Wheat-starch-based gluten-free products in the treatment of newly detected coeliac disease: prospective and randomized study

Authors

  • M. Peräaho,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • K. Kaukinen,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • K. Paasikivi,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • H. Sievänen,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • S. Lohiniemi,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • M. Mäki,

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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  • P. Collin

    1. Departments of *Medicine and §Paediatrics, Tampere University Hospital, Tampere (also *§Medical School, University of Tampere), †Bone Research Group, UKK Institute, Tampere, and ‡Finnish Coeliac Society, Tampere, Finland
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Dr P. Collin, Medical School, FIN-33014, University of Tampere, Finland.
E-mail: pekka.collin@uta.fi

Summary

Background : The safety of wheat-starch-based gluten-free products in the treatment of coeliac disease is debatable. Prospective studies are lacking.

Aim : To compare the clinical, histological and serological response to a wheat-starch-based or natural gluten-free diet in patients with newly detected coeliac disease.

Methods : Fifty-seven consecutive adults with untreated coeliac disease were randomized to a wheat-starch-based or natural gluten-free diet. Clinical response, small bowel mucosal morphology, CD3+, αβ+ and γδ+ intra-epithelial lymphocytes, mucosal human leucocyte antigen-DR expression and serum endomysial, transglutaminase and gliadin antibodies were investigated before and 12 months after the introduction of the gluten-free diet. Quality of life measurements were performed by standardized questionnaires and the bone mineral density was analysed.

Results : In both groups, abdominal symptoms were alleviated equally by a strict diet. There were no differences between the groups in mucosal morphology, the density of intra-epithelial lymphocytes, serum antibodies, bone mineral density or quality of life tests at the end of the study. Four patients on a natural gluten-free diet and two on a wheat-starch-based gluten-free diet had dietary lapses; as a result, inadequate mucosal, serological and clinical recovery was observed.

Conclusions : The dietary response to a wheat-starch-based gluten-free diet was as good as that to a natural gluten-free diet in patients with newly detected coeliac disease.

Introduction

A gluten-free diet affords clinical and small bowel mucosal recovery in coeliac disease. Patients who maintain a normal gluten-containing diet or have frequent dietary transgressions run an increased risk of osteoporosis1–3 and possibly of intestinal lymphoma.4, 5 However, it remains unproven whether trace amounts of gluten are detrimental, provided that the diet is, on the whole, strict.

Industrially purified wheat-starch-based gluten-free products meeting the current Codex Alimentarius Standard are allowed to contain up to 0.05 g nitrogen per 100 g of food product on a dry matter basis;6 therefore, these products may contain residual gluten up to 40–60 mg per 100 g of dry matter.7, 8

Theoretically, small amounts of gluten may be harmful in coeliac disease and, indeed, such products have evoked abdominal symptoms in an open challenge study.9 On the other hand, according to some studies, wheat-starch-based gluten-free flours are well tolerated and cause no small bowel mucosal deterioration.10–13 In some challenge studies, small bowel mucosal inflammation has appeared after the ingestion of 5 mg to 5 g of gluten,14, 15 whereas, in others, no significant mucosal changes have been observed.16–18 Due to this discrepancy, controversy persists as to the safety of wheat-starch-based gluten-free products: these have been used for over 30 years in the UK and Scandinavia, but their consumption is discouraged in the USA.19

All previous investigations concerning wheat-starch-based gluten-free products have been either open, short-term challenge studies9, 16, 17 or cross-sectional trials.10, 11, 13 In the event that wheat-starch-based gluten-free flours are harmful, one would expect mucosal recovery to be slower and inflammation to be sustained longer when these are used instead of natural gluten-free products. This prompted us to compare, in a randomized 1-year prospective study, the histological response to wheat-starch-based gluten-free and natural gluten-free diets in a group of patients with newly detected coeliac disease. We especially focused on minor mucosal inflammatory changes, and the clinical response to the diets was also evaluated.

Patients and methods

Subjects

Altogether, 65 adults were diagnosed as having coeliac disease at the Department of Medicine, Tampere University Hospital, between April 1998 and February 2000; eight refused to participate in the study. The remaining 57 were randomized to receive a wheat-starch-based or natural gluten-free diet. The randomization was carried out using random-number tables with permuted blocks.20 Each patient entered the trial before random treatment assignment was revealed. The diagnosis of all coeliac patients was based on severe partial or subtotal small bowel villous atrophy with crypt hyperplasia. The symptoms or signs leading to the diagnosis of coeliac disease, duration of symptoms, immunosuppressive medication, family history of coeliac disease and associated conditions were recorded. Clinical, histological and serological studies were carried out before and after the adoption of the gluten-free diet; the follow-up time was 1 year.

Fifty-nine adults who underwent upper gastrointestinal endoscopic examination due to indigestion or heartburn served as controls for histological analysis; all were consuming gluten and had no relatives with coeliac disease.

Dietary assessment

The dietitian provided advice on natural or wheat-starch-based gluten-free diets at the start of the study. A detailed dietary analysis was taken and a history of occasional or regular consumption of gluten-containing products was assessed by means of an interview and a 4-day record of food intake 3 and 9 months after adopting the diet. The dietitian evaluated the daily consumption of gluten-free flours (either natural or wheat-starch-based) in grams.

Small bowel biopsy

Small bowel biopsy specimens were taken by upper gastrointestinal endoscopy from the distal part of the duodenum at baseline and 12 months after commencing a gluten-free diet. Three to five biopsy specimens were processed and stained with haematoxylin–eosin and studied under light microscopy. Morphometric analysis, including the villous height to crypt depth ratio, enterocyte cell height and the density of intra-epithelial lymphocytes per 100 enterocytes, was performed for well-oriented biopsy samples, as described previously.17, 21 Poorly oriented sections were discarded; when necessary, the samples were dissected again until they were of good quality.

Frozen samples had not been taken routinely at the first endoscopy, but were still available in 20 of the 57 patients with newly detected coeliac disease. Immunohistochemical staining was performed in these 20 samples, and in all follow-up and control biopsy specimens. Two small bowel biopsy specimens were taken and freshly embedded in optimal temperature compound (Tissue-Tec, Miles Inc, Elkhart, IN, USA), snap frozen in liquid nitrogen and stored at − 70 °C. Immunohistochemical studies were carried out on 5-µm-thick frozen sections. The αβ+ intra-epithelial lymphocytes were stained with monoclonal βF1 antibody (Endogen, Woburn, MA, USA) and the γδ+ intra-epithelial lymphocytes with TcRγδ (Endogen). Intra-epithelial lymphocytes were counted with a × 100 flat-field light microscope objective in randomly selected surface epithelium; at least 30 fields with an epithelial length of 1.6 mm were counted, and the density of intra-epithelial lymphocytes was expressed as the number of cells per millimetre of epithelium.22, 23 Mucosal human leucocyte antigen (HLA)-DR expression was detected with monoclonal HLA-DR antibody (Becton Dickinson, San Jose, CA, USA) at a dilution of 1 : 1000. HLA-DR expression was considered to be enhanced (positive) when it was strong in the villous epithelium or expression was seen in the crypts; negative expression in the crypts and only slight to moderate staining in the villous epithelium was considered to be normal (negative).24

In our laboratory, the correlation coefficients for intra-observer variation for αβ+ and γδ+ intra-epithelial lymphocytes were 0.85 and 0.98, respectively, and those for inter-observer variation were 0.82 and 0.98, respectively. The intra-observer estimates of the enhanced up-regulation of small bowel HLA-DR expression were similar in 86% and the inter-observer estimates in 91%. In this study, all specimens were evaluated by the same investigator, who had no previous knowledge of the disease history or laboratory findings.

Serology and chemical analysis

Serum immunoglobulin A (IgA) class endomysial antibodies were determined by an indirect immunofluorescence method using human umbilical cord as a substrate;25 a dilution of 1 : ≥ 5 was considered to be positive. Serum IgA class tissue transglutaminase antibodies were investigated by enzyme-linked immunoabsorbent assay (ELISA) (Inova Diagnostics, San Diego, CA, USA), a unit value (U) of ≥ 20 being considered to be positive.26 Serum IgA class gliadin antibodies were measured by ELISA; the lower limit for positivity was 0.2 ELISA units per millilitre.27 Blood haemoglobin, serum iron, serum calcium, serum vitamin B12 and erythrocyte folic acid concentrations were measured using routine laboratory methods.

Clinical evaluation and bone mineral density

Gastrointestinal symptoms were evaluated by total score on the Gastrointestinal Symptom Rating Scale. This comprises 15 items describing abdominal pain (such as colicky pain, undefined pain), gastro-oesophageal reflux (epigastric pain, heartburn, acid regurgitation), indigestion (borborygmus, abdominal distension), diarrhoea and constipation (loose stools, hard stools, urgent need for defecation).28, 29 The quality of life was assessed by the Psychological General Well-Being Questionnaire.28, 29 The Psychological General Well-Being Index measures subjective well-being. It includes 22 items (anxiety, depressed mood, positive well-being, self-control, health and vitality); the total score gives a maximum value of 132 and a minimum of 22. The higher the score, the better the well-being. The body mass index was calculated using the formula: weight/height2 (kg/m2). The bone mineral density in the lumbar spine and the left femoral neck was measured by dual-energy X-ray absorptiometry (Norland XR26, Norland Corp, Fort Atkinson, WI, USA). The bone mineral density data were expressed as T scores with reference to data on sex-matched young individuals.

Statistics

Quantitative data were expressed as means and 95% confidence intervals (CI). A two-tailed t-test was used to compare the laboratory values between the groups. Cross-tabulations were carried out by Fisher's exact test. The required number of participants was determined to achieve a statistical power of 0.90 at a significance level of 0.05; differences of < 0.5 for the villous height to crypt depth ratio, ≥ 7/100 enterocytes for intra-epithelial lymphocytes or CD3+ cells, 3/mm for γδ+ cells and ≥ 0.5 for the Gastrointestinal Symptom Rating Scale were considered to be clinically relevant.

Ethical considerations

The study protocol was approved by the Ethical Committee of Tampere University Hospital. All subjects gave informed consent.

Results

Twenty-nine patients with newly detected coeliac disease were randomized to receive a natural gluten-free diet (group I) and 28 to receive a wheat-starch-based gluten-free diet (group II). The groups were similar with respect to age, gender and symptoms (Table 1). Two patients in group I decided to discontinue the study after 2 months, as they found the diet too convoluted. In addition, four patients in group I and two in group II did not follow a strict gluten-free diet. Thus, 23 in group I and 26 in group II completed the study with a proper diet. The mean consumption of gluten-free flours after 3 months of follow-up was 79 g/day (range, 20–186 g/day) in group I and 82 g/day (35–184 g/day) in group II; after 9 months, the corresponding values were 77 g/day (20–186 g/day) and 81 g/day (37–173 g/day), respectively.

Table 1.  Baseline characteristics of patients with newly diagnosed coeliac disease randomized to receive a natural or wheat-starch-based gluten-free diet (GFD)
 Natural GFD (n = 29)Wheat-starch-based GFD (n = 28)
  • IgA, immunoglobulin A.

  • First-degree relatives of patients with coeliac disease, diabetes mellitus, autoimmune thyroid disorders, Sjögren's syndrome, osteoporosis.

Female/male23/622/6
Median age (years) (range)47 (24–68)44 (22–69)
Symptoms or signs leading to the diagnosis of coeliac disease, n
 Abdominal symptoms1821
 Anaemia 5 2
 Arthritis/arthralgia, rash, dermatitis 3 1
 Screening in risk groups* 3 4
Median duration of symptoms (years) (range) 2 (0–30) 2 (0–20)
Other diseases, n
 Autoimmune thyroid disease 6 4
 Primary Sjögren's syndrome 2 1
 IgA glomerulonephritis 1 1
 Diabetes mellitus type I 1 0
 Sarcoidosis 0 1
 Collagenous colitis 0 1
Immunosuppressive medication, n 0 0
Family history of coeliac disease, n1310

The villous height to crypt depth ratio and enterocyte cell height increased and the density of intra-epithelial lymphocytes decreased equally in the groups on natural and wheat-starch-based gluten-free diets (Figure 1). On a gluten-free diet, small bowel mucosal αβ+ and γδ+ intra-epithelial lymphocytes and mucosal HLA-DR expression did not differ between the two study groups (Table 2). The density of γδ+ intra-epithelial lymphocytes and epithelial HLA-DR expression remained elevated in both groups compared with the values in non-coeliac controls. There were no significant differences in serum endomysial antibody, tissue transglutaminase antibody and gliadin antibody positivity, blood haemoglobin, serum iron, serum calcium, serum vitamin B12 and erythrocyte folic acid levels between the study groups (Table 3).

Figure 1.

Villous height to crypt depth ratio (a), density of intra-epithelial lymphocytes (IELs) (b) and enterocyte height (c) before (I) and after (II) 1 year of treatment in coeliac patients randomized to receive a natural or wheat-starch-based gluten-free diet (GFD). The differences before and after a gluten-free diet were statistically significant (P < 0.01), whereas the differences between treatment groups were not significant.

Table 2.  Small bowel mucosal intra-epithelial lymphocytes (IELs) and mucosal human leucocyte antigen (HLA) expression in coeliac disease patients adhering to a natural or wheat-starch-based gluten-free diet (GFD) for 1 year and in non-coeliac control subjects
 Before treatment
(n = 20)
Coeliac disease patientsNon-coeliac controls
(n = 59)
Natural GFD
(n = 23)
Wheat-starch-based GFD
(n = 26)
  • *

    P < 0.05 compared to non-coeliac controls.

αβ+ IELs (cells/mm), mean (95% CI)45 (38–53)*22 (18–26)21 (16–26)22 (18–25)
γδ+ IELs (cells/mm), mean (95% CI)13.2 (10.9–15.5)*8.9 (6.7–11.1)*9.4 (7.0–11.8)*2.3 (1.6–3.1)
Enhanced HLA-DR expression (n)15 (75%)*15 (60%)*16 (59%)*23 (39%)
Table 3.  Laboratory investigations before and after diet in coeliac disease patients randomized to receive a natural or wheat-starch-based gluten-free diet (GFD)
 ReferenceNatural GFD (n = 23)Wheat-starch-based GFD (n = 26)
Before GFDOn GFDBefore GFDOn GFD
  1. AGA, gliadin antibodies; EmA endomysial antibodies; tTg-ab, tissue transglutaminase antibodies.

  2. The differences between study groups were not statistically significant.

Haemoglobin (g/dL), mean (range)12–1812.6 (10.9–14.4)12.7 (11.1–14.5)12.9 (9.8–15.3)13.3 (10.8–14.9)
Serum calcium (mmol/L), mean (range)2.15–2.602.3 (2.12–2.45)2.3 (2.13–2.51)2.3 (2.08–2.54)2.3 (2.2–2.6)
Serum vitamin B12 (pmol/L),  mean (range)170–640301 (166–597)363 (191–661)271 (148–441)316 (127–602)
Erythrocyte folic acid (nmol/L),  mean (range)320–900398 (138–847)499 (293–1050)433 (120–632)585 (163–1200)
Serum iron (µmol/L), mean (range)6–3516.8 (3.6–46.2)19.0 (7.0–31.1)14.6 (3.3–22.8)17.4 (6.2–29.4)
Serum AGA, number of abnormal levels≤ 0.2 EU/L14/23 (61%)2/23 (9%)18/26 (70%)4/26 (15%)
Serum EmA, number of abnormal levels> 1 : 518/23 (78%)1/23 (4%)17/26 (65%)2/26 (8%)
Serum tTg-ab, number of abnormal levels≤ 20 U/L22/23 (96%)5/23 (22%)23/26 (88%)3/26 (12%)

Gastrointestinal symptoms were alleviated similarly in patients receiving a natural gluten-free diet or a wheat-starch-based gluten-free diet (Figure 2). Mean Gastrointestinal Symptom Rating Scale scores were 2.7 (95% CI, 2.4–3.0) in group I and 2.7 (2.4–2.9) in group II before treatment, and 1.8 (1.7–2.0) and 1.8 (1.6–2.0), respectively, at the end of the study. Psychological General Well-Being scores were 95.6 (90.4–100.8) in group I and 89.7 (83.4–95.4) in group II before treatment, and 109.9 (105.4–114.3) and 107.3 (103.2–111.4), respectively, at the end of the study (Figure 2). The body mass index was 24.8 in group I before treatment and 24.8 after treatment; in group II, the corresponding values were 26.5 and 26.8, respectively. At baseline and after 1 year on a gluten-free diet, there were no significant differences in bone mineral density (Table 4).

Figure 2.

Gastrointestinal symptoms (a) and general well-being (b) before (I) and after (II) 1 year of treatment in coeliac patients randomized to receive a natural or wheat-starch-based gluten-free diet (GFD). A higher score on the Gastrointestinal Symptom Rating Scale (GSRS) indicates more intestinal symptoms, and a higher score on the Psychological General Well-Being (PGWB) scale indicates a better quality of life. The differences before and after a gluten-free diet were statistically significant (P < 0.01), whereas the differences between treatment groups were not significant.

Table 4.  Bone mineral density (T score with 95% confidence intervals) before and after 1 year of treatment in patients randomized to receive a natural or wheat-starch-based gluten-free diet (GFD)
Diet groupLumbar spineFemoral neck
Before GFDAfter GFDBefore GFDAfter GFD
Natural GFD (n = 23)− 1.27; − 0.91, − 1.64− 0.72; − 0.29, − 1.14− 0.86; − 0.48, − 1.23− 0.85; − 0.52, − 1.17
Wheat-starch-based GFD (n = 26)− 1.01; − 0.58, − 1.45− 0.86; − 0.34, − 1.38− 0.43; − 0.12, − 0.74− 0.44; − 0.13, − 0.76

A slower response to treatment was not dependent on the flour intake in compliant patients (data not shown). Of the six non-compliant patients, two of three had villous atrophy at the end of the study; three patients refused biopsy, but had elevated antibody levels; five had anaemia.

Discussion

We have shown, for the first time, in a randomized prospective study that a natural gluten-free diet and a wheat-starch-based gluten-free diet produce a similar histological and clinical recovery in patients with newly detected coeliac disease. Full mucosal recovery may take more than 1 year,30 and the density of intra-epithelial lymphocytes has been considered to be the most sensitive gluten-induced intestinal mucosal change in coeliac disease.15, 18, 31, 32 In particular, the density of αβ+ intra-epithelial lymphocytes decreases when a gluten-free diet is adhered to, and increases again during gluten challenge.33, 34 It is therefore important that the densities of αβ+ intra-epithelial lymphocytes were similar in both groups. The γδ+ intra-epithelial lymphocytes remained at a high level, but decreased from baseline, which has been shown to occur previously by dietary treatment.35 Although mucosal recovery was not complete in all patients, all histological findings indicated that there were no differences in the healing rate between the study groups after 1 year on a gluten-free diet.

In this study, wheat-starch-based gluten-free products were well tolerated. This is in contrast with the open challenge study by Chartrand et al., where these products evoked more abdominal symptoms.9 In a study by Faulkner-Hogg et al., abdominal symptoms were alleviated when a wheat-starch-containing gluten-free diet was changed to a natural gluten-free diet.36 However, 36% of the patients had small bowel villous atrophy and there were no significant changes in mucosal morphology after switching to a natural gluten-free diet. It should be noted that neither of these two studies was randomized.9, 36

Due to the wide variety of products available, it was virtually impossible to evaluate the exact amounts of gluten consumed in the two diets during 1 year. We therefore assumed that the natural gluten-free diet contained no gluten, but it is possible that these products may also be gluten contaminated. This study does not answer the question of whether minor amounts of gluten produce inflammation, but it shows that wheat-starch-based gluten-free products are as safe and well tolerated as natural ones.

Repetitive dietary transgressions resulted in incomplete small bowel mucosal recovery, and this may clearly expose patients to health risks. It thus appears that the overall compliance with diet is much more important than the trace amounts of gluten possibly present in wheat-starch-based gluten-free products.

Acknowledgements

This study was supported by grants from the Yrjö Jahnsson Foundation and the Medical Research Fund of Tampere University Hospital.

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