In healthy volunteers, immunohistochemistry supports

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Mar 9, 2015 - The cardiac mucosa between gastric oxyntic mucosa and ... cells in glands; oxyntic, with parietal cells but no mucous cells in glands.12.

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In healthy volunteers, immunohistochemistry supports squamous to columnar metaplasia as mechanism of expansion of cardia, aggravated by central obesity Mohammad H Derakhshan,1 Elaine V Robertson,1 Yeong Yeh Lee,1,2 Tim Harvey,3 Rod K Ferrier,3 Angela A Wirz,1 Clare Orange,3 Stuart A Ballantyne,4 Scott L Hanvey,4 James J Going,3 Kenneth EL McColl1 1

Institute of Cardiovascular & Medical Sciences, University of Glasgow, Glasgow, UK 2 School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian, Kalantan, Malaysia 3 Institute of Cancer Sciences, University of Glasgow, Glasgow, UK 4 Gartnavel General Hospital, NHS Greater Glasgow & Clyde, Glasgow, UK Correspondence to Professor Kenneth E L McColl, Institute of Cardiovascular & Medical Sciences, 44 Church Street, Glasgow, G11 6NT, UK; [email protected] Received 28 November 2014 Revised 12 February 2015 Accepted 13 February 2015 Published Online First 9 March 2015

▸ gutjnl-2015-309407

To cite: Derakhshan MH, Robertson EV, Yeh Lee Y, et al. Gut 2015;64: 1705–1714.

ABSTRACT Introduction Recently, we showed that the length of cardiac mucosa in healthy volunteers correlated with age and obesity. We have now examined the immunohistological characteristics of this expanded cardia to determine whether it may be due to columnar metaplasia of the distal oesophagus. Methods We used the squamocolumnar junction (SCJ), antral and body biopsies from the 52 Helicobacter pylori-negative healthy volunteers who had participated in our earlier physiological study and did not have hiatus hernia, transsphincteric acid reflux, Barrett’s oesophagus or intestinal metaplasia (IM) at cardia. The densities of inflammatory cells and reactive atypia were scored at squamous, cardiac and oxyntocardiac mucosa of SCJ, antrum and body. Slides were stained for caudal type homeobox 2 (CDX-2), villin, trefoil factor family 3 (TFF-3) and liver–intestine (LI)-cadherin, mucin MUC1, Muc-2 and Muc-5ac. In addition, biopsies from 15 Barrett’s patients with/ without IM were stained and scored as comparison. Immunohistological characteristics were correlated with parameters of obesity and high-resolution pH metry recording. Results Cardiac mucosa had a similar intensity of inflammatory infiltrate to non-IM Barrett’s and greater than any of the other upper GI mucosae. The immunostaining pattern of cardiac mucosa most closely resembled non-IM Barrett’s showing only slightly weaker CDX-2 immunostaining. In distal oesophageal squamous mucosa, expression of markers of columnar differentiation (TFF-3 and LI-cadherin) was apparent and these correlated with central obesity (correlation coefficient (CC)=0.604, p=0.001 and CC=0.462, p=0.002, respectively). In addition, expression of TFF-3 in distal oesophageal squamous mucosa correlated with proximal extension of gastric acidity within the region of the lower oesophageal sphincter (CC=−0.538, p=0.001). Conclusions These findings are consistent with expansion of cardia in healthy volunteers occurring by squamo columnar metaplasia of distal oesophagus and aggravated by central obesity. This metaplastic origin of expanded cardia may be relevant to the substantial proportion of cardia adenocarcinomas unattributable to H. pylori or transsphincteric acid reflux.

Significance of this study What is already known on this subject?

▸ The length of the cardiac mucosa at the gastro-oesophageal junction is known to increase with age and central obesity. However, the mechanism of this and its relevance to the increasing incidence of adenocarcinoma at this site in subjects without reflux disease are unknown.

What are the new findings?

▸ In healthy volunteers without evidence of GORD, we have found that cardiac mucosa very closely resembles non-intestinal metaplasia Barrett’s mucosa immunohistochemically and with respect to intensity and nature of inflammatory infiltrate. ▸ We also found expression of markers of columnar differentiations (trefoil factor family 3 and liver–intestine cadherin) in most distal oesophageal squamous mucosa, which correlated with central obesity and proximal extension of gastric acidity within the lower oesophageal sphincter.

How might it impact on clinical practice in the foreseeable future?

▸ Our findings support expansion of cardia arising by squamocolumnar metaplasia of distal oesophagus and aggravated by central obesity. ▸ This metaplastic origin of the cardia may be relevant to the high incidence of adenocarcinoma at this anatomical site.

INTRODUCTION In the West, the gastric cardia and gastrooesophageal junction (GOJ) have a high incidence of adenocarcinoma, which is similar to or greater than the rest of the stomach and oesophagus.1 These junctional cancers share epidemiological characteristics with more proximal oesophageal adenocarcinoma, but have a weaker association

Derakhshan MH, et al. Gut 2015;64:1705–1714. doi:10.1136/gutjnl-2014-308914


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Oesophagus with GORD, the main aetiological factor in oesophageal adenocarcinoma.2 Only 29% of Western patients with junctional cancers have reflux symptoms compared with 16% of controls.3 The aetiology of junctional adenocarcinoma and the role of reflux in its development remain unclear. The cardiac mucosa between gastric oxyntic mucosa and oesophageal squamous mucosa is nearly always inflamed. This carditis has been attributed to either Helicobacter pylori infection or acid reflux4–6 but is also present in most people without H. pylori infection or symptoms or signs of reflux.4 Cardiac mucosa may also show intestinal metaplasia (IM) in the absence of H. pylori infection or GORD.7 IM at the cardia was observed in 16% of Caucasians having screening colonoscopy.7 Chandrasoma8 propose that cardiac mucosa is a response to reflux causing columnar metaplasia of the squamous mucosa of the most distal oesophagus, and is therefore a form of ultrashort Barrett’s oesophagus. In neonates, the cardiac mucosa is 2 mm of squamous mucosa, separate scores were assigned for proximal and squamous epithelium.

Inflammatory scoring of biopsies Biopsies were scored semiquantitatively for intensity of inflammation. For biopsies of the GOJ, each mucosal subtype present was analysed separately. Scores of 0–3 were given for inflammation, where 0=not inflamed, 1=mild inflammation, 2=moderate inflammation and 3=severe inflammation. Acute inflammation was defined by the neutrophil polymorphonuclear leucocytes and for chronic inflammation defined by mononuclear cell infiltration. Reactive atypia in squamous epithelium was defined as features associated with reflux oesophagitis, that is, basal layer thickening (>15%), connective tissue papillae >50% mucosal thickness, congestion and decreased surface maturation. In glandular mucosa, mucin depletion, reduced maturation towards the mucosal surface and increased cell proliferation are all reactive features, provided criteria for a diagnosis of dysplasia are not met. Scoring was performed independently by two persons (MD, JJG) blinded to clinical characteristics of the volunteers. Agreement was checked using a kappa test, which ranged between 0.74 and 0.89 ( p>0.01). In case of discrepancy between two observers, they reviewed the slides jointly for the second round and agreed on the final scores.

Statistical analysis Results are presented as either mean (+SD) or median (+IQR), based on symmetry of data distribution. Comparisons between groups were made using Mann–Whitney U test or independent groups T test, if appropriate. To compare two paired groups, we used related samples T test or related samples Wilcoxon signed-rank test, where appropriate. For correlations between continuous variables, Spearman’s rho bivariate correlations were used with significance taken at p

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