James J. Thornton

Keynote review: Epithelial and endothelial barriers in human disease

There is a spectrum of distinct disease states that have in common their effect of breaking down epithelial and/or endothelial barrier function. Fluid compartmentalization goes awry, with profound implications for epithelial and stromal homeostasis, fluid and/or electrolyte balance, generation of inflammatory states, and even tumor microenvironment. Specific effects on the tight junction are found to be integral to bacterial invasion and tumor progression.

Proton pump inhibitors: actions and reactions

Proton pump inhibitors are the second most commonly prescribed drug class in the United States. The increased utilization of PPIs parallels the rising incidence of reflux disease. Owing to their clinical efficacy and relative lack of tachyphylaxis, PPIs have largely displaced H-2 receptor antagonists in the treatment of acid peptic disorders. The elevation of intragastric pH and subsequent alterations of gastric physiology induced by PPIs may yield undesired effects within the upper GI tract. The ubiquity of the various types of H(+), K(+)-ATPase could also contribute to non-gastric effects. PPIs may influence physiology in other ways, such as inducing transepithelial leak.

Transepithelial Leak in Barrett’s Esophagus

Using orally administered sucrose as a probe of gastrointestinal permeability, this study focused on determining whether Barrett’s metaplasia exhibits a paracellular transepithelial leak to small nonelectrolytes. Subjects in five separate classes (nonendoscoped, asymptomatic controls; endoscoped, asymptomatic controls; gastroesophageal reflux disease without mucosal complications; grossly visible esophagitis; and Barrett’s esophagus) consumed a sucrose solution at bedtime and collected all overnight urine. Urine volume was measured and sucrose concentration was determined by high-performance liquid chromatography. Patients with Barrett’s were observed to exhibit a transepithelial leak to sucrose whose mean value was threefold greater than that seen in healthy control subjects or patients with reflux but without any mucosal defect. A parallel study of claudin tight junction proteins in endoscopy biopsy samples showed that whereas Barrett’s metaplasia contains dramatically more claudin-2 and claudin-3 than is found in normal esophageal mucosa, it is markedly lower in claudins 1 and 5, indicating very different tight junction barriers.

Tumor necrosis factor blockade for treatment of inflammatory bowel disease: efficacy and safety.

Inflammatory bowel disease (IBD), including Crohns Disease (CD) and Ulcerative Colitis (UC), is characterized by inflammation of the gastrointestinal tract. In UC, inflammation is confined to the mucosa, initially involving the rectum, and may extend proximally to involve the entire colon. In CD, transmural inflammation may affect any portion of the GI tract. The etiology of these disease processes has remained unclear. Therapies are aimed at reducing inflammation and thereby improving symptomatology and morbidity. Traditional medical therapies have included corticosteroids, aminosalicylates, and immunomodulators. Within the past decade, another class of medications has been utilized targeting Tumor Necrosis Factor (TNF), a key, early signaling molecule in the inflammatory cascade. Increased levels of TNF have been found in the blood, epithelial tissue, and stool of patients with active IBD. Anti-TNF medications can not only have direct effects on immune system components, but they also can ameliorate apoptotic cell death and tight junction compromise in the gastrointestinal epithelium. Several randomized, placebo controlled studies have demonstrated the efficacy of these medications in achieving induction and maintaining remission of disease. Their safety profile, however, remains a concern. There has been a reported association of biologic therapy and increased opportunistic infections. A link between biologic therapy and the development of certain malignancies has also been described. Despite these associations, TNF blockade remains an important therapeutic development in the modern therapy of IBD. The role of barrier breakdown at the tight junction level in IBD, and of TNF induction of barrier disruption, is also discussed.

Comparison of three integral tight junction barrier proteins in Barrett's epithelium versus normal esophageal epithelium.

rified DNA was used to determine the sex and to obtain a genetic fingerprint of the specimens. The cancer fragments showed a female phenotype, thereby excluding our patient as a possible source. The genetic fingerprint of the adenoma, however, was identical to the one of our patient (Table 1). Confounding of specimens is rare but cannot be completely avoided. As illustrated by our case, this can occur in the endoscopy room while tissue fragments are collected. In addition, specimens can be mislabeled, and during processing fragments from other specimens (so-called “floaters”) can be incorporated into the analysis (1). As soon as a mix-up is suspected by an unlikely finding, thorough investigation is critical, because misassignment of a diagnosis can delay appropriate treatment or lead to repeat procedures or unnecessary invasive treatment. The ideal test to determine the presence of a mix-up should be able to positively identify a person. Until recently, the identification of a person relied on the blood group and the human leukocyte antigens (2). Unfortunately, these tests lack the power to positively identify a person. For example, a discordance of the blood groups clearly indicates a different source, but the probability of different blood groups in two unrelated white persons is only approximately 60% (1). On the other hand, of course, the identification of the blood group does not prove the identity of the person(s). Almost unequivocal identification became possible only after the introduction of DNA fingerprinting (3). The probability of two unrelated persons having identical fingerprints is typically less than one in several millions. The only prerequisite to perform such a test is the availability of a tissue or blood sample to positively identify the person under question. It is almost always possible to extract enough DNA from tiny, even formalin-fixed and paraffin-embedded biopsy fragments, often even from individual pieces. There are few case reports in the literature in which this technology was used to correctly identify tissue, blood, and urine samples (2, 4, 5). We believe that this technology can be useful to all in the medical field who might be confronted with a possible tissue, blood, or urine mix-up. However, the analysis should only be performed in a laboratory with the necessary experience in forensic genetics.

Electrophysiological differences in normal colon mucosa from diverticular disease vs cancer

To The Editor: As part of a two-year study of tight-junction permeability of human colon, we have routinely obtained colon tissue from patients who have undergone colectomy for adenocarcinoma or diverticular disease. For studies involving, eg, effects of protein kinase C activators on transepithelial permeability, we had utilized grossly normal tissue that is at least 10 cm distant from the lesion, be it a tumor or a diverticulum. Such normal-appearing tissue from the diverticular disease and cancer patients was to be our “control” group for comparisons to epithelium from inflamed mucosa (inflammatory bowel disease patients) or from tumor itself (1). This practice appears to be followed in many laboratories. In the course of these studies, we began, however, to observe a distinct trend arguing that grossly normal colon epithelium from cancer vs diverticular disease colectomies is not equivalent. The result has been that we no longer use colectomy specimens from diverticular disease patients in certain studies. After mounting any colon tissue in Ussing chambers, there is a period during which transepithelial electrical parameters come to steady state, generally 20–45 min at 37°C in a bicarbonate-buffered saline. During this time, we have observed that transepithelial electrical resistance (Rt) will rise from 20 to as high as 120 ohms 3 cm, and short circuit current (Isc) will decrease from 250 mA/cm to 50 mA/cm or lower. Whereas differences in Rt between grossly normal tissue from patients with diverticular disease vs cancer were not observed, a sharp difference in the stability of Isc was seen as shown in Figure 1. Isc values from histologically normal colon tissue of diverticular disease patients often exhibit a very steep exponential drop over the initial 60 min, frequently falling below a level (,3 mA/cm) at which accurate readings (both Isc and potential difference) can be obtained. The Isc values also initially fell steeply using grossly normal tissue from cancer patients, but generally leveled off at values above 50 mA/cm. Figure 2 shows a scatter plot of Isc values from tissue obtained from left colectomy of these two patient groups, approximately 30

Drug delivery of zinc to Barrett’s metaplasia by oral administration to Barrett’s esophagus patients

BACKGROUND Delivery of a pharmacologically effective drug dosage to a target tissue is critical. Barretts epithelia are a unique challenge for drug delivery of orally administered zinc due to rapid transit down the esophageal lumen, incomplete absorptive differentiation of these epithelia, and the use of proton-pump inhibitor drugs abrogating intestinal uptake of supplemental zinc. METHODS Barretts esophagus patients were administered oral zinc gluconate (26 mg zinc twice daily) for 14 days prior to biopsy procurement. Barretts biopsies were analyzed for total zinc content by atomic absorption spectroscopy and by western immunoblot for cellular proteins known to be regulated by zinc. RESULTS Cellular levels of both the Znt-1 transport protein and the alpha isoform of PKC were over 50% lower in the zinc treatment group. CONCLUSION Oral zinc administration can result in effective delivery of zinc to Barretts epithelia with resulting effects on intracellular signal transduction.

Transepithelial leak in Barrett's esophagus patients: The role of proton pump inhibitors

AIM To determine if the observed paracellular sucrose leak in Barretts esophagus patients is due to their proton pump inhibitor (PPI) use. METHODS The in vivo sucrose permeability test was administered to healthy controls, to Barretts patients and to non-Barretts patients on continuous PPI therapy. Degree of leak was tested for correlation with presence of Barretts, use of PPIs, and length of Barretts segment and duration of PPI use. RESULTS Barretts patients manifested a near 3-fold greater, upper gastrointestinal sucrose leak than healthy controls. A decrease of sucrose leak was observed in Barretts patients who ceased PPI use for 7 d. Although initial introduction of PPI use (in a PPI-naïve population) results in dramatic increase in sucrose leak, long-term, continuous PPI use manifested a slow spontaneous decline in leak. The sucrose leak observed in Barretts patients showed no correlation to the amount of Barretts tissue present in the esophagus. CONCLUSION Although future research is needed to determine the degree of paracellular leak in actual Barretts mucosa, the relatively high degree of leak observed with in vivo sucrose permeability measurement of Barretts patients reflects their PPI use and not their Barretts tissue per se.

Letters to the editorComparison of three integral tight junction barrier proteins in Barrett's epithelium versus normal esophageal epithelium

Comparison of three integral tight junction barrier proteins in Barretts epithelium versus normal esophageal epithelium