Zhannat Z. Nurgalieva

B-Cell and T-Cell Immune Responses to Experimental Helicobacter pylori Infection in Humans

ABSTRACT The acute antibody and T-cell immune response to Helicobacter pylori infection in humans has not been studied systematically. Serum from H. pylori-naive volunteers challenged with H. pylori and cured after 4 or 12 weeks was tested by enzyme-linked immunosorbent assays for anti-H. pylori-specific immunoglobulin M (IgM) and IgA established using bacterial lysates from homologous (the infecting strain) and heterologous H. pylori. Proteins recognized by IgM antibody were identified by mass spectrometry of immunoreactive bands separated by two-dimensional gel electrophoresis. Mucosal T-cell subsets (CD4, CD8, CD3, and CD30 cells) were assessed by immunohistochemistry. All 18 infected volunteers developed H. pylori-specific IgM responses to both homologous or heterologous H. pylori antigens. H. pylori antigens reacted with IgM antibody at 4 weeks postinfection. IgM Western blotting showed immunoreactivity of postinfection serum samples to multiple H. pylori proteins with molecular weights ranging between 9,000 (9K) to 150K with homologous strains but only a 70K band using heterologous antigens. Two-dimensional electrophoresis demonstrated that production of H. pylori-specific IgM antibodies was elicited by H. pylori flagellins A and B, urease B, ABC transporter binding protein, heat shock protein 70 (DnaK), and alkyl hydroperoxide reductase. Mucosal CD3, CD4, and CD8 T-cell numbers increased following infection. IgM antibody responses were detected to a range of homologous H. pylori antigens 2 to 4 weeks postchallenge. The majority of H. pylori proteins were those involved in motility and colonization and may represent targets for vaccine development.

Correlation between Helicobacter pylori OipA Protein Expression and oipA Gene Switch Status

ABSTRACT Polyclonal antisera to either a synthetic OipA peptide or a recombinant OipA protein detected OipA expression in Helicobacter pylori and correlated with functional oipA status determined by PCR sequence (sensitivity and specificity of >94%). Immunoblotting is a simple and accurate method for detecting expression of the important virulence factor OipA.

A pilot study of Helicobacter pylori infection and risk of laryngopharyngeal cancer

Squamous cell carcinoma of the laryngopharynx has been linked to laryngopharyngeal reflux disease. Helicobacter pylori corpus gastritis decreases gastric acid secretion and provides some protection against complications of gastroesophageal reflux, including adenocarcinoma of the distal esophagus. The aim of this study was to investigate whether H. pylori infection also protects against laryngopharyngeal carcinoma.

Gene expression in Barrett's esophagus: Laser capture versus whole tissue

Objective. Diagnosis of Barretts esophagus (BE) is typically done through morphologic analysis of esophageal tissue biopsy. Such samples contain several cell types. Laser capture microdissection (LCM) allows the isolation of specific cells from heterogeneous cell populations. The purpose of this study was to determine the degree of overlap of the two sample types and to define a set of genes that might serve as biochemical markers for BE. Material and methods. Biopsies were obtained from regions of the glandular tissue of BE and normal esophagus from 9 subjects with BE. Samples from 5 subjects were examined as whole tissue (BE [whole]; E [whole]), and in 4 subjects the glandular epithelium of BE was isolated using LCM (BE [LCM]) and compared with the averaged values (E [LCM]) for both basal cell (B [LCM]) and squamous cell (S [LCM]) epithelium. Results. Gene expression revealed 1797 probe sets between BE [whole] and E [whole] (fold change > 2.0; p<0.001). Most of these genes (74%) were also differentially expressed between BE [LCM] and E [LCM], showing that there was high concordance between the two sampling methods. LCM provided a great deal of additional information (2113 genes) about the alterations in gene expression that may represent the BE phenotype. Conclusions. There are differences in gene expression profiles depending on whether specimens are whole tissue biopsies or LCM dissected. Whole tissue biopsies should prove satisfactory for diagnostic purposes. Because the data from LCM samples delineated many more Barretts-specific genes, this procedure might provide more information regarding pathogenesis than would whole tissue material.

The Use of Cytokeratin Stain to Distinguish Barrett’s Esophagus from Contiguous Tissues: A Systematic Review

Our objective was to systematically review the existing literature regarding the use of cytokeratin (CK) stain in differentiating Barrett’s esophagus (BE) from tissues of the gastric cardia, corpus, or antrum, with or without intestinal metaplasia (IM). Pubmed was searched for full publications in English (1983–2005) addressing the use of CK for differentiation of BE from contiguous tissues. Information was collected on the study sample, blinding, the methods used for CK staining, and for defining and applying the gold standard tests. Test characteristics were obtained or calculated. Sixteen studies (containing 46 comparisons) met the inclusion and exclusion criteria. Immunostaining for CK 7 and 20 was generally highly specific in distinguishing long-segment BE from antrum IM, fundus IM, or noncardiac gastric IM; 27 comparisons showed statistically significant differences. However, only 8 of 15 comparisons (6 of 12 studies) reported significant differences in CK staining patterns between BE and gastric cardia IM with a high sensitivity (89%–100%) and specificity (83%–100%) for long-segment BE and lower estimates for short-segment BE, while the other seven comparisons showed no significant differences and a very low sensitivity. Examination by a blinded pathologist was reported in five of six positive studies and in only one of six of the negative studies. In addition, variation in the patient populations, use of surgical resection versus endoscopic biopsies, and biopsy sampling technique in endoscopic studies may have accounted for these differences. Finally, two studies did not find significant differences in CK staining patterns between BE and normal cardiac mucosa. In conclusions, CK immunostaining has not performed well in differentiating BE, especially short-segment BE, from cardia IM. There seems to be a spectrum bias where the accuracy varies with different tested populations. CK immunostaining distinguished well between BE and IM in noncardiac segments of the stomach; however, these comparisons are not clinically relevant.

Phase I and II enzyme polymorphisms as risk factors for Barrett’s esophagus and esophageal adenocarcinoma: a systematic review and meta-analysis

Although several studies have examined the association between phase I/II enzyme polymorphisms and esophageal adenocarcinoma (EAC) and/or Barretts esophagus (BE), their overall findings remain unclear. We performed a systematic review and meta-analysis to determine whether phase I/II polymorphisms are independent risk factors for either BE or EAC. We employed keyword searches in multiple databases to identify studies published before October 1, 2007. Single-nucleotide polymorphisms (SNPs) examined in > or =3 studies were meta-analyzed to obtain a pooled estimate of effect. Meta-analysis suggested the minor allele for GSTP1 Val(105) conveys modest excess risk (odds ratio [OR](BE)= 1.50, 95% confidence interval [CI] 1.16-1.95; OR(EAC)= 1.20, 95% CI 0.94-1.54). No excess risk was observed with GSTM1 null (OR(BE)= 0.77, 95% CI: 0.56-1.08; OR(EAC)= 1.08, 95% CI: 0.79-1.48), GSTT1 null (OR(BE)= 1.35, 95% CI: 0.91-2.01; OR(EAC)= 0.84, 95% CI: 0.48-1.49), or CYP1A Val(462) (OR(EAC)= 0.89, 95% CI: 0.40-1.97). Insufficient data existed to meta-analyze remaining SNPs. Our review identified GSTP1(Ile105Val) as a possible risk factor for BE and EAC in Caucasian males. No excess risk was observed for other phase I/II polymorphisms with sufficient data to meta-analyze. Additional studies are needed to determine if GSTP1 conveys excess risk in females or non-Caucasians and to evaluate other phase I/II polymorphisms.

CagA in Barrett’s oesophagus in Colombia, a country with a high prevalence of gastric cancer

Background: In the USA, atrophic gastritis and gastric cancer are rare, whereas gastro-oesophageal reflux disease (GERD) is common. Infection with Helicobacter pylori, especially a CagA positive strain, is unusual in patients with GERD/Barrett’s oesophagus in the USA. Aim: To examine the relation between Barrett’s oesophagus and CagA positive H pylori in Colombia, a country with a high prevalence of CagA positive H pylori associated atrophic gastritis and gastric cancer. Methods:Helicobacter pylori and CagA status was determined among Colombian patients with long segment Barrett’s oesophagus and a control group with simple H pylori gastritis. Helicobacter pylori status was determined using a triple stain and CagA status was determined by immunohistochemistry using a specific rabbit anti-CagA serum. Results: Gastric and oesophageal mucosal biopsies were obtained from 51 patients—39 men (mean age, 57.8 years; SD, 13.1) and 12 women (mean age, 51.8 years; SD, 14.4)—with documented long segment Barrett’s oesophagus. The results were compared with 24 Colombian patients with H pylori gastritis without oesophageal disease. Thirty two patients with Barrett’s oesophagus had active H pylori infection. CagA status was evaluated in a subset of 23 H pylori infected patients with Barrett’s oesophagus, and was positive in eight of these patients compared with 19 of 24 controls (p  =  0.01). Conclusions: Although most Colombian patients with Barrett’s oesophagus had H pylori infection, CagA positive infections were unusual. These data illustrate how consistent corpus inflammation reduces acid secretion, which prevents Barrett’s oesophagus among those with abnormal gastro-oesophageal reflux barriers.

Use of a Dry-Plasma Collection Device to Overcome Problems with Storage and Transportation of Blood Samples for Epidemiology Studies in Developing Countries

ABSTRACT Studies are difficult in areas lacking modern facilities due to the inability to reliably collect, store, and ship samples. Thus, we sought to evaluate the use of a dry plasma collection device for seroepidemiology studies. Plasma was obtained by fingerstick using a commercial dry plasma collection device (Chemcard Plasma Collection Device) and serum (venipuncture) from individuals in Kazakhstan. Plasma samples were air dried for 15 min and then stored desiccated in foil zip-lock pouches at 4 to 6°C and subsequently shipped to the United States by air at ambient temperature. Serum samples remained frozen at −20°C until assayed. Helicobacter pylori status was determined by enzyme-linked immunosorbent assay (HM-CAP EIA) for the dry plasma and the serum samples. The results were concordant in 250 of the 289 cases (86.5%). In 25 cases (8.6%), the dry plasma samples gave indeterminate results and could not be retested because only one sample was collected. Five serum samples were positive, and the corresponding dry plasma samples were negative; one serum sample was negative, and the corresponding plasma sample was positive. The relative sensitivity and specificity of the Chemcard samples to serum were 97.6 and 97.9%, respectively, excluding those with indeterminate results. Repeated freeze-thawing had no adverse effect on the accuracy of the test. We found the dry plasma collection device to provide an accurate and practical alternative to serum when venipuncture may be difficult or inconvenient and sample storage and handling present difficulties, especially for seroepidemiologic studies in rural areas or developing countries and where freeze-thawing may be unavoidable.

Adherence to head and neck cancer guidelines: Multidisciplinary conference recommendations and care delivered.

202 Background: Adherence to clinical guidelines, such as those developed by the National Comprehensive Cancer Network (NCCN), can serve as a quality measure for delivered care. We aimed to determine NCCN guideline compliance of and concordance between multidisciplinary conference (MC) treatment recommendations and treatment received. METHODS A retrospective review of 71 consecutive previously untreated patients from 1/1 to 4/1/2006 evaluated whether MC recommendations were compliant with 2006 NCCN treatment guidelines, what treatment was actually provided, and whether this treatment was congruent with MC recommendations and/or adhered to 2006 NCCN guidelines. RESULTS TNM staging was documented in 94.4% of MC notes and treatment recommendations were documented for 98.6% of patients. Of enrolled patients, 53% had oropharyngeal cancer, 22.7% had oral cavity cancer, 18.2% had laryngeal cancer, and 6.1% had nasopharyngeal cancer. The mean age at presentation was 57.4 years. Forty-eight patients (72.7%) were men. MC recommendations were not compliant with NCCN guidelines in 7 (10.6%) patients. Reasons for non-compliance were recommended definitive radiation alone for T1/2N2 oropharyngeal cancer (3 patients), recommended over-use of induction chemotherapy (2 patients) or adjuvant therapy (1 patient), and incomplete documentation (1 patient). Actual treatment deviated from MC recommendations in 25 (38.5%) patients, although in 14 of these patients, treatment remained guideline-adherent. Actual treatment was not compliant with guidelines in 9 patients; 4 of these patients had treatment congruent with MC recommendations, which were not NCCN guideline-compliant. Reasons for guideline-discordance in the remaining patients were individual. CONCLUSIONS Discordance with NCCN guidelines for head and neck cancer treatment planning and delivery was observed in this cohort; while some deviations were due to incomplete documentation, areas for improvement were identified. Further study is warranted to determine whether treatment deviating from NCCN guidelines impacts patient outcome.