Reversible intestinal mucosal abnormality in acrodermatitis enteropathica.

In 3 cases of acrodermatitis enteropathica duodenal biopsy performed at the outset of treatment showed a similar abnormality of the intestinal mucosa. Further biopsies taken during treatment showed progressive improvement of the intestinal mucosa with subsequent complete restoration of the normal cellular and villous pattern. The initial treatment was with expressed human breast milk and oral di-iodohydroxyquinoline. The latter was continued alone and later replaced by zinc sulphate. Changes in the intestinal epithelial cells and inflammatory cell infiltration of the lamina propria still detectable on di-iodohydroxyquinoline therapy reverted to normal with oral zinc.     

Localization of transforming growth factor-beta receptor types I, II, and III in the postnatal rat small intestine

Transforming growth factor-beta2 (TGF-beta2) levels in rat milk are high in early lactation, whereas endogenous TGF-beta1 expression in the neonatal gut increases toward midweaning. Three types of transmembrane TGF-beta receptors have been identified in mammals. The receptor III (or betaglycan) binds and presents TGF-beta1 or beta2 to receptor II. Receptor I then interacts with receptor II, forming a signaling receptor complex, and propagates the signal. To determine whether TGF-beta receptor expression in the gut is also developmentally regulated, the present study assessed ontogeny of TGF-beta receptor expression in the postnatal rat small intestine. Jejunum and ileum tissues from rat pups at d 3, 10, 14, 21, and 28 of age were collected. Cryostat sections were stained with antibodies against TGF-bea receptors I, II, and III, and various cell markers by immunofluorescence. In both regions, receptor I staining was seen on apical and basolateral membranes of the villus and crypt epithelium at all ages, and staining on the apical membrane increased with age; receptor II was predominantly expressed in the crypt, and staining on the villi appeared after d 10; receptor III was distributed throughout the mucosa at early ages but diminished from the epithelium postweaning by d 28. T cells, B cells, and dendritic cells in the lamina propria expressed TGF-beta receptor III but lacked expression of receptor I and II. The pattern of TGF-beta receptor expression changes with age in a manner that may reflect the change in ligand from TGF-beta2 (milk-derived) to TGF-beta1 (endogenously produced).     

Intestinal mucosa in nephropathic cystinosis

The major manifestations of nephropathic cystinosis are renal tubular acidosis, vitamin D-resistant rickets, and dwarfism. Cystine crystals are deposited in a variety of cells, mainly phagocytic, including macrophages of the intestinal lamina propria. Previously, ultrastructural changes were suggested to occur in the absorptive epithelium as well, possibly as a result of local cystine toxicity. We report here on the light- and electron-microscopic findings in the jejunal mucosa of two patients, aged 4 and 9 years with nephropathic cystinosis. Cystine crystals were easily identified in semithin sections of plastic-embedded specimens as brick- and hexagon-shaped spaces in macrophages. Electron microscopy showed that all crystals were in single-membrane-limited bodies (lysosomes), within phagocytic cells, and exclusively located in the lamina propria. In contrast to previous findings, the absorptive epithelium showed no abnormalities. We conclude that the growth failure in cystinosis is not a consequence of morphological toxic alterations in the intestinal epithelium, but is related to the known metabolic abnormalities of this condition. The use of rectal suction biopsy as a means of diagnosing cystinosis is also suggested as an alternative to other diagnostic methods.     

Activation of the gut-associated lymphoid tissue with expression of interleukin-2 receptors that peaks during weaning in the rat.

BACKGROUND: Weaning exposes the intestinal mucosa to food and bacterial antigens at an age when the immune system is believed to be immature and functionally defective. The purpose of this study was to investigate changes in activation and phenotype of immune cells of the gut-associated lymphoid tissue during weaning. METHODS: Litters of infant rats were studied from pre- to postweaned life. The activation status, assessed by interleukin-2 receptor (IL-2R) expression, and phenotype of cells in the gut-associated lymphoid tissue were examined by immunostaining. RESULTS: Interleukin-2 receptor expression peaked two to four-fold at midweaning (day 21) in mesenteric lymph nodes, jejunal lamina propria, Peyer's patches, and intraepithelial lymphocytes, compared with adult animals (day 70). CD45+ cells expanded in the lamina propria, epithelium, and lymphocyte-filled villi. With CD45 as the denominator, 10% to 50% of lymphocytes in the lamina propria and epithelium were alphabetaT-cell receptor (TCR)+, but the remaining cells had a null phenotype, because there were low numbers of gammadeltaTCR+ T cells, B cells, and macrophages. Natural killer cells peaked at midweaning in the lamina propria (9%) and epithelium (20%) but were less than 5% of CD45+ cells after weaning. CONCLUSIONS: Rather than being immature or functionally inactive, the gut-associated lymphoid tissue reacts appropriately during weaning with expression of IL-2R and expansion of alphabetaTCR+ T-cells.     

RESPONSE OF THE JEJUNAL MUCOSA TO COW'S MILK IN THE MALABSORPTION SYNDROME WITH COW'S MILK INTOLERANCE A Light- and Electron-Microscopic Study

Three infants, in whom the malabsorption syndrome, small intestinal. mucosal damage and clinical cow's milk intolerance were found, were challenged with cow's milk after initial treatment with breast milk. The small intestinal mucosa was investigated with light and electron microscopy both before and after provocation. Clinically one patient reacted rapidly in a few hours and showed round cell infiltration in the surface epithelium and lamina propria 20 hours after a single challenge. Electron microscopy showed short microvilli, abnormal nuclei and thickened basement lamina in the surface epithelium. Two patients reacted slowly. One of them showed similar changes 4 days after commencement of the provocations, but the changes were much more evident 38 days later, when symptoms were also apparent. At this time large accumulations of lysosomes in the apical part of the surface epithelial cell and marked thickening of the basal lamina with accumulations of whirly collagen fibres were detected. The third patient reacted in a milder way, both clinically and morphologically. This study indicates that cow's milk, apparently through its protein fraction, may damage the surface epithelium of the small intestinal mucosa. These alterations during provocation periods resemble those found in coe-liac disease during gluten provocation.     

Value of antigliadin antibodies (AGA) in latent coeliac disease (CD)

BACKGROUND: The term latent coeliac disease (CD) is applied to patients who were previously shown to have a normal jejunal mucosa on a free diet. The aim of this study was to determine whether a high AGA value in the serum of patients with coeliac symptoms can also be regarded by itself, without typical mucosal atrophy, as a marker of latent CD, as some authors suggest in relatives of celiac patients. METHODS: We observed 31 patients with suspected CD and pathological values of serum IgA ang IgG AGA. In all we performed intestinal biopsy, assayed antiendomisium antibodies (AEA) in serum, AGA IgA, IgG, and IgM in duodenal jejunal fluid and in some of the lymphocytcs CD3+ gamma/delta+ in the lamina propria of the intestinal mucosa. RESULTS: In this study only pathological values of serum AGA without mucosa atrophy don't seem to be markers of latent CD, but an aspecific allergic response. CONCLUSIONS: As shown by other authors serum AEA, intestinal fluid AGA IgM and lamina propria lymphocytes CD3+ gamma/delta+ seem markers of latent CD.     

Immunohistochemical findings in the intestine of IgA-deficient persons: number of intraepithelial T lymphocytes is increased

We studied jejunal biopsy specimens of 13 IgA-deficient persons (IgAdp) and 12 controls. Four Ig-Adp had celiac disease, in the others the jejunal mucosa appeared normal. Monoclonal antibodies and the peroxidase technique were used to identify T lymphocytes, T-lymphocyte subsets, HLA-DR antigens, and IgE-containing cells in the lamina propria and epithelium. Intraepithelial lymphocytes (IEL); goblet cells; and IgA-, IgG-, and IgM-containing cells were counted in paraffin sections. Both IgAdp with normal jejunal structure and IgAd celiacs on gluten-free diet (p less than 0.001 and p less than 0.01 versus controls, respectively) had decreased numbers of IgA-containing cells, and an increased number of IgM-containing cells (p less than 0.01) was noted in the IgAdp with normal jejunal structure. The IgAdp with normal intestines had increased numbers of intraepithelial lymphocytes (mean 57 cells/mm versus 33 in controls, p less than 0.01) and so did the IgAd celiacs after gluten challenge (mean 74, p less than 0.001). The HLA-DR antibody stained the epithelial cells of the IgAd celiacs differently from those of controls and IgAdp with normal intestines: the whole cytoplasm was never stained in the celiacs, but in six of 12 controls (p less than 0.05) and during gluten challenge, the crypt cells of the IgAd celiacs showed strong staining, never seen in a normal intestine (p less than 0.05 compared with pre-challenge specimens). The increase in IEL number in the jejunal mucosa of IgAdp probably indicates ineffective antigen exclusion.     

Cytokines and adhesion molecules in duodenal mucosa of children with delayed-type food allergy

OBJECTIVES: The aim was to investigate the expression of cytokines, adhesion molecules, and activation and proliferation markers in duodenal biopsies from children with delayed-type food allergy (FA). METHODS: Seven children with untreated FA (uFA), seven children with treated FA (tFA) to cow milk and/or cereals, and five normal controls furnished duodenal biopsy specimens. Additionally, five pediatric patients with celiac disease were included, serving exclusively as positive controls for in situ hybridization. Interferon-gamma (IFN-gamma), interleukin-4 (IL-4), adhesion molecules, and activation markers were detected by immunohistochemistry, and expression of IFN-gamma and IL-4 messenger RNA was revealed by in situ hybridization. RESULTS: uFA patients had a higher density of IFN-gamma positive cells in the lamina propria than did tFA patients and controls (P = 0.053 and P = 0.018). Moreover, the uFA patients exhibited a higher proportion of crypt cells in mitosis than did tFA patients (P = 0.026), and stronger staining of HLA-DR in the crypts and increased density of gammadelta-T cell receptor-positive intraepithelial lymphocytes than did controls (P = 0.048 and P = 0.010). The densities of alpha(4)beta(7) positive cells in the lamina propria tended to be higher in controls than in uFA or tFA patients (P = 0.106, P = 0.073). Expression of IL-4 mRNA was significantly higher in celiac patients than in the other study groups (uFA P = 0.006, tFA P = 0.010; controls P = 0.029), and celiac patients showed higher expression of IFN-gamma mRNA than did tFA patients or controls (P = 0.017 and P = 0.016). CONCLUSIONS: As expected, Th1 dominance was present in the lamina propria of children with delayed-type FA. It may cause activation of epithelial cells and increase their turnover.     

Reaction of rectal mucosa of celiac patients to direct contact with gluten

It is assumed that the colonic mucosa of celiac patients is not sensitive to gluten. This assumption has been supported by the absence of clinical manifestations of colonic involvement in patients with active celiac disease which, by itself, does not confirm insensitivity to gluten. Eight children, aged from 11 to 25 months, with an initial diagnosis of celiac disease were studied: in five children a definite diagnosis has already been confirmed. Rectal gluten challenge was done by means of retention enemas. A volume of 100 ml of a 10% gluten suspension in water was introduced into the rectum three times per day for 8 days; each enema was retained at least 1 h. Rectosigmoidoscopies and rectal biopsies were done before and at the end of the challenge period. The endoscopic appearance of rectal mucosa was normal in all the children either before or after gluten challenge. The means of total mucosal thickness, intraepithelial lymphocyte counts, mitotic crypt cell activity, and cellular infiltration of lamina propria increased after challenge; the mean of goblet cell/epithelial cell ratio in the surface epithelium decreased. The differences between each pair of means (before and after challenge), however, were not significant except for total mucosal thickness (p less than 0.05), the meaning of which is unclear. This study did not definitely demonstrate that the rectal mucosa of celiacs is insensitive to gluten. For practical clinical purposes, however, it behaves as such. This makes the rectal mucosa a useless tool for the final diagnosis of celiac disease.     

The regulation of intestinal hypersensitivity reactions to ovalbumin by ω-3 fatty acid enriched diet: Studies of IEL and LPL in mucosal damage

In order to clarify the mechanisms of food-sensitive enteropathy, a food hypersensitive model was generated by feeding ovalbumin to female BALB/c mice after intraperitoneal injection of cyclophosphamide and morphological and immunological changes in the gut mucosa were investigated. Villus atrophy, crypt hyperplasia and increased numbers of intra-epithelial lymphocytes (IEL) were confirmed in this model, as seen in food-sensitive enteropathy in humans. Subpopulations of IEL and lamina propria lymphocytes were enumerated by immunohistochemical observation. CD8-positive cells were increased both in epithelium and lamina propria, whereas CD4-positive cells were decreased in lamina propria. We document here that orally administered food antigen actually induces food-sensitive enteropathy and mucosal damage is generated by lymphocytes that infiltrate the intestinal mucosa. We also investigated the effect of feeding an ω-3 fatty acid-enriched diet in this model and found that it was efficient in attenuating mucosal damage.     

Isolated deficient alpha6beta4 integrin expression in the gut associated with intractable diarrhea.

BACKGROUND: An infant born with pyloric atresia had development of intractable diarrhea and was found to have total epithelial detachment of gastric and small and large bowel mucosa. She had no skin abnormalities. Parental consanguinity and pyloric atresia in a sibling who died without autopsy suggest an inherited origin for this disorder. The purpose of this study was to examine defects in intestinal and skin cell adhesion. METHODS: Histologic, immunohistochemical, and ultrastructural characteristics of the skin and gut of the patient were compared with that of normal control subjects. Distribution of adhesion molecules was determined. RESULTS: Immunofluorescent analysis of the digestive mucosa showed alpha6beta4 integrin expression deficiency at the epithelial cell-lamina propria junction. Ultrastructural examination of the digestive mucosa revealed a complete epithelial detachment with a cleavage plane lying between the lamina densa and the basal pole of the enterocytes. Consistent with the absence of skin blistering, integrin alpha6beta4 was expressed at the dermal-epidermal junction. Electron micrographs of skin biopsy specimens showed the presence of normal hemidesmosomes and the absence of dermal-epidermal dysadhesion. CONCLUSION: It was postulated that this patient had protracted diarrhea related to epithelial detachment of the digestive mucosa as a consequence of a deficiency of an integrin alpha6beta4 isoform specific to the gut.     

Dietary nucleotides accelerate changes in intestinal lymphocyte maturation in weanling mice

OBJECTIVE: Nucleotides, the building blocks of nucleic acids, are normal components of the mammalian diet. These molecules have been implicated in biologic processes, such as the stimulation of the immunologic response. Nucleotides have also been considered as conditionally essential nutrients for infant formulas. The authors evaluated the influence of dietary nucleotides on the expression of several surface antigens by different intestinal lymphocyte populations in weanling mice. METHODS: Mice at weaning were fed a semipurified diet with or without 3 g/kg of each of the following nucleotides: adenosine monophosphate, cytosine monophosphate, guanosine monophosphate, and uridine monophosphate. Animals were killed at different times (0, 4, 7, 12, and 18 days) after weaning, and lymphocytes from intestinal Peyer's patches, epithelium, and lamina propria were isolated. The expression of different antigens (CD3, CD4, CD8alpha, CD8beta, TCRalphabeta, TCRgammadelta, CD5, CD22 and CD45R) was analyzed by flow cytometry. RESULTS: The expression of these antigens changed parallel to the maturation of the lymphocytes from Peyer's patches, epithelium, and lamina propria. However, developmental changes of expression for most of the antigens occurred sooner in the animals fed the diet supplemented with nucleotides. The expression of T and B antigens was different in the lymphocyte populations analyzed and also changed according to the diet within each population. In general, nucleotides promoted the expression of B- and T-helper cell antigens. CONCLUSIONS: The authors conclude that dietary nucleotides may affect the process of maturation and differentiation of intestinal lymphocytes, which usually takes place at weaning.     

In vitro activation of adenylate cyclase of atrophic celiac intestinal mucosa by wheat gliadin-derived peptides

In order to demonstrate that gliadin peptides may interact with cell membranes of celiac small intestinal mucosa, the capacity of these peptides to activate the cell membrane enzyme adenylate cyclase was tested. The addition of peptides from bread wheat purified A-gliadin and whole gliadin (proteins that are toxic for celiac patients) enhanced the adenylate cyclase activity of crude cell membrane preparations obtained from atrophic small intestinal mucosa of celiac patients. No activation of adenylate cyclase of this tissue was observed with peptides from proteins nontoxic for celiac patients (bread wheat albumin and maize prolamin). Gliadin peptides did not activate adenylate cyclase of morphologically normal small intestinal mucosa from normal subjects or from celiac patients in remission. These results, therefore, suggest that peptides from bread wheat gliadin may interact with cell membrane of atrophic small intestinal mucosa of celiac patients.     

Jejunal mucosa lymphoid cell subsets and the expression of major histocompatibility complex antigens in children

Using monoclonal antibodies with the immunoperoxidase technique the distribution pattern of class I and class II antigens of the major histocompatibility complex (MHC), and of the lymphocyte subsets have been studied in intestinal biopsies from children without mucosal lesions, from children with coeliac disease (CD) and from infants with cow's milk protein intolerance (CMPI). The staining of the intestinal mucosa for class I antigens is unaltered irrespective of the histological picture or the clinical diagnosis. Class II antigens are only partially or not expressed at all by epithelial cells in untreated cocliac disease and in some cases of cow's milk protein intolerance. The number and the composition of the lamina propria lymphocytes in both CD and CMPI are different from the normal situation. An increase of all lamina propria lymphocyte subsets is observed in untreated CD. A decrease of OKT4⁺ lymphocytes is observed in the lamina propria of CMPI patients. These changes may be involved in the pathogenesis of these diseases.Abbreviations MHC major histocompatibility complex - CD coeliac disease - CMPI cow's milk protein intolerance - IEL intra-epithelial lymphocytes     

Enteropathy in Niemann-Pick disease

Three consecutive cases of Niemann-Pick disease with predominant enteropathy were seen. The diagnosis was confirmed by the demonstration of typical foamy cells in the bone marrow, small intestinal mucosa, and liver. The enteropathy was apparent in steatorrhoea, xylose malabsorption, protein loss etc. The hypothesis of foamy cell infiltration of the lamina propria of the small intestinal mucosa as a cause of malabsorption is proposed.     

Expression of class II MHC antigens in the intestinal epithelium of pediatric celiac disease

Class II MHC antigen expression in the intestinal epithelium of 28 small bowel biopsies from 23 celiac patients were studied by means of indirect immunofluorescence and immunoperoxidase using monoclonal antibodies. Patients were divided on the basis of diet into two subgroups: 15 subjects on a gluten-containing diet (GCD) and 13 on a gluten-free diet (GFD). The control group included 10 pediatric subjects with normal intestinal mucosa who underwent intestinal biopsy for chronic diarrhea or short stature. DR antigens and invariant chain were expressed in all patients, regardless of the diet, as well as in the control subjects. DQ was found in one patient only on GCD. DP antigens were present in 12/15 patients on GCD, and in 2/13 on GFD (Fisher's exact test, p = 8.8 x 10(-4), as well as in 3/10 control subjects. In 4/5 celiac patients, DP antigens, which were undetectable on GFD, could be demonstrated after gluten challenge. The results of the study show that DR antigens are expressed by intestinal mucosa of celiac patients independently of their gluten exposure and that DQ antigens are consistently undetectable. Statistically significant differences in expression of DP antigens on enterocytes of celiac patients on GCD and their neoexpression after gluten challenge may represent a basis for further investigation.     

Intestinal gamma/delta receptor-bearing T lymphocytes in celiac disease and inflammatory bowel diseases in children. Constant increase in celiac disease

We studied the numbers of T-cell receptor alpha/beta- and gamma/delta-bearing lymphocytes in 27 jejunal specimens from 19 celiac patients, 27 rectal and colonic specimens from 14 ulcerative colitis patients and four patients with Crohn's disease, and 24 control specimens. MAb and a three-layer peroxidase staining method were used. Only low numbers of gamma/delta + cells were seen in normal jejunum and rectum of controls, as well as in the specimens of patients with inflammatory bowel diseases. In the lamina propria of celiac patients, the mean number of gamma/delta + cells was significantly higher than in the controls before treatment, during gluten-free diet, and after the gluten challenge. Within the jejunal epithelium, the number of gamma/delta + cells was elevated before and during gluten elimination and after the challenge test. The absolute number of intraepithelial gamma/delta + cells remained constant during gluten elimination and provocation. We infer that the constantly elevated population of gamma/delta + T cells in the epithelium of celiac patients may play an important role in the pathogenesis of celiac disease.     

Intestinal mast cells and neutrophil chemotactic activity of serum following a single challenge with gluten in celiac children on a gluten-free diet

The number of one subtype of mast cells (formalin fixation, toluidine blue staining), cells of the lamina propria, and intraepithelial lymphocytes were counted in the intestinal biopsy specimens of 14 children with treated celiac disease following a single challenge with gluten. The serum neutrophil chemotactic activity was measured at 0, 1, 3, 5, and 24 h after challenge. There was no significant change in the number of intraepithelial lymphocytes, but the biopsy samples obtained at 5 h showed a marked increase in the inflammatory cells of the lamina propria and a significant decrease in the number of mast cells. A pronounced decrease was present at 3-5 h in the number of eosinophil cells in the blood. The neutrophil chemotactic activity of sera showed a significant increment in 10 of 14 patients. The intestinal permeability of patients became abnormal, as detected by the increased absorption of lactulose. These findings suggest that degranulation of mast cells may be involved in the pathogenesis of the small intestinal mucosal injury in children with celiac disease.     

Experimentally induced gluten enteropathy and protective effect of epidermal growth factor in artificially fed neonatal rats

BACKGROUND: A protective effect of breast-feeding against the development of celiac disease has been described, but the nature and effects of the actual milk components have not been established. Epidermal growth factor (EGF), a milk cytokine affecting the proliferation and differentiation of mucosal epithelial cells, was studied as to its potential protective effect on the damage of intestinal mucosa by gliadin in a model system. METHODS: Enteropathy was induced by gliadin in inbred AVN strain rat pups delivered by cesarean section, breast-fed, or hand-fed a milk formula. All experimental groups were treated with interferon-gamma (1,000 U per animal, administered intraperitoneally) after birth. Gliadin (0.5 and 3 mg) was intragastrically administered to the pups on days 0 and 3, and a 30-mg challenge dose was given on day 20 (24 hours before the termination of the experiment). One group of artificially fed pups received EGF (100 ng/ml) continuously in the diet. RESULTS: Gliadin- and interferon-gamma-treated formula-fed rat pups showed villus atrophy, increase of inflammatory cells, including CD4+ T lymphocytes in the lamina propria, and damage to epithelial tight junctions and the enterocyte brush border. Morphometrically, the villus height was significantly less than in other groups. Recombinant EGF was markedly increased in the epithelial cells of injured jejunum. The intestinal mucosa of gliadin- and interferon-gamma-treated pups kept on a EGF-supplemented artificial diet resembled that of breast-fed pups. CONCLUSION: Pathologic changes in jejunal mucosa (villus atrophy and inflammation) resembling gliadin-induced atrophy appeared on administration of interferon-gamma and gliadin to rat pups fed an artificial milk diet immediately after birth. Addition of EGF to the diet protected the rats against pathologic mucosal changes.     

Assessment of the reparative capacity of the small intestine in celiac disease in children according to the DNA and RNA contents in enterocyte nuclei]

In 44 children suffering from celiac disease, cytophotometry was used to measure the content of DNA and RNA in enterocyte nuclei in the jejunum mucosa. The measurements were made after the diagnosis was established, 6-8 months and 1-1.5 years after administration of the agliadin diet. In celiac disease children, there was an increase of the content of DNA and RNA in enterocyte nuclei of the intestinal villi and cryptae and lack of gradient in the distribution of the amount of nucleic acids along the length of the villi, which characterized the hyper-regeneration type of small intestinal mucosa atrophy. The recovery of the crypta/villus gradient accompanied by the normalization of the content of nucleic acids, comparable with the analogous indicators of the unchanged mucosa of the small intestine, allows regarding the measurements of the content of DNA and RNA in the nuclei as an index of reparative capacity of enterocytes in children afflicted with celiac disease.