Food hypersensitivity is commonly reported, but often remains unexplained, despite extensive medical examinations. Patients with such subjective food hypersensitivity attribute a number of somatic health complaints to the ingestion of specific foods, most often cow’s milk, wheat products, egg, fruits, and vegetables.² The abdominal symptoms are typically consistent with irritable bowel syndrome.² Psychological disturbances are often associated,³ but do not seem to be major predictors of either gastrointestinal or extraintestinal symptom severity.⁴ Hence, the etiology of subjective food hypersensitivity remains obscure.

Alterations of the gastrointestinal enterochromaffin cell population has been described in patients with functional gastrointestinal disorders,⁵,⁶ and are conceivably implicated in the pathogenesis of subjective food hypersensitivity. In a preliminary study, we recently observed low serum levels of chromogranin A (CgA) in patients with subjective food hypersensitivity.⁷ CgA is stored and secreted together with amines and peptides from the diffuse neuroendocrine system, and serve as a precursor for several peptides with regulatory functions.⁸ Enterochromaffin cells are a major source for circulating CgA. In addition, enterochromaffin cells secrete 5-hydroxytryptamine (5-HT, serotonin), a signaling molecule regulating several gut functions, including secretion, motility, and sensitivity. Indeed, most of the body’s 5-HT is contained within enterochromaffin cells. High systemic levels of 5-HT may cause symptoms resembling hypersensitivity reactions to food, such as diarrhea,⁹,¹⁰ nausea,¹¹ or flushing and heart palpitations.¹²

5-HT is commonly measured in platelets or in plasma, the latter being a poor matrix due to rapid degradation of 5-HT into 5-hydroxyindolacetic acid. More than 95% of plasma 5-HT is present in platelets, and studies assessing platelet 5-HT content in patients with irritable bowel syndrome are somewhat conflicting.¹³–¹⁵ Alterations of 5-HT secretion within the gut are not necessarily reflected in the systemic circulation, and assessing 5-HT in gut tissue samples is often considered to be more accurate. However, the surface of the gastrointestinal tract is very large, and biopsies can only give information from a small fragment of this huge area. Indeed, enterochromaffin cell counts seem to vary a lot in different studies.¹⁶,¹⁷ Our group has developed a method for analyzing “whole gut 5-HT”, by analyzing gut lavage fluid.¹⁸ Enterochromaffin cells release 5-HT mainly from granules at the basal border, but studies have shown that enterochromaffin cells are bipolar and are able to secrete 5-HT both basally, into the systemic circulation, and apically, into the gut lumen.¹⁹–²¹ Our method quantifies luminal enterochromaffin cell secretion along the entire length of the small and large bowel.

The aim of the present study was to explore whether patients with subjective food hypersensitivity display abnormal enterochromaffin cell functions. To evaluate this, we extended our preliminary observations⁷ and employed our recently developed liquid chromatography tandem mass spectrometry (LCMS) method for determination of 5-HT in gut lavage fluid.¹⁸

Most intestinal enterochromaffin cells are of the “open” type with apical cytoplasmic extensions which project into the gut lumen and short microvilli,²⁶ where they sense and respond to the luminal contents by releasing regulatory compounds like 5-HT and CgA, both into the circulation and in the intestinal lumen. Hyperplasia of enterochromaffin cells together with an increase in 5-HT containing granules has been reported in several studies of both functional disorders and inflammatory diseases of the gut,¹⁴,²⁷,²⁸ and 5-HT is supposed to be involved in the pathogenesis of various gastrointestinal disorders.²⁹–³¹ However, in the present study, gut lavage 5-HT were found to be normal in patients with subjective food hypersensitivity. Patients with inflammatory bowel disease had slightly higher mean levels of 5-HT, but the difference was not significant. The high degree of variance in the 5-HT data may be due to high individual differences, as well as a consequence of the preanalytical sampling procedure. The use of gut lavage fluid for analytical purposes is unique. Based on our previous experience with the procedure,²²,²³ the first clear fluid was chosen because it was desirable to have as few particles as possible to avoid analytical problems, but we acknowledge that the time point of sampling was not necessarily the same for each participant.

Wang et al demonstrated an interesting immunoendocrine axis in the gut, where 5-HT production in enterochromaffin cells increases in response to secretory products from CD4⁺ T cells.³² Production and secretion of 5-HT from enterochromaffin cells may thus depend on the immunological profile of the immune response,³³ and there is also evidence that mucosal 5-HT modulates the immune response via serotonergic receptors expressed by immune cells.³⁴ Indeed, identification of multiple 5-HT receptor subtypes in the gut, especially 5-HT₃ and 5-HT₄ receptors, suggests multifaceted actions of 5-HT, and have led to the development of several therapeutic agents for functional gastrointestinal disorders.³⁵

Serum CgA was significantly lower in the patients with subjective food hypersensitivity compared with the healthy controls, which is consistent with our previous results.⁷ Sidhu et al have reported abnormally elevated levels of serum CgA in a small proportion of their patients with diarrhea-predominant irritable bowel syndrome.³⁶ However, they included no healthy controls, and in many of the patients with elevated CgA, the levels declined with time, which the authors explain as short-lived enterochromaffin cell hyperplasia, as previously reported by Dunlop et al.⁶ Thus, it would be of interest to perform serial measurements of CgA during a day, and also for several continuous days, in future studies to verify possible fluctuations in circulating CgA levels.

Although CgA is a major enteroendocrine secretion product, little is known about its potential role in gastrointestinal pathophysiology. CgA serves as a prohormone for shorter peptide fragments with regulatory properties.⁸ Fragments of CgA exert antimicrobial effects and may modulate gastrointestinal motility, sensitivity, and barrier function.³⁷,³⁸ The low levels of CgA demonstrated in the present study could be a consequence of alterations in the gut microbial flora in patients with subjective food hypersensitivity.³⁹,⁴⁰ Intriguingly, Dlugoz et al have recently demonstrated Chlamydia trachomatis antigens in enteroendocrine cells and macrophages of the small bowel in patients with irritable bowel syndrome.⁴¹ Although the clinical significance is yet unknown, this is an exciting finding that, together with the present finding of low serum levels of CgA, could imply impairment of enterochromaffin cell function in patients with subjective food hypersensitivity and irritable bowel syndrome. El-Salhy et al recently demonstrated decreased CgA-positive cells in biopsies from patients with irritable bowel syndrome.⁴² Thus, a decrease in numbers of CgA secretory cells could be an explanation for the low serum CgA found in the present study.

We conclude that measurements of systemic CgA reveal significantly lower levels in patients with subjective food hypersensitivity than in healthy controls, whereas “whole gut 5-HT” levels seem to be normal. Impaired secretion of CgA may play a role in functional gastrointestinal disorders, and warrants further investigation.

Disclosure

The authors report no conflicts of interest in this work.

KG, JV, KL, and GAL have research grants from the Western Norway Regional Health Authority and the University of Bergen. The authors are indebted to the patients for their participation.