Nancy Louis - Academia.edu (original) (raw)

Papers by Nancy Louis

CD44v6 mediates neutrophil clearance from the apical surface of the intestinal epithelium

The FASEB Journal, 2009

Targeting of Lewis‐a glycans accelerates chemotaxis and transepithelial migration by human PMNs (650.9)

The FASEB Journal, 2014

Pathogen-triggered neutrophil (PMN) recruitment is critical for innate immunity, but aberrant PMN... more Pathogen-triggered neutrophil (PMN) recruitment is critical for innate immunity, but aberrant PMN influx into intestinal crypts marks disease severity in inflammatory bowel disease (IBD). Fucosylated terminal glycans, such as Lewis-x (Lex) and Sialyl Lewis-x, have previously been implicated in the regulation of important PMN functions, including selectin-mediated PMN trafficking. While such glycans based on the type 2 sequence (Galβ1-4GlcNAc-R) are abundant on PMNs, it was previously thought that PMNs lack type 1 glycans (Galβ1-3GlcNAc-R) required for the expression of Lewis-a (Lea). Here, we demonstrate through immunoblot, flow cytometry, and immunohistochemical analyses of tissues from individuals with IBD, that Lea is in fact present on human PMNs. Further, immunoblotting revealed robust PMN expression of Fucosyltransferase 3/5, glycosyltransferases with α1/4 fucosyltransferase activity, required for the terminal step in Lea synthesis. We further report that treatment of PMN with Lea-specific Abs incre...

Targeting of Lewis‐X‐containing glycans blocks PMN transepithelial migration and increases phagocytosis and degranulation (488.3)

The FASEB Journal, 2014

The transepithelial migration (TEM) of neutrophils (PMN) is a histopathological hallmark of infla... more The transepithelial migration (TEM) of neutrophils (PMN) is a histopathological hallmark of inflammatory bowel disease (IBD), yet the mechanisms controlling PMN TEM remain poorly defined. The fucosylated, terminal glycan determinant Lewis X (Lex), expressed on the glycans of PMN surface glycoproteins (including MAC-1 and LFA-1), has previously been implicated in adhesive interactions between PMN and endothelium. However, little is known about the role of Lex in PMN function following extravasation. Flow cytometry analyses revealed increased surface expression of Lex on human PMN following TEM, while immunohistochemistry demonstrated robust expression of PMN-associated Lex within crypt abscesses in the colonic epithelium of individuals with IBD. Furthermore, engagement of Lex increased PMN adhesive interactions, both with other PMNs and with intestinal epithelial cells, blocking PMN chemotaxis and TEM. In addition to blocking PMN trafficking, targeting of Lex also altered post-migratory PMN functions, incr...

O‐Glycosylation sites on CD44v6 modulate PMN transepithelial migration

The FASEB Journal, 2010

P-319 Expressions of Lewis-A Glycans on PMN Augment Function by Increasing Transmigration

Inflammatory Bowel Diseases, 2017

TLR4, producing radical oxygen species after lipopolysaccharide (LPS) challenge. We hypothesized ... more TLR4, producing radical oxygen species after lipopolysaccharide (LPS) challenge. We hypothesized that TLR4 signaling differentially regulates Nox1 and Duox2 expression in the intestine under different pathophysiologic conditions. Methods: Duodenum, ileum and colon were collected from villin-TLR4 mice, which have constitutive activation of epithelial TLR4, and their wild-type (WT) littermates. Full-thickness samples, spontaneous duodenal adenomas (in villin-TLR4 mice) and isolated IEC were processed for quantitative PCR determinations. Using previously published Gene Expression Omnibus (GEO) datasets from NCBI (http://www.ncbi. nlm.nih.gov/gds), dataset searches containing keywords “inflammatory bowel disease” (IBD), “colon cancer” and “colorectal cancer” were performed. The Affymetrix Expression Console (EC) (build 1.4.1.46) and the Affymetrix Transcriptome Analysis Console 3.0 (TAC) software were used to obtain analysis of variance and false discovery rate P-values for Nox1, Duox2 ...

Control of Interferon‐alphaA by CD73: Implications for Mucosal Inflammation

The FASEB Journal, 2007

Journal of Leukocyte Biology, 2017

PMN-expressed fucosylated glycans from the Lewis glycan family, including Lewis-x (Lex) and sialy... more PMN-expressed fucosylated glycans from the Lewis glycan family, including Lewis-x (Lex) and sialyl Lewis-x (sLex), have previously been implicated in the regulation of important PMN functions, including selectin-mediated trafficking across vascular endothelium. Although glycans, such as Lex and sLex, which are based on the type 2 sequence (Galβ1-4GlcNAc-R), are abundant on PMNs, the presence of type 1 Galβ1-3GlcNAc-R glycans required for PMN expression of the closely related stereoisomer of Lex, termed Lewis-A (Lea), has not, to our knowledge, been reported. Here, we show that Lea is abundantly expressed by human PMNs and functionally regulates PMN migration. Using mAbs whose precise epitopes were determined using glycan array technology, Lea function was probed using Lea-selective mAbs and lectins, revealing increased PMN transmigration across model intestinal epithelia, which was independent of epithelial-expressed Lea. Analyses of glycan synthetic machinery in PMNs revealed expre...

Inflammatory Bowel Diseases, 2016

Soluble inhibitors revealed a critical role for MAPK signaling-specifically p38 MAPK-in mediating... more Soluble inhibitors revealed a critical role for MAPK signaling-specifically p38 MAPK-in mediating DR3 co-stimulation of ILC3. Conclusions: Collectively, these data highlight a key role for microbial regulation of MNP-derived TL1A in co-stimulating ILC3 effector cytokine production in vivo via p38 MAPK signaling. A mechanistic understanding of both the microbial components which induce TL1A in MNPs and the critical signaling pathways required for ILC3 co-stimulation will provide therapeutic targets to promote mucosal healing.

Journal of Clinical & Cellular Immunology, 2012

In Crohn's Disease (CD) and ulcerative colitis (UC), the major manifestations of inflammatory bow... more In Crohn's Disease (CD) and ulcerative colitis (UC), the major manifestations of inflammatory bowel disease (IBD), genetically predisposed individuals develop chronic intestinal inflammation in response to environmental stimuli, which are mainly derived from luminal flora. Intestinal responses to luminal flora breaching the intestinal barrier require cytokine-regulated activation of elements of innate and acquired immunity, leading to a targeted and contained inflammatory response. Recent population-based genetic analyses have identified polymorphisms in specific genes relevant to pathways critical for inflammatory signalling and cellular response to stress as carrying increased risk for the development of either CD or UC. Specifically, key mediators of apoptosis and autophagy are implicated in the genetic vulnerability to IBD. Patients with IBD have a compromise of their intestinal barrier integrity, as do their first-degree relatives even in the absence of clinical disease, underscoring the critical nature of barrier integrity in the prevention of aberrant immune responses to intestinal flora. Here we explore the relationships between two of the key proinflammatory cytokines mediating intestinal inflammation in IBD, TNF-α and IFNγ, and the mechanisms by which they regulate epithelial apoptosis and intestinal barrier. Specifically we review factors regulating the balance between pro-and antiapoptotic stimuli resulting from the activation of NF-κB and Aktdependent signalling by proinflammatory cytokines, as well as the influence of oxygen tension and nutritional factors on these pathways.

Mucosal Immunology, Nov 1, 2012

Proinflammatory cytokines induce Guanylate Binding Protein 1 (GBP-1) protein expression in intest... more Proinflammatory cytokines induce Guanylate Binding Protein 1 (GBP-1) protein expression in intestinal epithelial tissues. GBP-1 has been described as influencing a number of cellular processes important for epithelial homeostasis, including cell proliferation. However, many questions remain as to the role of GBP-1 in intestinal mucosal homeostasis. We therefore sought to investigate the function of proinflammatory cytokine induced GBP-1 during intestinal epithelial cell proliferation. Through the use of complementary GBP-1 overexpression and siRNA-mediated knockdown studies, we now show that GBP-1 acts to inhibit pro-mitogenic β-catenin/T cell factor (TCF) signaling. Interestingly, proinflammatory cytokine induced GBP-1 was found to be a potent suppressor of β-catenin protein levels and β-catenin serine 552 phosphorylation. Neither GSK3-β nor proteasomal inhibition alleviated GBP-1-mediated suppression of cell proliferation or βcatenin/TCF signaling, indicating a non-canonical mechanism of β-catenin inhibition. Together, these data show that cytokine-induced GBP-1 retards cell proliferation by forming a negative feedback loop that suppresses β-catenin/TCF signaling. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

The American Journal of Pathology, 2016

The Neutrophil

Mucosal Immunology, 2015

Neutrophils represent the first responders of the innate immune system. They possess an array of ... more Neutrophils represent the first responders of the innate immune system. They possess an array of powerful biological weapons including soluble mediators as well as a capacity for phagocytosis of pathogens and microbial killing. Thus, they are critical elements of the defense against the microbiota at mucosal surfaces and also serve as an indispensable bridge between the innate and the acquired immune system in the defense against infection. However, in the absence of appropriate regulation, neutrophils are also capable of significant tissue damage. Thus, mechanisms exist to regulate both the magnitude and the duration of their activity in response to infection and inflammation. Here, we review the basic structure, function, recruitment, and regulation of these important innate immune cells, along with their mechanisms of microbial killing. Finally, the roles of the neutrophil in interactions with mucosal tissues and other elements of the mucosal immune system are summarized.

The Scientific World JOURNAL, 2010

The ecto-5'-nucleotidase, CD73, catalyzes the rate-limiting step in the phosphohydrolysis of ... more The ecto-5'-nucleotidase, CD73, catalyzes the rate-limiting step in the phosphohydrolysis of ATP to adenosine, and is a critical regulator of the balance between adenosine and its nucleotide precursors. Each of these classes of mediators signal through their independent receptor families to regulate downstream inflammatory signaling. CD73 activity is primarily regulated at the level of transcription in response to the oxygen-sensing transcription factor HIF1, and its tissue-specific expression correlates negatively with oxygen tension. HIF1-dependent induction of CD73 contributes to the protective effects of hypoxia in the inflamed intestinal mucosa. These beneficial effects of CD73 have largely been attributed to downstream adenosine signaling through its tissue-specific receptors. In addition, adenosine signaling has been directly implicated in the protective effects of hypoxic preconditioning against acute hypoxic or ischemic insults. However, recent work has demonstrated tha...

Cancer research, Jan 15, 2002

The microenvironment of rapidly growing tumors is associated with increased energy demand and dim... more The microenvironment of rapidly growing tumors is associated with increased energy demand and diminished vascular supply, resulting in focal areas of prominent hypoxia. A number of hypoxia-responsive genes have been associated with growing tumors, and here we demonstrate that the multidrug resistance (MDR1) gene product P-glycoprotein, a Mr approximately 170,000 transmembrane protein associated with tumor resistance to chemotherapeutics, is induced by ambient hypoxia. Initial studies using quantitative microarray analysis of RNA revealed an approximately 7-fold increase in MDR in epithelial cells exposed to hypoxia (pO(2) 20 torr, 18 h). These findings were further confirmed at the mRNA and protein level. P-Glycoprotein function was studied by analysis of verapamil-inhibitable efflux of digoxin and rhodamine 123 in intact T84 cells and revealed that hypoxia enhances P-glycoprotein function by as much as 7 +/- 0.4-fold over normoxia. Subsequent studies confirmed hypoxia-elicited MDR1...

Cell-Cell Interactions on Solid Matrices

Cell-Cell Interactions in Health and Disease

Models to study molecular, biochemical, and functional responses in vitro generally incorporate a... more Models to study molecular, biochemical, and functional responses in vitro generally incorporate an individual cell type or group of cells organized in a random fashion. Normal physiological responses in vivo require that individual cell types be oriented in an organized fashion with three-dimensional architecture and appropriately positioned cellular interfaces. Much recent progress has been made in the development and implementation of models to study cell-cell contact using substrate grown cells. Here, we summarize the use of membrane permeable supports to study functional responses in appropriately positioned cell types. These models incorporate two or more different cells cultured in physiologically positioned locales on solid substrates. Models incorporating nonadherent cells (e.g., leukocytes) in co-culture with such models also are discussed. Such models have been used extensively to discovery both cell-bound as well as soluble mediators of physiological and pathophysiological processes.

Model Systems to Investigate Neutrophil Adhesion and Chemotaxis

Methods in Molecular Biology™, 2007

Polymorphonuclear neutrophil (PMN) recruitment from the blood stream into surrounding tissues, fo... more Polymorphonuclear neutrophil (PMN) recruitment from the blood stream into surrounding tissues, followed by migration through the tissue with triggered release of oxidative enzymes or eventual clearance from the epithelial surface, involves a regulated series of events central to acute responses in host defense. Accumulations of large numbers of neutrophils within mucosal tissues are pathognomonic features of both acute and chronic inflammatory conditions including sepsis and inflammatory bowel disease, but the precise signals governing neutrophil adhesion and transmigration remain to be fully characterized. Previous chapters examine methods employed for both neutrophil isolation and study of the mechanisms underlying regulation of PMN rolling behavior. Here, we describe in vitro experimental models for the examination of PMN adhesion to endothelial and epithelial monolayers as well as the characterization of signals influencing neutrophil migration, both along acellular matrices and across endothelial and epithelial monolayers, in the physiologically relevant directions. Studies employing these model systems allow further elucidation of the mechanisms governing PMN adhesion and transmigration.

The FASEB Journal, 2007

Migration of neutrophils (PMN) across epithelia is a pathological hallmark of numerous mucosal di... more Migration of neutrophils (PMN) across epithelia is a pathological hallmark of numerous mucosal diseases. Whereas lesions at mucosal surfaces are generally self-limiting, endogenous mechanisms of resolution are incompletely understood. Previous studies revealed that resolvins directly act on PMN to attenuate transendothelial migration, less is known about the influence of resolvins on PMN-epithelial interactions and whether they act on epithelia. We studied the dynamics of resolvin E1 (RvE1) actions on leukocyte transepithelial migration. PMN exposure to RvE1 or chemerin (peptide agonist of ChemR23) reduced transepithelial migration in a concentration-dependent manner. Conversely, activation of epithelial ChemR23 promoted apical clearance of PMN. A nonbiased screen of known PMN ligands expressed on epithelial cells in response to RvE1 revealed selective induction of CD55, an apically expressed antiadhesive molecule. CD55 promoter analysis demonstrated that both RvE1 and chemerin activate the CD55 promoter. Inhibition of CD55 by neutralizing antibody prevented RvE1-dependent augmentation of apical PMN clearance. Taken together these findings implicate a "two-hit" model of inflammatory resolution, whereby activation of the PMN RvE1 receptor attenuates transepithelial migration and subsequent actions on the epithelium promote CD55-dependent clearance of PMN across the epithelial cell surface promoting active inflammatory resolution.

The FASEB Journal, 2005

Sites of inflammation are associated with dramatic shifts in tissue metabolism. Inflammation can ... more Sites of inflammation are associated with dramatic shifts in tissue metabolism. Inflammation can result in significant tissue hypoxia, with resultant induction of hypoxia-responsive genes. Given this association, we hypothesized that neutrophil (PMN) ligands expressed on epithelial cells may be regulated by hypoxia. Initial studies confirmed earlier results that epithelial hypoxia enhances PMN transepithelial migration and promotes apical clearance of PMN from the epithelial surface. A screen of known PMN ligands revealed a surprisingly stable expression pattern in hypoxia. However, this screen identified one gene, CD55, as a highly hypoxia-inducible molecule expressed on the apical membrane of mucosal epithelia. Subsequent studies verified the induction of CD55 mRNA and protein expression by hypoxia. Overexpression of CD55 by transfection in nonhypoxic epithelia resulted in a similar pattern of apical PMN clearance, and peptide mimetics corresponding to the PMN binding site on DAF blocked such apical clearance of PMN. Studies directed at understanding molecular pathways of hypoxia inducibility revealed that a ϳ200 bp region of the CD55 gene conferred hypoxia inducibility for CD55. These studies identified a functional binding site for the transcriptional regulator hypoxia-inducible factor (HIF). Taken together, these results identify HIFdependent induction of epithelial CD55 in the resolution of ongoing inflammation through clearance of apical PMN.-Louis, N. A

Prostaglandins, Leukotrienes and Essential Fatty Acids, 2005

In intact tissues, vascular endothelial cells lie anatomically positioned as the central coordina... more In intact tissues, vascular endothelial cells lie anatomically positioned as the central coordinator of inflammation. Endothelia communicate with underlying cells (e.g. smooth muscle, fibroblasts, epithelia) in ways that both coordinate leukocyte trafficking, and control the composition of the inflammatory microenvironment. Such coordination occurs through both direct communication (e.g. cell adhesion) as well as via soluble mediators liberated at sites of inflammation (e.g. chemokines, cytokines, lipids). Locally generated mediators bind to surface receptors, and mediate both physiologic and pathophysiologic functional responses. Important in this regard, both endothelial and subendothelial cell populations express enzymes capable of utilizing arachidonic acid substrates to generate bioactive lipid mediators (e.g. lipoxygenases, cyclooxygenases). Such lipid mediators can signal via autocrine or paracrine pathways and, depending on the tissue microenvironment, can convey a pro-or anti-inflammatory message. This review will highlight recent studies characterizing inflammatory responses to lipid mediators liberated at sites of inflammation, with a particular emphasis on neutrophil (polymorphonuclear leukocyte or PMN) trafficking.

PLoS ONE, 2012

The intestinal microflora is critical for normal development, with aberrant colonization increasi... more The intestinal microflora is critical for normal development, with aberrant colonization increasing the risk for necrotizing enterocolitis (NEC). In contrast, probiotic bacteria have been shown to decrease its incidence. Multiple pro-and antiinflammatory cytokines have been identified as markers of intestinal inflammation, both in human patients with NEC and in models of immature intestine. Specifically, IL-10 signaling attenuates intestinal responses to gut dysbiosis, and disruption of this pathway exacerbates inflammation in murine models of NEC. However, the effects of probiotics on IL-10 and its signaling pathway, remain poorly defined. Real-time PCR profiling revealed developmental regulation of MIP-2, TNF-a, IL-12, IL-10 and the IL-10R2 subunit of the IL-10 receptor in immature murine colon, while the expression of IL-6 and IL-18 was independent of postnatal age. Enteral administration of the probiotic Lactobacillus rhamnosus GG (LGG) down-regulated the expression of TNF-a and MIP-2 and yet failed to alter IL-10 mRNA and protein expression. LGG did however induce mRNA expression of the IL-10R2 subunit of the IL-10 receptor. IL-10 receptor activation has been associated with signal transducer and activator of transcription (STAT) 3-dependent induction of members of the suppressors of cytokine signaling (SOCS) family. In 2 week-old mice, LGG also induced STAT3 phosphorylation, increased colonic expression of SOCS-3, and attenuated colonic production of MIP-2 and TNF-a. These LGG-dependent changes in phosphoSTAT3, SOCS3, MIP-2 and TNF-a were all inhibited by antibody-mediated blockade of the IL-10 receptor. Thus LGG decreased baseline proinflammatory cytokine expression in the developing colon through upregulation of IL-10 receptor-mediated signaling, most likely due to the combined induction of phospho-STAT3 and SOCS3. Furthermore, LGG-dependent increases in IL-10R2 were associated with reductions in TNF-a, MIP-2 and disease severity in a murine model of intestinal injury in the immature colon.

CD44v6 mediates neutrophil clearance from the apical surface of the intestinal epithelium

The FASEB Journal, 2009

Targeting of Lewis‐a glycans accelerates chemotaxis and transepithelial migration by human PMNs (650.9)

The FASEB Journal, 2014

Pathogen-triggered neutrophil (PMN) recruitment is critical for innate immunity, but aberrant PMN... more Pathogen-triggered neutrophil (PMN) recruitment is critical for innate immunity, but aberrant PMN influx into intestinal crypts marks disease severity in inflammatory bowel disease (IBD). Fucosylated terminal glycans, such as Lewis-x (Lex) and Sialyl Lewis-x, have previously been implicated in the regulation of important PMN functions, including selectin-mediated PMN trafficking. While such glycans based on the type 2 sequence (Galβ1-4GlcNAc-R) are abundant on PMNs, it was previously thought that PMNs lack type 1 glycans (Galβ1-3GlcNAc-R) required for the expression of Lewis-a (Lea). Here, we demonstrate through immunoblot, flow cytometry, and immunohistochemical analyses of tissues from individuals with IBD, that Lea is in fact present on human PMNs. Further, immunoblotting revealed robust PMN expression of Fucosyltransferase 3/5, glycosyltransferases with α1/4 fucosyltransferase activity, required for the terminal step in Lea synthesis. We further report that treatment of PMN with Lea-specific Abs incre...

Targeting of Lewis‐X‐containing glycans blocks PMN transepithelial migration and increases phagocytosis and degranulation (488.3)

The FASEB Journal, 2014

The transepithelial migration (TEM) of neutrophils (PMN) is a histopathological hallmark of infla... more The transepithelial migration (TEM) of neutrophils (PMN) is a histopathological hallmark of inflammatory bowel disease (IBD), yet the mechanisms controlling PMN TEM remain poorly defined. The fucosylated, terminal glycan determinant Lewis X (Lex), expressed on the glycans of PMN surface glycoproteins (including MAC-1 and LFA-1), has previously been implicated in adhesive interactions between PMN and endothelium. However, little is known about the role of Lex in PMN function following extravasation. Flow cytometry analyses revealed increased surface expression of Lex on human PMN following TEM, while immunohistochemistry demonstrated robust expression of PMN-associated Lex within crypt abscesses in the colonic epithelium of individuals with IBD. Furthermore, engagement of Lex increased PMN adhesive interactions, both with other PMNs and with intestinal epithelial cells, blocking PMN chemotaxis and TEM. In addition to blocking PMN trafficking, targeting of Lex also altered post-migratory PMN functions, incr...

O‐Glycosylation sites on CD44v6 modulate PMN transepithelial migration

The FASEB Journal, 2010

P-319 Expressions of Lewis-A Glycans on PMN Augment Function by Increasing Transmigration

Inflammatory Bowel Diseases, 2017

TLR4, producing radical oxygen species after lipopolysaccharide (LPS) challenge. We hypothesized ... more TLR4, producing radical oxygen species after lipopolysaccharide (LPS) challenge. We hypothesized that TLR4 signaling differentially regulates Nox1 and Duox2 expression in the intestine under different pathophysiologic conditions. Methods: Duodenum, ileum and colon were collected from villin-TLR4 mice, which have constitutive activation of epithelial TLR4, and their wild-type (WT) littermates. Full-thickness samples, spontaneous duodenal adenomas (in villin-TLR4 mice) and isolated IEC were processed for quantitative PCR determinations. Using previously published Gene Expression Omnibus (GEO) datasets from NCBI (http://www.ncbi. nlm.nih.gov/gds), dataset searches containing keywords “inflammatory bowel disease” (IBD), “colon cancer” and “colorectal cancer” were performed. The Affymetrix Expression Console (EC) (build 1.4.1.46) and the Affymetrix Transcriptome Analysis Console 3.0 (TAC) software were used to obtain analysis of variance and false discovery rate P-values for Nox1, Duox2 ...

Control of Interferon‐alphaA by CD73: Implications for Mucosal Inflammation

The FASEB Journal, 2007

Journal of Leukocyte Biology, 2017

PMN-expressed fucosylated glycans from the Lewis glycan family, including Lewis-x (Lex) and sialy... more PMN-expressed fucosylated glycans from the Lewis glycan family, including Lewis-x (Lex) and sialyl Lewis-x (sLex), have previously been implicated in the regulation of important PMN functions, including selectin-mediated trafficking across vascular endothelium. Although glycans, such as Lex and sLex, which are based on the type 2 sequence (Galβ1-4GlcNAc-R), are abundant on PMNs, the presence of type 1 Galβ1-3GlcNAc-R glycans required for PMN expression of the closely related stereoisomer of Lex, termed Lewis-A (Lea), has not, to our knowledge, been reported. Here, we show that Lea is abundantly expressed by human PMNs and functionally regulates PMN migration. Using mAbs whose precise epitopes were determined using glycan array technology, Lea function was probed using Lea-selective mAbs and lectins, revealing increased PMN transmigration across model intestinal epithelia, which was independent of epithelial-expressed Lea. Analyses of glycan synthetic machinery in PMNs revealed expre...

Inflammatory Bowel Diseases, 2016

Soluble inhibitors revealed a critical role for MAPK signaling-specifically p38 MAPK-in mediating... more Soluble inhibitors revealed a critical role for MAPK signaling-specifically p38 MAPK-in mediating DR3 co-stimulation of ILC3. Conclusions: Collectively, these data highlight a key role for microbial regulation of MNP-derived TL1A in co-stimulating ILC3 effector cytokine production in vivo via p38 MAPK signaling. A mechanistic understanding of both the microbial components which induce TL1A in MNPs and the critical signaling pathways required for ILC3 co-stimulation will provide therapeutic targets to promote mucosal healing.

Journal of Clinical & Cellular Immunology, 2012

In Crohn's Disease (CD) and ulcerative colitis (UC), the major manifestations of inflammatory bow... more In Crohn's Disease (CD) and ulcerative colitis (UC), the major manifestations of inflammatory bowel disease (IBD), genetically predisposed individuals develop chronic intestinal inflammation in response to environmental stimuli, which are mainly derived from luminal flora. Intestinal responses to luminal flora breaching the intestinal barrier require cytokine-regulated activation of elements of innate and acquired immunity, leading to a targeted and contained inflammatory response. Recent population-based genetic analyses have identified polymorphisms in specific genes relevant to pathways critical for inflammatory signalling and cellular response to stress as carrying increased risk for the development of either CD or UC. Specifically, key mediators of apoptosis and autophagy are implicated in the genetic vulnerability to IBD. Patients with IBD have a compromise of their intestinal barrier integrity, as do their first-degree relatives even in the absence of clinical disease, underscoring the critical nature of barrier integrity in the prevention of aberrant immune responses to intestinal flora. Here we explore the relationships between two of the key proinflammatory cytokines mediating intestinal inflammation in IBD, TNF-α and IFNγ, and the mechanisms by which they regulate epithelial apoptosis and intestinal barrier. Specifically we review factors regulating the balance between pro-and antiapoptotic stimuli resulting from the activation of NF-κB and Aktdependent signalling by proinflammatory cytokines, as well as the influence of oxygen tension and nutritional factors on these pathways.

Mucosal Immunology, Nov 1, 2012

Proinflammatory cytokines induce Guanylate Binding Protein 1 (GBP-1) protein expression in intest... more Proinflammatory cytokines induce Guanylate Binding Protein 1 (GBP-1) protein expression in intestinal epithelial tissues. GBP-1 has been described as influencing a number of cellular processes important for epithelial homeostasis, including cell proliferation. However, many questions remain as to the role of GBP-1 in intestinal mucosal homeostasis. We therefore sought to investigate the function of proinflammatory cytokine induced GBP-1 during intestinal epithelial cell proliferation. Through the use of complementary GBP-1 overexpression and siRNA-mediated knockdown studies, we now show that GBP-1 acts to inhibit pro-mitogenic β-catenin/T cell factor (TCF) signaling. Interestingly, proinflammatory cytokine induced GBP-1 was found to be a potent suppressor of β-catenin protein levels and β-catenin serine 552 phosphorylation. Neither GSK3-β nor proteasomal inhibition alleviated GBP-1-mediated suppression of cell proliferation or βcatenin/TCF signaling, indicating a non-canonical mechanism of β-catenin inhibition. Together, these data show that cytokine-induced GBP-1 retards cell proliferation by forming a negative feedback loop that suppresses β-catenin/TCF signaling. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

The American Journal of Pathology, 2016

The Neutrophil

Mucosal Immunology, 2015

Neutrophils represent the first responders of the innate immune system. They possess an array of ... more Neutrophils represent the first responders of the innate immune system. They possess an array of powerful biological weapons including soluble mediators as well as a capacity for phagocytosis of pathogens and microbial killing. Thus, they are critical elements of the defense against the microbiota at mucosal surfaces and also serve as an indispensable bridge between the innate and the acquired immune system in the defense against infection. However, in the absence of appropriate regulation, neutrophils are also capable of significant tissue damage. Thus, mechanisms exist to regulate both the magnitude and the duration of their activity in response to infection and inflammation. Here, we review the basic structure, function, recruitment, and regulation of these important innate immune cells, along with their mechanisms of microbial killing. Finally, the roles of the neutrophil in interactions with mucosal tissues and other elements of the mucosal immune system are summarized.

The Scientific World JOURNAL, 2010

The ecto-5'-nucleotidase, CD73, catalyzes the rate-limiting step in the phosphohydrolysis of ... more The ecto-5'-nucleotidase, CD73, catalyzes the rate-limiting step in the phosphohydrolysis of ATP to adenosine, and is a critical regulator of the balance between adenosine and its nucleotide precursors. Each of these classes of mediators signal through their independent receptor families to regulate downstream inflammatory signaling. CD73 activity is primarily regulated at the level of transcription in response to the oxygen-sensing transcription factor HIF1, and its tissue-specific expression correlates negatively with oxygen tension. HIF1-dependent induction of CD73 contributes to the protective effects of hypoxia in the inflamed intestinal mucosa. These beneficial effects of CD73 have largely been attributed to downstream adenosine signaling through its tissue-specific receptors. In addition, adenosine signaling has been directly implicated in the protective effects of hypoxic preconditioning against acute hypoxic or ischemic insults. However, recent work has demonstrated tha...

Cancer research, Jan 15, 2002

The microenvironment of rapidly growing tumors is associated with increased energy demand and dim... more The microenvironment of rapidly growing tumors is associated with increased energy demand and diminished vascular supply, resulting in focal areas of prominent hypoxia. A number of hypoxia-responsive genes have been associated with growing tumors, and here we demonstrate that the multidrug resistance (MDR1) gene product P-glycoprotein, a Mr approximately 170,000 transmembrane protein associated with tumor resistance to chemotherapeutics, is induced by ambient hypoxia. Initial studies using quantitative microarray analysis of RNA revealed an approximately 7-fold increase in MDR in epithelial cells exposed to hypoxia (pO(2) 20 torr, 18 h). These findings were further confirmed at the mRNA and protein level. P-Glycoprotein function was studied by analysis of verapamil-inhibitable efflux of digoxin and rhodamine 123 in intact T84 cells and revealed that hypoxia enhances P-glycoprotein function by as much as 7 +/- 0.4-fold over normoxia. Subsequent studies confirmed hypoxia-elicited MDR1...

Cell-Cell Interactions on Solid Matrices

Cell-Cell Interactions in Health and Disease

Models to study molecular, biochemical, and functional responses in vitro generally incorporate a... more Models to study molecular, biochemical, and functional responses in vitro generally incorporate an individual cell type or group of cells organized in a random fashion. Normal physiological responses in vivo require that individual cell types be oriented in an organized fashion with three-dimensional architecture and appropriately positioned cellular interfaces. Much recent progress has been made in the development and implementation of models to study cell-cell contact using substrate grown cells. Here, we summarize the use of membrane permeable supports to study functional responses in appropriately positioned cell types. These models incorporate two or more different cells cultured in physiologically positioned locales on solid substrates. Models incorporating nonadherent cells (e.g., leukocytes) in co-culture with such models also are discussed. Such models have been used extensively to discovery both cell-bound as well as soluble mediators of physiological and pathophysiological processes.

Model Systems to Investigate Neutrophil Adhesion and Chemotaxis

Methods in Molecular Biology™, 2007

Polymorphonuclear neutrophil (PMN) recruitment from the blood stream into surrounding tissues, fo... more Polymorphonuclear neutrophil (PMN) recruitment from the blood stream into surrounding tissues, followed by migration through the tissue with triggered release of oxidative enzymes or eventual clearance from the epithelial surface, involves a regulated series of events central to acute responses in host defense. Accumulations of large numbers of neutrophils within mucosal tissues are pathognomonic features of both acute and chronic inflammatory conditions including sepsis and inflammatory bowel disease, but the precise signals governing neutrophil adhesion and transmigration remain to be fully characterized. Previous chapters examine methods employed for both neutrophil isolation and study of the mechanisms underlying regulation of PMN rolling behavior. Here, we describe in vitro experimental models for the examination of PMN adhesion to endothelial and epithelial monolayers as well as the characterization of signals influencing neutrophil migration, both along acellular matrices and across endothelial and epithelial monolayers, in the physiologically relevant directions. Studies employing these model systems allow further elucidation of the mechanisms governing PMN adhesion and transmigration.

The FASEB Journal, 2007

Migration of neutrophils (PMN) across epithelia is a pathological hallmark of numerous mucosal di... more Migration of neutrophils (PMN) across epithelia is a pathological hallmark of numerous mucosal diseases. Whereas lesions at mucosal surfaces are generally self-limiting, endogenous mechanisms of resolution are incompletely understood. Previous studies revealed that resolvins directly act on PMN to attenuate transendothelial migration, less is known about the influence of resolvins on PMN-epithelial interactions and whether they act on epithelia. We studied the dynamics of resolvin E1 (RvE1) actions on leukocyte transepithelial migration. PMN exposure to RvE1 or chemerin (peptide agonist of ChemR23) reduced transepithelial migration in a concentration-dependent manner. Conversely, activation of epithelial ChemR23 promoted apical clearance of PMN. A nonbiased screen of known PMN ligands expressed on epithelial cells in response to RvE1 revealed selective induction of CD55, an apically expressed antiadhesive molecule. CD55 promoter analysis demonstrated that both RvE1 and chemerin activate the CD55 promoter. Inhibition of CD55 by neutralizing antibody prevented RvE1-dependent augmentation of apical PMN clearance. Taken together these findings implicate a "two-hit" model of inflammatory resolution, whereby activation of the PMN RvE1 receptor attenuates transepithelial migration and subsequent actions on the epithelium promote CD55-dependent clearance of PMN across the epithelial cell surface promoting active inflammatory resolution.

The FASEB Journal, 2005

Sites of inflammation are associated with dramatic shifts in tissue metabolism. Inflammation can ... more Sites of inflammation are associated with dramatic shifts in tissue metabolism. Inflammation can result in significant tissue hypoxia, with resultant induction of hypoxia-responsive genes. Given this association, we hypothesized that neutrophil (PMN) ligands expressed on epithelial cells may be regulated by hypoxia. Initial studies confirmed earlier results that epithelial hypoxia enhances PMN transepithelial migration and promotes apical clearance of PMN from the epithelial surface. A screen of known PMN ligands revealed a surprisingly stable expression pattern in hypoxia. However, this screen identified one gene, CD55, as a highly hypoxia-inducible molecule expressed on the apical membrane of mucosal epithelia. Subsequent studies verified the induction of CD55 mRNA and protein expression by hypoxia. Overexpression of CD55 by transfection in nonhypoxic epithelia resulted in a similar pattern of apical PMN clearance, and peptide mimetics corresponding to the PMN binding site on DAF blocked such apical clearance of PMN. Studies directed at understanding molecular pathways of hypoxia inducibility revealed that a ϳ200 bp region of the CD55 gene conferred hypoxia inducibility for CD55. These studies identified a functional binding site for the transcriptional regulator hypoxia-inducible factor (HIF). Taken together, these results identify HIFdependent induction of epithelial CD55 in the resolution of ongoing inflammation through clearance of apical PMN.-Louis, N. A

Prostaglandins, Leukotrienes and Essential Fatty Acids, 2005

In intact tissues, vascular endothelial cells lie anatomically positioned as the central coordina... more In intact tissues, vascular endothelial cells lie anatomically positioned as the central coordinator of inflammation. Endothelia communicate with underlying cells (e.g. smooth muscle, fibroblasts, epithelia) in ways that both coordinate leukocyte trafficking, and control the composition of the inflammatory microenvironment. Such coordination occurs through both direct communication (e.g. cell adhesion) as well as via soluble mediators liberated at sites of inflammation (e.g. chemokines, cytokines, lipids). Locally generated mediators bind to surface receptors, and mediate both physiologic and pathophysiologic functional responses. Important in this regard, both endothelial and subendothelial cell populations express enzymes capable of utilizing arachidonic acid substrates to generate bioactive lipid mediators (e.g. lipoxygenases, cyclooxygenases). Such lipid mediators can signal via autocrine or paracrine pathways and, depending on the tissue microenvironment, can convey a pro-or anti-inflammatory message. This review will highlight recent studies characterizing inflammatory responses to lipid mediators liberated at sites of inflammation, with a particular emphasis on neutrophil (polymorphonuclear leukocyte or PMN) trafficking.

PLoS ONE, 2012

The intestinal microflora is critical for normal development, with aberrant colonization increasi... more The intestinal microflora is critical for normal development, with aberrant colonization increasing the risk for necrotizing enterocolitis (NEC). In contrast, probiotic bacteria have been shown to decrease its incidence. Multiple pro-and antiinflammatory cytokines have been identified as markers of intestinal inflammation, both in human patients with NEC and in models of immature intestine. Specifically, IL-10 signaling attenuates intestinal responses to gut dysbiosis, and disruption of this pathway exacerbates inflammation in murine models of NEC. However, the effects of probiotics on IL-10 and its signaling pathway, remain poorly defined. Real-time PCR profiling revealed developmental regulation of MIP-2, TNF-a, IL-12, IL-10 and the IL-10R2 subunit of the IL-10 receptor in immature murine colon, while the expression of IL-6 and IL-18 was independent of postnatal age. Enteral administration of the probiotic Lactobacillus rhamnosus GG (LGG) down-regulated the expression of TNF-a and MIP-2 and yet failed to alter IL-10 mRNA and protein expression. LGG did however induce mRNA expression of the IL-10R2 subunit of the IL-10 receptor. IL-10 receptor activation has been associated with signal transducer and activator of transcription (STAT) 3-dependent induction of members of the suppressors of cytokine signaling (SOCS) family. In 2 week-old mice, LGG also induced STAT3 phosphorylation, increased colonic expression of SOCS-3, and attenuated colonic production of MIP-2 and TNF-a. These LGG-dependent changes in phosphoSTAT3, SOCS3, MIP-2 and TNF-a were all inhibited by antibody-mediated blockade of the IL-10 receptor. Thus LGG decreased baseline proinflammatory cytokine expression in the developing colon through upregulation of IL-10 receptor-mediated signaling, most likely due to the combined induction of phospho-STAT3 and SOCS3. Furthermore, LGG-dependent increases in IL-10R2 were associated with reductions in TNF-a, MIP-2 and disease severity in a murine model of intestinal injury in the immature colon.