Hernan Correa

Necrotising enterocolitis is characterised by disrupted immune regulation and diminished mucosal regulatory (FOXP3)/effector (CD4, CD8) T cell ratios

Background Necrotising enterocolitis (NEC) is the most common gastrointestinal emergency in premature infants. Immaturity of gastrointestinal immune regulation may predispose preterm infants to NEC as FOXP3 T regulatory cells (Treg) are critical for intestinal immune homoeostasis. Objective To investigate the hypothesis that abnormal developmental regulation of lamina propria Treg would define premature infants with NEC. Design Lamina propria mononuclear cell populations from surgically resected ileum from 18 patients with NEC and 30 gestational age-matched non-NEC surgical controls were prospectively isolated. Polychromatic flow cytometry was performed to phenotype and analyse lamina propria T cell populations. The cytokine gene expression profile in NEC tissue was compared with that of non-NEC controls. Results The total number of Treg, CD4, or CD8 T cells in each ileum section was independent of gestational age, age or postmenstrual age and similar between patients with NEC and controls. In contrast, the ratio of Treg to CD4 T cells or Treg to CD8 T cells was significantly lower in NEC ileum than in infants without NEC (medians 2.9% vs 6.6%, p=0.001 and medians 6.6% vs 25.9%, p<0.001, respectively). For any given number of CD4 or CD8 T cells, Treg were, on average, 60% lower in NEC ileum than in controls. NEC tissue cytokine gene expression profiles were characteristic of inhibited Treg development or function. Treg/CD4 and Treg/CD8 ratios recovered between initial resection for NEC and reanastomosis. Conclusion The proportion of lamina propria Treg is significantly reduced in the ileum of premature infants with NEC and may contribute to the excessive inflammatory state of this disease.

Eosinophils and mast cells in chronic gastritis: Possible implications in carcinogenesis

Eosinophils and mast cells participate in the immune response against Helicobacter pylori, but their involvement in the gastric precancerous process is unclear. This study aimed to estimate eosinophil and mast cell density in antral mucosa in subjects from 2 Colombian populations with contrasting gastric cancer risks. Gastric mucosa biopsies were collected from 117 adult males (72 from a high-risk area and 45 from a low-risk area). A histopathology score was used to quantify severity of the lesions. Quantitation of eosinophils in hematoxylin-eosin-stained sections and mast cells in immunostained sections for CD117/c-Kit was performed. Helicobacter pylori infection and genotyping were assessed in Steiner stain and polymerase chain reaction, respectively. Logistic regression models and semiparametric cubic smoothing splines were used for analysis of the results. Eosinophil density was significantly higher in subjects from the low-risk area as compared with subjects from the high-risk area. In both populations, eosinophil density increased with the histopathology score in the progression of lesions from normal morphology to multifocal atrophic gastritis. Intestinal metaplasia and dysplasia specimens showed further increase in eosinophil density in the high-risk area but an abrupt decrease in the low-risk area. Mast cell density increased in parallel to the histopathology score in both populations. Our results suggest that eosinophils play a dual role in chronic gastritis. In the low-risk area, elevated eosinophil density represents a T helper 2-biased response that may down-regulate the effects of proinflammatory cytokines preventing cancer development. In contrast, in the high-risk area, eosinophils might promote a T helper 1-type response leading to progression of precancerous lesions.

Ontogeny of FOXP3(+) regulatory T cells in the postnatal human small intestinal and large intestinal lamina propria.

FOXP3+ regulatory T cells (Treg) suppress innate and adaptive immune responses and are critical for intestinal immune homeostasis. Our objective was to define the postnatal developmental regulation of Treg in relationship to other T cells in the human intestinal tract. We analyzed 41 small and 18 large intestinal paraffin-embedded tissue samples from preterm and term infants with and without necrotizing enterocolitis (NEC) for the presence of CD3+, CD4+, CD8+, and FOXP3+ cells by immunohistochemistry. We compared labeled cells against age, gestational age (GA), or (corrected) postmenstrual age (PMA). The GA ranged from 23 to 40 weeks, with a mean of 32 (standard deviation, 4.7) weeks. Independent of age, GA, or PMA, the numbers of CD4+ cells were higher in the small intestine compared to the large intestine (P = 0.046), except in patients with NEC. FOXP3+ cells could be detected as early as 23 weeks in GA in both large and small bowel, and similar quantities were detected at the highest GA examined (40 weeks). We saw no statistically significant effect of GA, age, or PMA on total number of FOXP3+ cells or by comparing FOXP3+ to CD4+ or FOXP3+ to CD8+ ratios, indicating intact ontogeny of Treg in intestinal tissue early in gestation. Human infants exhibit presence of mucosal FOXP3+ cells in the small and large intestinal mucosa at birth and as early as 23 weeks GA. The frequency of FOXP3+ cells and the ratios of FOXP3+ to CD4+ or CD8+ cells do not change with increasing intrauterine development or postnatal age.

Early life establishment of site-specific microbial communities in the gut

Fecal sampling is widely utilized to define small intestinal tissue-level microbial communities in healthy and diseased newborns. However, this approach may lead to inaccurate assessments of disease or therapeutics in newborns because of the assumption that the taxa in the fecal microbiota are representative of the taxa present throughout the gastrointestinal tract. To assess the stratification of microbes in the newborn gut and to evaluate the probable shortcoming of fecal sampling in place of tissue sampling, we simultaneously compared intestinal mucosa and fecal microbial communities in 15 neonates undergoing intestinal resections. We report three key results. First, when the site of fecal and mucosal samples are further apart, their microbial communities are more distinct, as indicated by low mean Sørensen similarity indices for each patients fecal and tissue microbiota. Second, two distinct niches (intestinal mucosa and fecal microbiota) are evident by principal component analyses, demonstrating the critical role of sample source in defining microbial composition. Finally, in contrast to adult studies, intestinal bacterial diversity was higher in tissue than in fecal samples. This study represents an unprecedented map of the infant microbiota from intestinal mucosa and establishes discernable biogeography throughout the neonatal gastrointestinal tract. Our results question the reliance on fecal microbiota as a proxy for the developing intestinal microbiota. Additionally, the robust intestinal tissue-level bacterial diversity we detected at these early ages may contribute to the maturation of mucosal immunity.

Pediatric Patients With Dyspepsia Have Chronic Symptoms, Anxiety, and Lower Quality of Life as Adolescents and Adults

BACKGROUND & AIMS Little is known about long-term health outcomes of children with dyspeptic symptoms. We studied the natural history of pediatric patients with dyspeptic symptoms, with and without histologic reflux, compared with healthy controls. METHODS We performed a prospective study of consecutive new patients, ages 8-16 years, who underwent evaluation for dyspepsia, including upper endoscopy. Patients were assigned to groups with histologic evidence of reflux esophagitis (n = 50), or normal histology results (n = 53). Healthy children were followed up as controls (n = 143). Patients and controls were evaluated 5-15 years later. They provided self-reports on severity of dyspeptic symptoms, use of acid suppression, quality of life, anxiety, and depression. RESULTS When the study began, the groups with histologic evidence of esophagitis and normal histologies did not differ in severity of dyspeptic symptoms, functional disability, or depression. After a mean 7.6-year follow-up period, each group had significantly lower quality-of-life scores and more severe dyspeptic symptoms and functional disability than controls, but did not differ significantly from each other; both groups were significantly more likely than controls to meet criteria for an anxiety disorder. At time of follow-up evaluation, use of acid-suppression medication was significantly greater in the group with histologic evidence for esophagitis, compared with patients who had normal histology findings when the study began. CONCLUSIONS Among pediatric patients with dyspepsia evaluated by endoscopy and biopsy, those with histologic evidence for esophagitis or normal histology findings are at increased risk for chronic dyspeptic symptoms, anxiety disorder, and reduced quality of life in adolescence and young adulthood.

Apical sodium-dependent bile acid transporter upregulation is associated with necrotizing enterocolitis

Necrotizing enterocolitis (NEC) is the most common gastrointestinal emergency of premature infants. Previously, we showed that luminal bile acids (BAs) are increased and correlated with disease development and that the apical sodium-dependent BA transporter (ASBT), which transports BAs from the ileal lumen into enterocytes, is upregulated in rats with NEC. We hypothesized that intraenterocyte, rather than luminal, BAs are associated with NEC and that upregulation of ASBT may be a mechanism by which this occurs. Neonatal rats with or without the ASBT inhibitor SC-435, mice in which ASBT was knocked out, and mice that overproduce BAs were subjected to the NEC protocol. Disease development, ASBT, and the farnesoid X receptor protein, along with luminal and intraenterocyte BA levels, were assessed. In addition, ileal sections from premature infants with and without NEC were examined for ASBT via immunohistology and real-time PCR. When BAs were not transported into enterocytes (rats given SC-435 and ASBT knockout mice), severity and incidence of NEC were reduced. In contrast, in mice that overproduce BAs, ASBT was elevated, intraenterocyte BAs were increased, and disease development was increased. ASBT staining was more intense on the apical membrane of ileal enterocytes from premature infants with NEC than premature infants with non-NEC diagnoses. In addition, ASBT mRNA levels were significantly higher in infants with NEC. These data show that accumulation of intraenterocyte BAs contributes to disease development, elevated ASBT increases disease severity in experimental models of NEC, and ASBT is elevated in human NEC. These data confirm that BAs and upregulation of ASBT play a crucial role in NEC pathogenesis and suggest that inhibition of ASBT could be utilized as a therapeutic modality against this disease.

Absence of gastrointestinal pathogens in ileum tissue resected for necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is one of the most common gastrointestinal emergencies in premature infants and has been linked with viral antigens in as much as 40% of cases in single-center cohorts. We examined 28 tissue sections from surgically resected ileum from 27 preterm infants with NEC from 2 separate institutions for 15 common bacterial, viral, and parasitic gastrointestinal pathogens using multiplex reverse transcriptase polymerase chain reaction amplification and suspension array detection methods. We did not detect infectious enteritis pathogens in any of the NEC tissues and conclude that gastrointestinal pathogens are a rare cause of NEC.

Signal transduction pathway analysis in desmoid-type fibromatosis: transforming growth factor-β, COX2 and sex steroid receptors.

Despite reports of sex steroid receptor and COX2 expression in desmoid‐type fibromatosis, responses to single agent therapy with anti‐estrogens and non‐steroidal anti‐inflammatory drugs are unpredictable. Perhaps combination pharmacotherapy might be more effective in desmoid tumors that co‐express these targets. Clearly, further understanding of the signaling pathways deregulated in desmoid tumors is essential for the development of targeted molecular therapy. Transforming growth factor–β (TGFβ) and bone morphogenetic proteins (BMP) are important regulators of fibroblast proliferation and matrix deposition, but little is known about the TGFβ superfamily in fibromatosis. A tissue microarray representing 27 desmoid tumors was constructed; 14 samples of healing scar and six samples of normal fibrous tissue were included for comparison. Expression of selected receptors and activated downstream transcription factors of TGFβ family signaling pathways, β–catenin, sex steroid hormone receptors and COX2 were assessed using immunohistochemistry; patterns of co–expression were explored via correlational statistical analyses. In addition to β–catenin, immunoreactivity for phosphorylated SMAD2/3 (indicative of active TGFβ signaling) and COX2 was significantly increased in desmoid tumors compared with healing scar and quiescent fibrous tissue. Low levels of phosphorylated SMAD1/5/8 were detected in only a minority of cases. Transforming growth factor–β receptor type 1 and androgen receptor were expressed in both desmoid tumors and scar, but not in fibrous tissue. Estrogen receptor–β was present in all cases studied. Transforming growth factor–β signaling appears to be activated in desmoid‐type fibromatosis and phosphorylated SMAD2/3 and COX2 immunoreactivity might be of diagnostic utility in these tumors. Given the frequency of androgen receptor, estrogen receptor–β and COX2 co‐expression in desmoid tumors, further assessment of the efficacy of combination pharmacotherapy using hormonal agonists/antagonists together with COX2 inhibitors should be considered.

Laparoscopic nephron-sparing resection of synchronous Wilms tumors in a case of hyperplastic perilobar nephroblastomatosis.

Diffuse hyperplastic perilobar nephroblastomatosis (DHPLN) is a rare precursor lesion of Wilms tumor (WT). Because of the increased risk to develop WT in either kidney, current management algorithms of DHPLN merit nephron-sparing strategies, beginning with chemotherapy and close radiographic monitoring into late childhood. After resolution of DHPLN, subsequent detection of a renal nodule mandates resection to exclude WT. Here, we report the case of a 4-year-old girl who developed 2 synchronous nodules in the right kidney more than 2 years after completion of therapy for DHPLN. Because of the early detection and peripheral location of these 2 nodules, laparoscopic nephron-sparing resection of each was performed using ultrasonic dissection. Both nodules were determined on pathology to be favorable histology WT with negative surgical margins. The child was placed on vincristine and actinomycin D therapy for 18 weeks.

Unrelated stem cell transplant for infantile idiopathic myelofibrosis

Idiopathic myelofibrosis (IMF) is a rare disease in children that can present during infancy and have a protracted course. The only known curative approach for this disease in adult patients is allogeneic stem cell transplant. We present two cases of IMF during infancy that did not resolve with supportive care measures. Both patients underwent unrelated stem cell transplant with complete resolution of their hematologic manifestations and resolution of the bone marrow fibrosis. Pediatr Blood Cancer 2009;52:893–895.