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Background: Epithelial barrier defects are well known in coeliac disease, but the mechanisms are only poorly defined. It is unclear, whether barrier disturbance reflects upregulated epithelial transcytosis or paracellular leakage., Objective: To characterise the molecular structure and function of the epithelial tight junction (TJ) and mechanisms of its dysregulation., Methods: Molecular analysis of proteins involved in TJ assembly and their regulation was performed by western blotting and confocal microscopy correlated to electrophysiology., Results: A complex alteration of the composition of epithelial TJ proteins (with more pore-forming claudins like claudin-2 and a reduction in tightening claudins like claudin-3, -5 and -7) was found for protein expression and subcellular localisation, responsible for an increase in paracellular biotin-NHS uptake. In contrast, epithelial apoptosis was only moderately elevated (accounting for a minor portion of barrier defects) and epithelial gross lesions--for example, at cell extrusion zones, were absent. This TJ alteration was linked to an altered localisation/expression of proteins regulating TJ assembly, the polarity complex protein Par-3 and the serine-/threonine phosphatase PP-1., Conclusions: Changes in cell polarity proteins Par-3 and PP-1 are associated with altered expression and assembly of TJ proteins claudin-2, -3, -5 and -7 and ZO-1, causing paracellular leakage in active coeliac disease.

Background: Coeliac disease (gluten-sensitive enteropathy; GSE) and inflammatory bowel disease (IBD) are common gastrointestinal disorders. Both display enhanced intestinal permeability, initiated by gluten exposure (GSE) or bacterial interactions (IBD). Previous studies showed the association of both diseases with variants in MYO9B, presumably involved in epithelial permeability., Aim: It was hypothesised that genetic variants in tight junction genes might affect epithelial barrier function, thus contributing to a shared pathogenesis of GSE and IBD., Methods: This hypothesis was tested with a comprehensive genetic association analysis of 41 genes from the tight junction pathway, represented by 197 tag single nucleotide polymorphism (SNP) markers., Results: Two genes, PARD3 (two SNPs) and MAGI2 (two SNPs), showed weak association with GSE in a Dutch cohort. Replication in a British GSE cohort yielded significance for one SNP in PARD3 and suggestive associations for two additional SNPs, one each in PARD3 and MAGI2. Joint analysis of the British and Dutch data further substantiated the association for both PARD3 (rs10763976, p = 6.4 x 10(-5); OR 1.23, 95% CI 1.11 to 1.37) and MAGI2 (rs6962966, p = 7.6 x 10(-4); OR 1.19, 95% CI 1.08 to 1.32). Association was also observed in Dutch ulcerative colitis patients with MAGI2 (rs6962966, p = 0.0036; OR 1.26, 95% CI 1.08 to 1.47), and suggestive association with PARD3 (rs4379776, p = 0.068)., Conclusions: These results suggest that coeliac disease and ulcerative colitis may share a common aetiology through tight junction-mediated barrier defects, although the observations need further replication.

Background/aims: Whether a life-long gluten-free diet (GFD) is necessary in all children with diagnosed coeliac disease (CD) remains debated. To address this question, a retrospective analysis of the clinical and biological status of adult coeliac patients diagnosed in childhood, who remained on a normal diet after gluten challenge and were clinically silent, was carried out., Methods: Patients aged 18-65 years with CD diagnosed in childhood were included. Clinical status, gluten intake, biological parameters of malabsorption, bone mineral density, human leucocyte antigen (HLA) genotype, serological markers of CD, and histological and immunohistochemical parameters in duodenal biopsies were recorded., Results: Sixty-one patients had resumed a normal diet and were asymptomatic. Forty-eight showed different degrees of villous atrophy (silent CD), while 13 had no detectable atrophy (latent CD) on duodenal biopsies. Latent CD patients had significantly less osteopenia/osteoporosis (1/9 (11%) vs 23/33 (70%), p<0.001)), and lower T cell receptor (TCR) alphabeta+ intraepithelial T cell counts (38+/-20 vs 55+/-15, p<0.01) than silent CD patients. The mean age at diagnosis and first GFD was lower in latent than in silent patients (14.4+/-5 vs 40.1+/-47 months, p<0.05). Latent patients did not differ significantly from the seven control patients on a long-term GFD, except for a higher frequency of CD-specific serum antibodies. However, two latent patients relapsed clinically and histologically during subsequent follow-up., Conclusions: Long-term latency developed in about 20% of CD patients who remained symptom free after gluten reintroduction. This latency can be transient and thus a regular follow-up is mandatory. In silent patients, the increased risk of osteoporosis substantiates the need for a GFD.

Background: Villus atrophy is the most distinctive sign of untreated coeliac disease (CD) and epithelial apoptosis is considered to be involved in this stage of the coeliac lesion. The extent of villus atrophy is, however, not homogeneous and patients with patchy or mild lesions have been described., Aims: To address: (a) the degree of "patchiness" in untreated CD patients; and (b) to clarify if apoptosis, and eventually which trigger drives it, causes epithelial damage., Patients: Twenty of 40 untreated, 14 treated coeliac patients, and 15 controls received five or more multiple duodenal biopsies; the remaining 20 untreated CD patients had no more than three biopsies., Methods: All biopsies were analysed to monitor the presence of a "flat" mucosa. Biopsies of 14 untreated, 10 treated coeliacs, and seven controls were cultured with or without gliadin. DNA fragmentation was studied by terminal deoxynucleotidyl transferase (TdT) mediated dUTP digoxigenin nick end labelling (TUNEL), and FAS and Ki67 expression by immunohistochemistry. Antiendomysium antibodies (EMA) were surveyed in biopsy culture supernatants., Results: A pattern of patchy duodenal lesions was observed in all untreated CD patients biopsied up to five times. High enterocyte FAS expression, and a high number of TUNEL+ and Ki67+ enterocytes were detected in areas with villus atrophy but not in those with a normal morphology (p<0.001). Conversely, EMA in culture supernatants and signs of immunological activation were present in all untreated CD biopsies. In vitro gliadin challenge increased the number of TUNEL+ and Ki67+ enterocytes (p<0.001 v cultures with medium alone) only in "flat" biopsies. Neutralising anti-FAS monoclonal antibodies were found to control gliadin induced enterocyte apoptosis (p>0.01) while agonist anti-FAS monoclonal antibody increased it (p<0.001)., Conclusions: Patchy lesions are observed in untreated CD mucosa and epithelial FAS engagement is a key trigger in driving villus atrophy in CD.

Background: Coeliac disease (CD) mucosa is flattened despite epithelial hyperproliferation., Aims: To establish mechanisms of cell loss in CD., Patients: 14 controls, 17 active CD patients, and 16 maintained with gluten free diet., Methods: Programmed cell death was examined in small intestinal biopsy specimens by staining fragmented DNA using terminal uridine deoxynucleotidyl nick end labelling (TUNEL), in comparison with haematoxylin and eosin stained adjacent sections. Double staining with anti-CD45 antibodies determined the origin of apoptotic cells. Apoptosis was graded from 1-3 (< 5, 5-20, > 20% respectively). Proliferating cells, immunostained by Ki-67 (MIB-1) antibody, were counted., Results: Apoptotic cells were seen rarely by haematoxylin and eosin but more readily by TUNEL. In controls, 1.4 +/- 0.2% of epithelial cells were apoptotic (mean grade 1.1), mainly located in the upper villus. In active CD, frequent apoptotic cells were distributed throughout the crypt-villus unit (mean grade 2.4), decreasing after treatment to 1.1 (p < 0.001) even when still histologically abnormal. CD45 antibodies rarely stained apoptotic cells in active CD. The number of TUNEL positive cells correlated with proliferating cell number (p < 0.001)., Conclusion: Enterocyte apoptosis is greatly increased in untreated CD, correlates with proliferation, and falls to normal with a gluten free diet, before histological improvement. Increased apoptosis may be responsible for villous atrophy in CD.

Background: Current knowledge on splanchnic haemodynamics in coeliac disease is limited and incomplete., Aim: To evaluate splanchnic arterial and venous blood flow in coeliac disease., Methods: In 22 coeliac (13 untreated, nine treated) patients and in nine healthy subjects the following variables were assessed: vessel diameter and mean flow velocity in portal vein, splenic vein, superior mesenteric vein, and superior mesenteric artery. Peak systolic velocity, end diastolic velocity and pulsatility index were also determined in the superior mesenteric artery. Five patients of the untreated group were re-evaluated after nine months on a gluten free diet., Results: Significant differences in haemodynamic variables between the three groups were shown only in the superior mesenteric artery. An increase in both mean flow velocity and end diastolic velocity and a reduction in pulsatility index occurred in untreated patients compared with treated patients (p < 0.002; p < 0.04; p < 0.035) and with healthy controls (p < 0.001; p < 0.025; p < 0.0003). Similar results were obtained for the five patients evaluated before and after treatment (p < 0.03; p < 0.02; p < 0.03), in whom the mean flow velocity in the superior mesenteric vein also decreased after treatment (p < 0.05). No significant differences were noted between treated coeliac patients and healthy controls., Conclusions: In untreated coeliac disease there is a hyperdynamic mesenteric circulation that decreases after treatment.

To evaluate the effects of a gluten free diet on bone mineral density in untreated adult patients with coeliac disease, 63 patients (17-79 years, 35 women) were examined at diagnosis and after one year taking a gluten free diet. Bone mineral density was measured in the forearm using single photo absorptiometry and in the lumbar spine, femoral neck, and trochanter using dual energy x ray absorptiometry. The values for each patient were compared with those of 25 healthy controls, matched for sex, age, and menopausal state. Before being given a gluten free diet bone mineral density in the total group was reduced at all sites (p < 0.001). Age adjusted bone mineral density was inversely correlated with age. During the first year taking a gluten free diet bone mineral density increased at all sites (p < 0.01). This was seen in patients of all ages and in patients who were without symptoms of malabsorption (weight loss or diarrhoea) before treatment. Low bone mineral density in patients with untreated coeliac disease increases rapidly when treatment with a gluten free diet is followed. These findings emphasise the importance of early diagnosis and treatment in all patients with coeliac disease.

Epithelial barrier function and ion transport was studied in coeliac sprue using a miniaturised Ussing device for measurements on diagnostic aspiration biopsy specimens from the jejunum of untreated or gluten free nourished sprue patients, or from healthy controls. Pure epithelial resistance (Re) indicating epithelial barrier function was determined by transmural alternating current impedance analysis. It was reduced by 56% in acute sprue mean (SEM) (9 (1) omega.cm2) compared with controls (20(2) omega.cm2). In gluten free nourished sprue patients Re was only partly recovered (15 (1) omega.cm2). Subepithelial resistance (Rsub) was also changed from 28 (1) omega.cm2- in control to 17 (1) omega.cm2 in acute sprue because of the change in mucosal architecture, but was unchanged in gluten free nourished sprue patients (29 (4) omega.cm2). In acute sprue, unidirectional Na+ and Cl- fluxes were increased in both directions as a consequence of the decreased resistance. However, short circuit current (ISC) as well as Na+ and Cl- net fluxes were not significantly different from control. Subsequently, the electrogenic Cl- secretory system was investigated. After maximal stimulation with theophylline and prostaglandin E1, a Cl(-)-dependent increase in ISC was obtained in the sprue mucosa and control jejunum. It showed saturation characteristics and was blockable by serosal bumetanide. When compared with control, neither Km nor Vmax of this electrogenic Cl- secretion was significantly changed in coeliac sprue. In conclusion, a miniaturised Ussing device was used for transport measurements on intestinal biopsy specimens. In acute coeliac disease, the epithelial barrier of the jejunum was seriously disturbed. The active electrogenic Cl- secretory transport system was present in the sprue mucosa, but was not activated in the Ussing chamber in vitro when compared with control jejunum.

Intestinal function is poorly defined in patients with HIV infection. Absorptive capacity and intestinal permeability were assessed using 3-O-methyl-D-glucose, D-xylose, L-rhamnose, and lactulose in 88 HIV infected patients and the findings were correlated with the degree of immunosuppression (CD4 counts), diarrhoea, wasting, intestinal pathogen status, and histomorphometric analysis of jejunal biopsy samples. Malabsorption of 3-O-methyl-D-glucose and D-xylose was prevalent in all groups of patients with AIDS but not in asymptomatic, well patients with HIV. Malabsorption correlated significantly (r = 0.34-0.56, p < 0.005) with the degree of immune suppression and with body mass index. Increased intestinal permeability was found in all subgroups of patients. The changes in absorption-permeability were of comparable severity to those found in patients with untreated coeliac disease. Jejunal histology, however, showed only mild changes in the villus height/crypt depth ratio as compared with subtotal villus atrophy in coeliac disease. Malabsorption and increased intestinal permeability are common in AIDS patients. Malabsorption, which has nutritional implications, relates more to immune suppression than jejunal morphological changes.

A cross sectional study was undertaken to examine the relationship between coeliac disease and bone mineral density. The 135 female coeliac patients registered on the database of the Department of Gastroenterology at Hull Royal Infirmary were approached by letter, advising them of a potential risk of osteoporosis and inviting them to undergo bone densitometry. A total of 81 registered women (60%) attended the Osteoporosis Laboratory, Princess Royal Hospital and underwent dual energy x ray absorptiometry at the lumbar spine (L2-L4) and femoral neck. Historical data relating to the time of diagnosis and adherence to a gluten free diet were obtained. A control group was selected from the local normal population and was first matched for height, weight, and menopausal status. Postmenopausal patients were then further matched to controls of equivalent menopausal age. In coeliac patients, bone mineral density expressed in g/cm2 as mean (SD) was significantly lower at the lumbar spine (1.076 (0.186)) than in the control group (1.155 (0.143), p < 0.001). This was also the case at the femoral neck (0.887 (0.142) versus 0.965 (0.127), p < 0.001). When the coeliac patients were stratified by menopausal status, it was found that femoral neck bone mineral density was significantly below control values in both premenopausal and postmenopausal women. Spinal bone mineral density exhibited a significant decrement only in the postmenopausal group. The age at diagnosis of coeliac disease and adherence to a gluten free diet did not influence bone mineral density at either hip or spine. These results confirm coeliac patients' higher risk of osteopenia. Coeliac disease should be added to the list of medical conditions which constitute an indication for bone densitometry in order that the individual risk of osteoporosis related fracture may be determined.

Patients with coeliac disease may present with calcium malabsorption but it is unclear whether this results in longterm impairment of bone mineralisation. Dual energy x ray absorptiometry (DXA) was used to study bone mineral density in 34 asymptomatic coeliac disease patients, treated with a gluten free diet for at least two years, and also in 10 newly diagnosed or untreated patients. As expected, untreated patients had low bone mineral density in all regions. In the 29 treated female coeliac disease patients, overall mean values for age adjusted bone mineral density expressed as Z scores were normal although there were many patients with low values, particularly of the lumbar spine and total body. Scores in the postmenopausal patients were significantly worse than in the premenopausal patients and low mean Z scores were found in the five treated male patients. The subjects who had reduced bone mineral density could not be predicted clinically but, despite being asymptomatic, were more likely to have subtotal or partial villous atrophy on small intestinal biopsy (p < 0.0275). In conclusion, although many treated coeliac disease patients have normal bone mineral density, suboptimally treated and newly diagnosed or untreated patients have osteopenia. To reduce the risk of osteoporotic fractures, it is recommended that bone mineral density be measured in all treated coeliac disease patients and those with osteopenia have a repeat intestinal biopsy to assess disease activity.

Forty five women and 10 men with coeliac disease diagnosed in adult life, who were already on a gluten free diet, had serial bone mineral density measurements at the lumbar spine and femoral neck over 12 months. Osteoporosis, defined as a bone mineral density (BMD) < or = 2 SD below the normal peak bone mass was found in 50% of male and 47% of female coeliac patients. Patients with a BMD < or = 2 SD below age and sex matched normal subjects, had a significantly lower body mass index (21.3 kg.m-2 compared with 25.2 kg.m-2, p < 0.02 Wilcoxon rank sum test) and lower average daily calcium intake (860 mg/day compared with 1054 mg/day, p < 0.05 Wilcoxon rank sum test) than patients with normal bone mineral density. In postmenopausal women with coeliac disease there was a strong correlation between the age at menopause and BMD at both the lumbar spine (r = 0.681, p < 0.01, Spearman's rank correlation) and femoral neck (r = 0.632, p < 0.01). No overall loss of bone was shown over the 12 months of follow up, and relative to the reference population there was a significant improvement in BMD at the lumbar spine in women (p < 0.025, paired t test) and at the femoral neck in men (p < 0.05, paired t test). There was a significant negative correlation between the annual percentage change in BMD at the lumbar spine and the duration of gluten free diet (r = -0.429, p<0.01, Spearman's rank correlation), with the largest gain in BMD in patients with most recently diagnosed coeliac disease. Osteoporosis was shown in 47% of patients with treated adult coeliac disease. Recognised risk factors for osteoporosis in the general population including low body mass index, dietary calcium intake, and early menopause are particularly important in coeliac disease. Treatment of coeliac disease with a gluten free diet probably protects against further bone loss, and in the early stages is associated with a gain in bone mineral density.

Cholecystokinin (CCK) release and gall bladder emptying in response to a fatty meal are completely abolished in coeliac disease. To determine the effect of lipid digestion on CCK release and gall bladder motility, six patients with untreated coeliac disease and a flat jejunal mucosa were studied on two separate days. After an overnight fast, the plasma CCK concentration and gall bladder volume were measured before and at regular intervals after the intraduodenal instillation of 60 ml corn oil (triglycerides) incubated with 40 ml saline or with 40 ml bile and pancreatic juice. The mean (SEM) concentration of free fatty acids in the aqueous phase of corn oil after incubation with bile and pancreatic juice (predigested corn oil) was 78 (35) mM compared with 0.1 (0.1) mM in the aqueous phase of corn oil incubated with saline (undigested corn oil). Integrated plasma CCK in response to predigested corn oil was significantly greater than that in response to undigested corn oil (101 (18) pM. 80 min v-2 (9) pM.80 min; p < 0.005). Similarly, integrated gall bladder contraction in response to predigested corn oil was significantly larger than that after undigested corn oil (817 (210) ml. 80 min v-225 (243) ml. 80 min; p < 0.05). In contrast to undigested corn oil, corn oil that has been predigested with bile and pancreatic juice induces plasma CCK secretion and gall bladder contraction in patients with untreated coeliac disease, presumably by generating and rendering soluble lipolytic products.

This study was designed to determine the extent of pancreatic insufficiency in untreated coeliac disease and whether pancreatic secretion is impaired after a prolonged gluten free period. Three groups of patients were studied: group A comprised 44 patients, mean (SD) age 4.0 (3.1) years, with coeliac disease and total or subtotal atrophy of the intestinal mucosa; group B comprised 67 patients, mean age 4.4 (3.0) years, with coeliac disease but with normal morphology of the intestinal villi (after 12.9 months of a gluten free diet); group C comprised 49 control subjects, mean age 3.2 (3.0) years, with normal jejunal histology. In all subjects exocrine pancreatic function was determined by the secretin-caerulein test; bicarbonate concentration and lipase, phospholipase, and chymotrypsin activity were measured after an intravenous injection of secretin 1 clinical unit (CU) + caerulein 75 ng/kg body weight. Faecal chymotrypsin concentration was also assayed. No significant difference was found between values of the duodenal output of pancreatic enzymes and bicarbonate obtained in the three groups; however, 10 of 44 untreated coeliac patients showed tryptic or lipolytic activity, or both, below the normal limit for our laboratory. The mean value of the faecal chymotrypsin concentration was significantly lower in untreated than in treated coeliac patients (p less than 0.0001) or in control subjects (p less than 0.0001). It is concluded that untreated coeliac patients may have pancreatic deficiency independent of a decrease in enterohormone release. No primary or secondary pancreatic insufficiency was found in coeliac patients where the intestinal mucosa had returned to normal.

The possibility that malabsorbed fat passing through the human ileum exerts an inhibitory feedback control on jejunal motility has been investigated in 24 normal subjects by perfusing the ileum with a fat containing solution designed to produce ileal luminal fat concentrations similar to those in steatorrhoea (30-40 mg/ml). Mean transit times through a 30 cm saline perfused jejunal segment were measured by a dye dilution technique. Thirty minutes after ileal fat perfusion, mean transit times rose markedly to 18.9 +/- 2.5 minutes from a control value of 7.5 +/- 0.9 minutes (n = 5; p less than 0.05). This was associated with an increase in volume of the perfused segment which rose to 175.1 +/- 22.9 ml (control 97.6 +/- 10.3 ml, n = 5; p less than 0.05). Transit times and segmental volumes had returned towards basal values 90 minutes after completing the fat perfusion. Further studies showed that ileal fat perfusion produced a pronounced inhibition of jejunal pressure wave activity, percentage duration of activity falling from a control level of 40.3 +/- 5.0% to 14.9 +/- 2.8% in the hour after ileal perfusion (p less than 0.01). Ileal fat perfusion was associated with marked rises in plasma enteroglucagon and neurotensin, the peak values (218 +/- 37 and 68 +/- 13.1 pmol/l) being comparable with those observed postprandially in coeliac disease. These observations show the existence in man of an inhibitory intestinal control mechanism, whereby ileal fat perfusion inhibits jejunal motility and delays caudal transit of jejunal contents.

The prevalence of hypogonadism, sexual dysfunction and abnormalities of semen quality was determined in 28 consecutive males with coeliac disease. These observations were related to jejunal morphology and nutritional status, and were compared with findings in 19 men with Crohn's disease of similar age and nutritional status. Two of the 28 coeliacs (7%) had clinical evidence of hypogonadism but impotence and decreased sexual activity occurred more commonly, the latter apparently improving after gluten withdrawal. Of the married coeliacs, 19% had infertile marriages, a value greater than expected in the general population. Hypogonadism and sexual dysfunction were not detected in our patients with Crohn's disease. Seminal analysis in coeliacs revealed marked abnormalities of sperm morphology and motility, but only the former appeared to improve after gluten withdrawal. Similar abnormalities, however, were also detected in patients with Crohn's disease, although, unlike the coeliacs, 46% also had reduced concentrations of spermatozoa. Semen quality in coeliac disease could not be clearly related to general or specific (serum vitamin B(12) and red cell folate) nutritional deficiencies or to fertility, although sperm motility was markedly reduced in two of the three coeliacs with infertile marriages. The presence of antisperm antibodies did not appear to be an important aetiological factor in male infertility in coeliac disease. The pathogenesis of infertility and sexual dysfunction in coeliac disease remains unclear, suggesting that factors such as endocrine dysfunction or other specific nutritional deficiency may be involved.

We evaluated serum cationic trypsinogen as a marker of exocrine pancreatic function in children without cystic fibrosis. The ability of this test to determine steatorrhoea of pancreatic origin, and its relationship to a wide range of exocrine pancreatic function were assessed. Serum trypsinogen was measured in 32 children with steatorrhoea, 10 with pancreatic and 22 with non-pancreatic causes. In patients with pancreatic steatorrhoea, serum cationic trypsinogen was 4.9 +/- 4.9 micrograms/l (mean +/- SD), significantly below values in patients with non-pancreatic steatorrhoea (47.0 +/- 22.1 micrograms/l, p less than 0.001) and 50 control subjects (31.4 +/- 7.4 micrograms/l, p less than 0.001). Serum cationic trypsinogen values in patients with pancreatic steatorrhoea all fell below the lower limit of our control range and below all values for patients with non-pancreatic steatorrhoea. Serum cationic trypsinogen was also evaluated against pancreatic trypsin output in 47 patients (range 0.2-17.0 yr who underwent a hormonal pancreatic stimulation test. In 17 patients, serum cationic trypsinogen was low (less than -2SD or less than 16.6 micrograms/l), and associated with greatly impaired pancreatic trypsin output, ranging from 0-8% of mean normal trypsin output. Five of these 17 patients did not have steatorrhoea. In 30 patients with normal or raised serum cationic trypsinogen (greater than or equal to 16.6 micrograms/l), pancreatic trypsin output ranged from 15-183% of mean normal values. In conclusion, low serum cationic trypsinogen suggests severely impaired exocrine pancreatic function, with sensitivity extending above the steatorrhoeic threshold. In the presence of steatorrhoea, low serum cationic trypsinogen indicates a pancreatic aetiology. Normal serum cationic trypsinogen, however, does not exclude impaired pancreatic function, above the steatorrhoeic threshold.

Hypogonadism, infertility, and sexual dysfunction occur in some men with coeliac disease. We have measured plasma testosterone, dihydrotestosterone, sex-hormone binding globulin, oestradiol, and serum luteinising hormone in 41 men with coeliac disease and have related these findings to jejunal morphology, fertility, semen quality, and sexual function. To determine the specificity of these observations in coeliacs we also studied 19 nutritionally-matched men with Crohn's disease, and men with chronic ill-health due to rheumatoid arthritis and Hodgkin's disease. The most striking endocrine findings in untreated coeliacs were increased plasma testosterone and free testosterone index, reduced dihydrotestosterone (testosterone's potent peripheral metabolite), and raised serum luteinising hormone, a pattern of abnormalities indicative of androgen resistance. As jejunal morphology improved hormone levels appeared to return to normal. This specific combination of abnormalities was not present in any of the disease control groups and, to our knowledge, androgen resistance has not been described previously in any other non-endocrine disorder. Plasma oestradiol concentration was modestly raised in 10% of coeliacs and 11% of patients with Crohn's disease. Unlike plasma androgens and serum luteinising hormone in coeliacs, plasma oestradiol was not clearly related to jejunal morphology. Androgen resistance and associated hypothalamic-pituitary dysfunction appear to be relatively specific to coeliac disease and cannot be explained merely in terms of malnutrition or chronic ill-health. In addition, our findings suggest that this endocrine disturbance may be related to sexual dysfunction in coeliac disease but its relationship to disordered spermatogenesis in this condition has not been clearly established.

Intestinal permeability has been studied in 21 patients with coeliac disease in relapse and after gluten withdrawal using an oral test of intestinal permeability based on the simultaneous oral administration of two probe molecules. The increased absorption of the larger molecule (cellobiose) and the decreased absorption of the smaller (mannitol) found in untreated coeliac disease both returned to normal within five months of starting treatment, the abnormality in cellobiose absorption correcting more rapidly than that of mannitol. After exposure to a single oral dose of gluten, the intestinal permeability of six patients with treated coeliac disease became transiently abnormal with an increased absorption of cellobiose, returning to normal within one week. The possible structural and functional implications of these findings are discussed. The cellobiose/mannitol ratio appears to be of value in assessing the response to gluten withdrawal in coeliac disease, and also in monitoring patients who are already established on a gluten free diet by detecting dietary lapses and 'non-responding coeliac disease'. It may also offer an alternative to jejunal biopsy in patients subjected to gluten challenge.

Patients with ulcerative colitis and coeliac disease who had been shown by impaired clearance of heat damaged red cells to have diminished splenic phagocytosis, were examined for evidence of more generalised reticuloendothelial malfunction by measuring their circulatory clearance of micro-aggregated albumin. Although in animals micro-aggregated albumin is largely removed by Kupffer cells, we found impaired clearance in otherwise normal subjects who had previously had surgical splenectomy. In patients with hyposplenism because of bowel disease there was no additional impairment of micro-aggregated albumin clearance, indicating that their hyposplenism is an isolated phenomenon and not part of a generalised reticuloendothelial atrophy. Patients with coeliac disease and normal splenic function had increased reticuloendothelial catabolic activity; this was not present in patients with coeliac disease and abnormal splenic function.

We have investigated the possibility that the abnormally decreased gall bladder contraction after meals in patients with coeliac disease might result in part from an abnormality in the gall bladder response to endogenous cholecystokinetic hormones--for example, cholecystokinin and motilin--rather than solely from decreased secretion of such hormones. Eight patients with untreated coeliac disease and nine controls received intravenous infusions of the pure synthetic cholecystokinin analogue caerulein, 2-16 ng/kg/hour. Gall bladder emptying was measured on a minute-by-minute basis using 99mTc-HIDA scans. In the patients with coeliac disease, gall bladder emptying was greatly decreased (34.6 +/- 9.9 v 61.5 +/- 7.5% at 60 minutes, p less than 0.02), and a much greater dose of caerulein was needed to initiate gall bladder contraction (3.80 +/- 1.08 v 1.49 +/- 0.56 ng/kg, p less than 0.02). These results suggest that the abnormal gall bladder contraction in coeliac disease is not simply because of impaired release of cholecystokinin. Although mechanical factors secondary to the increased gall bladder size in patients with coeliac disease might to some extent account for the findings, the alternative explanation is that the gall bladder muscle is for some reason resistant to the action of cholecystokinetic agents. A similar phenomenon affecting the pancreas might contribute to the abnormally decreased pancreatic secretion found in coeliac disease.

In this investigation, the measurement of trypsin levels in duodenal juice following a standard test meal (Lundh test) was evaluated as a test of pancreatic function, and a study was made of diseases and other factors which may influence its diagnostic efficiency. The method of trypsin assay, which required only basic laboratory equipment, gave a linear concentration-activity curve, with a threshold at 50 mug of crystalline trypsin per ml. Intestinal juice could be frozen and stored for up to six weeks with no detectable loss of tryptic activity. The normal control values were very similar to those found by other workers and were unaffected by the sex or age of the subject. When used to assess 32 patients in whom the presence or absence of pancreatic disease had been clearly established, the test had a diagnostic success rate of 94%. Retrospective analysis of results from 98 patients showed that trypsin levels were generally grossly reduced in patients with chronic pancreatitis or carcinoma of the head of the pancreas. Trypsin levels were normal in most patients with steatorrhoea not of pancreatic origin. Levels were generally depressed to intermediate levels in patients with a diabetic glucose tolerance, with or without steatorrhoea, but no other sign of pancreatic insufficiency. Some lowering of trypsin levels was also noted in patients having an obstruction of the common bile duct. A small but significant depression of tryptic activity was noted in patients with villous atrophy and no pancreatic disease.

Splenic function was assessed using 'pitted' erythrocyte counts in 61 first degree relatives of patients with coeliac disease. 'Pitted' erythrocyte counts were normal in 12 parents, but were raised in 20% of 49 siblings and/or children of coeliac patients. First degree relatives had higher 'pitted' erythrocyte counts than normal controls (p = 0.002). The counts were lower in coeliac relatives than in age matched coeliacs (p = 0.0001), but no difference was present between the relatives and coeliac patients whose small bowel mucosa was morphologically normal. Considerable interfamily variation was found in 'pitted' erythrocyte counts, both in the coeliac patients and first degree relatives, and the pattern tended to 'run true' within families. The genetic factor influencing splenic function in coeliac disease is not HLA-linked but seems to be associated with a second, probably recessive, gene influencing the inheritance of coeliac disease.

Splenic function has been serially measured by counting pitted red cells in 15 coeliac patients, before and during a gluten-free diet. The basal percentage values of pitted cells decreased significantly during treatment but no correlation was observed between the duration of the gluten-free diet and the percentage of recovery of splenic function over basal values. Out of six coeliacs with pitted cell values consistent with splenic hypofunction, three showed a total recovery after gluten withdrawal. Our data suggest that, contrary to recent reports, hyposplenism in adult coeliac disease is improved by a gluten-free diet, and that environmental factors may be important in determining and maintaining this complication.

Mouth-caecum transit time (M-CTT) of a lactulose labelled liquid test meal has been measured in 27 coeliac patients and 10 healthy controls using the breath hydrogen technique. Although all patients were urged to maintain a gluten free diet, not all did, and there was, therefore, a wide range in the severity of fat malabsorption within the patient group. Gastric emptying of a 113Indium DTPA-labelled liquid test meal was also assessed in separate studies on six healthy controls and 11 of the coeliac patients. Fasting breath hydrogen concentrations and the response to lactulose, as assessed both by the rate of rise, and the peak breath hydrogen concentration reached, showed no difference between coeliacs and controls, regardless of the presence or absence of steatorrhoea. Mouth-caecum transit time in the 16 coeliac patients with steatorrhea (faecal fat greater than 7 g/24 h) was, however, significantly prolonged being 158 +/- 18 minutes (mean +/- SEM), compared with 70 +/- 9 minutes for the controls (p less than 0.02), and 83 +/- 15 minutes for the 11 coeliacs without steatorrhoea (p less than 0.002). Mouth-caecum transit time in the coeliac patients was linearly related to the 24 hour faecal fat excretion, r = 0.55, n = 27, p less than 0.01. Slow mouth-caecum transit in the coeliacs with steatorrhoea was not caused by delayed gastric emptying as the t1/2 for coeliacs with steatorrhoea was within the normal range. Coeliacs with delayed mouth-caecum transit had impaired insulin release but the postprandial profiles of the other peptides measured (cholecystokinin, GIP, secretin, motilin, neurotensin, enteroglucagon, and peptide YY) were all within the normal range in this group of partially treated coeliac patients.

Pancreatic polypeptide (PP) is a newly discovered hormonal peptide localised in a distinct endocrine cell type in the pancreas. PP circulates in plasma and in normal subjects levels rise substantially on the ingestion of food (mean rise 138 pmol/l). In 10 patients with chronic pancreatitis with exocrine deficiency the PP response to a test breakfast was greatly reduced (mean rise 20 pmol/l, P less than 0.001). PP response to the meal was normal in 10 patients with active coeliac disease and 12 patients with acute tropical sprue with steatorrhoea.

Subfractions of fraction 9, obtained from a peptic-tryptic-pancreatinic digest of wheat gliadin, were subjected to in vitro mucosal digestion and the filtrates examined for residual peptides. Small-intestinal mucosa from four groups of individuals were studied-eight patients with coeliac disease in remission; eight healthy controls; nine first degree relatives of patients with coeliac disease, and six children with recurrent diarrhoea investigated for possible coeliac disease, but in whom the diagnosis was excluded. The highest amounts of residual peptides (measured by scanning densitometer) were detected after digestion with mucosa from patients with coeliac disease and the lowest amounts with the control groups. The results obtained with the group of relatives fell between those of the coeliac disease and control groups, while the recurrent diarrhoea group overlapped the relatives and controls. The residual peptides were derived chiefly from the B-type subfractions of subfractions 1 and 2, obtained by ion-exchange chromatography of fraction 9. These subfractions are rich in glutamine/glutamic acid and proline and have a molecular weight (apparent) of not greater than 1500 Daltons. The results lend further support to the hypothesis of an enzyme deficiency in coeliac disease. A partial enzyme deficiency may exist in some first-degree relatives and in some children with recurrent diarrhoea but with histology of the small intestine within normal limits. HLA-B8 antigen is not correlated with this deficiency, but, when the two factors are associated, they could be related to the manifestation and severity of coeliac disease.

We measured splenic function using a simple, non-isotopic method in childhood coeliac disease. No patients were shown to have hyposplenism. This has important clinical and therapeutic implications.

The uptake of (75)Se Selenomethionine by the pancreas has been evaluated in 102 patients and compared with the secretin-pancreozymin test of pancreatic function. In groups of patients with chronic pancreatitis and cancer of the pancreas abnormal scans closely parallel the diminished exocrine secretion, especially bicarbonate output, following a submaximal dose of secretin. Thirty per cent of the group with no pancreatic abnormality have abnormal scans, though the secretinpancreozymin test is normal. Though a normal scan excludes the presence of chronic pancreatitis and cancer of the pancreas with a probability greater than 90%, an abnormal scan is found so frequently in normal subjects that it does not provide a reliable index of impaired pancreatic function.