Edith Porter

Publications of Edith Porter

• Expansion of Paneth cell population in response to enteric Salmonella enterica serovar Typhimurium infection.

  Authors: Nadine R Martinez Rodriguez, Marjannie D Eloi, Alexandria Huynh, Teresa Dominguez, Annie H Cheung Lam, Dayana Carcamo-Molina, Zeina Naser, Robert Desharnais, Nita H Salzman, Edith Porter

  Infection and immunity. 01/2012; 80(1):266-75.

  Paneth cells residing at the base of the small intestinal crypts contribute to the mucosal intestinal first line defense by secreting granules filled with antimicrobial polypeptides includingPaneth cells residing at the base of the small intestinal crypts contribute to the mucosal intestinal first line defense by secreting granules filled with antimicrobial polypeptides including lysozyme. These cells derive from the columnar intestinal stem cell located at position 0 and the transit amplifying cell located at position +4 in the crypts. We have previously shown that Salmonella enterica serovar Typhimurium (ST), a leading cause of gastrointestinal infections in humans, effects an overall reduction of lysozyme in the small intestine. To extend this work, we examined small-intestinal tissue sections at various time points after ST infection to quantify and localize expression of lysozyme and assess Paneth cell abundance, apoptosis, and the expression of Paneth cell differentiation markers. In response to infection with ST, the intestinal Paneth cell-specific lysozyme content, the number of lysozyme-positive Paneth cells, and the number of granules per Paneth cell decreased. However, this was accompanied by increases in the total number of Paneth cells and the frequency of mitotic events in crypts, by increased staining for the proliferation marker PCNA, primarily at the crypt side walls where the transit amplifying cell resides and not at the crypt base, and by apoptotic events in villi. Furthermore, we found a time-dependent upregulation of first β-catenin, followed by EphB3, and lastly Sox9 in response to ST, which was not observed after infection with a Salmonella pathogenicity island 1 mutant deficient in type III secretion. Our data strongly suggest that, in response to ST infection, a Paneth cell differentiation program is initiated that leads to an expansion of the Paneth cell population and that the transit amplifying cell is likely the main progenitor responder. Infection-induced expansion of the Paneth cell population may represent an acute intestinal inflammatory response similar to neutrophilia in systemic infection.

• Evolving neural network optimization of cholesteryl ester separation by reversed-phase HPLC.

  Authors: Michael A Jansen, Jacqueline Kiwata, Jennifer Arceo, Kym F Faull, Grady Hanrahan, Edith Porter

  Analytical and bioanalytical chemistry. 07/2010; 397(6):2367-74.

  Cholesteryl esters have antimicrobial activity and likely contribute to the innate immunity system. Improved separation techniques are needed to characterize these compounds. In this study,Cholesteryl esters have antimicrobial activity and likely contribute to the innate immunity system. Improved separation techniques are needed to characterize these compounds. In this study, optimization of the reversed-phase high-performance liquid chromatography separation of six analyte standards (four cholesteryl esters plus cholesterol and tri-palmitin) was accomplished by modeling with an artificial neural network-genetic algorithm (ANN-GA) approach. A fractional factorial design was employed to examine the significance of four experimental factors: organic component in the mobile phase (ethanol and methanol), column temperature, and flow rate. Three separation parameters were then merged into geometric means using Derringer's desirability function and used as input sources for model training and testing. The use of genetic operators proved valuable for the determination of an effective neural network structure. Implementation of the optimized method resulted in complete separation of all six analytes, including the resolution of two previously co-eluting peaks. Model validation was performed with experimental responses in good agreement with model-predicted responses. Improved separation was also realized in a complex biological fluid, human milk. Thus, the first known use of ANN-GA modeling for improving the chromatographic separation of cholesteryl esters in biological fluids is presented and will likely prove valuable for future investigators involved in studying complex biological samples.

• Antimicrobial lipids: novel innate defense molecules are elevated in sinus secretions of patients with chronic rhinosinusitis.

  Authors: Jivianne T Lee, Mike Jansen, Abebayehu N Yilma, Angels Nguyen, Robert Desharnais, Edith Porter

  American journal of rhinology & allergy. 03/2010; 24(2):99-104.

  Airway secretions possess intrinsic antimicrobial properties that contribute to the innate host defense of the respiratory tract. These microbicidal capabilities have largely been attributed to theAirway secretions possess intrinsic antimicrobial properties that contribute to the innate host defense of the respiratory tract. These microbicidal capabilities have largely been attributed to the presence of antibacterial polypeptides. However, recent investigation has indicated that host-derived lipids including cholesteryl esters also exhibit antimicrobial properties. The purpose of this study was to determine whether sinus secretions contain such antimicrobial lipids and to compare the lipid composition in patients with and without chronic rhinosinusitis (CRS). Maxillary sinus fluid was obtained via antral lavage from subjects with (seven patients) and without (nine patients) a history of CRS. After specimen collection, total lipid was extracted according to Bligh and Dyer (Bligh EG and Dyer WJ, A rapid method of total lipid extraction and purification, Can J Biochem Physiol 37:911-918, 1959) and lipid profiles were obtained by reverse phase high-performance liquid chromatography on an amide-embedded C18 column. In addition, the neutrophil-specific antimicrobial peptides human neutrophil peptides 1-3 (HNP1-3) were quantified by Western immunoblotting. Lipids, including cholesteryl esters, were identified in the maxillary sinus secretions of patients with and without CRS. However, levels of lipid composition differed between the two groups with CRS patients exhibiting greater amounts of all classes of lipids, reaching over 10-fold higher concentration when compared with non-CRS patients. This increase was independent of HNP1-3 content. Sinus secretions of patients with CRS appear to show elevated levels of antimicrobial lipids compared with controls independent from neutrophil influx. This up-regulation suggests that host-derived lipids act as mediators of mucosal immunity in CRS. Further study is necessary to determine if such antimicrobial lipids function alone or synergistically with antibacterial peptides in conferring such inherent microbicidal properties.

• Antibacterial activity of human defensins on anaerobic intestinal bacterial species: a major role of HBD-3.

  Authors: Sabine Nuding, Lutz T Zabel, Corinne Enders, Edith Porter, Klaus Fellermann, Jan Wehkamp, Holger A G Mueller, Eduard F Stange

  Microbes and infection / Institut Pasteur. 02/2009;

  Defensins are natural mucosal antimicrobial peptides and their broad spectrum activity against aerobic or facultative anaerobic bacteria has been well investigated. The aim of this study was toDefensins are natural mucosal antimicrobial peptides and their broad spectrum activity against aerobic or facultative anaerobic bacteria has been well investigated. The aim of this study was to systematically examine the antibacterial activity of the small intestinal Paneth cell-derived a-defensin HD5 and the major colonic beta-defensins HBD-1 - 3 against strict anaerobic intestinal bacteria. The antibacterial activity was assessed with a flow cytometric assay employing a membrane potential sensitive dye as marker for loss of cell viability. The majority of the tested strains belonging to the dominant anaerobe genera of the gut, Bacteroides and Parabacteroides, were only minimally affected by the constitutively expressed defensins HD5 and HBD1. The inducible defensin HBD-2 had a limited antibacterial effect, whereas the inducible HBD-3 exhibited potent activity against most strains. The effect of HBD-3 on Bacteroides sp. appeared to be dependent on the presence of oxygen. Bacteroides fragilis strains isolated from blood during bacteraemia or from extraintestinal infections were more resistant to HBD-3 than strains from the physiological gut flora. Thus, defensin resistance is not only species but also strain specific and may be clinically relevant in the host-bacterial interaction influencing mucosal translocation and systemic infection.

• Membrane-targeted synergistic activity of docosahexaenoic acid and lysozyme against P. aeruginosa.

  Authors: Jose Martinez, Michael Waldon, Qiyu Huang, Sandra Alvarez, Ami Oren, Natalie Sandoval, Ming Du, Feimeng Zhou, Alexandra Zenz, Karl Lohner, Robert Desharnais, Edith Porter

  The Biochemical journal. 01/2009;

  Antimicrobial polypeptides including lysozymes (Ly) have membrane perturbing activity and are well documented effector molecules of innate immunity. In cystic fibrosis, a hereditary disease withAntimicrobial polypeptides including lysozymes (Ly) have membrane perturbing activity and are well documented effector molecules of innate immunity. In cystic fibrosis, a hereditary disease with frequent lung infection with Pseudomonas aeruginosa, the free fatty acid docosahexaenoic acid (DA), but not oleic acid (OA), is decreased and DA supplementation has been shown to improve the clinical condition in these patients. We hypothesized that DA may alone, or in conjunction with Ly, exert antibacterial action against P. aeruginosa. We found that DA and Ly synergistically inhibit the metabolic activity of P. aeruginosa, in contrast to OA. Electron microscopy and equilibrium dialysis suggest that DA accumulates in the bacterial membrane in the presence of Ly. Surface plasmon resonance with live bacteria and differential scanning calorimetry studies with bacterial model membranes reveal that, initially, DA facilitates lysozyme incorporation into the membrane, which in turn allows influx of more DA leading to bacterial cell death. Our study elucidates a molecular basis for the synergistic action of free fatty acids and antimicrobial polypeptides, which may be dysfunctional in cystic fibrosis.

• Lipids including cholesteryl linoleate and cholesteryl arachidonate contribute to the inherent antibacterial activity of human nasal fluid.

  Authors: Thai Q Do, Safiehkhatoon Moshkani, Patricia Castillo, Suda Anunta, Adelina Pogosyan, Annie Cheung, Beth Marbois, Kym F Faull, William Ernst, Su Ming Chiang, Gary Fujii, Catherine F Clarke, Krishna Foster, Edith Porter

  Journal of immunology (Baltimore, Md. : 1950). 10/2008; 181(6):4177-87.

  Mucosal surfaces provide first-line defense against microbial invasion through their complex secretions. The antimicrobial activities of proteins in these secretions have been well delineated, butMucosal surfaces provide first-line defense against microbial invasion through their complex secretions. The antimicrobial activities of proteins in these secretions have been well delineated, but the contributions of lipids to mucosal defense have not been defined. We found that normal human nasal fluid contains all major lipid classes (in micrograms per milliliter), as well as lipoproteins and apolipoprotein A-I. The predominant less polar lipids were myristic, palmitic, palmitoleic, stearic, oleic, and linoleic acid, cholesterol, and cholesteryl palmitate, cholesteryl linoleate, and cholesteryl arachidonate. Normal human bronchioepithelial cell secretions exhibited a similar lipid composition. Removal of less-polar lipids significantly decreased the inherent antibacterial activity of nasal fluid against Pseudomonas aeruginosa, which was in part restored after replenishing the lipids. Furthermore, lipids extracted from nasal fluid exerted direct antibacterial activity in synergism with the antimicrobial human neutrophil peptide HNP-2 and liposomal formulations of cholesteryl linoleate and cholesteryl arachidonate were active against P. aeruginosa at physiological concentrations as found in nasal fluid and exerted inhibitory activity against other Gram-negative and Gram-positive bacteria. These data suggest that host-derived lipids contribute to mucosal defense. The emerging concept of host-derived antimicrobial lipids unveils novel roads to a better understanding of the immunology of infectious diseases.

• Reduced Paneth cell alpha-defensins in ileal Crohn's disease.

  Authors: Jan Wehkamp, Nita H Salzman, Edith Porter, Sabine Nuding, Michael Weichenthal, Robert E. Petras, Bo Shen, Elke Schaeffeler, Matthias Schwab, Rose Linzmeier, Ryan W Feathers, Hiutung Chu, Heriberto Lima, Klaus Fellermann, Tomas Ganz, Eduard F Stange, Charles L Bevins

  Proceedings of the National Academy of Sciences of the United States of America. 01/2006; 102(50):18129-34.

  The pathogenesis of Crohn's disease (CD), an idiopathic inflammatory bowel disease, is attributed, in part, to intestinal bacteria that may initiate and perpetuate mucosal inflammation in geneticallyThe pathogenesis of Crohn's disease (CD), an idiopathic inflammatory bowel disease, is attributed, in part, to intestinal bacteria that may initiate and perpetuate mucosal inflammation in genetically susceptible individuals. Paneth cells (PC) are the major source of antimicrobial peptides in the small intestine, including human alpha-defensins HD5 and HD6. We tested the hypothesis that reduced expression of PC alpha-defensins compromises mucosal host defenses and predisposes patients to CD of the ileum. We report that patients with CD of the ileum have reduced antibacterial activity in their intestinal mucosal extracts. These specimens also showed decreased expression of PC alpha-defensins, whereas the expression of eight other PC products either remained unchanged or increased when compared with controls. The specific decrease of alpha-defensins was independent of the degree of inflammation in the specimens and was not observed in either CD of the colon, ulcerative colitis, or pouchitis. The functional consequence of alpha-defensin expression levels was examined by using a transgenic mouse model, where we found changes in HD5 expression levels, comparable to those observed in CD, had a pronounced impact on the luminal microbiota. Thus, the specific deficiency of PC defensins that characterizes ileal CD may compromise innate immune defenses of the ileal mucosa and initiate and/or perpetuate this disease.

• Distinct defensin profiles in Neisseria gonorrhoeae and Chlamydia trachomatis urethritis reveal novel epithelial cell-neutrophil interactions.

  Authors: Edith Porter, Huixia Yang, Sujata Yavagal, Gloria C Preza, Omar Murillo, Heriberto Lima, Sheila Greene, Laily Mahoozi, Marcia Klein-Patel, Gill Diamond, Sunita Gulati, Tomas Ganz, Peter A Rice, Alison J Quayle

  Infection and immunity. 08/2005; 73(8):4823-33.

  Defensins are key participants in mucosal innate defense. The varied antimicrobial activity and differential distribution of defensins at mucosal sites indicate that peptide repertoires are tailoredDefensins are key participants in mucosal innate defense. The varied antimicrobial activity and differential distribution of defensins at mucosal sites indicate that peptide repertoires are tailored to site-specific innate defense requirements. Nonetheless, few studies have investigated changes in peptide profiles and function after in vivo pathogen challenge. Here, we determined defensin profiles in urethral secretions of healthy men and men with Chlamydia trachomatis- and Neisseria gonorrhoeae-mediated urethritis by immunoblotting for the epithelial defensins HBD1, HBD2, and HD5 and the neutrophil defensins HNP1 to -3 (HNP1-3). HBD1 was not detectable in secretions, and HBD2 was only induced in a small proportion of the urethritis patients; however, HD5 and HNP1-3 were increased in C. trachomatis infection and significantly elevated in N. gonorrhoeae infection. When HNP1-3 levels were low, HD5 appeared mostly as the propeptide; however, when HNP1-3 levels were >10 microg/ml, HD5 was proteolytically processed, suggesting neutrophil proteases might contribute to HD5 processing. HD5 and HNP1-3 were bactericidal against C. trachomatis and N. gonorrhoeae, but HD5 activity was dependent upon N-terminal processing of the peptide. In vitro proteolysis of proHD5 by neutrophil proteases and analysis of urethral secretions by surface-enhanced laser desorption ionization substantiated that neutrophils contribute the key convertases for proHD5 in the urethra during these infections. This contrasts with the small intestine, where Paneth cells secrete both proHD5 and its processing enzyme, trypsin. In conclusion, we describe a unique defensin expression repertoire in response to inflammatory sexually transmitted infections and a novel host defense mechanism wherein epithelial cells collaborate with neutrophils to establish an antimicrobial barrier during infection.

• Use of an efflux pump inhibitor to determine the prevalence of efflux pump-mediated fluoroquinolone resistance and multidrug resistance in Pseudomonas aeruginosa.

  Authors: Jane Kriengkauykiat, Edith Porter, Olga Lomovskaya, Annie Wong-Beringer

  Antimicrobial agents and chemotherapy. 03/2005; 49(2):565-70.

  Fluoroquinolone-resistance in Pseudomonas aeruginosa may be due to efflux pump overexpression (EPO) and/or target mutations. EPO can result in multidrug resistance (MDR) due to broad substrateFluoroquinolone-resistance in Pseudomonas aeruginosa may be due to efflux pump overexpression (EPO) and/or target mutations. EPO can result in multidrug resistance (MDR) due to broad substrate specificity of the pumps. MC-04,124, an efflux pump inhibitor (EPI) shown to significantly potentiate activity of levofloxacin in P. aeruginosa, was used to examine the prevalence of EPO in clinical isolates. MICs were determined for ciprofloxacin, levofloxacin, moxifloxacin, and gatifloxacin with or without EPI and for other antipseudomonal agents by using broth microdilution against P. aeruginosa isolates from adults (n = 119) and children (n = 24). The prevalence of the EPO phenotype (>/=8-fold MIC decrease when tested with EPI) was compared among subgroups with different resistance profiles. The EPO phenotype was more prevalent among levofloxacin-resistant than levofloxacin-sensitive strains (61%, 48/79 versus 9%, 6/64). EPO was present in 60% of fluoroquinolone-resistant strains without cross-resistance, while it was present at variable frequencies among strains with cross-resistance to other agents: piperacillin-tazobactam (86%), ceftazidime (76%), cefepime (65%), imipenem (56%), gentamicin (55%), tobramycin (48%), and amikacin (27%). The magnitude of MIC decrease with an EPI paralleled the frequency of which the EPO phenotype was observed in different subgroups. EPI reduced the levofloxacin MIC by as much as 16-fold in eight strains for which MICs were 128 microg/ml. Efflux-mediated resistance appears to contribute significantly to fluoroquinolone resistance and MDR in P. aeruginosa. Our data support the fact that increased fluoroquinolone usage can negatively impact susceptibility of P. aeruginosa to multiple classes of antipseudomonal agents.

• Paneth cell trypsin is the processing enzyme for human defensin-5.

  Authors: Dipankar Ghosh, Edith Porter, Bo Shen, Sarah K Lee, Dennis Wilk, Judith Drazba, Satya P Yadav, John W Crabb, Tomas Ganz, Charles L Bevins

  Nature immunology. 07/2002; 3(6):583-90.

  The antimicrobial peptide human alpha-defensin 5 (HD5) is expressed in Paneth cells, secretory epithelial cells in the small intestine. Unlike other characterized defensins, HD5 is stored inThe antimicrobial peptide human alpha-defensin 5 (HD5) is expressed in Paneth cells, secretory epithelial cells in the small intestine. Unlike other characterized defensins, HD5 is stored in secretory vesicles as a propeptide. The storage quantities of HD5 are approximately 90 450 microg per cm2 of mucosal surface area, which is sufficient to generate microbicidal concentrations in the intestinal lumen. HD5 peptides isolated from the intestinal lumen are proteolytically processed forms--HD5(56-94) and HD5(63-94)--that are cleaved at the Arg55-Ala56 and Arg62-Thr63 sites, respectively. We show here that a specific pattern of trypsin isozymes is expressed in Paneth cells, that trypsin colocalizes with HD5 and that this protease can efficiently cleave HD5 propeptide to forms identical to those isolated in vivo. By acting as a prodefensin convertase in human Paneth cells, trypsin is involved in the regulation of innate immunity in the small intestine.

• Antibacterial activity of human defensins on anaerobic intestinal bacterial species: a major role of HBD-3

  Authors: Sabine Nuding, Lutz T. Zabel, Corinne Enders, Edith Porter, Klaus Fellermann, Jan Wehkamp, Holger A.G. Mueller, Eduard F. Stange

  Microbes and Infection.

  Defensins are natural mucosal antimicrobial peptides and their broad spectrum activity against aerobic or facultative anaerobic bacteria has been well investigated. The aim of this study was toDefensins are natural mucosal antimicrobial peptides and their broad spectrum activity against aerobic or facultative anaerobic bacteria has been well investigated. The aim of this study was to systematically examine the antibacterial activity of the small intestinal Paneth cell derived α-defensin HD5 and the major colonic β-defensins HBD-1–3 against strict anaerobic intestinal bacteria. The antibacterial activity was assessed with a flow cytometric assay employing a membrane potential sensitive dye as marker for loss of cell viability. The majority of the tested strains belonging to the dominant anaerobe genera of the gut, Bacteroides and Parabacteroides, were only minimally affected by the constitutively expressed defensins HD5 and HBD-1. The inducible defensin HBD-2 had a limited antibacterial effect, whereas the inducible HBD-3 exhibited potent activity against most strains. The effect of HBD-3 on Bacteroides sp. appeared to be dependent on the presence of oxygen. Bacteroides fragilis strains isolated from blood during bacteremia or from extraintestinal infections were more resistant to HBD-3 than strains from the physiological gut flora. Thus, defensin resistance is not only species- but also strain-specific and may be clinically relevant in the host–bacteria interaction influencing mucosal translocation and systemic infection.