Nita H Salzman

Publications of Nita H Salzman

• A Novel IL-10-Independent Regulatory Role for B Cells in Suppressing Autoimmunity by Maintenance of Regulatory T Cells via GITR Ligand.

  Authors: Avijit Ray, Sreemanti Basu, Calvin B Williams, Nita H Salzman, Bonnie N Dittel

  Journal of immunology (Baltimore, Md. : 1950). 02/2012; 188(7):3188-98.

  B cells are important for the regulation of autoimmune responses. In experimental autoimmune encephalomyelitis (EAE), B cells are required for spontaneous recovery in acute models. Production ofB cells are important for the regulation of autoimmune responses. In experimental autoimmune encephalomyelitis (EAE), B cells are required for spontaneous recovery in acute models. Production of IL-10 by regulatory B cells has been shown to modulate the severity EAE and other autoimmune diseases. Previously, we suggested that B cells regulated the number of CD4(+)Foxp3(+) T regulatory cells (Treg) in the CNS during EAE. Because Treg suppress autoimmune responses, we asked whether B cells control autoimmunity by maintenance of Treg numbers. B cell deficiency achieved either genetically (μMT) or by depletion with anti-CD20 resulted in a significant reduction in the number of peripheral but not thymic Treg. Adoptive transfer of WT B cells into μMT mice restored both Treg numbers and recovery from EAE. When we investigated the mechanism whereby B cells induce the proliferation of Treg and EAE recovery, we found that glucocorticoid-induced TNF ligand, but not IL-10, expression by B cells was required. Of clinical significance is the finding that anti-CD20 depletion of B cells accelerated spontaneous EAE and colitis. Our results demonstrate that B cells play a major role in immune tolerance required for the prevention of autoimmunity by maintenance of Treg via their expression of glucocorticoid-induced TNFR ligand.

• 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.

• The potter's wheel: the host's role in sculpting its microbiota.

  Authors: Charles L Bevins, Nita H Salzman

  Cellular and molecular life sciences : CMLS. 11/2011; 68(22):3675-85.

  Animals, ranging from basal metazoans to primates, are host to complex microbial ecosystems; engaged in a symbiotic relationship that is essential for host physiology and homeostasis. EpithelialAnimals, ranging from basal metazoans to primates, are host to complex microbial ecosystems; engaged in a symbiotic relationship that is essential for host physiology and homeostasis. Epithelial surfaces vary in the composition of colonizing microbiota as one compares anatomic sites, developmental stages and species origin. Alterations of microbial composition likely contribute to susceptibility to several distinct diseases. The forces that shape the colonizing microbial composition are the focus of much current investigation, and it is evident that there are pressures exerted both by the host and the external environment to mold these ecosystems. The focus of this review is to discuss recent studies that demonstrate the critical importance of host factors in selecting for its microbiome. Greater insight into host-microbiome interactions will be essential for understanding homeostasis at mucosal surfaces, and developing useful interventions when homeostasis is disrupted.

• A requisite role for induced regulatory T cells in tolerance based on expanding antigen receptor diversity.

  Authors: Dipica Haribhai, Jason B Williams, Shuang Jia, Derek Nickerson, Erica G Schmitt, Brandon Edwards, Jennifer Ziegelbauer, Maryam Yassai, Shun-Hwa Li, Lance M Relland, Petra M Wise, Andrew Chen, Yu-Qian Zheng, Pippa M Simpson, Jack Gorski, Nita H Salzman, Martin J Hessner, Talal A Chatila, Calvin B Williams

  Immunity. 06/2011; 35(1):109-22.

  Although both natural and induced regulatory T (nTreg and iTreg) cells can enforce tolerance, the mechanisms underlying their synergistic actions have not been established. We examined the functionsAlthough both natural and induced regulatory T (nTreg and iTreg) cells can enforce tolerance, the mechanisms underlying their synergistic actions have not been established. We examined the functions of nTreg and iTreg cells by adoptive transfer immunotherapy of newborn Foxp3-deficient mice. As monotherapy, only nTreg cells prevented disease lethality, but did not suppress chronic inflammation and autoimmunity. Provision of Foxp3-sufficient conventional T cells with nTreg cells reconstituted the iTreg pool and established tolerance. In turn, acute depletion of iTreg cells in rescued mice resulted in weight loss and inflammation. Whereas the transcriptional signatures of nTreg and in vivo-derived iTreg cells were closely matched, there was minimal overlap in their T cell receptor (TCR) repertoires. Thus, iTreg cells are an essential nonredundant regulatory subset that supplements nTreg cells, in part by expanding TCR diversity within regulatory responses.

• Targeted intestinal epithelial deletion of the chemokine receptor CXCR4 reveals important roles for extracellular-regulated kinase-1/2 in restitution.

  Authors: Noah P Zimmerman, Rebecca A Vongsa, Sheena L Faherty, Nita H Salzman, Michael B Dwinell

  Laboratory investigation; a journal of technical methods and pathology. 05/2011; 91(7):1040-55.

  Barrier defects and/or alterations in the ability of the gut epithelium to repair itself are critical etiological mechanisms of gastrointestinal disease. Our ongoing studies indicate that theBarrier defects and/or alterations in the ability of the gut epithelium to repair itself are critical etiological mechanisms of gastrointestinal disease. Our ongoing studies indicate that the chemokine receptor CXCR4 and its cognate ligand CXCL12 regulate intestinal-epithelial barrier maturation and restitution in cell culture models. Gene-deficient mice lacking CXCR4 expression specifically by the cells of the intestinal epithelium were used to test the hypothesis that CXCR4 regulates mucosal barrier integrity in vivo. Epithelial expression of CXCR4 was assessed by RT-PCR, Southern blot, immunoblot and immunohistochemistry. In vivo wounding assays were performed by addition of 3% dextran sodium sulfate (DSS) in drinking water for 5 days. Intestinal damage and DAI scores were assessed by histological examination. Extracellular-regulated kinase (ERK) phosphorylation was assessed in vivo by immunoblot and immunofluorescence. CXCR4 knockdown cells were established using a lentiviral approach and ERK phosphorylation was assessed. Consistent with targeted roles in restitution, epithelium from patients with inflammatory bowel disease indicated that CXCR4 and CXCL12 expression was stable throughout the human colonic epithelium. Conditional CXCR4-deficient mice developed normally, with little phenotypic differences in epithelial morphology, proliferation or migration. Re-epithelialization was absent in CXCR4 conditional knockout mice following acute DSS-induced inflammation. In contrast, heterozygous CXCR4-depleted mice displayed significant improvement in epithelial ulcer healing in acute and chronic inflammation. Mucosal injury repair was correlated with ERK1/2 activity and localization along the crypt-villus axis, with heterozygous mice characterized by increased ERK1/2 activation. Lentiviral depletion of CXCR4 in IEC-6 cells similarly altered ERK1/2 activity and prevented chemokine-stimulated migration. Taken together, these data indicate that chemokine receptors participate in epithelial barrier responses through coordination of the ERK1/2 signaling pathway.

• Paneth cells, antimicrobial peptides and maintenance of intestinal homeostasis.

  Authors: Charles L Bevins, Nita H Salzman

  Nature reviews. Microbiology. 03/2011; 9(5):356-68.

  Building and maintaining a homeostatic relationship between a host and its colonizing microbiota entails ongoing complex interactions between the host and the microorganisms. The mucosal immuneBuilding and maintaining a homeostatic relationship between a host and its colonizing microbiota entails ongoing complex interactions between the host and the microorganisms. The mucosal immune system, including epithelial cells, plays an essential part in negotiating this equilibrium. Paneth cells (specialized cells in the epithelium of the small intestine) are an important source of antimicrobial peptides in the intestine. These cells have become the focus of investigations that explore the mechanisms of host-microorganism homeostasis in the small intestine and its collapse in the processes of infection and chronic inflammation. In this Review, we provide an overview of the intestinal microbiota and describe the cell biology of Paneth cells, emphasizing the composition of their secretions and the roles of these cells in intestinal host defence and homeostasis. We also highlight the implications of Paneth cell dysfunction in susceptibility to chronic inflammatory bowel disease.

• Microbiota-immune system interaction: an uneasy alliance.

  Authors: Nita H Salzman

  Current opinion in microbiology. 10/2010; 14(1):99-105.

  An estimated 100 trillion microbes colonize human beings, with the majority of organisms residing in the intestines. This microbiota impacts host nutrition, protection, and gut development.An estimated 100 trillion microbes colonize human beings, with the majority of organisms residing in the intestines. This microbiota impacts host nutrition, protection, and gut development. Alterations in microbiota composition are associated with susceptibility to various infectious and inflammatory gut diseases. The mucosal surface is not a static barrier that simply prevents microbial invasion but a critical interface for microbiota-immune system interactions. Recent work suggests that dynamic interactions between microbes and the host immune system at the mucosal surface inform immune responses both locally and systemically. This review focuses on intestinal microbiota-immune interactions leading to intestinal homeostasis, and show that these interactions at the GI mucosal surface are critical for driving both protective and pathological immune responses systemically.

• Induction and rescue of Nod2-dependent Th1-driven granulomatous inflammation of the ileum.

  Authors: Amlan Biswas, Yuen-Joyce Liu, Liming Hao, Atsushi Mizoguchi, Nita H Salzman, Charles L Bevins, Koichi S Kobayashi

  Proceedings of the National Academy of Sciences of the United States of America. 08/2010; 107(33):14739-44.

  Mutations in the NOD2 gene are strong genetic risk factors for ileal Crohn's disease. However, the mechanism by which these mutations predispose to intestinal inflammation remains a subject ofMutations in the NOD2 gene are strong genetic risk factors for ileal Crohn's disease. However, the mechanism by which these mutations predispose to intestinal inflammation remains a subject of controversy. We report that Nod2-deficient mice inoculated with Helicobacter hepaticus, an opportunistic pathogenic bacterium, developed granulomatous inflammation of the ileum, characterized by an increased expression of Th1-related genes and inflammatory cytokines. The Peyer's patches and mesenteric lymph nodes were markedly enlarged with expansion of IFN-gamma-producing CD4 and CD8 T cells. Rip2-deficient mice exhibited a similar phenotype, suggesting that Nod2 function likely depends on the Rip2 kinase in this model. Transferring wild-type bone marrow cells into irradiated Nod2-deficient mice did not rescue the phenotype. However, restoring crypt antimicrobial function of Nod2-deficient mice by transgenic expression of alpha-defensin in Paneth cells rescued the Th1 inflammatory phenotype. Therefore, through the regulation of intestinal microbes, Nod2 function in nonhematopoietic cells of the small intestinal crypts is critical for protecting mice from a Th1-driven granulomatous inflammation in the ileum. The model may provide insight into Nod2 function relevant to inflammation of ileal Crohn's disease.

• Paneth cell defensins and the regulation of the microbiome: détente at mucosal surfaces.

  Authors: Nita H Salzman

  Gut microbes. 01/2010; 1(6):401-6.

  Recently, our laboratory demonstrated that Paneth cell defensins, innate antimicrobial peptides that contribute to mucosal host defense, are able to regulate the composition of the intestinalRecently, our laboratory demonstrated that Paneth cell defensins, innate antimicrobial peptides that contribute to mucosal host defense, are able to regulate the composition of the intestinal bacterial microbiome. Using complementary mouse models of defensin deficiency (MMP7(-/-)) and surplus (HD5(+/+)), we noted defensin-dependent reciprocal shifts in the dominant bacterial species of the small intestine, without changes in total bacterial numbers. In addition, mice that expressed HD5 showed a significant loss of segemented filamentous bacteria (SFB), resulting in reduced numbers of Th17 cells in the lamina propria. This data showed a novel role for PC defensins in intestinal homeostasis, by regulation of the small intestinal microbiome. The microbiome plays an essential role in mediating host physiology, metabolism and immune response. The ability of PC defensins to regulate the composition of the biome suggests a much broader importance of these innate immune effectors than previously considered. In this addendum, the role of PC defensins in the regulation of the intestinal microbiome is reviewed, and discussed in the context of recent evidence that highlights the important role of PCs and defensins in the pathophysiology of inflammatory bowel disease.

• Enteric defensins are essential regulators of intestinal microbial ecology.

  Authors: Nita H Salzman, Kuiechun Hung, Dipica Haribhai, Hiutung Chu, Jenny Karlsson-Sjöberg, Elad Amir, Paul Teggatz, Melissa Barman, Michael Hayward, Daniel Eastwood, Maaike Stoel, Yanjiao Zhou, Erica Sodergren, George M Weinstock, Charles L Bevins, Calvin B Williams, Nicolaas A Bos

  Nature immunology. 10/2009;

  Antimicrobial peptides are important effectors of innate immunity throughout the plant and animal kingdoms. In the mammalian small intestine, Paneth cell alpha-defensins are antimicrobial peptidesAntimicrobial peptides are important effectors of innate immunity throughout the plant and animal kingdoms. In the mammalian small intestine, Paneth cell alpha-defensins are antimicrobial peptides that contribute to host defense against enteric pathogens. To determine if alpha-defensins also govern intestinal microbial ecology, we analyzed the intestinal microbiota of mice expressing a human alpha-defensin gene (DEFA5) and in mice lacking an enzyme required for the processing of mouse alpha-defensins. In these complementary models, we detected significant alpha-defensin-dependent changes in microbiota composition, but not in total bacterial numbers. Furthermore, DEFA5-expressing mice had striking losses of segmented filamentous bacteria and fewer interleukin 17 (IL-17)-producing lamina propria T cells. Our data ascribe a new homeostatic role to alpha-defensins in regulating the makeup of the commensal microbiota.

• Prolonged Impact of Antibiotics on Intestinal Microbial Ecology and Susceptibility to Enteric Salmonella Infection.

  Authors: Amy Croswell, Elad Amir, Paul Teggatz, Melissa Barman, Nita H Salzman

  Infection and immunity. 05/2009;

  The impact of antibiotics on the host's protective microbiota, and resulting increased susceptibility to mucosal infection are poorly understood. In this study, antibiotic regimens commonly appliedThe impact of antibiotics on the host's protective microbiota, and resulting increased susceptibility to mucosal infection are poorly understood. In this study, antibiotic regimens commonly applied in murine enteritis models are used to examine the impact of antibiotics on the intestinal microbiota, the time course of recovery of the biota, and the resulting susceptibility to enteric Salmonella infection. Molecular analysis of the microbiota showed that antibiotic treatment has impact on the colonization of the murine gut that is site and antibiotic dependent. While combinations of antibiotics were able to eliminate culturable bacteria, none of the antibiotic treatments were effective at sterilizing the intestinal tract. Recovery of total bacterial numbers occurs within one week after antibiotic withdrawal, but alterations in specific bacterial groups persist for several weeks. Increased Salmonella translocation associated with antibiotic pre-treatment corrects rapidly in association with the recovery of the most dominant bacterial group, which parallels recovery of total bacterial numbers. However susceptibility to intestinal colonization and mucosal inflammation persists when mice are infected several weeks after withdrawal of antibiotics, correlating with subtle alterations in the intestinal microbiome involving alteration of specific bacterial groups. These results show that the colonizing microbiota are integral to mucosal host protection, that specific features of the microbiome impact different aspects of enteric Salmonella pathogenesis, and that antibiotics can have prolonged deleterious effects on intestinal colonization resistance.

• A central role for induced regulatory T cells in tolerance induction in experimental colitis.

  Authors: Dipica Haribhai, Wen Lin, Brandon Edwards, Jennifer Ziegelbauer, Nita H Salzman, Marc R Carlson, Shun-Hwa Li, Pippa M Simpson, Talal A Chatila, Calvin B Williams

  Journal of immunology (Baltimore, Md. : 1950). 04/2009; 182(6):3461-8.

  In addition to thymus-derived or natural T regulatory (nT(reg)) cells, a second subset of induced T regulatory (iT(reg)) cells arises de novo from conventional CD4(+) T cells in the periphery. TheIn addition to thymus-derived or natural T regulatory (nT(reg)) cells, a second subset of induced T regulatory (iT(reg)) cells arises de novo from conventional CD4(+) T cells in the periphery. The function of iT(reg) cells in tolerance was examined in a CD45RB(high)CD4(+) T cell transfer model of colitis. In situ-generated iT(reg) cells were similar to nT(reg) cells in their capacity to suppress T cell proliferation in vitro and their absence in vivo accelerated bowel disease. Treatment with nT(reg) cells resolved the colitis, but only when iT(reg) cells were also present. Although iT(reg) cells required Foxp3 for suppressive activity and phenotypic stability, their gene expression profile was distinct from the established nT(reg) "genetic signature," indicative of developmental and possibly mechanistic differences. These results identified a functional role for iT(reg) cells in vivo and demonstrated that both iT(reg) and nT(reg) cells can act in concert to maintain tolerance.

• Negative interactions with the microbiota: IBD.

  Authors: Nita H Salzman, Charles L Bevins

  Advances in experimental medicine and biology. 02/2008; 635:67-78.

  Mucosal surfaces are colonized by a complex microbiota that provides beneficial functions under normal physiological conditions, but is capable of contributing to chronic inflammatory disease inMucosal surfaces are colonized by a complex microbiota that provides beneficial functions under normal physiological conditions, but is capable of contributing to chronic inflammatory disease in susceptible individuals. Of the mucosal tissues, the mammalian intestine harbors an especially high number of microbes with a remarkable diversity. Inflammatory bowel disease (IBD) is a group of chronic relapsinginflammatory disorders of the intestinal mucosa. Evidence from human studies and animal models provides compelling support that intestinal microbes play a key role in disease pathogenesis. While the existence a specific causative pathogen is possible, it appears more likely that intestinal microbes normally present as commensal microbiota may trigger inflammation and perpetuate disease in genetically susceptible individuals. There may be also a shift in the makeup of the commensal flora to a nonphysiologic composition that is more prone to disease (termed dysbiosis). Evidence supports that genetic susceptibility stems from one or more defects in mucosal immune functions, including microbe recognition, barrier function, intercellular communication and antimicrobial effector mechanisms. It is quite plausible to imagine that the chronic inflammation of IBD may in some cases be a normal immune response to an abnormal adherent invasive microbiota and in other cases an over exuberant immune response to an otherwise normal commensal microbiota.

• A Francisella tularensis Schu S4 purine auxotroph is highly attenuated in mice but offers limited protection against homologous intranasal challenge.

  Authors: Roger D Pechous, Travis R McCarthy, Nrusingh P Mohapatra, Shilpa Soni, Renee M Penoske, Nita H Salzman, Dara W Frank, John S Gunn, Thomas C Zahrt

  PLoS ONE. 02/2008; 3(6):e2487.

  BACKGROUND: Francisella tularensis is a gram-negative coccobacillus that causes the febrile illness tularemia. Subspecies that are pathogenic for humans include those comprising the type ABACKGROUND: Francisella tularensis is a gram-negative coccobacillus that causes the febrile illness tularemia. Subspecies that are pathogenic for humans include those comprising the type A (subspecies tularensis) or type B (subspecies holarctica) biovars. An attenuated live vaccine strain (LVS) developed from a type B isolate has previously been used to vaccinate at-risk individuals, but offers limited protection against high dose (>1000 CFUs) challenge with type A strains delivered by the respiratory route. Due to differences between type A and type B F. tularensis strains at the genetic level, it has been speculated that utilization of an attenuated type A strain as a live vaccine might offer better protection against homologous respiratory challenge compared with LVS. Here, we report the construction and characterization of an unmarked Delta purMCD mutant in the highly virulent type A strain Schu S4. METHODOLOGY/PRINCIPAL FINDINGS: Growth of Schu S4 Delta purMCD was severely attenuated in primary human peripheral blood monocyte-derived macrophages and in the A549 human lung epithelial cell line. The Schu S4 Delta purMCD mutant was also highly attenuated in mice when delivered via either the intranasal or intradermal infection route. Mice vaccinated intranasally with Schu S4 Delta purMCD were well protected against high dose intradermal challenge with virulent type A or type B strains of F. tularensis. However, intranasal vaccination with Schu S4 Delta purMCD induced tissue damage in the lungs, and conferred only limited protection against high dose Schu S4 challenge delivered by the same route. The level of protection observed was similar to that conferred following vaccination with wild-type LVS or the analogous LVS Delta purMCD mutant. CONCLUSIONS/SIGNIFICANCE: Collectively, these results argue that development of the next generation live attenuated vaccine for Francisella should be based on use of the less pathogenic type B biovar rather than the more reactogenic type A biovar.

• Paneth cells, defensins, and the commensal microbiota: a hypothesis on intimate interplay at the intestinal mucosa.

  Authors: Nita H Salzman, Mark A Underwood, Charles L Bevins

  Seminars in immunology. 05/2007; 19(2):70-83.

  Mucosal surfaces are colonized by a diverse and dynamic microbiota. Much investigation has focused on bacterial colonization of the intestine, home to the vast majority of this microbiota.Mucosal surfaces are colonized by a diverse and dynamic microbiota. Much investigation has focused on bacterial colonization of the intestine, home to the vast majority of this microbiota. Experimental evidence has highlighted that these colonizing microbes are essential to host development and homeostasis, but less is known about host factors that may regulate the composition of this ecosystem. While evidence shows that IgA has a role in shaping this microbiota, it is likely that effector molecules of the innate immune system are also involved. One hypothesis is that gene-encoded antimicrobial peptides, key elements of innate immunity throughout nature, have an essential role in this regulation. These effector molecules characteristically have activity against a broad spectrum of bacteria and other microbes. At mucosal surfaces, antimicrobial peptides may affect the numbers and/or composition of the colonizing microbiota. In humans and other mammals, defensins are a predominant class of antimicrobial peptides. In the small intestine, Paneth cells (specialized secretory epithelial cells) produce high quantities of defensins and several other antibiotic peptides and proteins. Data from murine models indicate that Paneth cell defensins play a pivotal role in defense from food and water-borne pathogens in the intestinal lumen. Recent studies in humans provide evidence that reduced Paneth cell defensin expression may be a key pathogenic factor in ileal Crohn's disease, a subgroup of inflammatory bowel disease (IBD), and changes in the colonizing microbiota may mediate this pathogenic mechanism. It is also possible that low levels of Paneth cell defensins, characteristic of normal intestinal development, may predispose premature neonates to necrotizing enterocolitis (NEC) through similar close links with the composition of the intestinal microbiota. Future studies to further define mechanisms by which defensins and other host factors regulate the composition of the intestinal microbiota will likely provide new insights into intestinal homeostasis and new therapeutic strategies for inflammatory and infectious diseases of the bowel.

• 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.

• Protection against enteric salmonellosis in transgenic mice expressing a human intestinal defensin.

  Authors: Nita H Salzman, Dipankar Ghosh, Kenneth M Huttner, Yvonne Paterson, Charles L Bevins

  Nature. 05/2003; 422(6931):522-6.

  Genetically encoded antibiotic peptides are evolutionarily ancient and widespread effector molecules of immune defence. Mammalian defensins, one subset of such peptides, have been implicated in theGenetically encoded antibiotic peptides are evolutionarily ancient and widespread effector molecules of immune defence. Mammalian defensins, one subset of such peptides, have been implicated in the antimicrobial defence capacity of phagocytic leukocytes and various epithelial cells, but direct evidence of the magnitude of their in vivo effects have not been clearly demonstrated. Paneth cells, specialized epithelia of the small intestinal crypt, secrete abundant alpha-defensins and other antimicrobial polypeptides including human defensin 5 (HD-5; also known as DEFA5). Although antibiotic activity of HD-5 has been demonstrated in vitro, functional studies of HD-5 biology have been limited by the lack of in vivo models. To study the in vivo role of HD-5, we developed a transgenic mouse model using a 2.9-kilobase HD-5 minigene containing two HD-5 exons and 1.4 kilobases of 5'-flanking sequence. Here we show that HD-5 expression in these mice is specific to Paneth cells and reflects endogenous enteric defensin gene expression. The storage and processing of transgenic HD-5 also matches that observed in humans. HD-5 transgenic mice were markedly resistant to oral challenge with virulent Salmonella typhimurium. These findings provide support for a critical in vivo role of epithelial-derived defensins in mammalian host defence.

• Enteric salmonella infection inhibits Paneth cell antimicrobial peptide expression.

  Authors: Nita H Salzman, Margaret M Chou, Hendrik de Jong, Lide Liu, Edith M Porter, Yvonne Paterson

  Infection and immunity. 04/2003; 71(3):1109-15.

  Paneth cells, highly secretory epithelial cells found at the bases of small intestinal crypts, release a variety of microbicidal molecules, including alpha-defensins and lysozyme. The secretion ofPaneth cells, highly secretory epithelial cells found at the bases of small intestinal crypts, release a variety of microbicidal molecules, including alpha-defensins and lysozyme. The secretion of antimicrobials by Paneth cells is thought to be important in mucosal host defense against invasion by enteric pathogens. We explored whether enteric pathogens can interfere with this arm of defense. We found that oral inoculation of mice with wild-type Salmonella enterica serovar Typhimurium decreases the expression of alpha-defensins (called cryptdins in mice) and lysozyme. Oral inoculation with Salmonella serovar Typhimurium strains that are heat killed, lack the PhoP regulon, and lack the SPI1 type III secretion system or with Listeria monocytogenes does not have this effect. Salmonella may gain a specific survival advantage in the intestinal lumen by decreasing the expression of microbicidal peptides in Paneth cells through direct interactions between Salmonella and the small intestinal epithelium.

• Analysis of 16S libraries of mouse gastrointestinal microflora reveals a large new group of mouse intestinal bacteria.

  Authors: Nita H Salzman, Hendrik de Jong, Yvonne Paterson, Hermie J M Harmsen, Gjalt W Welling, Nicolaas A Bos

  Microbiology (Reading, England). 12/2002; 148(Pt 11):3651-60.

  Total genomic DNA from samples of intact mouse small intestine, large intestine, caecum and faeces was used as template for PCR amplification of 16S rRNA gene sequences with conserved bacterialTotal genomic DNA from samples of intact mouse small intestine, large intestine, caecum and faeces was used as template for PCR amplification of 16S rRNA gene sequences with conserved bacterial primers. Phylogenetic analysis of the amplification products revealed 40 unique 16S rDNA sequences. Of these sequences, 25% (10/40) corresponded to described intestinal organisms of the mouse, including Lactobacillus spp., Helicobacter spp., segmented filamentous bacteria and members of the altered Schaedler flora (ASF360, ASF361, ASF502 and ASF519); 75% (30/40) represented novel sequences. A large number (11/40) of the novel sequences revealed a new operational taxonomic unit (OTU) belonging to the Cytophaga-Flavobacter-Bacteroides phylum, which the authors named 'mouse intestinal bacteria'. 16S rRNA probes were developed for this new OTU. Upon analysis of the novel sequences, eight were found to cluster within the Eubacterium rectale-Clostridium coccoides group and three clustered within the Bacteroides group. One of the novel sequences was distantly related to Verrucomicrobium spinosum and one was distantly related to Bacillus mycoides. Oligonucleotide probes specific for the 16S rRNA of these novel clones were generated. Using a combination of four previously described and four newly designed probes, approximately 80% of bacteria recovered from the murine large intestine and 71% of bacteria recovered from the murine caecum could be identified by fluorescence in situ hybridization (FISH).

• Paneth cells, defensins, and the commensal microbiota: A hypothesis on intimate interplay at the intestinal mucosa

  Authors: Nita H. Salzman, Mark A. Underwood, Charles L. Bevins

  Seminars in Immunology.

  Mucosal surfaces are colonized by a diverse and dynamic microbiota. Much investigation has focused on bacterial colonization of the intestine, home to the vast majority of this microbiota.Mucosal surfaces are colonized by a diverse and dynamic microbiota. Much investigation has focused on bacterial colonization of the intestine, home to the vast majority of this microbiota. Experimental evidence has highlighted that these colonizing microbes are essential to host development and homeostasis, but less is known about host factors that may regulate the composition of this ecosystem. While evidence shows that IgA has a role in shaping this microbiota, it is likely that effector molecules of the innate immune system are also involved. One hypothesis is that gene-encoded antimicrobial peptides, key elements of innate immunity throughout nature, have an essential role in this regulation. These effector molecules characteristically have activity against a broad spectrum of bacteria and other microbes. At mucosal surfaces, antimicrobial peptides may affect the numbers and/or composition of the colonizing microbiota. In humans and other mammals, defensins are a predominant class of antimicrobial peptides. In the small intestine, Paneth cells (specialized secretory epithelial cells) produce high quantities of defensins and several other antibiotic peptides and proteins. Data from murine models indicate that Paneth cell defensins play a pivotal role in defense from food and water-borne pathogens in the intestinal lumen. Recent studies in humans provide evidence that reduced Paneth cell defensin expression may be a key pathogenic factor in ileal Crohn's disease, a subgroup of inflammatory bowel disease (IBD), and changes in the colonizing microbiota may mediate this pathogenic mechanism. It is also possible that low levels of Paneth cell defensins, characteristic of normal intestinal development, may predispose premature neonates to necrotizing enterocolitis (NEC) through similar close links with the composition of the intestinal microbiota. Future studies to further define mechanisms by which defensins and other host factors regulate the composition of the intestinal microbiota will likely provide new insights into intestinal homeostasis and new therapeutic strategies for inflammatory and infectious diseases of the bowel.