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.