Regional variations in Paneth cell antimicrobial peptide expression along the mouse intestinal tract

<p>Abstract</p> <p>Background</p> <p>Enteric antimicrobial peptides secreted from Paneth cells, including α-defensins (in mice named cryptdins), are key effector molecules of innate immunity in the small intestine. The importance of Paneth cells α-defensins emerged from studies of enteric bacterial infection in genetically modified mice, as well as from recent studies linking reduced levels of these α-defensins to Crohn's disease localized to the ileum. However, analysis of expression of Paneth cell α-defensins is incomplete. We therefore performed a comprehensive evaluation of the distribution of antimicrobial molecules along the mouse small intestinal tract to identify potential variations in regional expression.</p> <p>Results</p> <p>In conventionally reared mice, the repertoire of Paneth cell antimicrobials differs between duodenum and ileum. In contrast to the uniform expression of most Paneth cell antimicrobials, both cryptdin 4 and cryptdin-related sequences (CRS) 4C peptides were expressed at progressively increasing amounts (10¹- and 10⁴-fold, respectively) comparing duodenum and ileum. In tissues other than the small intestine, expression of CRS peptides was noted in thymus and caecum. Most Paneth cell products were also produced in the small intestine of germ-free mice at levels similar to those in controls, however CRS4C and RegIIIγ had reduced levels in the former (3- and 8-fold, respectively). No significant changes in expression levels of Paneth cell antimicrobial peptides was observed after oral challenge with either <it>Salmonella enterica </it>serovar typhimurium or <it>Listeria monocytogenes</it>, supporting current notions on the constitutive nature of this defensive system.</p> <p>Conclusion</p> <p>The repertoire of antimicrobial peptides changes along the small intestinal tract, and a subset of these molecules are up-regulated upon colonization, but not in response to enteric bacterial pathogens. The changes detected upon colonization suggest that Paneth cell antimicrobial peptides may play an important role in commensal microbial homeostasis, in addition to their proposed role in protection against infection. In addition, the differential expression of CRS4C along the small intestine suggests mechanisms of regulation that are distinct from other Paneth cell derived antimicrobial peptides.</p

The human cathelicidin hCAP-18 in serum of children with haemato-oncological diseases

The human cathelicidin hCAP-18 (pro-LL-37) is the pro-protein of the antimicrobial peptide LL-37. hCAP-18 can be produced by many different cell types; bone marrow neutrophil precursors are the main source of hCAP-18 in the circulation. Neutrophil count is used as a marker for myelopoiesis but does not always reflect neutrophil production in the bone marrow, and thus additional markers are needed. In this study, we established the reference interval of serum hCAP-18 level in healthy children and compared serum hCAP-18 levels between different diagnostic groups of children with haemato-oncological diseases, at diagnosis. We found that children with diseases that impair myelopoiesis, such as acute leukaemia, aplastic anaemia, or myelodysplastic syndrome, presented with low hCAP-18 levels, whereas patients with non-haematological malignancies displayed serum hCAP-18 levels in the same range as healthy children. Children with chronic myeloid leukaemia presented with high circulating levels of hCAP-18, probably reflecting the high number of all differentiation stages of myeloid cells. We suggest that analysis of serum hCAP-18 provides additional information regarding myelopoiesis in children with haemato-oncological diseases, which may have future implications in assessment of myelopoiesis in clinical management.Peer reviewe

Developmental switch of intestinal antimicrobial peptide expression

Paneth cell–derived enteric antimicrobial peptides provide protection from intestinal infection and maintenance of enteric homeostasis. Paneth cells, however, evolve only after the neonatal period, and the antimicrobial mechanisms that protect the newborn intestine are ill defined. Using quantitative reverse transcription–polymerase chain reaction, immunohistology, reverse-phase high-performance liquid chromatography, and mass spectrometry, we analyzed the antimicrobial repertoire in intestinal epithelial cells during postnatal development. Surprisingly, constitutive expression of the cathelin-related antimicrobial peptide (CRAMP) was observed, and the processed, antimicrobially active form was identified in neonatal epithelium. Peptide synthesis was limited to the first two weeks after birth and gradually disappeared with the onset of increased stem cell proliferation and epithelial cell migration along the crypt–villus axis. CRAMP conferred significant protection from intestinal bacterial growth of the newborn enteric pathogen Listeria monocytogenes. Thus, we describe the first example of a complete developmental switch in innate immune effector expression and anatomical distribution. Epithelial CRAMP expression might contribute to bacterial colonization and the establishment of gut homeostasis, and provide protection from enteric infection during the postnatal period

Mutations in the ELANE Gene are Associated with Development of Periodontitis in Patients with Severe Congenital Neutropenia

# The Author(s) 2011. This article is published with open access at Springerlink.com Background Patients with severe congenital neutropenia (SCN) often develop periodontitis despite standard medical and dental care. In light of previous findings that mutations in the neutrophil elastase gene, ELANE, are associated with more severe neutropenic phenotypes, we hypothesized an association between the genotype of SCN and development of periodontitis. Methods Fourteen Swedish patients with SCN or cyclic neutropenia harboring different genetic backgrounds were recruited for periodontal examination. Peripheral blood, gingival crevicular fluid (GCF), and subgingival bacterial Thomas Modéer and Katrin Pütsep have contributed equally to the study

Benzyl Isothiocyanate, a Major Component from the Roots of Salvadora Persica Is Highly Active against Gram-Negative Bacteria

Plants produce a number of antimicrobial substances and the roots of the shrub Salvadora persica have been demonstrated to possess antimicrobial activity. Sticks from the roots of S. persica, Miswak sticks, have been used for centuries as a traditional method of cleaning teeth. Diverging reports on the chemical nature and antimicrobial repertoire of the chewing sticks from S. persica led us to explore its antibacterial properties against a panel of pathogenic or commensal bacteria and to identify the antibacterial component/s by methodical chemical characterization. S. persica root essential oil was prepared by steam distillation and solid-phase microextraction was used to sample volatiles released from fresh root. The active compound was identified by gas chromatography-mass spectrometry and antibacterial assays. The antibacterial compound was isolated using medium-pressure liquid chromatography. Transmission electron microscopy was used to visualize the effect on bacterial cells. The main antibacterial component of both S. persica root extracts and volatiles was benzyl isothiocyanate. Root extracts as well as commercial synthetic benzyl isothiocyanate exhibited rapid and strong bactericidal effect against oral pathogens involved in periodontal disease as well as against other Gram-negative bacteria, while Gram-positive bacteria mainly displayed growth inhibition or remained unaffected. The short exposure needed to obtain bactericidal effect implies that the chewing sticks and the essential oil may have a specific role in treatment of periodontal disease in reducing Gram-negative periodontal pathogens. Our results indicate the need for further investigation into the mechanism of the specific killing of Gram-negative bacteria by S. persica root stick extracts and its active component benzyl isothiocyanate

On the control of the microflora in the gastro intestinal tract : functional examples of antibacterial peptides from Helicobacter pylori and mouse small intestine

The human microflora contains at least 400 different species mainly localized to the gastro-intestinal tract. This indigenous flora is controlled by host factors as well as by the interference within the microbial community through the action of antimicrobial components. These components are in the focus of this thesis. A microprotocol was developed for biochemical analysis of antimicrobial peptides in tiny tissue material. The protocol was based on an one extraction step, followed by separation on HPLC and mass-spectrometry analysis. This method was sensitive enough for the detection of antimicrobial substances of the small intestine from a single mouse. Ribosomal proteins with antibacterial activity contributed substantially to the microbicidal activity, in addition to already known components such as defensins, lysozyme and phospholipase A, Using this protocol we could demonstrate that the processing of prodefensins to mature microbicidal defensin peptides involves at lest two-steps. No major impact of the microflora on the production of antimicrobial components in the small intestine could be found except for the production of the peptide CRS4-4C. We found that the gastro-intestinal bacteria Helicobacter pylori possessed antibacterial activity. The origin of this activity was traced to cecropin like N-terminal fragments of ribosomal protein L1 (RpL1). Synthetic peptides based on the H. pylori N-terminal sequence were antibacterial but lacked cytolytic as well as hemolytic properties. To H. pylori this antibacterial activity may confer survival advantage during the colonisation phase of a new host when the pH is raised and H. pylori may have to face other faster growing bacteria. The antibacterial activity from ribosomal proteins was found to be a common theme for both the mouse small intestine and H. pylori. The impact of microbicidal substances on host microbe interactions is an important factor for the understanding of the dynamics of the normal microflora. By the use of the microprotocol described here it may become possible to biochemically analyse these substances from human biopsies

Studies on the antibacterial effects of statins--in vitro and in vivo.

BACKGROUND: Statin treatment has been associated with a beneficial outcome on respiratory tract infections. In addition, previous in vitro and in vivo experiments have indicated favorable effects of statins in bacterial infections. AIM: The aim of the present study was to elucidate possible antibacterial effects of statins against primary pathogens of the respiratory tract. METHODS: MIC-values for simvastatin, fluvastatin and pravastatin against S. pneumoniae, M. catarrhalis and H. influenzae were determined by traditional antibacterial assays. A BioScreen instrument was used to monitor effects of statins on bacterial growth and to assess possible synergistic effects with penicillin. Bacterial growth in whole blood and serum from healthy volunteers before and after a single dose of simvastatin, fluvastatin and penicillin (positive control) was determined using a blood culture system (BactAlert). FINDINGS: The MIC-value for simvastatin against S pneumoniae and M catarrhalis was 15 µg/mL (36 mmol/L). Fluvastatin and Pravastatin showed no antibacterial effect in concentrations up to 100 µg/mL (230 µmol/L). Statins did not affect growth or viability of H influenzae. Single doses of statins given to healthy volunteers did not affect growth of pneumococci, whereas penicillin efficiently killed all bacteria. CONCLUSIONS: Simvastatin at high concentrations 15 µg/mL (36 µmol/L) rapidly kills S pneumoniae and M catarrhalis. However, these concentrations by far exceed the concentrations detected in human blood during simvastatin therapy (1-15 nmol/L) and single doses of statins given to healthy volunteers did not improve antibacterial effects of whole blood. Thus, a direct bactericidal effect of statins in vivo is probably not the mechanism behind the observed beneficial effect of statins against various infections