Laboratory Personnel
Research Fellows
Eric Campbell
Ph.D. 2005 University College Dublin, Ireland
Thomas Weissmuller
M.D. 2003 University of Hamburg, Germany
Simon Keely
Ph.D. 2006 University College, Dublin, Ireland
Karina Irizarry
M.D. 2001 Ponce School of Medicine, Puerto Rico
Chris MacManus
Ph.D. 2005 Queen’s University of Belfast, Northern Ireland
Technical Staff
Melanie Scully
B.A. 2005 Boston University
Topics:
Barrier Function and Disease
Epithelial cells and vascular endothelial cells provide excellent models to study dynamic organization of tissues and organs. In particular, both endothelia and epithelia function as barrier cell types to partition tissue spaces and to allow the regulated transport of solutes and cells. Our laboratory has a particular interest in understanding the contribution of barrier function to disease processes. Recent studies, for example, indicate that during periods of modeled inflammation, epithelial cells lose many classic epithelial features (e.g. barrier function, ion secretion) and garner many qualities indicative of immune accessory cells (e.g. MHC class expression, chemokine / cytokine synthesis). This epithelial "phenotypic switch" contributes to the pathophysiologic mechanisms of intestinal diseases such as ulcerative colitis and Crohn's disease and current studies are directed at developing models to examine potential therapeutic approaches to treat such disorders.
Proposed model of PMN-derived 5’-AMP-mediated decrease in endothelial permeability. PMN-derived 5’-AMP undergoes conversion to adenosine via endothelial 5’-ectonucleotidase (CD73). Adenosine activates the endothelial A2B receptor (A2BR), and via elevated intracellular cAMP levels, decreases endothelial permeability through actions at the endothelial intercellular junction.
Vasodilator-stimulated phosphoprotein (VASP) is associated with zonula occludens-1 (ZO-1) at epithelial tight junctions. Confluent T84 monolayers were stained for VASP (green) and ZO-1 (red) to determine if VASP is localized to the region of the tight junction. Stained monolayers were analyzed at the level of the tight junction by confocal laser scanning microscopy. In T84 cells staining at the level of the tight junction for ZO-1 was observed as the characteristic "chicken-wire" staining pattern. Staining for VASP resulted in a nearly identical staining pattern. Indeed, a computer generated image of specific colocalized areas (yellow) supports VASP localization to the tight junction.
Selected Publications
- Colgan SP, Parkos CA, Matthews JB, D'Andrea L, Awtrey CS, Lichtman A, Delp C, Madara JL: IFN_ induces a cell surface phenotype switch in intestinal epithelia. Am J Physiol 1994; 267: 402-10.
- Zünd G, Madara JL, Dzus AL, Awtrey CS, and Colgan SP: Interleukin 4 and interleukin 13 differentially regulate epithelial chloride secretion. J. Biol. Chem. 1996; 271: 7460-7464.
- Lennon PF, Taylor CT, Stahl GL, Colgan SP: Neutrophil-derived 5’AMP promotes endothelial barrier function via CD73-mediated conversion to adenosine and endothelial A2b receptor activation. J Ex Med 188: 1433-1443, 1998. (download PDF)
- Comerford KM, Lawrence DL, Synnestvedt K, Levi B, and Colgan SP: Role of vasodilator-stimulated phosphoprotein (VASP) in PKA-induced changes in endothelial junctional permeability. FASEB J. 16: 583-585, 2002 (full text published at http://www.fasebj.org/cgi/doi/10.1096/fj.01-0739fje).
- Collard CD, Park KA, Montalto MC, Alipati S, Buras, JA, Stahl GL, and Colgan SP: Neutrophil-derived glutamate regulates vascular endothelial barrier. J. Biol. Chem. 277: 14801-14811, 2002. (download PDF)
Leukocyte Cell-Cell Interactions
The pathological hallmark of many mucosal infectious diseases is the accumulation of inflammatory cells such as neutrophils, macrophages and lymphocytes. At present, the signals and events which elicit recruitment of leukocytes to mucosal sites are not fully understood. Ongoing studies are aimed at determining the cellular and acellular components which contribute to trafficking of leukocytes across cell surfaces such as the intestine during infectious and inflammatory events. Specifically, we have focused on defining epithelial adhesion molecules important in neutrophil-epithelial interactions, and how such interactions might be regulated by soluble mediators present in the tissue microenvironment (chemokines, cytokines, lipids). Understanding basic mechanisms of these events provides the basis for development of specific therapies aimed at treating such inflammatory conditions.
Phases of cytokine actions on acute epithelial infection. Epithelial cells are central to the coordinated response to microorganism infection (phase 1). Rapid induction and release of PMN chemokines (phase 2) elicits the accumulation of PMN within and across the epithelial surface. PMN at the lumenal surface of initiates a Cl- secretiory response and fluid transport which functionally flushes the surface of the epithelium. In phase three of the response, epithelial-derived cytokines (e.g. TNFa) synergize with cytokines liberated in the intraepithelial and lamina propria compartments (e.g. IFNg) to attenuate fluid transport. In phase 4, intraepithelial and lamina propria lymphocyte-derived cytokines (e.g. IFNg, IL-4 and IL-13) induce epithelial lipoxin receptors, the ligation of which results in attenuation of PMN accumulation. In phase 5, lipoxin-induced epithelial BPI provides a protective anti-microbial action for the resolution of acute inflammation.
Multi-step model of neutrophil emigration into and across mucosal epithelial cells. This pathway can be conceptually divided into 5 interdependent events: (a) microvascular emigration, (b) transmigration across the lamina propria matrix, (c) initial interaction with the basal pole of epithelia, (d) transepithelial migration and (e) functional interaction with the apical epithelial membrane. As indicated, these steps are dependent on surface or soluble molecules. A partial list of regulatory mechanisms are also listed.
Selected Publications:
- Colgan SP, Parkos CA, Delp C, Arnaout MA, Madara JL: Neutrophil migration across cultured intestinal epithelial monolayers is modulated by epithelial exposure to IFNg in a highly polarized fashion. J Cell Biol 1993; 120: 785-798.
- Colgan SP, Serhan CN, Parkos CA, Delp C, Madara JL: Lipoxin A4 modulates transmigration of human neutrophils across intestinal epithelial monolayers. J Clin Invest 1993; 92: 75-82.
Colgan SP, Parkos CA, McGuirk DK, Brady HR, Papayianni AA, Frendl G and Madara JL: Receptors involved in carbohydrate-binding modulate epithelial-PMN intxs. J. Biol. Chem. 1995; 270: 10531-10539. - Parkos CA, Colgan SP, Liang T, Nusrat A, Bacarra AE, Carnes DK, Madara JL: CD47 mediates post-adhesive events required for neutrophil migration across polarized intestinal epithelia. J. Cell Biol. 1996; 132: 437-450.
- Bruyninckx WJ, Comerford K, Lawrence DW and Colgan SP: Phosphoinositide 3-kinase modulation of b3-integrin represents an endogenous "braking mechanism" during neutrophil transmatrix migration. Blood 97: 3251-3258, 2001
- Canny G, Levy O, Furuta GT, Narravula S, Sisson RB, Serhan CN, and Colgan SP: Lipid mediator induced expression of bactericidal / permeability-increasing protein (BPI) in human mucosal epithelia. Proc. Nat. Acad. Sci (USA) 99: 3902-7, 2002. (download PDF)
- Lawrence DW, Bruyninckx WJ, Louis NA, Lublin DM, Stahl GL, Parkos CA, and Colgan SP: Anti-adhesive role of apical decay-accelerating factor (DAF, CD55) in human neutrophil transmigration across mucosal epithelia. J. Exp. Med 198: 999-1010, 2003 (Cover image).
Transcriptional Signaling by Hypoxia
Diminished oxygen tension (hypoxia) is a contributing factor to developmental pathways and to a number of disease processes. Basic mechanisms of eukaryotic oxygen sensing and the transcriptional profile of genes induced by cellular hypoxia are not fully elucidated. Current studies are aimed at understanding basic cellular signaling pathways which contribute to transcriptional regulation of gene expression by hypoxia, particularly those associated with the establishment and resolution of inflammation. Specifically, we employ the use of differential mRNA display technologies to profile transcriptional regulation of eukaryotic genes during episodes of diminished oxygen tension. Such studies are aimed at understanding novel crosstalk pathways between different cell types, how such crosstalk may be important during episodes of inflammation, and strategies for development of putative therapeutics.
Model of barrier protection during hypoxia: Epithelial cells form the predominant barrier to the outside world. A number of studies have indicated that hypoxia elicits a transcriptional response associated with the induction of barrier-protective genes, including intestinal trefoil factor (ITF), ecto-5’-nucleotidase (CD73), and MDR1 (for which the gene product if P-glycoprotein). In each case, transcriptional responses are associated with induction of hypoxia-inducible factor-1 (HIF-1). Interestingly, a number of these genes encode proteins targeted to the apical plasma membrane, and indicative of a protective "apical cluster".
Hypothesis: Hypoxia and inflammation are concomitant events. Mucosal organs such as the lung, intestine and kidney are supported by a rich underlying vasculature, and as such, are primary targets for diminished blood flow and hypoxia. Ongoing studies in the laboratory indicate that hypoxia and inflammation are concurrent events. An understanding of the pathways important to hypoxia may lead to the development of novel therapeutics for a number of inflammatory diseases, including those at the mucosal surface.
Selected Publications:
- Colgan SP, Dzus AL, Parkos CA: Hypoxia modulates neutrophil transepithelial migration. J. Ex. Med. 1996; 184: 1003-1015.
- Taylor CT, Furuta GT, Synnestvedt K, and Colgan SP: Phosphorylation-dependent targeting of cAMP response element binding protein (CREB) to the ubiquitin/proteasome pathway in hypoxia. Proc Nat Acad Sci (USA) 97: 12091-12096, 2000. (download PDF)
- Furuta GT, Turner JR, Taylor CT, Hershberg RS, Comerford KM, Narravula S, Podolsky DK and Colgan SP: Hypoxia-inducible factor-1 (HIF-1)-dependent induction of intestinal trefoil factor (ITF) protects barrier function during hypoxia. J Ex Med. 193: 1027-1034, 2001. (download PDF)
- Comerford KM, Wallace TJ, Karhausen J, Louis NA, Montalto MC and Colgan SP: Hypoxia-inducible factor-1 regulation of the multidrug resistance 1(MDR1) gene. Canc. Res. 62: 3387-3394, 2002.
- Synnestvedt K, Furuta GT, Comerford KM, Louis N, Karhausen J, Eltzschig HK, Hansen KR, Thompson LF, Colgan SP: Ecto-5’-nucleotidase (CD73) regulation by hypoxia-inducible factor-1 (HIF-1) mediates permeability changes in intestinal epithelia. J. Clin. Invest. 110: 993-1002, 2002.
- Eltzschig HK, Ibla JC, Furuta GT, Leonard MO, Jacobson KA, Enjyoji K, Robson SC and Colgan SP: Coordinated adenine nucleotide phosphohydrolysis and nucleoside signaling in post-hypoxic endothelium: role of ectonucleotidases and adenosine A2B-receptors. J. Exp. Med. 198: 783-796, 2003. (download PDF)