Matthew R. Banks

Image cytometry accurately detects DNA ploidy abnormalities and predicts late relapse to high-grade dysplasia and adenocarcinoma in Barrett's oesophagus following photodynamic therapy

Background and aims:DNA ploidy abnormalities (aneuploidy/tetraploidy) measured by flow cytometry (FC) are strong predictors of future cancer development in untreated Barretts oesophagus, independent of histology grade. Image cytometric DNA analysis (ICDA) is an optical technique allowing visualisation of abnormal nuclei that may be undertaken on archival tissue. Our aim was to determine the accuracy of ICDA vs FC, and evaluate DNA ploidy as a prognostic biomarker after histologically successful treatment with photodynamic therapy (PDT).Methods:Nuclei were extracted from 40 μm sections of paraffin-embedded biopsies and processed for ICDA at UCL and FC at UW using standardised protocols. Subsequently, DNA ploidy was evaluated by ICDA on a cohort of 30 patients clear of dysplasia 1 year after aminolaevulinic acid PDT for high-grade dysplasia (HGD). The results were correlated with long-term outcome.Results:In the comparative study, 93% (41 out of 44) of cases were classified identically. Errors occurred in the near-diploid region by ICDA and the tetraploid region by FC. In the cohort study, there were 13 cases of late relapse (7 cancer, 6 HGD) and 17 patients who remained free of dysplasia after a mean follow-up of 44 months. Aneuploidy post-PDT was highly predictive for recurrent HGD or cancer with a hazard ratio of 8.2 (1.8–37.8) (log-rank P=0.001).Conclusions:ICDA is accurate for the detection of DNA ploidy abnormalities when compared with FC. After histologically successful PDT, patients with residual aneuploidy are significantly more likely to develop HGD or cancer than those who become diploid. DNA ploidy by ICDA is a valuable prognostic biomarker after ablative therapy.

Antisecretory actions of a novel vasoactive intestinal polypeptide (VIP) antagonist in human and rat small intestine

1 Vasoactive intestinal peptide (VIP) has been demonstrated in intestinal mucosal neurones and elicits chloride secretion from enterocytes. These findings have led to the proposal that VIP is a secretomotor neurotransmitter. Confirmation of such a role may now be possible with the development of PG 97–269, a high‐affinity, selective antagonist of VIP type 1 (VPAC1) receptor, which is expressed by gut epithelial cells. We have evaluated the VIP antagonism and antisecretory potential of this novel compound using in vitro and in vivo models of intestinal secretion. 2 Monolayers of the human colonic cell line (T84) and muscle‐stripped preparations of rat jejunum and human ileum were set up in Ussing chambers for recording of transepithelial resistance and short‐circuit current. Ussing chambers were modified to allow electrical stimulation of mucosal neurones. Effects of PG 97–269 on enterotoxin‐induced secretion were investigated in perfused rat jejunum in vivo. 3 PG 97–269 competitively antagonised VIP in T84 monolayers. In rat jejunum and human ileum, responses to VIP were inhibited as were responses of rat jejunum to electrical stimulation of mucosal neurons. 4 In perfused rat jejunum, PG 97–269 abolished the effects of VIP on fluid and electrolyte transport and attenuated cholera toxin and Escherichia coli heat labile toxin‐induced net fluid and electrolyte secretion. 5 PG 97–269 is a competitive antagonist of enterocyte VIP receptors and effectively inhibits responses of rat and human intestinal mucosa to VIP. Antagonism of secretory responses to electrical stimulation of mucosal neurons and lumenal application of enterotoxins imply a secretory role for VIP in these processes.

Multicenter, randomized, controlled trial of confocal laser endomicroscopy assessment of residual metaplasia after mucosal ablation or resection of GI neoplasia in Barrett's esophagus.

BACKGROUND Endoscopic ablation is an accepted standard for neoplasia in Barretts esophagus (BE). Eradication of all glandular mucosa in the distal esophagus cannot be reliably determined at endoscopy. OBJECTIVE To assess if use of probe-based confocal laser endomicroscopy (pCLE) in addition to high-definition white light (HDWL) could aid in determination of residual BE. DESIGN Prospective, multicenter, randomized, clinical trial. SETTING Academic medical centers. PATIENTS Patients with Barretts esophagus undergoing ablation. INTERVENTION After an initial attempt at ablation, patients were followed-up either with HDWL endoscopy or HDWL plus pCLE, with treatment of residual metaplasia or neoplasia based on endoscopic findings and pCLE used to avoid overtreatment. MAIN OUTCOME MEASUREMENTS The proportion of optimally treated patients, defined as those with residual BE who were treated and had complete ablation plus those without BE who were not treated and had no evidence of disease at follow-up. RESULTS The study was halted at the planned interim analysis based on a priori criteria. After enrollment was halted, all patients who had been randomized were followed to study completion. Among the 119 patients with follow-up, there was no difference in the proportion of patients achieving optimal outcomes in the two groups (15/57, 26% for HDWL; 17/62, 27% with HDWL + pCLE). Other outcomes were similar in the two groups. LIMITATIONS The study was closed after the interim analysis due to low conditional power resulting from lack of difference between groups as well as higher-than-expected residual Barretts esophagus in both arms. CONCLUSION This study yields no evidence that the addition of pCLE to HDWL imaging for detection of residual Barretts esophagus or neoplasia can provide improved treatment.

Radiofrequency ablation for early oesophageal squamous neoplasia: Outcomes form United Kingdom registry

AIM To report outcomes on patients undergoing radiofrequency ablation (RFA) for early oesophageal squamous neoplasia from a National Registry. METHODS A Prospective cohort study from 8 tertiary referral centres in the United Kingdom. Patients with squamous high grade dysplasia (HGD) and early squamous cell carcinoma (ESCC) confined to the mucosa were treated. Visible lesions were removed by endoscopic mucosal resection (EMR) before RFA. Following initial RFA treatment, patients were followed up 3 monthly. Residual flat dysplasia was treated with RFA until complete reversal dysplasia (CR-D) was achieved or progression to invasive Squamous cell cancer defined as infiltration into the submucosa layer or beyond. The main outcome measures were CR-D at 12 mo from start of treatment, long term durability, progression to cancer and adverse events. RESULTS Twenty patients with squamous HGD/ESCC completed treatment protocol. Five patients (25%) had EMR before starting RFA treatment. CR-D was 50% at 12 mo with a median of 1 RFA treatment, mean 1.5 (range 1-3). Two further patients achieved CR-D with repeat RFA after this time. Eighty per cent with CR-D remain dysplasia free at latest biopsy, with median follow up 24 mo (IQR 17-54). Six of 20 patients (30%) progressed to invasive cancer at 1 year. Four patients (20%) required endoscopic dilatations for symptomatic structuring after treatment. Two of these patients have required serial dilatations thereafter for symptomatic dysphagia with a median of 4 dilatations per patient. The other 2 patients required only a single dilatation to achieve an adequate symptomatic response. One patient developed cancer during follow up after end of treatment protocol. CONCLUSION The role of RFA in these patients remains unclear. In our series 50% patients responded at 12 mo. These figures are lower than limited published data.

Fluid and electrolyte transport in the small intestine.

The small intestine is in a dynamic state of secretion and absorption, the sum of which results in net absorption. Secretion is principally the result of chloride and bicarbonate extrusion through apical chloride channels after the activation of the second messengers cAMP, cGMP, and calcium. In addition to the cystic fibrosis transmembrane conductance regulator, several other candidate chloride channels have been identified and proposed to play a role in intestinal secretion, including the calcium-dependent chloride channel hCLCA1. Pathways leading to the negative control of secretion have been described that use cellular messengers, including inositol (3,4,5,6) tetrakisphosphate and phosphatidylinositol 3-kinase, which may act via basolateral potassium channels. The control of ion transport can also be viewed in terms of the enteric nervous system. The reflex neural pathways involved in enterotoxin-induced secretion have been substantiated and shown to involve 5-hydroxytryptamine, substance P, and the neurokinin 1 and 2 receptors in the sensory arm, and vasoactive intestinal peptide in the secretomotor efferents. Absorption of glucose in addition to active cotransport with sodium via the Na/glucose cotransporter protein has also been shown to occur passively through a carrier-mediated mechanism, using the membrane protein glucose transporter protein 2.

Pulse Oximetry Saturation Levels during Routine Unsedated Diagnostic Upper Gastrointestinal Endoscopy

BACKGROUND Diagnostic procedures account for over 90% of all upper gastrointestinal endoscopies. Pulse oximetry saturation (SpO2) levels were assessed in patients attending for routine unsedated diagnostic gastroscopy to identify factors associated with oxygen desaturation. METHODS Three hundred and thirty unsedated patients were monitored with continuous pulse oximetry. A further 154 patients who requested sedation prior to endoscopy were studied as a positive control group. RESULTS SpO2 levels were lower in sedated compared to unsedated patients (P < 0.0001). Six unsedated patients (2%) desaturated to 90% or less during endoscopy compared to 32 sedated patients (21%) (P < 0.0001). SpO2 levels in unsedated patients were not related to patient sex, age, cigarette smoking, endoscope diameter, basal SpO2 levels or duration of endoscopy. In contrast, examination of the pharyngeal area and epiglottis (P = 0.0002) and a longer intubation time (P = 0.0002) were associated with lower SpO2 levels. The lowest SpO2 level recorded during unsedated endoscopy was 88%. CONCLUSION Pulse oximetry is not a prerequisite to performing routine unsedated diagnostic gastroscopy in patients without severe systemic disease.Background: Diagnostic procedures account for over 90% of all upper gastrointestinal endoscopies. Pulse oximetry saturation (SpO2) levels were assessed in patients attending for routine unsedated diagnostic gastroscopy to identify factors associated with oxygen desaturation. Methods: Three hundred and thirty unsedated patients were monitored with continuous pulse oximetry. A further 154 patients who requested sedation prior to endoscopy were studied as a positive control group. Results: SpO2 levels were lower in sedated compared to unsedated patients (P < 0.0001). Six unsedated patients (2%) desaturated to 90% or less during endoscopy compared to 32 sedated patients (21%) (P < 0.0001). SpO2 levels in unsedated patients were not related to patient sex, age, cigarette smoking, endoscope diameter, basal SpO2 levels or duration of endoscopy. In contrast, examination of the pharyngeal area and epiglottis (P = 0.0002) and a longer intubation time (P = 0.0002) were associated with lower SpO2 levels. The lowest SpO2 level recorded during unsedated endoscopy was 88%. Conclusion: Pulse oximetry is not a prerequisite to performing routine unsedated diagnostic gastroscopy in patients without severe systemic disease.

Intracellular potentiation between two second messenger systems may contribute to cholera toxin induced intestinal secretion in humans

Background: Cholera toxin (CT) acts on intestinal epithelial cells both directly and indirectly via activation of a secretory neural reflex. The reflex may release acetylcholine as one of its final neurotransmitters. This opens up the possibility of a third mechanism of action for CT, namely a synergistic interaction between two secretagogues acting on different second messenger systems within the epithelial cell. Aims: To establish evidence for cholinergic innervation to human ileal epithelial cells and to investigate whether CT potentiates the action of acetylcholine on human intestinal epithelial cells. Methods: Transverse sections of human ileum were examined for mucosal cholinergic nerves and M3 muscarinic receptors using antibodies raised to choline acetyltransferase and M3 receptors. Short circuit current (Isc) responses and ion flux movements were elicited from T84 epithelial cell monolayers set up in Ussing chambers. Results: Immunohistochemistry of native human ileal mucosa revealed the presence of both cholinergic nerves and muscarinic M3 receptors located to the basolateral domain of epithelial cells. Secretory responses of T84 cell monolayers to acetylcholine were greatly potentiated in the presence of CT. This effect, substituting forskolin for CT, was mirrored by increases in basolateral 86Rb and apical 125I efflux. Charybdotoxin plus apamin reduced both Isc and 86Rb efflux evoked by acetylcholine, in the presence of forskolin. Conclusions: Human ileal mucosa receives a direct cholinergic innervation to its epithelial cells. Secretory effects of acetylcholine on epithelial cells are augmented in the presence of CT. Such a synergistic response is dependent on optimum opening of basolateral potassium channels by acetylcholine and apical chloride channels by CT. The interaction may contribute to the mechanism of action of cholera toxin induced secretory diarrhoea.

Radiofrequency ablation is effective for the treatment of high-grade dysplasia in Barrett's esophagus after failed photodynamic therapy.

Endoscopic radiofrequency ablation (RFA) is an effective treatment for high-grade dysplasia in Barretts esophagus in ablation-naïve patients, but no studies have evaluated its use in patients in whom ablative therapy has previously failed. We describe 14 patients with residual high-grade dysplasia following aminolevulinic acid or Photofrin (porfimer sodium) photodynamic therapy (PDT). An overall complete reversal of dysplasia was achieved in 86 % with a combination of RFA and rescue endoscopic mucosal resection. The median total follow-up is 19 months. The rate of strictures was 7 % (1/14) and there was a low rate of buried glands (0.5 % follow-up biopsies). These data suggest RFA is both safe and effective for eradication of high-grade dysplasia in patients in whom PDT has failed.

High resolution colonoscopy in a bowel cancer screening program improves polyp detection

AIM To compare high resolution colonoscopy (Olympus Lucera) with a megapixel high resolution system (Pentax HiLine) as an in-service evaluation. METHODS Polyp detection rates and measures of performance were collected for 269 colonoscopy procedures. Five colonoscopists conducted the study over a three month period, as part of the United Kingdom bowel cancer screening program. RESULTS There were no differences in procedure duration (χ² P = 0.98), caecal intubation rates (χ² P = 0.67), or depth of sedation (χ² P = 0.64). Mild discomfort was more common in the Pentax group (χ² P = 0.036). Adenoma detection rate was significantly higher in the Pentax group (χ² test for trend P = 0.01). Most of the extra polyps detected were flat or sessile adenomas. CONCLUSION Megapixel definition colonoscopes improve adenoma detection without compromising other measures of endoscope performance. Increased polyp detection rates may improve future outcomes in bowel cancer screening programs.

Pharmacological Management of Diarrhea

According to the World Health Organization, there are approximately 2 billion annual cases of diarrhea worldwide. Diarrhea is the leading cause of death in children younger than 5 years and kills 1.5 million children each year. It is especially prevalent in the developing world, where mortality is related to dehydration, electrolyte disturbance, and the resultant acidosis, and in 2001, it accounted for 1.78 million deaths (3.7% of total deaths) in low- and middle-income countries. However, diarrhea is also a common problem in the developed world, with 211 million to 375 million episodes of infectious diarrheal illnesses in the United States annually, resulting in 73 million physician consultations, 1.8 million hospitalizations, and 3100 deaths. Furthermore, 4% to 5% of the Western population suffers from chronic diarrhea. Given the high prevalence of diarrhea, research has been directed at learning more about the cellular mechanisms underlying diarrheal illnesses in order to develop new medications directed at novel cellular targets. These cellular mechanisms and targets are discussed in this article.