Katunori Saigenji

Antimicrobial host defense in the upper gastrointestinal tract

Background With the exception of fungi, microbial infections are rare in the oesophagus. Herein, we aimed to systematically assess the distribution and quantity of different antimicrobial host factors as well as, for thefirst time, functional mucosal antimicrobial activity in the upper gastrointestinal tract. Methods We investigated biopsies from the healthy oesophagus, three different locations in the stomach and the duodenum in a total of 12 individuals. Using real-time PCR with external standards, we compared absolute expression of mRNA encoding antimicrobial peptides including defensins, cathelicidin, bactericidal/permeability-increasing protein, psoriasin, and elafin. In addition, we performed immunostaining for human-&bgr;-defensin-1 (HBD1), elafin, and psoriasin. To test functional relevance, we assessed antimicrobial as well as antifungal activity of cationic extracts from biopsies against E. coli ATCC 25922 and a clinical isolate of Candida albicans. Results In contrast to HBD1 which was similarly expressed in all tissues, inducible &bgr;-defensins in the healthy oesophagus were much higher compared with the stomach and duodenum (for HBD2–4: P<0.01). In addition, the antiproteases elafin and psoriasin were also predominantly expressed in the oesophagus (P<0.005). In contrast, LL-37 and bactericidal/permeability-increasing protein were only marginally expressed. Cationic tissue extracts from both the oesophagus as well as the stomach showed potent antibacterial activity against E. coli. Consistent with susceptibility to Candida infection, the esophageal extracts exhibited a weaker activity against C. albicans (P=0.026). Conclusion Despite dominant expression of antimicrobial host peptides, oesophageal tissue shows a weakened potency to kill C. albicans. These data suggest an important role of yet unknown antimicrobial molecules.

Gastric Antimicrobial Peptides Fail to Eradicate Helicobacter pylori Infection Due to Selective Induction and Resistance

Background Although antimicrobial peptides protect mucus and mucosa from bacteria, Helicobacter pylori is able to colonize the gastric mucus. To clarify in which extend Helicobacter escapes the antimicrobial defense, we systematically assessed susceptibility and expression levels of different antimicrobial host factors in gastric mucosa with and without H. pylori infection. Materials and Methods We investigated the expression levels of HBD1 (gene name DEFB1), HBD2 (DEFB4A), HBD3 (DEFB103A), HBD4 (DEFB104A), LL37 (CAMP) and elafin (PI3) by real time PCR in gastric biopsy samples in a total of 20 controls versus 12 patients colonized with H. pylori. Immunostaining was performed for HBD2 and HBD3. We assessed antimicrobial susceptibility by flow cytometry, growth on blood agar, radial diffusion assay and electron microscopy. Results H. pylori infection was associated with increased gastric levels of the inducible defensin HBD2 and of the antiprotease elafin, whereas the expression levels of the constitutive defensin HBD1, inducible HBD3 and LL37 remained unchanged. HBD4 was not expressed in significant levels in gastric mucosa. H. pylori strains were resistant to the defensins HBD1 as well as to elafin, and strain specific minimally susceptible to HBD2, whereas HBD3 and LL37 killed all H. pylori strains effectively. We demonstrated the binding of HBD2 and LL37 on the surface of H. pylori cells. Comparing the antibacterial activity of extracts from H. pylori negative and positive biopsies, we found only a minimal killing against H. pylori that was not increased by the induction of HBD2 in H. pylori positive samples. Conclusion These data support the hypothesis that gastric H. pylori evades the host defense shield to allow colonization.

Characterization of pKU701, a 2.5-kb plasmid, in a Japanese Helicobacter pylori isolate.

A cryptic plasmid of Helicobacter pylori, pKU701 (accession number AB078638), was isolated and the complete nucleotide sequence was determined. No drug resistance properties were mediated by pKU701. The 2454b pKU701 sequence, which had a 38% content of G-C residues, generated one polypeptide from a single open reading frame (ORF1). Extensive sequence homology was evident between pKU701 and ORF1 of H. pylori plasmid pHPO100 (repA, 88.8% identity) as well as ORF3 of plasmid pHPS1 (repB, 80.2% identity), but pKU701 showed only 46.3% homology with ORF1 of plasmid pHPK255 (repA). Tandem direct repeats of a 33-bp segment were found in pKU701 outside ORF1, but there were no inverted repeat ends such as those found in typical insertion sequences. The ability of drug resistance plasmids to replicate in H. pylori is probably limited, so chromosomal mutation may be a more likely cause of resistance.

Use of the restriction enzyme EcoRI for pulsed-field gel electrophoretic analysis of Helicobacter pylori.

ABSTRACT Pulsed-field gel electrophoretic (PFGE) analysis of Helicobacter pylori isolates is not commonly employed because of the inability to compare the typing with other typing systems. We adapted the PFGE analysis for H. pylori by using EcoRI and slightly modified our laboratory methods to improve the typing of isolates (typeability was 97%).

Study on Small Colorectal Polyps and Early Cancers

Abstract: We investigated the histopathological features of polyps with a diameter of less than 5 mm that had been resected by a snare or hot‐biopsy (1, 357 lesions in 712 patients), and considered the problems associated with these techniques. 67.7% of the polyps were adenomas, 15.5% were metaplastic polyps, 0.6% were colon cancers, and 0.1% were carcinoids. Eighty percent of the polyps situated on the oral side of the descending colon were adenomas. Although adenomas occurred somewhat more frequently in the sigmoid colon, they tended to be distributed evenly throughout the entire colon. The reddened color of the surface of the polyps tended to accompany adenomas, while a whitish surface color was frequently associated with metaplastic polyps. Multiple polyps occurred in 57% of the patients and 33.7% of the patients had a large polyp with a diameter of more than 6 mm. In six out of eight cases of cancer the cancers were limited to the mucosa (m), the other 2 were submucosal invaded carcinomas. One of these cases was a depressd type of cancer and the other lesions were classified as being elevated type polyps. The elevated type of diminutive early colorectal carcinoma with a diameter of less than 5 mm was difficult to distinguish endoscopically from benign polyps, so we recommended that small colonic polyps should be removed when encountered during a colonoscopy.

Traitement endoscopique du cancer colorectal précoce

RésuméLe présent article décrit le traitement endoscopique du cancer colorectal précoce. La polypectomie endoscopique a été réalisée en utilisant trois techniques: la pince chaude, la polypectomie conventionnelle à l’anse diathermique et la résection muqueuse. Les cancers colorectaux précoces de type saillant ont été réséqués par la polypectomie conventionnelle à l’anse diathermique et les cancers de type superficiel par la technique de la résection muqueuse. Les cancers précoces avec envahissement limité à la muqueuse ont été définis comme cancers met les cancers avec envahissement de la sous-muqueuse mais en deçà de celle-ci ont été définis sous le terme de cancers sm. Les cancers sm ont été classés en fonction de la profondeur de l’envahissement en sm1, sm2 ou sm3. La polypectomie endoscopique est indiquée pour les cancers précoces colorectaux met sm1, du fait que les cancers sm1 sont habituellement exempts de métastases lymphatiques. La colectomie est indiquée après polypectomie chez les patients qui présentent: (1) une marge de résection positive, (2) un envahissement vasculaire ou (3) le diagnostic de cancer peu différencié. Nous rappelons que la polypectomie s’accompagne d’une faible incidence de complications telles que les hémorragies et les perforations.En conclusion, la polypectomie à l’anse diathermique et la résection muqueuse constituent des techniques de choix du traitement des cancers précoces colorectaux.SummaryThis article describes endoscopic treatment of early colo-rectal cancers. Endoscopic polypectomy was performed by three procedures: hot biopsy, conventional snare polypectomy, and mucosal resection. Protruded types of early colorectal cancers were resected by conventional snare polypectomy and superficial type were resected by mucosal resection technique. Early cancers with invasion limited to mucosa were defined as m cancer, and cancers that invaded the submucosa, but beyond, were defined as sm cancers. Sm cancers were classified according to the depth of invasion as sm1, sm2, or sm3. Endoscopic polypectomy was indicated for m and sm1 early colorectal cancers, because sm1 cancers usually have no lymph node metastasis. Colectomy should be indicated after polypectomy in these patients because of: 1) a positive resection margin, 2) evidence of vascular invasion, or 3) a diagnosis of poorly differentiated type cancer. We should mention polypectomy was associated with a low incidence of complications such as bleeding and perforation.It is concluded that snare polypectomy and mucosal resection were very useful for the treatment of early colo-rectal cancers.