ABSTRACTObjectives:The aim of this study was to assess common laboratory tests in identifying severe ulcerative colitis in children at diagnosis.Methods:A cohort of 427 children 4 to 17 years of age newly diagnosed with ulcerative colitis (UC) was prospectively enrolled. Boosted classification trees were used to characterize predictive ability of disease attributes based on clinical disease severity using Pediatric Ulcerative Colitis Activity Index (PUCAI), severe (65+) versus not severe (<65) and total Mayo score, severe (10–12) versus not severe (<10); mucosal disease by Mayo endoscopic subscore, severe (3) versus not severe (<3); and extensive disease versus not extensive (left‐sided and proctosigmoiditis).Results:Mean age was 12.7 years; 49.6% (n = 212) were girls, and 83% (n = 351) were Caucasian. Severe total Mayo score was present in 28% (n = 120), mean PUCAI score was 49.8 ± 20.1, and 33% (n = 142) had severe mucosal disease with extensive involvement in 82% (n = 353). Classification and regression trees identified white blood cell count, erythrocyte sedimentation rate, and platelet count (PLT) as the set of 3 best blood laboratory tests to predict disease extent and severity. For mucosal severity, albumin (Alb) replaced PLT. Classification models for PUCAI and total Mayo provided sensitivity of at least 0.65 using standard clinical cut‐points with misclassification rates of approximately 30%.Conclusions:A combination of the white blood cell count, erythrocyte sedimentation rate, and either PLT or albumin is the best predictive subset of standard laboratory tests to identify severe from nonsevere clinical or mucosal disease at diagnosis in relation to objective clinical scores.

BACKGROUND:Statural growth impairment is more common in males with Crohn's disease (CD). We assessed sex differences in height Z score differences and bone age (BA) Z scores and characterized age of menarche in a novel contemporary cohort of pediatric CD patients undergoing screening for enrollment in the multicenter longitudinal Growth Study.METHODS:Crohn's disease patients (females with chronological age [CA] 5 years and older and younger than 14 years; males with CA 6 years and older and younger than 16 years) participated in a screening visit for the Growth Study. Height BA-Z scores are height Z scores calculated based on BA. Height CA-Z scores are height Z scores calculated based on CA. The height Z score difference equals height CA-Z score minus height BA-Z score.RESULTS:One hundred seventy-one patients (60% male) qualified for this analysis. Mean CA was 12.2 years. Mean height CA-Z score was -0.4, and mean height BA-Z score was 0.4 in females. Mean height CA-Z score was -0.1, and mean height BA-Z score was 0.2 in males. The absolute value of the mean height Z score difference was significantly greater in females (0.8) than males (0.3; P = 0.005). The mean BA-Z score in females (-1.0) was significantly lower than in males (-0.2; P = 0.002). The median CA at menarche was 13.6 (95% CI, 12.6-14.6) years.CONCLUSIONS:Our screening visit data suggest that standardized height gain is lower in males with skeletal maturation and delayed puberty is common in females in CD. We are investigating these findings in the ongoing Growth Study.

Statural growth impairment is both a marker and complication of poorly controlled Crohn’s disease (CD), and occurs more commonly in males than females. The specific molecular mechanisms responsible for this sex difference in growth impairment in CD remain poorly characterized. We are conducting a prospective multicenter longitudinal study (Growth Study) examining sex differences in statural growth impairment in school-age children with CD with growth potential based on bone age (BA) [females (BA 4 yrs, 2 mos to 12 years, 0 months) and males (BA 5 years, 0 months to 14 yrs, 0 mos)]. We have enrolled 118 (65% male) patients. Variable Z scores calculated based on BA are denoted as Variable BA-Z scores; those calculated based on chronological age (CA) are denoted as Variable CA-Z scores. Height Z score difference was calculated as Height CA-Z score minus Height BA-Z score. We analyzed the concentration of selected cytokines in serum in 54 patients (56% male), using a V-Plex kit (Meso Scale Discovery, Rockville, MD). Serum hormone levels were analyzed at Esoterix Endocrinology (Calabasas Hills, CA). We used t-test and Chi-squared test to examine the sex difference for continuous variables and categorical variables respectively. We assessed the association between cytokines and hormones via linear regression. The mean height Z score difference was -0.4 ± 1.0 (SD) [range: -3.9 to 2.0] in males and -1.1 ± 1.1 [-3.9 to 0.9] in females (Figure). The absolute value of the mean height Z score difference was significantly lower in males (P= 0.021). The mean BA Z score was (0.0 ± .04 [-0.7 to 0.9]) in males and (0.0 ± 0.2 [-0.4 to 0.5]) in females (P= 0.952). Race/ethnicity [76% White], Tanner stage [63% pre/early puberty; 37% mid/late puberty], mean disease duration [2.3 ± 1.9 (SD) (range: 0.1 to 8.3) years], medication use [65% infliximab; 39% methotrexate; 13% azathioprine/6-mercaptopurine; 9% adalimumab], and mean disease activity indices (indicated remission) did not differ by sex (P= NS for all). TNF-α, IL-4, IL-6, IL-10, and IL-12 were significantly negatively associated with hormones important in growth in males, while IL-13 was significantly negatively associated with hormones important in growth in females (Table). Statural growth remains compromised in males compared with females in a novel contemporary cohort of children with CD. Our early data suggest that the higher frequency of growth impairment in males appears to be due to less standardized height gain with skeletal maturation, rather than advanced BA progression. BA does not appear to be significantly delayed in this cohort. Furthermore, our preliminary data suggest that systemic inflammation exerts a greater negative impact on hormone levels important in growth in males, and that different molecular pathways lead to growth impairment in males versus females. We are further investigating these preliminary findings in the ongoing prospective multicenter longitudinal Growth Study.

endoscopists differentiated CD from ITB with a NPV of 100% (NPV rages from 100%-100%).DNN-CNN model differentiated CD from ITB with perfect intra-observer agreement ( kappa score of 1).There was a low level of intra-observer and inter -observer agreement in differentiated CD from ITB. Conclusions: The experimental results suggest that DNN-CNN model could achieve better discriminative results and hold promise in discrimination CD from ITB lesions in the region especially where tuberculosis expert resources are limited.Deep learning with the larger data sets of colonoscopic images are needed in future.

Age-of-diagnosis associated variation in disease location and antimicrobial sero-reactivity has suggested fundamental differences in pediatric Crohn Disease (CD) pathogenesis. This variation may be related to pubertal peak incidence of ileal involvement and Peyer’s patches maturation, represented by IFNγ-expressing Th1 cells. However, direct mucosal evidence is lacking. We characterize the global pattern of ileal gene expression and microbial communities in 525 treatment-naive pediatric CD patients and controls (Ctl), stratifying samples by their age-of-diagnosis. We show a robust ileal gene signature notable for higher expression of specific immune genes including GM-CSF and INFγ, and reduced expression of antimicrobial Paneth cell α-defensins, in older compared to younger patients. Reduced α-defensin expression in older patients was associated with higher IFNγ expression. By comparison, the CD-associated ileal dysbiosis, characterized by expansion of Enterobacteriaceae and contraction of Lachnospiraceae and Ruminococcaceae, was already established within the younger group and did not vary systematically with increasing age-of-diagnosis. Multivariate analysis considering individual taxa, however did demonstrate negative associations between Lachnospiraceae and IFNγ, and positive associations between Bacteroides and α-defensin expression. These data provide evidence for maturation of mucosal Th1 immune responses and loss of epithelial antimicrobial α-defensins which are associated with specific taxa with increasing age-of-diagnosis in pediatric CD.

BACKGROUND:Variation in care is common in medical practice. Reducing variation in care is shown to improve quality and increase favorable outcomes in chronic diseases. We sought to identify factors associated with variation in care in children with newly diagnosed Crohn's disease (CD). METHODS:Prospectively collected data from a 28-site multicenter inception CD cohort were analyzed for variations in diagnostic modalities, treatment, and follow-up monitoring practices, along with complicated disease outcomes over 3 years in 1046 children. Generalized linear mixed effects models were used to investigate the intercenter variations in each outcome variable. RESULTS:The mean age at diagnosis was 12 years, and 25.9% were nonwhite. The number of participants ranged from 5 to 112 per site. No variation existed in the initial diagnostic approach. When medication exposure was analyzed, steroid exposure varied from 28.6% to 96.9% (P < 0.01) within 90 days, but variation was not significant over a 3-year period (P = 0.13). Early anti-tumor necrosis factor (anti-TNF) exposure (within 90 days) varied from 2.1% to 65.7% (P < 0.01), but variation was not significant over a 3-year period (P > 0.99). Use of immunomodulators (IMs) varied among centers both within 90 days (P < 0.01) and during 3 years of follow-up (P < 0.01). A significant variation was seen at the geographic level with follow-up small bowel imaging and colonoscopy surveillance after initial therapy. CONCLUSIONS:Intercenter variation in care was seen with the initial use of steroids and anti-TNF, but there was no difference in total 3-year exposure to these drugs. Variation in the initiation and long-term use of IMs was significant among sites, but further research with objective measures is needed to explain this variation of care. Small bowel imaging or repeat colonoscopy in CD patients was not uniformly performed across sites. As our data show the widespread existence of variation in care and disease monitoring at geographic levels among pediatric CD patients, future implementation of various practice strategies may help reduce the variation in care.

in brain and peripheral blood monocytes, respectively.At least one evaluated signal was enriched in each of the 17 tissue types, highlighting the diversity of relevant tissues in IBD.Three regions showed widespread enrichment in active enhancer elements across >25 of 74 assessed tracks, while a fourth PTGER4 risk region displayed ubiquitous enrichment in active promoter elements across all tissues.Conclusion: In silico functional annotation is the first step towards a better understanding of the biological implications of non-coding intergenic and intronic IBD-associated variants.Utilizing funciVar and StateHub, we identified tissuespecific regulatory enrichment of CD, UC and shared IBD associations in immune, gastrointestinal, and brain-derived tissues, respectively, and successfully annotated 23 fine-mapped risk regions for which no previous annotation was available.Functional annotation of a comprehensive collection of fine-mapped IBD associations is underway.

Background Lack of evidence-based outcomes data leads to uncertainty in developing treatment regimens in children who are newly diagnosed with ulcerative colitis. We hypothesised that pretreatment clinical, transcriptomic, and microbial factors predict disease course. Methods In this inception cohort study, we recruited paediatric patients aged 4-17 years with newly diagnosed ulcerative colitis from 29 centres in the USA and Canada. Patients initially received standardised mesalazine or corticosteroids, with pre-established criteria for escalation to immunomodulators (ie, thiopurines) or anti-tumor necrosis factor-alpha (TNF alpha) therapy. We used RNA sequencing to define rectal gene expression before treatment, and 16S sequencing to characterise rectal and faecal microbiota. The primary outcome was week 52 corticosteroid-free remission with no therapy beyond mesalazine. We assessed factors associated with the primary outcome using logistic regression models of the per-protocol population. This study is registered with ClinicalTrials. gov, number NCT01536535. Findings Between July 10, 2012, and April 21, 2015, of 467 patients recruited, 428 started medical therapy, of whom 400 (93%) were evaluable at 52 weeks and 386 (90%) completed the study period with no protocol violations. 150 (38%) of 400 participants achieved week 52 corticosteroid-free remission, of whom 147 (98%) were taking mesalazine and three (2%) were taking no medication. 74 (19%) of 400 were escalated to immunomodulators alone, 123 (31%) anti-TNF alpha therapy, and 25 (6%) colectomy. Low baseline clinical severity, high baseline haemoglobin, and week 4 clinical remission were associated with achieving week 52 corticosteroid-free remission (n=386, logistic model area under the curve [AUC] 0.70, 95% CI 0.65-0.75; specificity 77%, 95% CI 71-82). Baseline severity and remission by week 4 were validated in an independent cohort of 274 paediatric patients with newly diagnosed ulcerative colitis. After adjusting for clinical predictors, an antimicrobial peptide gene signature (odds ratio [OR] 0.57, 95% CI 0.39-0.81; p=0.002) and abundance of Ruminococcaceae (OR 1.43, 1.02-2.00; p=0.04), and Sutterella (OR 0.81, 0.65-1.00; p=0.05) were independently associated with week 52 corticosteroid-free remission. Interpretation Our findings support the utility of initial clinical activity and treatment response by 4 weeks to predict week 52 corticosteroid-free remission with mesalazine alone in children who are newly diagnosed with ulcerative colitis. The development of personalised clinical and biological signatures holds the promise of informing ulcerative colitis therapeutic decisions.

Background: The genetic contributions to pediatric onset ulcerative colitis (UC), characterized by severe disease and extensive colonic involvement, are largely unknown. In adult onset UC, Genome Wide Association Study (GWAS) has identified numerous loci, most of which have a modest susceptibility risk (OR 0.84-1.14), with the exception of the human leukocyte antigen (HLA) region on Chromosome 6 (OR 3.59). Method: To study the genetic contribution to exclusive pediatric onset UC, a GWAS was performed on 466 cases with 2099 healthy controls using UK Biobank array. SNP2HLA was used to impute classical HLA alleles and their corresponding amino acids, and the results are compared with adult onset UC. Results: HLA explained the almost entire association signal, dominated with 191 single nucleotide polymorphisms (SNPs) (p = 5 x 10(-8) to 5 x 10(-10)). Although very small effects, established SNPs in adult onset UC loci had similar direction and magnitude in pediatric onset UC. SNP2HLA imputation identified HLA-DRB1*0103 (odds ratio [OR] = 6.941, p = 1.92*10(-13)) as the most significant association for pediatric UC compared with adult onset UC (OR = 3.59). Further conditioning showed independent effects for HLA-DRB1* 1301 (OR = 2.25, p = 7.92*10(-9)) and another SNP rs17188113 (OR = 0.48, p = 7.56*10(-9)). Two HLA-DRB1 causal alleles are shared with adult onset UC, while at least 2 signals are unique to pediatric UC. Subsequent stratified analyses indicated that HLA-DRB1*0103 has stronger association for extensive disease (E4: OR = 8.28, p = 4.66x10(-10)) and female gender (OR = 8.85, p = 4.82x10(-13)). Conclusion: In pediatric onset UC, the HLA explains almost the entire genetic associations. In addition, the HLA association is approximately twice as strong in pediatric UC compared with adults, due to a combination of novel and shared effects. We speculate the paramount importance of antigenic stimulation either by infectious or noninfectious stimuli as a causal event in pediatric UC onset.

Background:In contrast to pediatric Crohn's disease (CD), little is known in pediatric ulcerative colitis (UC) about the relationship between disease phenotype and serologic reactivity to microbial and other antigens.Aim:The aim of this study was to examine disease phenotype and serology in a well-characterized inception cohort of children newly diagnosed with UC during the PROTECT Study (Predicting Response to Standardized Pediatric Colitis Therapy).Methods:Patients were recruited from 29 participating centers. Demographic, clinical, laboratory, and serologic (pANCA, ASCA IgA/IgG, Anti-CBir1, and Anti-OmpC) data were obtained from children 4-17 years old with UC.Results:Sixty-five percent of the patients had positive serology for pANCA, with 62% less than 12 years old and 66% 12 years old or older. Perinuclear anti-neutrophil cytoplasmic antibodies did not correspond to a specific phenotype though pANCA ≥100, found in 19%, was strongly associated with pancolitis (P = 0.003). Anti-CBir1 was positive in 19% and more common in younger children with 32% less than 12 years old as compared with 14% 12 years old or older (P < 0.001). No association was found in any age group between pANCA and Anti-CBir1. Relative rectal sparing was more common in +CBir1, 16% versus 7% (P = 0.02). Calprotectin was lower in Anti-CBir1+ (Median [IQR] 1495 mcg/g [973-3333] vs 2648 mcg/g [1343-4038]; P = 0.04). Vitamin D 25-OH sufficiency was associated with Anti-CBir1+ (P = 0.0009).Conclusions:The frequency of pANCA in children was consistent with adult observations. High titer pANCA was associated with more extensive disease, supporting the idea that the magnitude of immune reactivity may reflect disease severity. Anti-CBir1+ was more common in younger ages, suggesting host-microbial interactions may differ by patient age.

BIOAVAILABLE SERUM VITAMIN D AND RECTAL VITAMIN D RECEPTOR EXPRESSION AT DIAGNOSIS IN PEDIATRIC ULCERATIVE COLITIS: ASSOCIATIONS WITH DISEASE SEVERITY, CLINICAL OUTCOMES, AND RECTAL PATTERNS OF GENE EXPRESSION Laura E. Bauman, Yael Haberman, Li Hao, Shiven Patel, Vin Tangpricha, Rebekah Karns, Phillip J. Dexheimer, Margaret H. Collins, Michael J. Rosen, Erin Bonkowski, Alison Marquis, Nathan Gotman, Paul A. Rufo, Susan S. Baker, Cary G. Sauer, James Markowitz, Marian D. Pfefferkorn, Joel R. Rosh, Brendan M. Boyle, David R. Mack, Robert N. Baldassano, David J. Keljo, Neal S. Leleiko, Melvin B. Heyman, Anne M. Griffiths, Ashish S. Patel, Joshua D. Noe, Bruce J. Aronow, Subra Kugathasan, Thomas D. Walters, Sonia Davis, Jeffrey S. Hyams, Lee A. Denson

Background: Alterations in the intestinal microbiome are an emerging environmental factor in the pathogenesis of inflammatory bowel disease (IBD).Dysfunction of the IBD candidate gene, protein tyrosine phosphatase non-receptor type 2 ( PTPN2), which encodes the T-cell protein tyrosine phosphatase (TCPTP) protein, contributes to alterations in the intestinal microbiome and the onset of chronic intestinal inflammation in vivo.Expansion of intestinal pathobionts, such as adherent-invasive Escherichia coli (AIEC), can increase susceptibility to colitis in a genetically-susceptible host, and is implicated in IBD.The AIEC binding protein, carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6), is increased in IBD, and promotes AIEC pathogenesis.The aim of this study was to identify if loss of TCPTP promotes pathobiont expansion by altering expression of CEACAM6.Methods: Mouse AIEC (mAIEC) isolated from distal colon of Ptpn2-deficient mice was used to infect HT-29 intestinal epithelial cells (IEC) for 3hrs.Anti-CEACAM6 blocking antibody was used to interrupt mAIEC binding (1hr pretreatment at 1:100).Caco-2 cells stably expressing scrambled shRNA and shRNA against PTPN2 were used for bacterial adherence-invasion (3hr infection), RNAisolation, and protein isolation.RNA-Seq libraries were generated by depleting ribosomal RNA from total RNA and sequenced on the Illumina NextSeq2000 platform.Analysis was performed with systemPipeR.Reads were aligned to the human genome (hg38) using Tophat and differentially expressed genes (DEGs) were identified using EdgeR.DEGs were defined as those with a fold change $ 2 and FDR-adjusted P value # 0.05.CEACAM expression in isolated mouse IEC was determined by RT-PCR and Western blot.Results: In PTPN2-KD Caco-2 cells, mAIEC showed significantly increased invasion (40±0.4% above control cells; n=6, P<0.001), despite reduced adherence (39±14% above control cells; n=6, P=0.009).PTPN2-KD cells showed a 2.8-fold increase ( n=2, P<0.001) in mRNA and 3.3-fold increase (n=4, P<0.001) in protein expression of CEACAM6 vs. control cells.Blocking CEACAM6 reduced adherence of mAIEC (by 47±0.4% of untreated cells; n=2-3, P<0.05) compared with untreated HT-29 IEC.While CEACAM6 is not expressed in mice, KO mouse IEC showed increased CEACAM1 protein in the proximal colon (1.5-fold increase) and distal colon (2.1-fold increase) vs. WT mice, and increased CEACAM20 mRNA in the proximal colon (1.6-fold increase; n=3-4, P<0.05) and distal colon (1.5-fold increase; n=3-4, P<0.05) vs. WT mice.Conclusion: PTPN2 loss increases expression of epithelial binding proteins that elevate host susceptibility to AIEC invasion.This identifies a mechanism by which IBD-associated PTPN2 loss of function mutations contribute to IBD.

OBJECTIVES: Environmental factors play an important role in the pathogenesis of Crohn's Disease (CD). In particular, by virtue of the instability of the microbiome and development of immunologic tolerance, early life factors may exert the strongest influence on disease risk and phenotype. METHODS: We used data from 1119 CD subjects recruited from RISK inception cohort to examine the impact of early life environment on disease progression. Our primary exposures of interest were breastfeeding in infancy and exposure to maternal, active, or passive smoke. Our primary outcomes were development of complicated (stricturing or penetrating) disease, and need for CD-related hospitalization, and surgery. Multivariable logistic regression models were used to define independent associations, adjusting for relevant covariates. RESULTS: Our study cohort included 1119 patients with CD among whom 15% had stricturing (B2) or penetrating disease (B3) by 3 years. 331 patients (35%) and 95 patients (10.6%) required CD-related hospitalizations and surgery respectively. 74.5% were breastfed in infancy and 31% were exposed to smoking among whom 7% were exposed to maternal smoke. On multivariable analysis, a history of breastfeeding was inversely associated with complicated (B2/B3 disease) 0.65, CI 95% 0.44-96; P = 0.03) in pediatric CD. Maternal smoking during pregnancy was associated with increased risk of hospitalization during the 3-year follow-up period (OR 1.75, CI 95% 1.05-2.89; P = 0.03). CONCLUSIONS: Early life environmental factors influence the eventual phenotypes and disease course in CD.

Background Vitamin D regulates intestinal epithelial and immune functions, and vitamin D receptor deficiency increases the severity of murine colitis. Bioavailable 25-hydroxyvitamin D (25(OH)D) is available to target tissues and may be a driver of immune function. The aim is to evaluate the relationship of bioavailable 25(OH)D to the clinical expression of treatment naive pediatric ulcerative colitis (UC). Methods The PROTECT (Predicting Response to Standardized Pediatric Colitis Therapy) study enrolled children ≤17 years newly diagnosed with UC. Free and total 25(OH)D were directly measured and 25(OH)D fractions were compared with disease activity measures. Results Data were available on 388 subjects, mean age 12.7 years, 49% female, 84% with extensive/pancolitis. The median (IQR) total 25(OH)D concentration was 28.5 (23.9, 34.8) ng/mL, and 57% of subjects demonstrated insufficient vitamin D status (25(OH)D < 30 ng/mL). We found no evidence of association between total 25(OH)D and disease activity. Regression models adjusted for age, sex, race, and ethnicity demonstrated that an increase from 25th to 75th percentile for bioavailable and free 25(OH)D were associated with a mean (95th CI) decrease in the Pediatric Ulcerative Colitis Activity Index (PUCAI) of -8.7 (-13.7, -3.6) and -3.1 (-5.0, -1.2), respectively. No associations were detected between 25(OH)D fractions and fecal calprotectin or Mayo endoscopy score. Conclusions Vitamin D insufficiency is highly prevalent in children with newly diagnosed UC. We found associations of free and bioavailable, but not total 25(OH)D, with PUCAI. Bioavailable vitamin D may contribute to UC pathophysiology and clinical activity.