Plasminogen Activator Inhibitor-1 as a Marker of Esophageal Functional Changes in Pediatric Eosinophilic Esophagitis.

BACKGROUND & AIMS: Esophageal remodeling in eosinophilic esophagitis (EoE) can lead to esophageal rigidity with eventual luminal compromise and stenoses. Gauging esophageal functional alterations in EoE is challenging. An epithelial marker of functional remodeling would impact EoE management. METHODS: Esophageal biopsy specimens from children with and without EoE and primary human esophageal epithelial cells were used for PAI-1 immunohistochemistry, and cell proliferation experiments. PAI-1 immunostaining and basal cell hyperplasia were assessed in the context of concurrently obtained esophageal compliance measures on endoscopic functional lumen imaging probe (EndoFLIP). RESULTS: EndoFLIPs were performed in 45 children (32 with and 13 without EoE). Epithelial PAI-1 was increased in patients with active EoE versus inactive or control patients (P < .01). Esophageal compliance was lower in EoE patients versus controls, particularly in the proximal esophagus (P < .001). Proximal compliance was the strongest predictor of EoE (AUROC 0.88, 95% CI 0.77, 0.98) with esophageal compliance of less than 2.6%mL/mmHg demonstrating 82% sensitivity and 84% specificity for EoE. PAI-1 inhibition significantly diminished esophageal epithelial cell proliferation, suggesting PAI-1 could trigger basal cell hyperplasia. A composite mid-esophageal BZH + PAI-1 score was the strongest predictor of altered compliance (P = .02, AUROC 0.89 (95% CI 0.80, 0.99). CONCLUSIONS: PAI-1 is significantly elevated in pediatric EoE and distinguishes altered compliance in children. PAI-1 may be a novel disease marker and therapeutic target.

Congenital cutaneous non-neural granular cell tumor mimicking herpetic infection: Case report and review of the literature.

We present a case of a female neonate with a cluster of six skin colored to yellowish pseudovesicular papules on her right forearm present since birth, initially thought to be a herpes simplex virus infection. Punch biopsy with immunostaining revealed a diagnosis of S100-negative, CD163-positive congenital cutaneous non-neural granular cell tumor. Only four other reports are presented in the literature of this entity, three of which also presented on the arm with somewhat similar clinical findings. We briefly reviewed the subtypes of classic and S100-negative non-neural granular cell tumors.

Immune Dysregulation in the Tonsillar Microenvironment of Periodic Fever, Aphthous Stomatitis, Pharyngitis, Adenitis (PFAPA) Syndrome.

Periodic Fever, Aphthous stomatitis, Pharyngitis and Adenitis (PFAPA) syndrome is an inflammatory disorder of childhood classically characterized by recurrent fevers, pharyngitis, stomatitis, cervical adenitis, and leukocytosis. While the mechanism is unclear, previous studies have shown that tonsillectomy can be a therapeutic option with improvement in quality of life in many patients with PFAPA, but the mechanisms behind surgical success remain unknown. In addition, long-term clinical follow-up is lacking. In our tertiary care center cohort, 62 patients with PFAPA syndrome had complete resolution of symptoms after surgery (95.3%). Flow cytometric evaluation demonstrates an inflammatory cell population, distinct from patients with infectious pharyngitis, with increased numbers of CD8+ T cells (5.9% vs. 3.8%, p < 0.01), CD19+ B cells (51% vs. 35%, p < 0.05), and CD19+CD20+CD27+CD38-memory B cells (14% vs. 7.7%, p < 0.01). Cells are primed at baseline with increased percentage of IL-1ß positive cells compared to control tonsil-derived cells, which require exogenous LPS stimulation. Gene expression analysis demonstrates a fivefold upregulation in IL1RN and TNF expression in whole tonsil compared to control tonsils, with persistent activation of the NF-κB signaling pathway, and differential microbial signatures, even in the afebrile period. Our data indicates that PFAPA patient tonsils have localized, persistent inflammation, in the absence of clinical symptoms, which may explain the success of tonsillectomy as an effective surgical treatment option. The differential expression of several genes and microbial signatures suggests the potential for a diagnostic biomarker for PFAPA syndrome.

Variation in Endoscopic Activity Assessment and Endoscopy Score Validation in Adults With Eosinophilic Esophagitis.

BACKGROUND & AIMS: Eosinophilic esophagitis (EoE) is assessed endoscopically (endoscopic activity), based on grades of edema, rings, exudates, furrows, and strictures (EREFS). We examined variations in endoscopic assessments of severity, developed and validated 3 EREFS-based scoring systems, and assessed responsiveness of these systems using data from a randomized placebo-controlled trial of patients with EoE. METHODS: For the development set, 5 gastroenterologists reviewed EREFS findings from 266 adults with EoE and provided endoscopist global assessment scores (EndoGA, scale of 0 to 10); variation (ΔEndoGA) was assessed using linear regression. We evaluated simple scores (features given arbitrary values from 0 to 3) and developed 2 scoring systems (adjusted score range, 0-100). We then fitted our linear regression model with mean EndoGA to data from 146 adults recruited in centers in Switzerland and the United States between April 2011 and December 2012. For the validation set, we collected data from 120 separate adults (recruited in centers in Switzerland and the United States between May 2013 and July 2014), assessing regression coefficient-based scores using Bland-Altman method. We assessed the responsiveness of our scoring systems using data from a randomized trial of patients with EoE given fluticasone (n=16) or placebo (n=8). RESULTS: The distribution of EndoGA values differed among endoscopists (mean ΔEndoGA, 2.6±1.8; range 0-6.6). We developed 2 regression-based scoring systems to assess overall and proximal and distal esophageal findings; variation in endoscopic features accounted for more than 90% of the mean EndoGA variation. In the validation group, differences between mean EndoGA and regression-based scores were small (ranging from -4.70 to 2.03), indicating good agreement. In analyses of data from the randomized trial, the baseline to end of study change in patients given fluticasone was a reduction of 24.3 in simple score (reduction of 4.6 in patients given placebo, P=.052); a reduction of 23.5 in regression-based overall score (reduction of 6.56 in patients given placebo, P=.12), and a reduction of 23.8 (reduction of 8.44 in patients given placebo, P=.11). CONCLUSION: Assessments of endoscopic activity in patients with EoE vary among endoscopists. In an analysis of data from a randomized controlled trial, we found that newly developed scoring systems are no better than simple scoring system in detecting changes in endoscopic activity. These results support the use of a simple scoring system in evaluation of endoscopic activity in patients with EoE. clinicaltrials.gov no: NCT00939263 and NCT01386112.

Non-alcoholic fatty liver disease in pediatric type 2 diabetes: Metabolic and histologic characteristics in 38 subjects.

BACKGROUND: Obesity and type 2 diabetes (T2D) is risk factors for non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH). In children with T2D and liver biopsies, we investigated correlations between NAFLD/NASH and transaminase activity, A1c, lipids, and histologic changes in repeat biopsies. METHODS: Liver histology of children with T2D was evaluated using the NASH CRN scoring system and NAFLD Activity Score (NAS). We included results ≤6 months from biopsy and A1c nearest biopsy. RESULTS: Thirty-eight subjects (21 females, 17 males, 63.2% Hispanic, 15.8% Caucasian) had T2D diagnosed at 13.4 ± 2.7 years, 78.9% using metformin and 50% on insulin. Histological diagnosis of NAFLD occurred at mean age 14.3 ± 2.3 years, notable for NASH in 61%. Steatosis grade was higher in children with NASH than those without (mean 2.6 ± 0.7 vs 2.1 ± 0.5 (P < 0.001). Stage 3 fibrosis was noted only in subjects with NASH (26%). ALT was higher in NASH vs those without (112 ± 56 vs 85 ± 112, P = 0.016). NAS correlated with A1c (r = 0.51, P < 0.01) and triglycerides (r = 0.5, P < 0.01), and inversely with high-density lipoprotein (HDL) (r = -0.42, P = 0.04). Males had lower HDL and higher triglycerides (P < 0.04). In eight subjects with repeat biopsies, NAS was equal (37.5%) or improved (62.5%), and steatosis decreased (68.1% to 32.8%, P = 0.027). CONCLUSIONS: In children with T2D and NAFLD, NASH is common. Having advanced fibrosis in 26% of NASH cases at this age is concerning. Better control of lipids, weight, and diabetes may help avoid worsening in NAS.

Esophageal Compliance Quantifies Epithelial Remodeling in Pediatric Patients With Eosinophilic Esophagitis.

BACKGROUND: The management of eosinophilic esophagitis (EoE) relies on the severity of esophageal eosinophilia, yet there is poor evidence of its prediction of esophageal fibrotic remodeling and subsequent complications such as dysphagia, food impactions, or strictures. Functional luminal imaging planimetry (FLIP) has had limited use in pediatric patients to evaluate esophageal tissue mechanics. We aimed to standardize the FLIP technique and to measure esophageal compliance in children with EoE in comparison to controls. METHODS: Subjects were enrolled into a prospective observational study and had FLIP performed at the time of endoscopy. We calculated esophageal distensibility and compliance for the total and segmental esophagus independently (ie, proximal, middle, and distal esophageal segments). We evaluated esophageal biopsies for eosinophilia and epithelial remodeling, calculated endoscopy scores, and documented patient symptoms. RESULTS: We enrolled 11 EoE and 12 controls subjects, aged 5 to 18 years old. While EoE subjects had lower esophageal compliance (P = 0.004) than controls, the difference in distensibility did not reach significance (P = 0.151). Epithelial remodeling severity was more strongly correlated with compliance than with distensibility. Epithelial remodeling scores ≥2 had a significant association with lower compliance both segmentally and in the entire esophagus (P = 0.029), but not with distensibility. Compliance measures were more sensitive in detecting subjects with remodeling score ≥2 than distensibility (79% vs 64%). CONCLUSIONS: Compliance is a more sensitive measure of esophageal epithelial remodeling in children compared to distensibility, and a more appropriate measure of esophageal tissue mechanics. Standardized placement of the FLIP catheter is important to accurately assess esophageal compliance.

Interleukin 9 Alters Epithelial Barrier and E-cadherin in Eosinophilic Esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is a chronic TH2-assocated inflammatory condition accompanied by substantial impairments in epithelial barrier function and increased numbers of interleukin 9 (IL-9) expressing inflammatory cells. While IL-9 is known to affect barrier function in the intestine, the functional effects of IL-9 on the esophagus are unclear. Herein we aimed to understand the expression of the IL-9 receptor and effects of IL-9 on the epithelium in EoE. METHODS: We used esophageal biopsies from pediatric EoE patients with active and inactive disease to analyze the expression of the IL-9 receptor, the adherens junction protein E-cadherin and the tight junction protein claudin-1. We treated primary human esophageal epithelial cells with IL-9 to understand its effects on E-cadherin expression and function. RESULTS: Active EoE subjects had increased epithelial expression of IL-9 receptor mRNA and protein (P < 0.05) and decreased membrane bound E-cadherin (P < 0.01) and claudin-1 (P < 0.05) expression. IL-9 receptor expression and mislocalized claudin-1 positively correlated and while membrane bound E-cadherin expression negatively correlated with the degree of histologic epithelial remodeling (P < 0.05). IL-9 decreased epithelial resistance in stratified primary human esophageal epithelial cells (P < 0.01) and membrane bound E-cadherin in epithelial cell monolayers (P < 0.01). CONCLUSIONS: These data suggest that IL-9, its receptor, and its effects on E-cadherin may be important mechanisms for epithelial barrier disruption in EoE.

Reactivation of TWIST1 contributes to Ewing sarcoma metastasis.

BACKGROUND: Ewing sarcoma is a cancer of bone and soft tissue. Despite aggressive treatment, survival remains poor, particularly in patients with metastatic disease. Failure to treat Ewing sarcoma is due to the lack of understanding of the molecular pathways that regulate metastasis. In addition, no molecular prognostic markers have been identified for Ewing sarcoma to risk stratify patients. PROCEDURE: Ewing sarcoma patients were divided into high or low Twist1 gene expression and survival curves were generated using the R2 microarray-based Genomic Analysis platform (http://r2.amc.nl). Tumors from Ewing sarcoma patients were also evaluated for TWIST1 expression by immunohistochemistry. Ewing sarcoma xenografts were established to evaluate the role of TWIST1 in metastasis. The effects of Twist1 on migration and invasion were evaluated using migration and invasion assays in A673 and RDES cells. RESULTS: Twist1 expression was a negative prognostic marker for overall survival in a public Ewing sarcoma patient data set based on Twist1 mRNA levels and in patient tumor samples based on Twist1 immunohistochemistry. TWIST1 is detected in significantly higher percentage of patients with metastatic diseases than localized disease. Using Ewing sarcoma tumor xenografts in mice, we found that suppressing TWIST1 levels suppressed metastasis without affecting primary tumor development. Knockdown of Twist1 inhibited the migration and invasion capability, while overexpression of Twist1 promoted migration and invasion in Ewing sarcoma cells. CONCLUSION: These results suggest that TWIST1 promotes metastasis in Ewing sarcoma and could be used as a prognostic marker for treatment stratification; however, further validation is required in a larger cohort of patients.

Non-anastomotic failure of woven Dacron tube grafts in the thoracic aorta in young adults.

BACKGROUND: Dacron tube grafts have been used in the surgical management of cardiovascular disease since the 1970s. Complications at the site of the anastomosis have been well described. Non-anastomotic failure is far less common. We present a series of four patients who presented with complications of non-anastomotic failure of woven Dacron tube grafts. METHODS: A retrospective chart review of four patients who presented to our institution between March 2014 and March 2017 with clinical complications of a Dacron tube graft was conducted. RESULTS: All four patients underwent a staged surgical repair for an interrupted aortic arch between the years of 1988 and 2001. All four patients underwent revision of their original interposition graft (Gore-Tex, W.L. Gore & Associates, Flagstaff, AZ) with implantation of a Hemashield woven Dacron tube graft (Maquet, Rastatt, Germany). From 13 to 22 years postimplant of the Dacron tube graft, all patients presented with symptoms or clinical evidence of primary graft failure. Two patients underwent urgent surgical intervention and did not survive. One patient underwent attempted surgical intervention, which was aborted in the setting of profuse bleeding, and ultimately had an endovascular rescue of the tube graft with a Zenith Alpha endograft (Cook Medical, Bloomington, IN). One patient underwent elective endovascular intervention prior to onset of symptoms. CONCLUSION: Non-anastomotic failure of woven Dacron tube grafts can occur in the thoracic aorta in young adults and may be managed with endovascular techniques.

Symptoms Have Modest Accuracy in Detecting Endoscopic and Histologic Remission in Adults With Eosinophilic Esophagitis.

BACKGROUND & AIMS: It is not clear whether symptoms alone can be used to estimate the biologic activity of eosinophilic esophagitis (EoE). We aimed to evaluate whether symptoms can be used to identify patients with endoscopic and histologic features of remission. METHODS: Between April 2011 and June 2014, we performed a prospective, observational study and recruited 269 consecutive adults with EoE (67% male; median age, 39 years old) in Switzerland and the United States. Patients first completed the validated symptom-based EoE activity index patient-reported outcome instrument and then underwent esophagogastroduodenoscopy with esophageal biopsy collection. Endoscopic and histologic findings were evaluated with a validated grading system and standardized instrument, respectively. Clinical remission was defined as symptom score <20 (range, 0-100); histologic remission was defined as a peak count of <20 eosinophils/mm(2) in a high-power field (corresponds to approximately <5 eosinophils/median high-power field); and endoscopic remission as absence of white exudates, moderate or severe rings, strictures, or combination of furrows and edema. We used receiver operating characteristic analysis to determine the best symptom score cutoff values for detection of remission. RESULTS: Of the study subjects, 111 were in clinical remission (41.3%), 79 were in endoscopic remission (29.7%), and 75 were in histologic remission (27.9%). When the symptom score was used as a continuous variable, patients in endoscopic, histologic, and combined (endoscopic and histologic remission) remission were detected with area under the curve values of 0.67, 0.60, and 0.67, respectively. A symptom score of 20 identified patients in endoscopic remission with 65.1% accuracy and histologic remission with 62.1% accuracy; a symptom score of 15 identified patients with both types of remission with 67.7% accuracy. CONCLUSIONS: In patients with EoE, endoscopic or histologic remission can be identified with only modest accuracy based on symptoms alone. At any given time, physicians cannot rely on lack of symptoms to make assumptions about lack of biologic disease activity in adults with EoE. ClinicalTrials.gov, Number: NCT00939263.

Long-term assessment of esophageal remodeling in patients with pediatric eosinophilic esophagitis treated with topical corticosteroids.

BACKGROUND: Eosinophilic esophagitis (EoE) is a chronic TH2 inflammatory disease characterized by tissue remodeling that leads to esophageal strictures and food impactions. Effects of therapy on long-term remodeling in patients with pediatric eosinophil-associated diseases have not been previously described. OBJECTIVE: We sought to understand the long-term control of esophageal remodeling in patients with EoE. METHODS: We assessed endoscopic and histologic remodeling and TGF-ß1 expression in esophageal biopsy specimens from children (n = 32) with EoE treated with topical corticosteroids (TCSs) over 10 years (mean, 4.5 years). We used standardized EoE scoring tools to gauge endoscopic and symptom features. RESULTS: Seven hundred thirty-eight biopsy specimens from 246 endoscopic procedures were evaluated over 10 years. Four hundred eighty-six biopsy specimens had adequate lamina propria for evaluation of subepithelial remodeling. The severity of epithelial esophageal eosinophilia correlated with epithelial remodeling (basal zone hyperplasia, desquamation, and dilated intercellular spaces; P < .0001), lamina propria eosinophilia (P < .0001), and fibrosis (P < .0001). Sixteen subjects were initial responders (<15 eosinophils/high-power field) to TCSs. Responders and nonresponders spent 54% and 97% of their total disease duration with active EoE (P < .001) and 23% and 53% (P < .02) with maximal fibrosis scores, respectively. Responders had lower endoscopy scores during their disease duration (P = .013). Having less than 15 eosinophils/high-power field at any time correlated with lower fibrosis and endoscopic severity. TGF-ß1(+) cell counts decreased in responders at the first biopsy, but this was not sustained. Symptoms did not correlate with other disease features. CONCLUSIONS: Children with EoE have substantial esophageal remodeling, which associates with inflammation and can improve in a sustainable manner with TCSs. Although endoscopic features correspond to histologic features, symptoms did not correlate with inflammation or fibrosis.

TGF-ß1-induced PAI-1 contributes to a profibrotic network in patients with eosinophilic esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is an allergic disease of increasing worldwide incidence. Complications are due to tissue remodeling and involve TGF-ß1-mediated fibrosis. Plasminogen activator inhibitor 1 (PAI-1/serpinE1) can be induced by TGF-ß1, but its role in EoE is not known. OBJECTIVE: We sought to understand the expression and role of PAI-1 in patients with EoE. METHODS: We used esophageal biopsy specimens and plasma samples from control subjects and patients with EoE, primary human esophageal epithelial cells, and fibroblasts from patients with EoE in immunohistochemistry, quantitative PCR, and immunoassay experiments to understand the induction of PAI-1 by TGF-ß1, the relationship between PAI-1 and esophageal fibrosis, and the role of PAI-1 in fibrotic gene expression. RESULTS: PAI-1 expression was significantly increased in epithelial cells of biopsy specimens from patients with active EoE compared with that seen in biopsy specimens from patients with inactive EoE or control subjects (P < .001). Treatment of primary esophageal epithelial cells with recombinant TGF-ß1 increased PAI-1 transcription, intracellular protein expression, and secretion. Esophageal PAI-1 expression correlated with basal zone hyperplasia, fibrosis, and markers of esophageal remodeling, including vimentin, TGF-ß1, collagen I, fibronectin, and matrix metalloproteases, and plasma PAI-1 levels correlated with plasma TGF-ß1 levels. PAI-1 inhibition significantly decreased baseline and TGF-ß1-induced fibrotic gene expression. CONCLUSIONS: PAI-1 expression is significantly increased in the epithelium in patients with EoE and reflects fibrosis, and its inhibition decreases TGF-ß1-induced gene expression. Epithelial PAI-1 might serve as a marker of EoE severity and form part of a TGF-ß1-induced profibrotic network.

How do gastroenterologists assess overall activity of eosinophilic esophagitis in adult patients?

OBJECTIVES: There is no "gold standard" for assessing disease activity in patients with eosinophilic esophagitis (EoE). We aimed to compare physicians' judgment of EoE activity with patients' judgment of symptom severity. We also aimed to examine the relative contribution of symptoms as well as endoscopic and histologic findings in shaping physicians' judgment of EoE activity. METHODS: Six gastroenterologists (all EoE experts) assessed EoE-associated symptoms in adult patients. Patients completed a symptom instrument and provided global assessment of EoE symptom severity (PatGA) (Likert scale: 0 (inactive) to 10 (most active)). Following esophagogastroduodenoscopy with biopsy sampling, gastroenterologists provided a global assessment of EoE activity (PhysGA) (Likert scale from 0 to 10) based on patient history and endoscopic and histologic findings. Linear regression and analysis of variance was used to quantify the extent to which variations in severity of EoE symptoms and endoscopic and histologic findings explain variations in PhysGA. RESULTS: A total of 149 EoE patients were prospectively included (71.8% male, median age at inclusion 38 years, 71.8% with concomitant allergies). A moderate positive correlation between PhysGA and PatGA (rho=0.442, P<0.001) was observed and the mean difference in the Bland-Altman plot was 1.77. Variations in severity of endoscopic findings, symptoms, and histologic findings alone explained 53%, 49%, and 30%, of the variability in PhysGA, respectively. Together, these findings explained 75% of variability in PhysGA. CONCLUSIONS: Gastroenterologists rate EoE activity mainly on the basis of endoscopic findings and symptoms and, to a lesser extent, on histologic findings.

MMPs-2 and -14 Are Elevated in Eosinophilic Esophagitis and Reduced Following Topical Corticosteroid Therapy.

OBJECTIVES: Eosinophilic esophagitis (EoE) is a chronic, antigen-mediated disease in children and adults associated with substantial esophageal remodeling and fibrosis. The expression of the remodeling-associated matrix metalloproteinases (MMPs) has not been previously detailed in EoE. METHODS: MMP-2 and -14 expression and cellular localization were assessed using real-time quantitative polymerase chain reaction and immunohistochemistry/immunofluorescence in EoE fibroblasts, active and inactive pediatric EoE biopsies, and nondiseased control biopsies. The effect of transforming growth factor (TGF)-ß1 treatment on MMP-2 expression in cultured esophageal epithelial (HET1A) cells was analyzed. RESULTS: MMP-2 and -14 mRNA were expressed in EoE fibroblasts and biopsies. Proliferating epithelial cells produced MMP-14 more abundantly in EoE than in controls (P < 0.001) and the degree of epithelial MMP-14 expression correlated positively with basal zone hyperplasia (r = 0.65, P = 0.002). EoE lamina propria had higher numbers of MMP-2- and -14-positive cells (906 ±â€Š167 and 701 ±â€Š93 cells/mm²) as compared with controls (258 ±â€Š93 cells/mm², P < 0.01 and 232 ±â€Š54 cells/mm², P < 0.01), and MMP-14 expression correlated with the severity of fibrosis. Following therapy with topical corticosteroids, MMP-14 and -2 were significantly diminished (P < 0.01). TGF-ß1 increased the expression and secretion of MMP-2 from esophageal epithelial HET1A cells. CONCLUSIONS: MMP-2 and -14 are elevated in pediatric patients with EoE and significantly decrease following topical corticosteroid therapy. TGF-ß1 increases MMP-2 in esophageal epithelial cells. This alludes to previously unappreciated role for MMPs in EoE-associated esophageal remodeling and a potential positive feedback loop via TGF-ß1.

TGF-ß1-induced phospholamban expression alters esophageal smooth muscle cell contraction in patients with eosinophilic esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is a chronic antigen-mediated disease characterized by esophageal eosinophilia, remodeling, and fibrosis. TGF-ß1 is a central regulator of EoE remodeling and increases esophageal smooth muscle (ESM) cell contraction. OBJECTIVE: In this study we aimed to understand the molecular mechanisms by which TGF-ß1 could induce ESM cell contraction. METHODS: We used primary human ESM cells and esophageal myofibroblasts (EMFs) to assess the mechanisms of TGF-ß1-induced contraction. We analyzed the expression, phosphorylation, and function of phospholamban (PLN), a sarcoendoplasmic reticulum regulatory protein induced by TGF-ß1. Expression of PLN, phospho-PLN, and its regulatory pathway was analyzed in the ESM of biopsy specimens from patients with EoE and control subjects. Gene silencing in EMFs from patients with EoE was used to understand the role of PLN in contraction. RESULTS: TGF-ß1 induced and phosphorylated PLN in primary human ESM cells and EMFs from patients with EoE. PLN and phospho-PLN levels were increased in smooth muscle from patients with EoE compared with that seen in smooth muscle from control subjects in vivo. PLN inhibition significantly diminished TGF-ß1-induced EMF contraction in patients with EoE. PLN expression and ESM/EMF contraction depended on TGF-ß receptor I signals. CONCLUSION: We describe a previously unrecognized mechanism for ESM cell contraction that depends on TGF-ß1, its receptors, and PLN. Because PLN levels are increased in smooth muscle from patients with EoE and PLN silencing diminishes contraction, we provide a novel potential mechanistic framework and therapeutic target for ESM dysfunction in patients with EoE.

Glioblastoma multiforme arising from dysembryoplastic neuroepithelial tumor in a child in the absence of therapy.

Dysembryoplastic neuroepithelial tumors (DNETs) are considered as low-grade tumors commonly associated with intractable seizures. We report a case of an unusual hemispheric DNET in a young child presenting with new-onset focal seizures. The tumor was notable for its atypical neuroimaging features and very rapid malignant transformation into a glioblastoma multiforme in the absence of radiation or chemotherapy, 1-year postdiagnosis. Our case highlights the malignant potential of atypical DNETs in the absence of therapy.

Anti-IL-5 therapy reduces mast cell and IL-9 cell numbers in pediatric patients with eosinophilic esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is a clinicopathologic entity of increasing worldwide prevalence. IL-5 is essential for eosinophil trafficking, and anti-IL-5 therapy decreases esophageal eosinophilia. EoE is associated with prominent mast cell infiltration. OBJECTIVE: We investigated whether anti-IL-5 (mepolizumab) treatment reduced esophageal mast cell accumulation in biopsy specimens from pediatric patients with EoE from a previous randomized anti-IL-5 trial. METHODS: A subanalysis was completed for children treated with 0.55, 2.5, or 10 mg/kg mepolizumab monthly for 12 weeks followed by no treatment until week 24. Quantitative immunochemistry was used to assess the numbers of eosinophils, tryptase-positive mast cells, IL-9(+) cells, and mast cell-eosinophil couplets before and after treatment. RESULTS: Forty-three biopsy specimens had adequate tissue for paired analysis. Forty percent of subjects responded to anti-IL-5 (defined as <15 eosinophils per high-power field [hpf] after mepolizumab therapy), and 77% of all subjects had decreased numbers of mast cells after anti-IL-5. In responders epithelial mast cell numbers decreased from 62 to 19 per hpf (P < .001), were significantly lower than in nonresponders after therapy (P < .05), and correlated with eosinophil numbers (r = 0.75, P < .0001). Mast cells and eosinophils were found in couplets before therapy, and these were significantly decreased only in responders after anti-IL-5 (P < .001). Esophageal eosinophils comprised the majority of cells that made the mast cell growth factor IL-9. IL-9(+) cell numbers decreased from 102 to 71 per hpf (P < .001) after anti-IL-5. CONCLUSIONS: Pediatric patients with EoE had significantly fewer mast cells, IL-9(+) cells, and mast cell-eosinophil couplets in the esophageal epithelium after anti-IL-5 therapy. Because eosinophils were one source of IL-9, they might support esophageal mastocytosis.

Mutations of complement factor I and potential mechanisms of neuroinflammation in acute hemorrhagic leukoencephalitis.

PURPOSE: Acute Hemorrhagic Leukoencephalitis (AHLE) is a rare demyelinating disorder of acute onset, rapid deterioration and significant morbidity and mortality. Most often described as a post-infectious complication of an upper respiratory illness, its precise pathophysiology remains unclear. We describe two pediatric patients with AHLE with partial complement factor I (FI) deficiency whose successful treatment included the interleukin-1 (IL-1) receptor antagonist, anakinra, implicating a role for FI and IL-1 in this disorder. METHODS: Extensive clinical workup of two patients presenting with AHLE revealed complement abnormalities, specifically related to the alternative pathway and its regulator, FI. Aggressive management with steroids, immunoglobulin, and anakinra ultimately led to improvement of clinical status and near return to neurologic baseline in both patients. Genetic sequencing of the FI coding regions of the patients and their families was performed. In vitro protein expression studies and immunohistochemistry of fixed brain tissue was used to investigate pathogenic mechanisms. RESULTS: Two novel mutations in FI were identified in our patients, which result in failure to secrete FI. Immunohistochemical evaluation of brain tissue demonstrated positive staining for C3, membrane attack complex (MAC) and IL-1. CONCLUSIONS: We propose AHLE is an unreported, rare phenotype for partial FI deficiency. The upregulation of C3, MAC and IL-1 with subsequent demyelination support a pathologic role for complement activation in AHLE, and suggest anakinra as an important adjunctive therapy in this disease.

Genomic and genic deletions of the FOX gene cluster on 16q24.1 and inactivating mutations of FOXF1 cause alveolar capillary dysplasia and other malformations.

Alveolar capillary dysplasia with misalignment of pulmonary veins (ACD/MPV) is a rare, neonatally lethal developmental disorder of the lung with defining histologic abnormalities typically associated with multiple congenital anomalies (MCA). Using array CGH analysis, we have identified six overlapping microdeletions encompassing the FOX transcription factor gene cluster in chromosome 16q24.1q24.2 in patients with ACD/MPV and MCA. Subsequently, we have identified four different heterozygous mutations (frameshift, nonsense, and no-stop) in the candidate FOXF1 gene in unrelated patients with sporadic ACD/MPV and MCA. Custom-designed, high-resolution microarray analysis of additional ACD/MPV samples revealed one microdeletion harboring FOXF1 and two distinct microdeletions upstream of FOXF1, implicating a position effect. DNA sequence analysis revealed that in six of nine deletions, both breakpoints occurred in the portions of Alu elements showing eight to 43 base pairs of perfect microhomology, suggesting replication error Microhomology-Mediated Break-Induced Replication (MMBIR)/Fork Stalling and Template Switching (FoSTeS) as a mechanism of their formation. In contrast to the association of point mutations in FOXF1 with bowel malrotation, microdeletions of FOXF1 were associated with hypoplastic left heart syndrome and gastrointestinal atresias, probably due to haploinsufficiency for the neighboring FOXC2 and FOXL1 genes. These differences reveal the phenotypic consequences of gene alterations in cis.

A potential new method to estimate tissue cystine content in nephropathic cystinosis.

OBJECTIVES: To evaluate intestinal mucosal cystine crystal (CC) load as a way to estimate tissue cystine content in children with cystinosis. STUDY DESIGN: Intestinal mucosal biopsies were obtained endoscopically from children (ages 2-18 years) with cystinosis. Using a special processing technique, CC within histiocytes were easily visible and enumerable in the mucosal tissue. Mean CC counts, calculated from stomach and duodenum combined (CC-GD), were correlated with duration of cysteamine treatment, estimated glomerular filtration rate (eGFR), and mean white blood cells (WBC) cystine levels. RESULTS: Seventeen subjects (6 male) were enrolled in 2 studies from 2001 and 2003. The CC-GD count (mean 12.5 ± 1.41 crystals/histiocyte) was lower than the colonic crystal count (mean 23.6 ± 3.38, P = .0031). Nine of 17 subjects underwent repeated endoscopy 2 years later and the trend for CC-GD was to decrease over time (P = .065). Biopsies, however, were never completely depleted of CC. In subjects who were diagnosed before age 18 months, the percent change from baseline of both eGFR and CC-GD were inversely correlated (P = .026). Mean WBC cystine levels were positively correlated with CC-GD (P = .023). CONCLUSIONS: CC are easily visible in the intestinal mucosa. CC-GD counts appear to correlate with eGFR and may help monitor response to treatment. Even when mean WBC cystine levels are low, the mucosal CC are not depleted suggesting that tissue cysteamine levels may not achieve therapeutic efficacy.