UNC45A deficiency causes microvillus inclusion disease-like phenotype by impairing myosin VB-dependent apical trafficking.

Variants in the UNC45A cochaperone have been recently associated with a syndrome combining diarrhea, cholestasis, deafness, and bone fragility. Yet the mechanism underlying intestinal failure in UNC45A deficiency remains unclear. Here, biallelic variants in UNC45A were identified by next-generation sequencing in 6 patients with congenital diarrhea. Corroborating in silico prediction, variants either abolished UNC45A expression or altered protein conformation. Myosin VB was identified by mass spectrometry as client of the UNC45A chaperone and was found misfolded in UNC45AKO Caco-2 cells. In keeping with impaired myosin VB function, UNC45AKO Caco-2 cells showed abnormal epithelial morphogenesis that was restored by full-length UNC45A, but not by mutant alleles. Patients and UNC45AKO 3D organoids displayed altered luminal development and microvillus inclusions, while 2D cultures revealed Rab11 and apical transporter mislocalization as well as sparse and disorganized microvilli. All those features resembled the subcellular abnormalities observed in duodenal biopsies from patients with microvillus inclusion disease. Finally, microvillus inclusions and shortened microvilli were evidenced in enterocytes from unc45a-deficient zebrafish. Taken together, our results provide evidence that UNC45A plays an essential role in epithelial morphogenesis through its cochaperone function of myosin VB and that UNC45A loss causes a variant of microvillus inclusion disease.

Aberrant Epithelial Differentiation Contributes to Pathogenesis in a Murine Model of Congenital Tufting Enteropathy.

BACKGROUND & AIMS: Congenital tufting enteropathy (CTE) is an intractable diarrheal disease of infancy caused by mutations of epithelial cell adhesion molecule (EpCAM). The cellular and molecular basis of CTE pathology has been elusive. We hypothesized that the loss of EpCAM in CTE results in altered lineage differentiation and defects in absorptive enterocytes thereby contributing to CTE pathogenesis. METHODS: Intestine and colon from mice expressing a CTE-associated mutant form of EpCAM (mutant mice) were evaluated for specific markers by quantitative real-time polymerase chain reaction, Western blotting, and immunostaining. Body weight, blood glucose, and intestinal enzyme activity were also investigated. Enteroids derived from mutant mice were used to assess whether the decreased census of major secretory cells could be rescued. RESULTS: Mutant mice exhibited alterations in brush-border ultrastructure, function, disaccharidase activity, and glucose absorption, potentially contributing to nutrient malabsorption and impaired weight gain. Altered cell differentiation in mutant mice led to decreased enteroendocrine cells and increased numbers of nonsecretory cells, though the hypertrophied absorptive enterocytes lacked key features, causing brush border malfunction. Further, treatment with the Notch signaling inhibitor, DAPT, increased the numbers of major secretory cell types in mutant enteroids (graphical abstract 1). CONCLUSIONS: Alterations in intestinal epithelial cell differentiation in mutant mice favor an increase in absorptive cells at the expense of major secretory cells. Although the proportion of absorptive enterocytes is increased, they lack key functional properties. We conclude that these effects underlie pathogenic features of CTE such as malabsorption and diarrhea, and ultimately the failure to thrive seen in patients.

Loss of HAI-2 in mice with decreased prostasin activity leads to an early-onset intestinal failure resembling congenital tufting enteropathy.

Prostasin (CAP1/PRSS8) is a glycosylphosphatidylinositol (GPI)-anchored serine protease that is essential for epithelial development and overall survival in mice. Prostasin is regulated primarily by the transmembrane serine protease inhibitor, hepatocyte growth factor activator inhibitor (HAI)-2, and loss of HAI-2 function leads to early embryonic lethality in mice due to an unregulated prostasin activity. We have recently reported that critical in vivo functions of prostasin can be performed by proteolytically-inactive or zymogen-locked variants of the protease. Here we show that the zymogen form of prostasin does not bind to HAI-2 and, as a result, loss of HAI-2 does not affect prenatal development and survival of mice expressing only zymogen-locked variant of prostasin (Prss8 R44Q). Indeed, HAI-2-deficient mice homozygous for R44Q mutation (Spint2-/-;Prss8R44Q/R44Q) are born in the expected numbers and do not exhibit any obvious developmental abnormality at birth. However, postnatal growth in these mice is severely impaired and they all die within 4 to 7 days after birth due to a critical failure in the development of small and large intestines, characterized by a widespread villous atrophy, tufted villi, near-complete loss of mucin-producing goblet cells, loss of colonic crypt structure, and bleeding into the intestinal lumen. Intestines of Spint2-/-;Prss8R44Q/R44Q mice showed altered expression of epithelial junctional proteins, including reduced levels of EpCAM, E-cadherin, occludin, claudin-1 and -7, as well as an increased level of claudin-4, indicating that the loss of HAI-2 compromises intestinal epithelial barrier function. Our data indicate that the loss of HAI-2 in Prss8R44Q/R44Q mice leads to development of progressive intestinal failure that at both histological and molecular level bears a striking resemblance to human congenital tufting enteropathy, and may provide important clues for understanding and treating this debilitating human disease.

In Vivo DNA Re-replication Elicits Lethal Tissue Dysplasias.

Mammalian DNA replication origins are "licensed" by the loading of DNA helicases, a reaction that is mediated by CDC6 and CDT1 proteins. After initiation of DNA synthesis, CDC6 and CDT1 are inhibited to prevent origin reactivation and DNA overreplication before cell division. CDC6 and CDT1 are highly expressed in many types of cancer cells, but the impact of their deregulated expression had not been investigated in vivo. Here, we have generated mice strains that allow the conditional overexpression of both proteins. Adult mice were unharmed by the individual overexpression of either CDC6 or CDT1, but their combined deregulation led to DNA re-replication in progenitor cells and lethal tissue dysplasias. This study offers mechanistic insights into the necessary cooperation between CDC6 and CDT1 for facilitation of origin reactivation and describes the physiological consequences of DNA overreplication.

Contractile forces at tricellular contacts modulate epithelial organization and monolayer integrity.

Monolayered epithelia are composed of tight cell assemblies that ensure polarized exchanges. EpCAM, an unconventional epithelial-specific cell adhesion molecule, is assumed to modulate epithelial morphogenesis in animal models, but little is known regarding its cellular functions. Inspired by the characterization of cellular defects in a rare EpCAM-related human intestinal disease, we find that the absence of EpCAM in enterocytes results in an aberrant apical domain. In the course of this pathological state, apical translocation towards tricellular contacts (TCs) occurs with striking tight junction belt displacement. These unusual cell organization and intestinal tissue defects are driven by the loss of actomyosin network homoeostasis and contractile activity clustering at TCs, yet is reversed by myosin-II inhibitor treatment. This study reveals that adequate distribution of cortical tension is crucial for individual cell organization, but also for epithelial monolayer maintenance. Our data suggest that EpCAM modulation protects against epithelial dysplasia and stabilizes human tissue architecture.

The Association Between Fecal Biomarkers of Environmental Enteropathy and Rotavirus Vaccine Response in Nicaraguan Infants.

BACKGROUND: Environmental enteropathy (EE) is a common intestinal condition among children living in low- and middle-income countries and is associated with diminished enteric immunity to gastrointestinal pathogens, and possibly to oral vaccine antigens. The goal of this study was to examine associations between biomarkers of EE and immunogenicity to the pentavalent rotavirus vaccine (RV5). METHODS: Infants were recruited 1 day before their first RV5 immunization in León, Nicaragua, from public health rosters. Infants provided a preimmunization blood and stool sample, and a second blood sample 1 month after receipt of RV5. We measured immunoglobin A (IgA) seroconversion to the first dose of RV5 and concentrations of 4 previously identified fecal biomarkers of EE (alpha-1 antitrypsin, neopterin, myeloperoxidase and calprotectin). We then assessed associations between concentrations of these biomarkers, both individually and as combined scores, and seroconversion to the first dose of RV5. RESULTS: Of the 43 enrolled infants, 24 (56%) seroconverted after the first dose of RV5. As compared with infants who seroconverted, those who did not seroconvert had higher median concentrations of both myeloperoxidase (3.1 vs. 1.1 µg/mL, P = 0.002) and calprotectin (199.1 vs. 156.2 µg/mL, P = 0.03). Further, those who did not seroconvert had a higher median combined score of the 4 biomarkers as compared with those who seroconverted (6.5 vs. 4.5, P = 0.017). CONCLUSIONS: We found an association between biomarkers of EE and seroconversion to the first dose of RV5. It is possible that interventions that prevent or ameliorate EE may also improve oral rotavirus vaccine response.

IBD-Like Features in Syndromic Diarrhea/Trichohepatoenteric Syndrome.

BACKGROUND: Very early onset inflammatory bowel disease (VEOIBD) (inflammatory bowel disease [IBD] before 6 years of age) may manifest as a monogenic disease affecting the gastrointestinal tract. Syndromic diarrhea/trichohepatoenteric syndrome (SD/THE), a rare disorder caused by alteration of a complex involved in RNA degradation, has been reported to present with some degree of colitis and in some cases an IBD-like presentation. METHODS: We reviewed clinical and biological data of 4 previously published cases and added detailed data of 2 new cases of SD/THE with an IBD-like presentation. RESULTS: All the 6 patients presented with typical intractable diarrhea and hair abnormalities. The colon was affected in all of the patients: 1 had ileitis, 2 had panenteritis, and 2 presented with perianal disease. Fecal calprotectin level and erythrosedimentation rate were elevated in 2 cases each. All the therapeutic classes of IBD treatment (mesalazine, steroids, immunomodulators, and biological therapy) were used in the 6 cases. In 2 patients, treatment had no effect. Three showed a partial effect, and 1 patient sustained only a transient effect. CONCLUSIONS: SD/THE can have a similar presentation as VEOIBD, often as pancolitis. IBD treatments appear to have little efficacy for SD/THE, suggesting a different pathogenesis for the IBD-like features in SD/THE compared with classical IBD.

Tricho-hepato-enteric syndrome (THE-S): two cases and review of the literature.

UNLABELLED: Tricho-hepato-enteric syndrome (THE-S) is characterized by severe infantile diarrhea, failure to thrive, dysmorphism, woolly hair, and immune or hepatic dysfunction. We report two cases of East Asian descent with THE-S who had remained undiagnosed despite extensive investigations but were diagnosed on whole exome sequencing (WES). Both cases presented with chronic diarrhea, failure to thrive, and recurrent infections. Case 1 had posteriorly rotated low set ears, mild retrognathia, and fine curly hypopigmented hair. She was managed with prolonged total parenteral nutrition and intravenous immunoglobulin infusions. Case 2 had sparse coarse brown hair as well as multiple lentigines and café-au-lait macules. She was managed with amino acid-based formula. For both cases, routine investigations were inconclusive. WES in both cases showed biallelic truncating mutations in TTC37 (c.3507T>G;p.Y1169X and c.3601C>T;p.R1201X in case 1 and c.3507T>G;p.Y1169X and c.154G>T;p.E52X in case 2), suggesting a diagnosis of THE-S. CONCLUSION: We present novel mutations in the TTC37 gene in two individuals of East Asian descent with the rare THE-S, detected by WES. Future identification of patients with THE-S and establishing genotype-phenotype correlations will aid in counseling the patients and their families. WHAT IS KNOWN: • Tricho-Hepato-Enteric syndrome (THE-S) is characterized by severe infantile diarrhea, failure to thrive, dysmorphism, woolly hair, and immune or hepatic dysfunction. • Complex patients with diagnostic dilemmas undergo extensive investigations. What is New: • This is a report of novel mutations in TTC37 in individuals of East Asian descent. • Whole exome sequencing (WES) can be useful in certain complex cases with diagnostic dilemmas.

Evaluation of intestinal biopsies for pediatric enteropathy: a proposed immunohistochemical panel approach.

Congenital enteropathies are rare disorders with significant clinical consequences; however, definitive diagnosis based on morphologic assessment of duodenal biopsies with routine stains alone is often impossible. To determine the role of immunohistochemistry (IHC) in the evaluation for microvillous inclusion disease, congenital tufting enteropathy (intestinal epithelial dysplasia), and enteroendocrine cell dysgenesis, a series of duodenal biopsies from 26 pediatric patients with chronic/intractable diarrhea was retrospectively reviewed. IHC stains for CD10, EpCAM, chromogranin, and villin were performed on all biopsies, and the results were correlated with hematoxylin and eosin and ultrastructural findings using electron microscopy, when available. Biopsies from 2 patients diagnosed with microvillous inclusion disease at the time of original biopsy demonstrated diffuse CD10-positive cytoplasmic inclusions within enterocytes and normal expression of EpCAM and chromogranin. Biopsies from 3 patients, including 2 siblings with confirmed EPCAM mutations, demonstrated complete loss of EpCAM expression and normal expression of CD10 and chromogranin; electron microscopic evaluation revealed characteristic ultrastructural findings of tufting enteropathy. Biopsies from 1 patient with a confirmed NEUROG3 mutation demonstrated an absence of intestinal enteroendocrine cells by chromogranin staining, consistent with enteroendocrine cell dysgenesis. Four patients' biopsies displayed nonspecific staining patterns for CD10 and/or EpCAM with normal expression of chromogranin, and 16 patients' biopsies exhibited normal expression for all 3 markers. Villin stains demonstrated heterogenous brush border labeling with nonspecific cytoplasmic reactivity, a pattern variably present throughout the biopsy series. In conclusion, the routine use of an IHC panel of CD10, EpCAM, and chromogranin is warranted in patients meeting specific age and/or clinical criteria, as the morphologic findings of congenital enteropathies may be subtle, focal, or inapparent on routine stains.

Tufting enteropathy revisited: the utility of MOC31 (EpCAM) immunohistochemistry in diagnosis.

Tufting enteropathy (TE) is an uncommon disease causing intractable diarrheas starting in early childhood and resulting in failure to thrive, dependence on total parenteral nutrition, and eventually requiring transplantation for treatment. The diagnosis has been based on histology showing the presence of epithelial "tufts" in the small bowel and colonic mucosa and variable villus alterations with mild to no inflammatory changes and preserved brush border. The gene for TE has been identified to be the EpCAM gene on chromosome 2p21. With Institutional Review Board approval, all cases of intractable diarrhea in children in whom TE was suspected or diagnosed were retrieved from the pathology files (17 patients). Other cases of infantile, neonatal, and childhood diarrhea were also retrieved to serve as controls for the staining studies (total 37 patients). EpCAM/MOC31 antibody staining was performed on all cases. The study cohort comprised 17 patients (13 boys, 4 girls) with a diagnosis of TE ranging in age at diagnosis from 3 months to 9 years, all presenting with protracted diarrhea and/or failure to thrive, usually since birth. Staining with MOC31 was carried out in all but 2 patients (both consults) and was completely negative in the epithelium irrespective of the site of biopsy or resection. In contrast, MOC31 was positive in all other cases tested, giving a sensitivity and specificity of 100% for loss of staining. MOC31 is a diagnostic stain for TE and should be included in the panel in any case of prolonged diarrhea in children to exclude this possibility.

MYO5B and bile salt export pump contribute to cholestatic liver disorder in microvillous inclusion disease.

UNLABELLED: Microvillous inclusion disease (MVID) is a congenital disorder of the enterocyte related to mutations in the MYO5B gene, leading to intractable diarrhea often necessitating intestinal transplantation (ITx). Among our cohort of 28 MVID patients, 8 developed a cholestatic liver disease akin to progressive familial intrahepatic cholestasis (PFIC). Our aim was to investigate the mechanisms by which MYO5B mutations affect hepatic biliary function and lead to cholestasis in MVID patients. Clinical and biological features and outcome were reviewed. Pretransplant liver biopsies were analyzed by immunostaining and electron microscopy. Cholestasis occurred before (n = 5) or after (n = 3) ITx and was characterized by intermittent jaundice, intractable pruritus, increased serum bile acid (BA) levels, and normal gamma-glutamyl transpeptidase activity. Liver histology showed canalicular cholestasis, mild-to-moderate fibrosis, and ultrastructural abnormalities of bile canaliculi. Portal fibrosis progressed in 5 patients. No mutation in ABCB11/BSEP or ATP8B1/FIC1 genes were identified. Immunohistochemical studies demonstrated abnormal cytoplasmic distribution of MYO5B, RAB11A, and BSEP in hepatocytes. Interruption of enterohepatic BA cycling after partial external biliary diversion or graft removal proved the most effective to ensure long-term remission. CONCLUSION: MVID patients are at risk of developing a PFIC-like liver disease that may hamper outcome after ITx. Our results suggest that cholestasis in MVID patients results from (1) impairment of the MYO5B/RAB11A apical recycling endosome pathway in hepatocytes, (2) altered targeting of BSEP to the canalicular membrane, and (3) increased ileal BA absorption. Because cholestasis worsens after ITx, indication of a combined liver ITx should be discussed in MVID patients with severe cholestasis. Future studies will need to address more specifically the effect of MYO5B dysfunction in BA homeostasis.

Functional consequences of EpCam mutation in mice and men.

Congenital tufting enteropathy (CTE) is a severe diarrheal disease of infancy characterized by villous changes and epithelial tufts. We previously identified mutations in epithelial cell adhesion molecule (EpCAM) as the cause of CTE. We developed an in vivo mouse model of CTE based on EpCAM mutations found in patients with the aim to further elucidate the in vivo role of EpCAM and allow for a direct comparison to human CTE. Using Cre-LoxP recombination technology, we generated a construct lacking exon 4 in Epcam. Epcam(Δ4/Δ4) mice and CTE patient intestinal tissue integrity was analyzed by histology using both light immunohistochemistry and electron microscopy. Epcam(Δ4/Δ4) mice demonstrate neonatal lethality and growth retardation with pathological features, including epithelial tufts, enterocyte crowding, altered desmosomes, and intercellular gaps, similar to human CTE patients. Mutant EpCAM protein is present at low levels and is mislocalized in the intestine of Epcam(Δ4/Δ4) mice and CTE patients. Deletion of exon 4 was found to decrease expression of both EpCAM and claudin-7 causing a loss of colocalization, functionally disrupting the EpCAM/claudin-7 complex, a finding for the first time confirmed in CTE patients. Furthermore, compared with unaffected mice, mutation of Epcam leads to enhanced permeability and intestinal cell migration, uncovering underlying disease mechanisms.

A novel nonsense mutation in the EpCAM gene in a patient with congenital tufting enteropathy.

OBJECTIVES: Tufting enteropathy (TE) is a classical congenital disorder of the intestinal mucosa causing protracted diarrhea in infancy as a result of a dysfunctional epithelial cell barrier, which is mainly caused by mutations in the EpCAM gene and expression of a nonfunctional epithelial cell adhesion molecule in the intestine. We report here a novel nonsense mutation in a patient suspected of having TE, resulting in a complete absence of EpCAM in duodenal enterocytes. METHODS: A patient presenting with congenital diarrhea and suspected of having TE was screened for EpCAM mutations, and duodenal biopsies were stained for EpCAM using immunohistochemistry analysis. RESULTS: We identified a novel homozygous nonsense mutation in the EpCAM gene in a patient suspected of having TE, causing a complete loss of EpCAM expression in duodenal enterocytes. CONCLUSIONS: With screening analysis for EpCAM mutations and immunohistochemistry for EpCAM expression in duodenal enterocytes, we found a novel homozygous mutation in a patient with classical protracted diarrhea in infancy finally diagnosed as TE, which results in a complete absence of EpCAM and in dysfunctional barrier formation in duodenal enterocytes.

mTrop1/Epcam knockout mice develop congenital tufting enteropathy through dysregulation of intestinal E-cadherin/ß-catenin.

Congenital tufting enteropathy (CTE) is a life-threatening hereditary disease that is characterized by enteric mucosa tufting degeneration and early onset, severe diarrhea. Loss-of-function mutations of the human EPCAM gene (TROP1, TACSTD1) have been indicated as the cause of CTE. However, loss of mTrop1/Epcam in mice appeared to lead to death in utero, due to placental malformation. This and indications of residual Trop-1/EpCAM expression in cases of CTE cast doubt on the role of mTrop1/Epcam in this disease. The aim of this study was to determine the role of TROP1/EPCAM in CTE and to generate an animal model of this disease for molecular investigation and therapy development. Using a rigorous gene-trapping approach, we obtained mTrop1/Epcam -null (knockout) mice. These were born alive, but failed to thrive, and died soon after birth because of hemorrhagic diarrhea. The intestine from the mTrop1/Epcam knockout mice showed intestinal tufts, villous atrophy and colon crypt hyperplasia, as in human CTE. No structural defects were detected in other organs. These results are consistent with TROP1/EPCAM loss being the cause of CTE, thus providing a viable animal model for this disease, and a benchmark for its pathogenetic course. In the affected enteric mucosa, E-cadherin and ß-catenin were shown to be dysregulated, leading to disorganized transition from crypts to villi, with progressive loss of membrane localization and increasing intracellular accumulation, thus unraveling an essential role for Trop-1/EpCAM in the maintenance of intestinal architecture and functionality.Supporting information is available for this article.

Effect on infant illness of maternal supplementation with 400 000 IU vs 200 000 IU of vitamin A.

BACKGROUND AND OBJECTIVE: Postpartum vitamin A supplementation is a strategy used to combat vitamin A deficiency and seems to reduce maternal/infant morbidity and mortality. However, studies have shown that a dose of 200 000 IU (World Health Organization [WHO] protocol) does not seem to provide adequate retinol levels in maternal breast milk, infant serum, and infant tissue. The objective of this study was to compare the effect of postpartum maternal supplementation with 400 000 IU (International Vitamin A Consultative Group protocol) compared with 200 000 IU of vitamin A on infant morbidity. METHODS: This was a randomized controlled, triple-blinded clinical trial conducted at 2 public maternity hospitals in Recife in northeastern Brazil. There were 276 mother-child pairs that were allocated to 2 treatment groups: 400 000 IU or 200 000 IU of vitamin A. They were followed up for >6 months to evaluate infant morbidity. RESULTS: Fever (rate ratio [RR]: 0.92 [95% confidence interval (CI): 0.75-1.14]), diarrhea (RR: 0.96 [95% CI: 0.72-1.28]), otitis (RR: 0.94 [95% CI: 0.48-1.85]), acute respiratory infection (RR: 1.03 [95% CI: 0.88-1.21]), the need for intravenous rehydration (RR: 2.08 [95% CI: 0.64-2.07]), and the use of antibiotic treatment (RR: 0.80 [95% CI: 0.43-1.47]) did not differ significantly between the 2 treatment groups. CONCLUSIONS: Our findings suggest that postpartum maternal supplementation with 400 000 IU of vitamin A does not provide any additional benefits in the reduction of illness in children aged <6 months; therefore, we do not support the proposal to increase the standard vitamin A dose in the existing WHO protocol.

Microvillous inclusion disease: a clinicopathologic study of 17 cases from the UK.

BACKGROUND: Microvillous inclusion disease (MVID) is a rare congenital disease producing intractable secretory diarrhea in early infancy. It is characterized by diffuse intestinal villous atrophy with no inflammatory reaction. Ultrastructural identification of apical microvillous inclusions in the surface enterocytes is diagnostic. However, there is difficulty in the diagnosis of MVID due to the existence of variants (e.g., microvillous dystrophy), possible disease resolution, and tissue orientation for electron microscopy (EM). The authors analyzed materials from 4 patients with MVID from a single institution. The morphologic features, distribution of lesions, biomarkers, and complementary ultrastructural characteristics were studied. DESIGN: Materials of MVID cases were collected from 6 different hospitals in the United Kingdom between 1990 and 2008. Epidemiological data, including age range, median, mode, sex ratios, and follow-up, were retrieved. All intestinal biopsy specimens were analyzed histologically, histochemically (for PAS, n = 17), immunohistochemically (for CD10, n = 2 and polyclonal CEA, n = 4), and ultrastructurally (n = 9). RESULTS: Ultrastructurally, apical microvillous inclusions in surface enterocytes in duodenal biopsies were identified in all cases, while 1 case had variant morphology (microvillous dystrophy and very occasionally atypical microvillous inclusions). Tissue orientation for EM was supportive for identification of inclusions in apical enterocytes. Morphologically, a bubbly vacuolated appearance of the apical cytoplasm with extensive or patchy absence of the brush border with occasional cytoplasm inclusions was observed in the enterocytes. Some of these changes vaguely resembled gastric mucin cell metaplasia. Architecturally, villous blunting with either crypt hypoplasia or hyperplasia and absence of inflammation were common findings. The epithelial changes were also found in colon biopsies. PAS, CD10, and p-CEA showed a bright apical cytoplasmic blush/staining, which correlated ultrastructurally with apical granules with inclusions of variable electron density in all cases. These stains also highlighted the targetoid inclusions. CONCLUSION: Besides electron microscopy identification of inclusions, the light microscopic morphological features together with the biomarker studies highlighting the apical cytoplasmic blush are quite unique and diagnostic of MVID. Furthermore, it is the opinion of the authors that a diagnosis of MVID can be made without electron microscopy.

Light microscopic diagnosis of microvillus inclusion disease on colorectal specimens using CD10.

Microvillus inclusion disease (MID) is a rare neonatal enteropathy that is typically diagnosed using electron microscopy to show characteristic inclusions in conjunction with light microscopy and periodic acid-Schiff staining to show lack of the normal brush border on biopsies obtained endoscopically from the small bowel. MID has also been diagnosed using CD10 immunoreactivity that shows abnormal intense cytoplasmic staining in absorptive small bowel enterocytes. As ultrastructural studies also show abnormal microvillus inclusions in absorptive colonocytes in these patients, we investigated the use of CD10 immunoreactivity on colon specimens. We studied the CD10 staining pattern in 4 colon specimens from patients with MID and in colon biopsy specimens from pediatric control patients with and without histopathologic abnormalities. All MID cases had cytoplasmic CD10 staining in absorptive colonocytes in contrast to the control patients who did not show any epithelial CD10 staining. We conclude that abnormal cytoplasmic CD10 staining of absorptive colonocytes can aid in the diagnosis of MID, which may be invaluable in the situations where only colon biopsy specimens are available for examination.

ApoE polymorphisms and diarrheal outcomes in Brazilian shanty town children.

A series of studies have shown that the heavy burdens of diarrheal diseases in the first 2 formative years of life in children living in urban shanty towns have negative effects on physical and cognitive development lasting into later childhood. We have shown that APOE4 is relatively common in shanty town children living in Brazil (13.4%) and suggest that APOE4 has a protective role in cognitive development as well as weight-for-height in children with heavy burdens of diarrhea in early childhood (64/123; 52%), despite being a marker for cognitive decline with Alzheimer's and cardiovascular diseases later in life. APOE2 frequency was higher among children with heaviest diarrhea burdens during the first 2 years of life, as detected by PCR using the restriction fragment length polymorphism method, raising the possibility that ApoE-cholesterol balance might be critical for growth and cognitive development under the stress of heavy diarrhea burdens and when an enriched fat diet is insufficient. These findings provide a potential explanation for the survival advantage in evolution of genes, which might raise cholesterol levels during heavy stress of diarrhea burdens and malnutrition early in life.

ApoE polymorphisms and diarrheal outcomes in Brazilian shanty town children

A series of studies have shown that the heavy burdens of diarrheal diseases in the first 2 formative years of life in children living in urban shanty towns have negative effects on physical and cognitive development lasting into later childhood. We have shown that APOE4 is relatively common in shanty town children living in Brazil (13.4 percent) and suggest that APOE4 has a protective role in cognitive development as well as weight-for-height in children with heavy burdens of diarrhea in early childhood (64/123; 52 percent), despite being a marker for cognitive decline with Alzheimer’s and cardiovascular diseases later in life. APOE2 frequency was higher among children with heaviest diarrhea burdens during the first 2 years of life, as detected by PCR using the restriction fragment length polymorphism method, raising the possibility that ApoE-cholesterol balance might be critical for growth and cognitive development under the stress of heavy diarrhea burdens and when an enriched fat diet is insufficient. These findings provide a potential explanation for the survival advantage in evolution of genes, which might raise cholesterol levels during heavy stress of diarrhea burdens and malnutrition early in life.