mucus, EDL933 is found both in mucus and closely associated with intestinal epithelial cells. The data are discussed with respect to competition for nutrients and to the protection that some intestinal commensal E. coli strains might afford against infection by O157:H7 strains.Escherichia coli strains of serotype O157:H7 cause outbreaks of hemorrhagic colitis and hemolytic uremic syndrome in humans (reviewed in reference 14). E. coli O157:H7 initiates infection by binding to intestinal epithelial cells and producing Shiga toxins Stx1 and/or Stx2, depending on the strain (reviewed in reference 14). Stx1 and Stx2 depurinate a critical residue in the eucaryotic 28S rRNA of 60S ribosomes, resulting in the inhibition of protein synthesis and consequent cell death (33). E. coli EDL933, an O157:H7 strain, does not normally kill streptomycin-treated mice and appears to colonize the mouse intestine by growing in intestinal mucus (38, 40), but little is known about the nutrients that are utilized for growth or the metabolic pathways involved. If these pathways were defined, it is likely that preventative measures or more effective treatments for patients infected with O157:H7 strains could be developed. With this goal in mind, we isolated an EDL933 ⌬ppsA ⌬pckA mutant, which grows normally on glycolytic substrates but is unable to utilize tricarboxylic acid (TCA) cycle intermediates and gluconeogenic substrates for growth, and tested its ability to colonize the m...

Although replete with cytotoxic machinery, uterine NK (uNK) cells remain tolerant at the maternal-fetal interface. The mechanisms that facilitate the uNK cell tolerance are largely unknown. Here we demonstrate that VEGF C, a pro-angiogenic factor produced by uNK cells, is responsible for their non-cytotoxic activity. VEGF C-producing uNK cells support endovascular processes as demonstrated in a three dimensional co-culture model of capillary tube formation on matrigel. Peripheral blood NK cells fail to produce VEGF C and remain cytotoxic. This response can be reversed by exogenous VEGF C. We show that cytoprotection by VEGF C can be related to induction of the “transporter associated with antigen processing (TAP)-1” expression and MHC class I assembly in target cells. siRNA-mediated silencing of TAP-1 expression abolished the VEGF C-imparted protection. Overall, these results demonstrate that empowerment of uNK cells with angiogenic factors keeps them non-cytotoxic. This phenotype is critical to their pregnancy compatible immuno-vascular role during placentation and fetal development.

Preeclampsia is a major pregnancy complication with potential short- and long-term consequences for both mother and fetus. Understanding its pathogenesis and causative biomarkers is likely to yield insights for prediction and treatment. Herein, we provide evidence that transthyretin, a transporter of thyroxine and retinol, is aggregated in preeclampsia and is present at reduced levels in sera of preeclamptic women, as detected by proteomic screen. We demonstrate that transthyretin aggregates form deposits in preeclampsia placental tissue and cause apoptosis. By using in vitro approaches and a humanized mouse model, we provide evidence for a causal link between dysregulated transthyretin and preeclampsia. Native transthyretin inhibits all preeclampsia-like features in the humanized mouse model, including new-onset proteinuria, increased blood pressure, glomerular endotheliosis, and production of anti-angiogenic factors. Our findings suggest that a focus on transthyretin structure and function is a novel strategy to understand and combat preeclampsia.

Early diagnosis and treatment of preeclampsia would significantly reduce maternal and fetal morbidity and mortality. However, its etiology and prediction have remained elusive. Based on the hypothesis that sera from patients with preeclampsia could function as a "blueprint" of causative factors, we describe a serum-based pregnancy-specific mouse model that closely mirrors the human condition as well as an in vitro predictive assay. We show that a single administration of human preeclampsia serum in pregnant IL-10-/- mice induced the full spectrum of preeclampsia-like symptoms, caused hypoxic injury in uteroplacental tissues, and elevated soluble fms-like tyrosine kinase 1 and soluble endoglin, markers thought to be related to the disease. The same serum sample(s) induced a partial preeclampsia phenotype in wild-type mice. Importantly, preeclampsia serum disrupted cross talk between trophoblasts and endothelial cells in an in vitro model of endovascular activity. Disruption of endovascular activity could be documented in serum samples as early as 12 to 14 weeks of gestation from patients who subsequently developed preeclampsia. These results indicate that preeclampsia patient sera can be used to understand the pregnancy-specific disease pathology in mice and can predict the disorder.

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Human chorionic gonadotropin (hCG) is crucial for successful pregnancy. Its many functions include angiogenesis and immune regulation. Despite years of research, the etiology of preeclampsia remains unknown. Marked by insufficient trophoblast invasion and poor spiral artery remodeling, preeclampsia has also been linked to immune dysregulation. Here we discuss the roles of hCG in the context of endovascular cross-talk between trophoblasts and endothelial cells and immune tolerance. We propose that functional and glycosylation modifications of hCG may contribute to the pathogenesis of preeclampsia.

IL-10 is a pregnancy compatible cytokine that plays a vital role in maintaining balance of anti-inflammatory and pro-inflammatory milieu at the maternal-fetal interface. Recent evidence now suggests that IL-10 is a potent vascular cytokine that can blunt hypertension and inflammation-mediated vascular dysfunction. Thus, a re-evaluation of IL-10 as a cytokine supporting endovascular interactions and angiogenesis as well as blunting hypoxic-injury and preeclampsia-like features is warranted. In this review, we will highlight these novel functions of IL-10 and propose that its immunomodulatory and vascular functions are mutually inclusive, particularly in the context of normal gestation.