Single-Cell Analysis of Crohn's Disease Lesions Identifies a Pathogenic Cellular Module Associated with Resistance to Anti-TNF Therapy.

Clinical benefits of cytokine blockade in ileal Crohn's disease (iCD) are limited to a subset of patients. Here, we applied single-cell technologies to iCD lesions to address whether cellular heterogeneity contributes to treatment resistance. We found that a subset of patients expressed a unique cellular module in inflamed tissues that consisted of IgG plasma cells, inflammatory mononuclear phagocytes, activated T cells, and stromal cells, which we named the GIMATS module. Analysis of ligand-receptor interaction pairs identified a distinct network connectivity that likely drives the GIMATS module. Strikingly, the GIMATS module was also present in a subset of patients in four independent iCD cohorts (n = 441), and its presence at diagnosis correlated with failure to achieve durable corticosteroid-free remission upon anti-TNF therapy. These results emphasize the limitations of current diagnostic assays and the potential for single-cell mapping tools to identify novel biomarkers of treatment response and tailored therapeutic opportunities.

Biopsy and blood-based molecular biomarker of inflammation in IBD.

OBJECTIVE: IBD therapies and treatments are evolving to deeper levels of remission. Molecular measures of disease may augment current endpoints including the potential for less invasive assessments. DESIGN: Transcriptome analysis on 712 endoscopically defined inflamed (Inf) and 1778 non-inflamed (Non-Inf) intestinal biopsies (n=498 Crohn's disease, n=421 UC and 243 controls) in the Mount Sinai Crohn's and Colitis Registry were used to identify genes differentially expressed between Inf and Non-Inf biopsies and to generate a molecular inflammation score (bMIS) via gene set variance analysis. A circulating MIS (cirMIS) score, reflecting intestinal molecular inflammation, was generated using blood transcriptome data. bMIS/cirMIS was validated as indicators of intestinal inflammation in four independent IBD cohorts. RESULTS: bMIS/cirMIS was strongly associated with clinical, endoscopic and histological disease activity indices. Patients with the same histologic score of inflammation had variable bMIS scores, indicating that bMIS describes a deeper range of inflammation. In available clinical trial data sets, both scores were responsive to IBD treatment. Despite similar baseline endoscopic and histologic activity, UC patients with lower baseline bMIS levels were more likely treatment responders compared with those with higher levels. Finally, among patients with UC in endoscopic and histologic remission, those with lower bMIS levels were less likely to have a disease flare over time. CONCLUSION: Transcriptionally based scores provide an alternative objective and deeper quantification of intestinal inflammation, which could augment current clinical assessments used for disease monitoring and have potential for predicting therapeutic response and patients at higher risk of disease flares.

Integrative Analysis of the Inflammatory Bowel Disease Serum Metabolome Improves Our Understanding of Genetic Etiology and Points to Novel Putative Therapeutic Targets.

BACKGROUND & AIMS: Polygenic and environmental factors are underlying causes of inflammatory bowel disease (IBD). We hypothesized that integration of the genetic loci controlling a metabolite's abundance, with known IBD genetic susceptibility loci, may help resolve metabolic drivers of IBD. METHODS: We measured the levels of 1300 metabolites in the serum of 484 patients with ulcerative colitis (UC) and 464 patients with Crohn's disease (CD) and 365 controls. Differential metabolite abundance was determined for disease status, subtype, clinical and endoscopic disease activity, as well as IBD phenotype including disease behavior, location, and extent. To inform on the genetic basis underlying metabolic diversity, we integrated metabolite and genomic data. Genetic colocalization and Mendelian randomization analyses were performed using known IBD risk loci to explore whether any metabolite was causally associated with IBD. RESULTS: We found 173 genetically controlled metabolites (metabolite quantitative trait loci, 9 novel) within 63 non-overlapping loci (7 novel). Furthermore, several metabolites significantly associated with IBD disease status and activity as defined using clinical and endoscopic indexes. This constitutes a resource for biomarker discovery and IBD biology insights. Using this resource, we show that a novel metabolite quantitative trait locus for serum butyrate levels containing ACADS was not supported as causal for IBD; replicate the association of serum omega-6 containing lipids with the fatty acid desaturase 1/2 locus and identify these metabolites as causal for CD through Mendelian randomization; and validate a novel association of serum plasmalogen and TMEM229B, which was predicted as causal for CD. CONCLUSIONS: An exploratory analysis combining genetics and unbiased serum metabolome surveys can reveal novel biomarkers of disease activity and potential mediators of pathology in IBD.

Molecular Characterization of Limited Ulcerative Colitis Reveals Novel Biology and Predictors of Disease Extension.

BACKGROUND AND AIMS: Disease extent varies in ulcerative colitis (UC) from proctitis to left-sided colitis to pancolitis and is a major prognostic factor. When the extent of UC is limited there is often a sharp demarcation between macroscopically involved and uninvolved areas and what defines this or subsequent extension is unknown. We characterized the demarcation site molecularly and determined genes associated with subsequent disease extension. METHODS: We performed RNA sequence analysis of biopsy specimens from UC patients with endoscopically and histologically confirmed limited disease, of which a subset later extended. Biopsy specimens were obtained from the endoscopically inflamed upper (proximal) limit of disease, immediately adjacent to the uninvolved colon, as well as at more proximal, endoscopically uninflamed colonic segments. RESULTS: Differentially expressed genes were identified in the endoscopically inflamed biopsy specimens taken at each patient's most proximal diseased site relative to healthy controls. Expression of these genes in the more proximal biopsy specimens transitioned back to control levels abruptly or gradually, the latter pattern supporting the concept that disease exists beyond the endoscopic disease demarcation site. The gradually transitioning genes were associated with inflammation, angiogenesis, glucuronidation, and homeodomain pathways. A subset of these genes in inflamed biopsy specimens was found to predict disease extension better than clinical features and were responsive to biologic therapies. Network analysis revealed critical roles for interferon signaling in UC inflammation and poly(ADP-ribose) polymerase 14 (PARP14) was a predicted key driver gene of extension. Higher PARP14 protein levels were found in inflamed biopsy specimens of patients with limited UC that subsequently extended. CONCLUSION: Molecular predictors of disease extension reveal novel strategies for disease prognostication and potential therapeutic targeting.

Intestinal Inflammation Modulates the Expression of ACE2 and TMPRSS2 and Potentially Overlaps With the Pathogenesis of SARS-CoV-2-related Disease.

BACKGROUND AND AIMS: The presence of gastrointestinal symptoms and high levels of viral RNA in the stool suggest active severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication within enterocytes. METHODS: Here, in multiple, large cohorts of patients with inflammatory bowel disease (IBD), we have studied the intersections between Coronavirus Disease 2019 (COVID-19), intestinal inflammation, and IBD treatment. RESULTS: A striking expression of ACE2 on the small bowel enterocyte brush border supports intestinal infectivity by SARS-CoV-2. Commonly used IBD medications, both biologic and nonbiologic, do not significantly impact ACE2 and TMPRSS2 receptor expression in the uninflamed intestines. In addition, we have defined molecular responses to COVID-19 infection that are also enriched in IBD, pointing to shared molecular networks between COVID-19 and IBD. CONCLUSIONS: These data generate a novel appreciation of the confluence of COVID-19- and IBD-associated inflammation and provide mechanistic insights supporting further investigation of specific IBD drugs in the treatment of COVID-19. Preprint doi: https://doi.org/10.1101/2020.05.21.109124.

Relationship Between Combined Histologic and Endoscopic Endpoints and Efficacy of Ustekinumab Treatment in Patients With Ulcerative Colitis.

BACKGROUND & AIMS: Ustekinumab induces and maintains histologic improvement in patients with ulcerative colitis (UC). The clinical relevance of this endpoint alone, and in combination with endoscopic improvement, is unknown. METHODS: Histologic disease activity was evaluated in 2630 colonic biopsy samples from patients with UC treated in the UNIFI phase 3 UC clinical studies of ustekinumab. We evaluated associations between histologic improvement (defined as the composite of neutrophil infiltration in less than 5% of crypts and no crypt destruction, erosions, ulcerations, or granulation tissue) and clinical endpoints at the end of induction (week 8 and 16) and maintenance (week 44) periods. We assessed the validity of a combined histologic and endoscopic (Mayo endoscopy subscore, 0 or 1) improvement endpoint, which we called histo-endoscopic mucosal healing (or histo-endoscopic mucosal improvement). RESULTS: Histologic improvement was significantly (P < .0001) associated with clinical remission, lower mean disease activity scores, and greater improvement in disease activity at the end of induction and maintenance studies. Ustekinumab induced and maintained significantly higher rates of histologic improvement at induction week 8 and maintenance week 44 than placebo when more stringent definitions of histologic improvement were used. Histologic improvement and endoscopic improvement following induction were associated with 10% to 20% higher rates of histo-endoscopic mucosal healing, clinical remission, and corticosteroid-free remission at week 44 (all P < .05) in patients who received ustekinumab maintenance therapy. At week 44, 61% of patients (56/92) with histo-endoscopic mucosal healing after induction therapy achieved clinical remission, versus 39% of patients (9/23, P = .0983) and 34% of patients (24/71, P = .0009) with endoscopic or histologic improvement alone after induction, respectively. CONCLUSION: Data from the UNIFI program of ustekinumab in patients with UC treated with ustekinumab indicated the achievement of histo-endoscopic mucosal healing after induction therapy is associated with lower disease activity at the end of maintenance therapy than either histologic or endoscopic improvement alone. ClinicalTrials.gov number: NCT02407236.

Effects of Ustekinumab on Histologic Disease Activity in Patients With Crohn's Disease.

BACKGROUND & AIMS: Although ustekinumab is an effective therapy for moderate to severe Crohn's disease (CD), its effects on the microscopic manifestations of CD are unknown. METHODS: We evaluated the effects of ustekinumab on histologic CD activity in an analysis of data from 251 participants in phase 3 induction and maintenance studies. Two endoscopic biopsy samples were collected at weeks 0, 8, and 44 from the ileum, splenic flexure, and rectum (18 biopsy samples from each patient). Histologic activity was assessed based on global histology activity scores (GHASs). RESULTS: At week 8, the mean GHAS was significantly reduced after ustekinumab induction treatment (from 10.4 ± 7.0 to 7.1 ± 5.9; P < .001) but not in patients who received placebo (from 9.2 ± 6.4 to 7.8 ± 6.2). At week 44 in the randomized maintenance therapy population, the mean GHAS remained reduced from week 8 in patients who received subcutaneous ustekinumab (90 mg every 8 weeks; from 7.4 ± 7.7 to 6.1 ± 4.7) but not every 12 weeks (from 5.3 ± 3.9 to 8.7 ± 4.1) or placebo (from 9.2 ± 3.8 to 10.9 ± 7.1). In the pooled (randomized and nonrandomized) maintenance therapy population, histologic improvement continued in patients given ustekinumab every 8 weeks (from 7.1 ± 6.2 to 5.2 ± 4.2; P < .0001) but not in those given ustekinumab every 12 weeks (from 6.1 ± 5.7 to 7.2 ± 5.1) or placebo (from 8.2 ± 4.2 to 8.9 ± 6.8). A significantly greater proportion of patients achieved histologic response (≥50% decrease in GHAS from baseline) at week 44 if they received ustekinumab every 8 weeks (50% in the randomized maintenance population and 54% in the pooled maintenance population) compared with every 12 weeks (17% and 39% in the randomized and pooled populations, respectively) or placebo (0% and 22% in the randomized and pooled populations, respectively) (P = .0137 for every 8 weeks vs placebo and P = .3529 for every 12 weeks vs placebo in the randomized population; P = .0168 for every 8 weeks vs placebo and P = .3069 for every 12 weeks vs placebo in the pooled population). Regional and overall mean GHASs correlated with the simple endoscopic score for CD (r = .6255, P < .0001). Multivariate analysis found an association between histologic improvement and endoscopic or histologic burden at baseline. CONCLUSIONS: In an analysis of data from participants in phase 3 induction and maintenance trials, we found histologic improvement in a greater proportion of patients given ustekinumab vs placebo. The largest improvements occurred in patients who received ustekinumab maintenance therapy every 8 weeks. ClinicalTrials.gov nos. NCT01369329, NCT01369342, and NCT01369355.

Ustekinumab as Induction and Maintenance Therapy for Crohn's Disease.

BACKGROUND: Ustekinumab, a monoclonal antibody to the p40 subunit of interleukin-12 and interleukin-23, was evaluated as an intravenous induction therapy in two populations with moderately to severely active Crohn's disease. Ustekinumab was also evaluated as subcutaneous maintenance therapy. METHODS: We randomly assigned patients to receive a single intravenous dose of ustekinumab (either 130 mg or approximately 6 mg per kilogram of body weight) or placebo in two induction trials. The UNITI-1 trial included 741 patients who met the criteria for primary or secondary nonresponse to tumor necrosis factor (TNF) antagonists or had unacceptable side effects. The UNITI-2 trial included 628 patients in whom conventional therapy failed or unacceptable side effects occurred. Patients who completed these induction trials then participated in IM-UNITI, in which the 397 patients who had a response to ustekinumab were randomly assigned to receive subcutaneous maintenance injections of 90 mg of ustekinumab (either every 8 weeks or every 12 weeks) or placebo. The primary end point for the induction trials was a clinical response at week 6 (defined as a decrease from baseline in the Crohn's Disease Activity Index [CDAI] score of ≥100 points or a CDAI score <150). The primary end point for the maintenance trial was remission at week 44 (CDAI score <150). RESULTS: The rates of response at week 6 among patients receiving intravenous ustekinumab at a dose of either 130 mg or approximately 6 mg per kilogram were significantly higher than the rates among patients receiving placebo (in UNITI-1, 34.3%, 33.7%, and 21.5%, respectively, with P≤0.003 for both comparisons with placebo; in UNITI-2, 51.7%, 55.5%, and 28.7%, respectively, with P<0.001 for both doses). In the groups receiving maintenance doses of ustekinumab every 8 weeks or every 12 weeks, 53.1% and 48.8%, respectively, were in remission at week 44, as compared with 35.9% of those receiving placebo (P=0.005 and P=0.04, respectively). Within each trial, adverse-event rates were similar among treatment groups. CONCLUSIONS: Among patients with moderately to severely active Crohn's disease, those receiving intravenous ustekinumab had a significantly higher rate of response than did those receiving placebo. Subcutaneous ustekinumab maintained remission in patients who had a clinical response to induction therapy. (Funded by Janssen Research and Development; ClinicalTrials.gov numbers, NCT01369329 , NCT01369342 , and NCT01369355 .).

Abnormal Small Intestinal Epithelial Microvilli in Patients With Crohn's Disease.

BACKGROUND & AIMS: Crohn disease (CD) presents as chronic and often progressive intestinal inflammation, but the contributing pathogenic mechanisms are unclear. We aimed to identify alterations in intestinal cells that could contribute to the chronic and progressive course of CD. METHODS: We took an unbiased system-wide approach by performing sequence analysis of RNA extracted from formalin-fixed paraffin-embedded ileal tissue sections from patients with CD (n = 36) and without CD (controls; n = 32). We selected relatively uninflamed samples, based on histology, before gene expression profiling; validation studies were performed using adjacent serial tissue sections. A separate set of samples (3 control and 4 CD samples) was analyzed by transmission electron microscopy. We developed methods to visualize an overlapping modular network of genes dysregulated in the CD samples. We validated our findings using biopsy samples (110 CD samples for gene expression analysis and 54 for histologic analysis) from the UNITI-2 phase 3 trial of ustekinumab for patients with CD and healthy individuals (26 samples used in gene expression analysis). RESULTS: We identified gene clusters that were altered in nearly all CD samples. One cluster encoded genes associated with the enterocyte brush border, leading us to investigate microvilli. In ileal tissues from patients with CD, the microvilli were of decreased length and had ultrastructural defects compared with tissues from controls. Microvilli length correlated with expression of genes that regulate microvilli structure and function. Network analysis linked the microvilli cluster to several other down-regulated clusters associated with altered intracellular trafficking and cellular metabolism. Enrichment of a core microvilli gene set also was lower in the UNITI-2 trial CD samples compared with controls; expression of microvilli genes was correlated with microvilli length and endoscopy score and was associated with response to treatment. CONCLUSIONS: In a transcriptome analysis of formalin-fixed and paraffin-embedded ileal tissues from patients with CD and controls, we associated transcriptional alterations with histologic alterations, such as differences in microvilli length. Decreased microvilli length and decreased expression of the microvilli gene set might contribute to epithelial malfunction and the chronic and progressive disease course in patients with CD.

Molecular Comparison of Adult and Pediatric Ulcerative Colitis Indicates Broad Similarity of Molecular Pathways in Disease Tissue.

BACKGROUND: Efficacy data from adult ulcerative colitis (UC) clinical trials are often extrapolated for pediatric prescribing. Consequently, it is important to understand similarities/differences in pediatric and adult UC. Pediatric UC tends to have more extensive disease at presentation, yet genetic studies have not detected pathways that distinguish the populations, and differences in mucosal gene expression between adult and pediatric UC are not well characterized. METHODS: Using colonic microarray data from a phase 3 trial of golimumab in adult UC (87 UC; 21 healthy), the GSE10616 pediatric dataset (10 UC; 11 healthy), and a phase 1B trial of golimumab in pediatric UC (n = 19), UC expression profiles were compared and unique genes were defined as those with significant changes (|FC|>2×, adjusted P < 0.05) in one population, but not the other (|FC| < 1.2×, adjusted P > 0.05). Pathway and upstream regulator analyses were performed. Profiles by disease extent (extensive [pancolitis] vs limited [left-sided] involvement) were compared within each population. RESULTS: Pediatric and adult disease profiles overlapped substantially, with ∼50% to 75% overlap, depending on the fold-change cutoff used. Conversely, <10% of the disease profiles were unique to each population. Similar canonical pathways were enriched in both datasets. Predicted upstream regulators were also concordant, including lipopolysaccharide, interleukin-1ß, and tumor necrosis factor-α. Expression profiles of extensive UC were indistinguishable from those of patients with limited involvement in each population. CONCLUSIONS: The UC gene expression landscape is shared by adults and children, independent of disease extent. This supports extrapolation of efficacy from adults to children in developing new therapies for UC.

MicroRNA-122 regulates polyploidization in the murine liver.

UNLABELLED: A defining feature of the mammalian liver is polyploidy, a numerical change in the entire complement of chromosomes. The first step of polyploidization involves cell division with failed cytokinesis. Although polyploidy is common, affecting ∼90% of hepatocytes in mice and 50% in humans, the specialized role played by polyploid cells in liver homeostasis and disease remains poorly understood. The goal of this study was to identify novel signals that regulate polyploidization, and we focused on microRNAs (miRNAs). First, to test whether miRNAs could regulate hepatic polyploidy, we examined livers from Dicer1 liver-specific knockout mice, which are devoid of mature miRNAs. Loss of miRNAs resulted in a 3-fold reduction in binucleate hepatocytes, indicating that miRNAs regulate polyploidization. Second, we surveyed age-dependent expression of miRNAs in wild-type mice and identified a subset of miRNAs, including miR-122, that is differentially expressed at 2-3 weeks, a period when extensive polyploidization occurs. Next, we examined Mir122 knockout mice and observed profound, lifelong depletion of polyploid hepatocytes, proving that miR-122 is required for complete hepatic polyploidization. Moreover, the polyploidy defect in Mir122 knockout mice was ameliorated by adenovirus-mediated overexpression of miR-122, underscoring the critical role miR-122 plays in polyploidization. Finally, we identified direct targets of miR-122 (Cux1, Rhoa, Iqgap1, Mapre1, Nedd4l, and Slc25a34) that regulate cytokinesis. Inhibition of each target induced cytokinesis failure and promoted hepatic binucleation. CONCLUSION: Among the different signals that have been associated with hepatic polyploidy, miR-122 is the first liver-specific signal identified; our data demonstrate that miR-122 is both necessary and sufficient in liver polyploidization, and these studies will serve as the foundation for future work investigating miR-122 in liver maturation, homeostasis, and disease. (Hepatology 2016;64:599-615).

Pediatric eosinophilic esophagitis is associated with changes in esophageal microRNAs.

The incidence of eosinophilic esophagitis (EoE) has increased in the past several years, yet our understanding of its pathogenesis remains limited. To test the hypothesis that microRNAs (miRNAs) are altered in children with EoE, miRNAs were profiled in esophageal mucosa biopsies obtained from patients with active disease (n = 5) and healthy control subjects (n = 6). Fourteen miRNAs were significantly altered between groups; four of these miRNAs were decreased in EoE patients. A panel of five miRNAs (miR-203, miR-375, miR-21, miR-223, and miR-142-3p) were selected for validation in an independent set of samples from control (n = 22), active disease (n = 22), inactive disease (n = 22), and gastroesophageal reflux disease (n = 6) patients. Each panel miRNA was significantly altered among groups. miRNA changes in esophageal biopsies were not reflected in the circulating RNA pool, as no differences in panel miRNA levels were observed in sera collected from the four patient groups. In addition, in contrast to previous studies, no change in esophageal miRNA levels was detected following treatment that resolved esophageal eosinophilia. In an effort to identify the ramifications of reduced esophageal miR-203, miR-203 activity was inhibited in cultured epithelial cells via expression of a tough decoy miRNA inhibitor. Luciferase reporter assays demonstrated that miR-203 does not directly regulate human IL-15 through targeting of the IL-15 3'-untranslated region. From these experiments, it is concluded that miRNAs are perturbed in the esophageal mucosa, but not the serum, of pediatric EoE patients. Further investigation is required to decipher pathologically relevant consequences of miRNA perturbation in this context.

Dynamic recruitment of microRNAs to their mRNA targets in the regenerating liver.

BACKGROUND: Validation of physiologic miRNA targets has been met with significant challenges. We employed HITS-CLIP to identify which miRNAs participate in liver regeneration, and to identify their target mRNAs. RESULTS: miRNA recruitment to the RISC is highly dynamic, changing more than five-fold for several miRNAs. miRNA recruitment to the RISC did not correlate with changes in overall miRNA expression for these dynamically recruited miRNAs, emphasizing the necessity to determine miRNA recruitment to the RISC in order to fully assess the impact of miRNA regulation. We incorporated RNA-seq quantification of total mRNA to identify expression-weighted Ago footprints, and developed a microRNA regulatory element (MRE) prediction algorithm that represents a greater than 20-fold refinement over computational methods alone. These high confidence MREs were used to generate candidate 'competing endogenous RNA' (ceRNA) networks. CONCLUSION: HITS-CLIP analysis provide novel insights into global miRNA:mRNA relationships in the regenerating liver.

Extrahepatic cholangiocyte cilia are abnormal in biliary atresia.

OBJECTIVES: Biliary atresia (BA) is a rapidly progressive form of biliary fibrosis affecting neonates. We previously reported that primary cilia on the intrahepatic cholangiocytes of patients with both syndromic and nonsyndromic BA were structurally abnormal. Our objective was to determine whether extrahepatic cholangiocytes in human biliary atresia, intrahepatic and extrahepatic cholangiocytes of rhesus rotavirus (RRV)-infected neonatal mice, and RRV-infected primary neonatal extrahepatic cholangiocytes also demonstrate ciliary abnormalities. METHODS: The livers of neonatal BALB/c mice injected with RRV that developed jaundice, human extrahepatic bile duct samples obtained at time of hepatoportoenterostomy, and RRV-infected primary neonatal cholangiocytes were stained with antibodies against acetylated α tubulin to identify primary cilia. RESULTS: Extrahepatic cholangiocytes from RRV-treated mice demonstrated minimal loss of primary cilia at day 3 but almost complete loss at day 8 and partial loss at day 12. No changes were seen in mouse intrahepatic bile ducts at any of the time points. In the human BA samples, primary cilia were almost completely absent from extrahepatic duct cholangiocytes. There were, however, abundant cilia in the peribiliary glands adjacent to extrahepatic ducts in the BA sample. Cilia in RRV-infected primary neonatal cholangiocytes were significantly decreased compared with controls. CONCLUSIONS: Primary cilia are selectively lost from neonatal extrahepatic but not intrahepatic cholangiocytes after RRV infection in BALB/c mice. The cilia are also decreased in RRV-infected primary cholangiocytes and the extrahepatic ducts from human patients with BA. This suggests that ciliary abnormalities are part of the pathophysiology of BA.

Deep learning models capture histological disease activity in Crohn's Disease and Ulcerative Colitis with high fidelity.

BACKGROUND AND AIMS: Histologic disease activity in Inflammatory Bowel Disease (IBD) is associated with clinical outcomes and is an important endpoint in drug development. We developed deep learning models for automating histological assessments in IBD. METHODS: Histology images of intestinal mucosa from phase 2 and phase 3 clinical trials in Crohn's disease (CD) and Ulcerative Colitis (UC) were used to train artificial intelligence (AI) models to predict the Global Histology Activity Score (GHAS) for CD and Geboes histopathology score for UC. Three AI methods were compared. AI models were evaluated on held-back testing sets and model predictions were compared against an expert central reader and five independent pathologists. RESULTS: The model based on multiple instance learning and the attention mechanism (SA-AbMILP) demonstrated the best performance among competing models. AI modeled GHAS and Geboes sub-grades matched central readings with moderate to substantial agreement, with accuracies ranging from 65% to 89%. Furthermore, the model was able to distinguish the presence and absence of pathology across four selected histological features with accuracies for colon, in both CD and UC, ranging from 87% to 94% and, for CD ileum, ranging from 76% to 83%. For both CD and UC, and across anatomical compartments (ileum and colon) in CD, comparable accuracies against central readings were found between the model assigned scores and scores by an independent set of pathologists. CONCLUSIONS: Deep learning models based upon GHAS and Geboes scoring systems were effective at distinguishing between the presence and absence of IBD microscopic disease activity.

A temporal classifier predicts histopathology state and parses acute-chronic phasing in inflammatory bowel disease patients.

Previous studies have conducted time course characterization of murine colitis models through transcriptional profiling of differential expression. We characterize the transcriptional landscape of acute and chronic models of dextran sodium sulfate (DSS) and adoptive transfer (AT) colitis to derive temporal gene expression and splicing signatures in blood and colonic tissue in order to capture dynamics of colitis remission and relapse. We identify sub networks of patient-derived causal networks that are enriched in these temporal signatures to distinguish acute and chronic disease components within the broader molecular landscape of IBD. The interaction between the DSS phenotype and chronological time-point naturally defines parsimonious temporal gene expression and splicing signatures associated with acute and chronic phases disease (as opposed to ordinary time-specific differential expression/splicing). We show these expression and splicing signatures are largely orthogonal, i.e. affect different genetic bodies, and that using machine learning, signatures are predictive of histopathological measures from both blood and intestinal data in murine colitis models as well as an independent cohort of IBD patients. Through access to longitudinal multi-scale profiling from disease tissue in IBD patient cohorts, we can apply this machine learning pipeline to generation of direct patient temporal multimodal regulatory signatures for prediction of histopathological outcomes.

Circulating microRNA is a biomarker of biliary atresia.

OBJECTIVE: The lack of reliable noninvasive diagnostic biomarkers of biliary atresia (BA) results in delayed diagnosis and worsened patient outcome. Circulating microRNAs (miRNAs) are a new class of noninvasive biomarkers with encouraging diagnostic utility. METHODS: We examined the ability of serum miRNAs to distinguish BA from other forms of neonatal hyperbilirubinemia. BA-specific serum miRNAs were identified using a microfluidic array platform and validated in a larger, independent sample set. RESULTS: The miR-200b/429 cluster was significantly increased in the sera of patients with BA relative to infants with non-BA cholestatic disorders. CONCLUSIONS: Circulating levels of the miR-200b/429 cluster are elevated in infants with BA and have promising diagnostic clinical performance.

MicroRNA profiling identifies miR-29 as a regulator of disease-associated pathways in experimental biliary atresia.

Biliary atresia (BA) is a pediatric liver disease of unknown underlying etiology, in which fibroinflammatory destruction of the extrahepatic biliary system leads to obstructive cholestasis. MicroRNAs are a class of short (18-23 nucleotide), noncoding RNA molecules, which act as negative regulators of target mRNA stability and translation. The importance of these molecules in normal and diseased liver has been demonstrated, but their potential role in the pathogenesis of BA has not been addressed. We have profiled changes in liver microRNA levels in an established mouse model of the disease, identified significantly altered transcripts, and defined the spatial expression patterns of selected microRNAs. Two of these, miR-29a/29b1, are upregulated in experimental BA. Using antisense oligonucleotide-mediated inhibition in mice, we have delineated the full set of hepatic genes regulated by miR-29 and identified 2 mRNA targets of potential pathological relevance in experimental BA, Igf1 and Il1RAP. We have used reporter assays to confirm that Igf1 and Il1RAP are direct targets of miR-29.

A role for microRNA in cystic liver and kidney diseases.

The polycystic liver and kidney diseases are a family of disorders with heterogeneous etiologies. Proposed mechanisms of disease include ciliary dysfunction, excess cell proliferation, and altered cell-cell or cell-matrix interactions. In this issue of the JCI, Lee and colleagues provide data to support a novel mechanism for cystogenesis involving microRNA (miRNA) (see the related article beginning on page 3714). They demonstrate that levels of the miRNA miR15a are decreased in livers of patients with autosomal recessive and autosomal dominant polycystic kidney disease (ARPKD and ADPKD, respectively) and congenital hepatic fibrosis as well as in the PKC rat model of ARPKD. This results in increased expression of the cell-cycle regulator Cdc25A, which is a direct target of miR15a, and increased cellular proliferation and cystogenesis in vitro. These findings suggest that other miRNAs may also participate in the molecular pathogenesis of cystic liver and kidney diseases.

MicroRNAs control intestinal epithelial differentiation, architecture, and barrier function.

BACKGROUND & AIMS: Whereas the importance of microRNA (miRNA) for the development of several tissues is well established, its role in the intestine is unknown. We aimed to quantify the complete miRNA expression profile of the mammalian intestinal mucosa and to determine the contribution of miRNAs to intestinal homeostasis using genetic means. METHODS: We determined the miRNA transcriptome of the mouse intestinal mucosa using ultrahigh throughput sequencing. Using high-throughput sequencing of RNA isolated by cross-linking immunoprecipitation (HITS-CLIP), we identified miRNA-messenger RNA target relationships in the jejunum. We employed gene ablation of the obligatory miRNA-processing enzyme Dicer1 to derive mice deficient for all miRNAs in intestinal epithelia. RESULTS: miRNA abundance varies dramatically in the intestinal mucosa, from 1 read per million to 250,000. Of the 453 miRNA families identified, mmu-miR-192 is the most highly expressed in both the small and large intestinal mucosa, and there is a 53% overlap in the top 15 expressed miRNAs between the 2 tissues. The intestinal epithelium of Dicer1(loxP/loxP);Villin-Cre mutant mice is disorganized, with a decrease in goblet cells, a dramatic increase in apoptosis in crypts of both jejunum and colon, and accelerated jejunal cell migration. Furthermore, intestinal barrier function is impaired in Dicer1-deficient mice, resulting in intestinal inflammation with lymphocyte and neutrophil infiltration. Our list of miRNA-messenger RNA targeting relationships in the small intestinal mucosa provides insight into the molecular mechanisms behind the phenotype of Dicer1 mutant mice. CONCLUSIONS: We have identified all intestinal miRNAs and shown using gene ablation of Dicer1 that miRNAs play a vital role in the differentiation and function of the intestinal epithelium.