Analysis and expansion of the eosinophilic esophagitis transcriptome by RNA sequencing.

Eosinophilic esophagitis (EoE) is an allergic inflammatory disorder of the esophagus that is compounded by genetic predisposition and hypersensitivity to environmental antigens. Using high-density oligonucleotide expression chips, a disease-specific esophageal transcript signature was identified and was shown to be largely reversible with therapy. In an effort to expand the molecular signature of EoE, we performed RNA sequencing on esophageal biopsies from healthy controls and patients with active EoE and identified a total of 1607 significantly dysregulated transcripts (1096 upregulated, 511 downregulated). When clustered by raw expression levels, an abundance of immune cell-specific transcripts are highly induced in EoE but expressed at low (or undetectable) levels in healthy controls. Moreover, 66% of the gene signature identified by RNA sequencing was previously unrecognized in the EoE transcript signature by microarray-based expression profiling and included several long non-coding RNAs (lncRNA), an emerging class of transcriptional regulators. The lncRNA BRAF-activated non-protein coding RNA (BANCR) was upregulated in EoE and induced in interleukin-13 (IL-13)-treated primary esophageal epithelial cells. Repression of BANCR significantly altered the expression of IL-13-induced proinflammatory genes. Together, these data comprise new potential biomarkers of EoE and demonstrate a novel role for lncRNAs in EoE and IL-13-associated responses.

A gene encoding a liver-specific ABC transporter is mutated in progressive familial intrahepatic cholestasis.

The progressive familial intrahepatic cholestases (PFIC) are a group of inherited disorders with severe cholestatic liver disease from early infancy. A subgroup characterized by normal serum cholesterol and gamma-glutamyltranspeptidase (gammaGT) levels is genetically heterogeneous with loci on chromosomes 2q (PFIC2) and 18q. The phenotype of the PFIC2-linked group is consistent with defective bile acid transport at the hepatocyte canalicular membrane. The PFIC2 gene has now been identified by mutations in a positional candidate, BSEP, which encodes a liver-specific ATP-binding cassette (ABC) transporter, sister of p-glycoprotein (SPGP). The product of the orthologous rat gene has been shown to be an effective bile acid transporter in vitro. These data provide evidence that SPGP is the human bile salt export pump (BSEP).

Bacteremia after intestinal transplantation in children correlates temporally with rejection or gastrointestinal lymphoproliferative disease.

BACKGROUND: Bacteremia occurs frequently after intestinal transplantation (ITx) in children. During our initial experience with this procedure, we noted that bacteremic episodes tended to occur simultaneously with the presence of rejection and/or gastrointestinal (GI) posttransplant lymphoproliferative disease (PTLD). AIM: To document the association of bacteremia with rejection and GI PTLD in pediatric ITx recipients. METHODS: Retrospective analysis of all medical records from 62 children who underwent ITx between July 1990 and January 1998 at Children's Hospital of Pittsburgh. A bacteremic episode was defined as two positive blood cultures from different sites at the same time or from the same site at different times. Rejection and PTLD were defined using previously published criteria. RESULTS: A total of 39/62 ITx recipients had 133 blood stream infections (2.1 episodes/patient) including 121 episodes of bacteremia and 12 of fungemia. Enteric organisms were the most frequently recovered pathogens (Gram negative rods, n=76; enterococci, n=36). Enteric organisms were recovered as a single organism (n=57), with another enteric bacteria (n=23), or with coagulase negative staphylococci (CONS) (n=24). CONS were recovered as a single organism on 21 occasions. An obvious source of bacteremia was not found for 115/121 episodes. Endoscopy was performed for 107 of the 115 bacteremia episodes; an abnormal histology was identified in 74 revealing rejection (n=36), GI PTLD (n=21), or both (n=17). When endoscopy showed GI pathology, enteric organisms alone or in combination with CONS were recovered on 63/107 occasions, although CONS were recovered alone only 11 times. CONCLUSIONS: Bacteremia accompanies GI rejection and intestinal PTLD in ITx recipients. Endoscopy should be performed to inspect the allograft when bacteremia occurs without an obvious source in these patients. This is especially true for patients with bacteremia due to enteric organisms.

Cricopharyngeal dysfunction associated with Chiari malformations.

UNLABELLED: Dysphagia due to upper esophageal sphincter (UES) dysfunction can be a manifestation of Chiari malformation. We evaluated five young children with dysphagia and a Chiari malformation before and after craniocervical decompression. Preoperatively, esophageal manometry with a multilumen perfused catheter revealed failure of complete relaxation of the UES in three patients, pharyngo-UES incoordination in one patient, and both abnormalities in the last patient. Preoperative barium esophagograms were obtained in four of the patients and were normal in two. One patient had nasal regurgitation of barium and delayed passage of barium through the UES. One patient had a posterior pharyngeal impression (bar) at the level of the UES and delayed transit of barium. All patients had clinical and manometric resolution of UES dysfunction following surgical decompression of the Chiari malformation. All swallows were coordinated, and UES relaxations were complete. However, the posterior pharyngeal bar persisted on postoperative esophagogram in the only patient who had had the abnormality preoperatively, although it no longer interfered with passage of barium. Another patient had a narrowed UES with decreased relaxation. Swallowing was radiographically normal in three patients postoperatively. CONCLUSION: Surgical decompression of Chiari malformation may lead to complete clinical and manometric resolution of dysphagia due to upper esophageal sphincter dysfunction. Esophageal manometry is more likely than barium swallow to demonstrate the abnormality, and correlates better with symptomatic improvement postoperatively.

Rectal bleeding in Prader-Willi syndrome.

Individuals with Prader-Willi syndrome manifest severe skin picking behavior. We report three patients with this syndrome in whom an extension of this behavior to rectal picking resulted in significant lower gastrointestinal bleeding and anorectal disease. The recognition of this behavior is important to avoid misdiagnosing inflammatory bowel disease in this group of patients.

Determinants of depression in recent onset pediatric inflammatory bowel disease.

Thirteen children with new-onset inflammatory bowel disease were given a Kiddie-SADS-E interview. Three were depressed at diagnosis. Six mothers had a history of depression based on Adult-SADS-L interviews. The depressed children were less severely ill than the nondepressed children and had not required steroids. However, the depressed children had significantly more life events and their families showed more evidence of conflict than did the nondepressed children.

Stridor and gastroesophageal reflux: diagnostic use of intraluminal esophageal acid perfusion (Bernstein test).

Five pediatric patients with stridor were evaluated to determine whether gastroesophageal reflux (GER) contributed to their stridor. Intraluminal esophageal acid perfusion (Bernstein test), pH probe, radiographic studies, laryngobronchoscopy, and esophageal histology were utilized. Thereafter, three of the five patients responded to therapy for GER; two did not. The Bernstein test proved to have greater diagnostic reliability than any other test employed, using response to antireflux therapy as the "gold standard" for diagnosing reflux-provoked stridor.

Sodium salicylate and bile acid-induced colonic secretion in the rat.

Previous studies by us suggested that luminally administered sodium salicylate blocks dihydroxy-bile acid-induced colonic secretion in the rat. In the present study, an in vivo rat cecal loop technique is employed to compare the effects of luminally administered and parenterally administered sodium salicylate upon chenodeoxycholic acid-induced colonic secretion. In our experiment, inoculation of four mM chenodeoxycholic acid into the rat cecum produced net secretion of water and sodium which was not reversed by preincubation of this bile acid with eight mM of sodium salicylate. Similarly, an intravenous bolus of either five mg of sodium salicylate per kg of body weight or 50 mg of sodium salicylate per kg of body weight failed to block salt or water secretion. Furthermore, 30 minute incubation of chenodeoxycholic acid with sodium salicylate produced neither reduction of in vitro aqueous bile acid concentration nor inhibition of ex vivo bile acid-facilitated hypotonic red cell hemolysis. These data suggest that sodium salicylate fails to sequester bile acids from aqueous solution and fails to block bile acid-mediated colonic secretion in the rat.

Characterization of late abdominal accumulation of 99Tcm-HMPAO leukocytes in a large population of children.

We retrospectively evaluated the incidence of late accumulation of 99Tcm-HMPAO leukocytes (99Tcm-WBC) in the right lower quadrant of a large population of children and characterized some predictive patterns that would enable differentiation of active inflammation from this late occasional accumulation of 99Tcm-WBC. We reviewed the charts of 211 children. The first group evaluated consisted of 79 controls: 30 normal children with no gastrointestinal disease, but who underwent 99Tcm-WBC scanning for other medical problems, and 49 children who had non-specific gastrointestinal (GI) complaints, but had no demonstrable inflammatory bowel disease by conventional diagnostic methods. The second group consisted of 132 children with inflammatory bowel disease: 80 children with Crohn's disease (CD), 34 with ulcerative colitis (UC) and 18 with indeterminate colitis (IC). Children were imaged at 30 min and 3 h. Fifteen (19%) of the 79 controls scanned showed accumulation of 99Tcm-WBC in the right lower quadrant at 3 h and none at 30 min. Of those 15, 8 were from the control population and 7 from the group with non-specific GI complaints and negative work-ups. There was no uptake in other segments of the bowel. The accumulation was faint, of lesser intensity than in the iliac wing, and diffuse, such that identification of a specific loop of involved bowel was not possible. Migration of the 99Tcm-WBC distal to the terminal ileum was demonstrated. The other 64 children in the control group showed no accumulation of 99Tcm-WBC at any time during their scans. All 79 scans were blindly interpreted as normal studies. There were no false-positive readings encountered in the 132 children with inflammatory bowel disease (80 CD, 34 UC, 18 IC) when the aforementioned characteristics of the late accumulation of 99Tcm were used to differentiate inflammation from this physiological excretion. In conclusion, the late accumulation of 99Tcm-WBC in the right lower quadrant is characterized by (1) accumulation at no less that 3 h, (2) no accumulation in other segments of the bowel, (3) faint accumulation of lesser intensity than in the iliac wing, (4) a diffuse accumulation pattern and (5) migration of the 99Tcm-WBC into the caecum and ascending colon over time. Recognition of this excretion pattern enables differentiation of active Crohn's disease of the small bowel from migration and accumulation of 99Tcm-WBC in the right lower quadrant of the abdomen.

Pediatric liver transplantation. History, recent innovations, and outlook for the future.

Pediatric liver transplantation has advanced remarkably over the past three decades. One-year survival has progressively increased to nearly 90% in patients transplanted for most forms of liver disease. Parallel advances in organ procurement, operative technique, immunosuppression, and infection control are responsible for improved patient survival. Among the most important advances are use of the University of Wisconsin (UW) organ preservation solution, the employment of venovenous bypass and/or "piggyback" operative technique, the development of cyclosporine A (CyA) and FK506, and the emergence of acyclovir, ganciclovir, foscarnet, and alpha interferon to combat life-threatening viral infections. The current organ shortage is being addressed by "cutdown" liver transplantation, "split liver" transplantation, and living-related donations. The next decade is likely to see advances in multi-visceral transplantation, induction of chimerism by simultaneous bone marrow-solid organ transplantation, and performance of cross-species xenografting.

Retrospective analysis of alternate-day prednisone maintenance therapy for Crohn's disease.

We reviewed the medical records of 98 children with Crohn's disease followed at Children's Hospital of Pittsburgh from 1983 to 1993 to evaluate the merits of alternate-day prednisone (AD) maintenance therapy once initial remission was achieved. Of the 98 children, 35 had adequate data recorded for eligibility to the study. Of these, 11 were in the AD group and 24 were in a group whose maintenance regimen did not include prednisone (NO). The dependent variables were frequency of flares and linear growth over time. AD therapy reduced mean symptomatic flares (0.23 +/- 0.1 vs 0.69 +/- 0.14 flares/patient/year; p = 0.04) over a 2-year follow-up period but did not delay significantly the onset of a flare after remission was achieved (16.5 +/- 3.4, vs 13.4 +/- 1.8 months; p = 0.4). Site of disease involvement had no impact on frequency of flares. Fewer patients in the AD group experienced flares, but this finding did not achieve statistical significance (4/11, 36%, vs 17/24, 71%; p = 0.07). Linear growth, measured in height percentile and growth velocity (cm/year), was not significantly reduced by the second year of either therapy. This small retrospective study suggests that AD prednisone therapy may be effective in reducing symptomatic flares in Crohn's patients without a resultant inhibition of linear growth.

Prevalence and characteristics of Blastocystis hominis infection in children.

Blastocystis hominis, a protozoan whose pathogenicity has been questioned, is sometimes found in the human gastrointestinal tract. We sought to determine the prevalence of Blastocystis in stool and to characterize clinical features of infection with Blastocystis in children. Forty-six (3%) of 1,736 patients undergoing fecal microscopy at Children's Hospital of Pittsburgh between January 1, 1985, and December 31, 1988, harbored Blastocystis. Of these 46 children, 75% had exposure to well water or had been in developing countries. Thirty-nine of the 46 (85%) experienced gastrointestinal symptoms, such as abdominal pain, diarrhea, vomiting, and weight loss. Blastocystis was the only parasite found in 35 of those 39 symptomatic children. Symptoms resolved within one month in 90% of patients receiving antiparasitic pharmacotherapy, but in only 58% (P < .04) of those receiving no therapy. We conclude that children infected with Blastocystis often experience gastrointestinal symptoms and that treatment increases the rate of symptomatic improvement. We speculate that Blastocystis is a human pathogen.

Diagnosis and treatment of cryptosporidiosis in children.

Clinicians have only recently become aware that Cryptosporidium is a major human pathogen, causing a self-limited gastroenteritis in immunocompetent individuals and profound enteropathy in the immunocompromised. Among the immunologically intact, children appear more prone to clinically significant cryptosporidiosis than do adults. Diagnosis is most easily made by a specialized three-step stool examination, but mucosal biopsy may also be necessary to establish the diagnosis. Although no satisfactory antibiotic therapy exists today, cases of cryptosporidiosis in the community and hospital should be identified in order to contain human transmission. Only then can spread from immunocompetent individuals to immunodeficient individuals be avoided.

Small bowel transplantation in infants and children.

Multivisceral transplantation, combined liver-intestine transplantation, and isolated small bowel transplantation are very similar procedures that were first developed in the 1950s. If the viscera can be conceptualized as a cluster of grapes hanging from its arterial stems, the three procedures are characterized by virtually identical vascular anastomoses, with exclusion or inclusion of as many viscera (grapes) as necessary; however, these procedures languished for nearly four decades because of the imperfect immunosuppressive regimens of the 1960s, 1970s, and 1980s. Finally, after the development of FK506, pediatric patients may undergo intestinal transplantation with the hope for long-term survival. These procedures are reserved for TPN-dependent children with permanent intestinal insufficiency. Candidacy for transplantation is also predicated on development of potentially fatal TPN complications such as cholestasis, recurrent sepsis, or thrombosis of access sites. Since 1990, 32 pediatric patients have undergone intestinal transplantation at the University of Pittsburgh, with an overall survival of 65%. Immunosuppression has been accomplished with a combination of corticosteroids, FK506, and prostaglandin E1. Although GVHD has not been a major problem, most patients have experienced rejection episodes requiring intensification of immunosuppression with a steroid bolus, a steroid recycle, an increase in FK506 dosage, or addition of OKT3. CMV has caused little morbidity, but EBV-related PTLD has affected 20% of all patients. It has not been possible to discontinue immunosuppression in the face of PTLD without engendering severe small intestinal rejection. Other problems have included recurrent sepsis, intestinal dysmotility, and persistent food avoidance. Future therapeutic trends are likely to include the performance of combined bone marrow-visceral transplant to induce a chimeric tolerogenic state and to lessen the need for long-term immunosuppression.

Failure of Saccharomyces boulardii to hydrolyse bile acids in vitro.

The bile acid deconjugating capacity of Saccharomyces boulardii was tested in isolated culture and in co-culture upon the bile acid metabolizing capacity of Bacteroides fragilis. S. boulardii was grown over 60 h in plain brain heart infusion (BHI) broth and in the presence of either 2 mM glycocholic or glycochenodeoxycholic acid under both aerobic and anaerobic conditions. S. boulardii did not metabolize either glycocholic acid or glycochenodeoxycholic acid. S. boulardii and B. fragilis were grown anaerobically together in BHI broth supplemented with either 0.1 mM glycholic acid or 0.1 mM glycochenodeoxycholic acid. When S. boulardii and B. fragilis were anaerobically co-cultured, S. boulardii did not alter the bile acid metabolism of B. fragilis. Specifically primary and conjugated bile acids decreased over time, while keto bile acids and free bile acids increased. S. boulardii not only fails to metabolize primary conjugated bile acids but it does not enhance or inhibit the bile acid metabolism of B. fragilis when the organisms are co-cultured in anaerobic medium.

Patient and procedure variables associated with complications following variceal sclerotherapy in children.

Variceal sclerotherapy has been performed in the pediatric population, but techniques and dosages of sclerosant recommended in the literature are largely empirical. Having accumulated much data through experience with sclerotherapy in children, we have identified patient and procedure variables associated with early and late complications. We reviewed our experience with 37 pediatric patients, ages 1-18, who underwent 150 sclerotherapy sessions. Sclerotherapy was associated with 12 early complications in 11 patients. Early complications were bleeding (five), respiratory problems (three), false channel formation (two), and gross hematuria (two). Four patients developed strictures. We identified the following variables associated with complications: for bleeding, platelet count less than 100,000/mm3; for respiratory complications and false channel formation, weight less than 12 kg; for false channel formation, dosage of sclerosant/kg/session greater than 1.75 ml/kg; for gross hematuria, total sclerosant/session greater than 20 ml. A need for more than six sclerotherapy sessions for obliteration of varices was associated with a greater frequency of stricture formation. Sclerotherapy can be performed safely in children. Patients less than 12 kg or with platelet counts less than 100,000/mm3 should be monitored carefully for respiratory complications and postprocedure bleeding. Sclerosant dosages greater than 1.75 ml/kg, or 20 ml total sclerosant, should be given with caution.