title: THBS2Is a Candidate Modifier of Liver Disease Severity in Alagille Syndrome.
abstract: OBJECTIVE
  Alagille syndrome is an autosomal-dominant, multisystem disorder caused primarily by mutations inJAG1, resulting in bile duct paucity, cholestasis, cardiac disease, and other features. Liver disease severity in Alagille syndrome is highly variable, however, factors influencing the hepatic phenotype are unknown. We hypothesized that genetic modifiers may contribute to the variable expressivity of this disorder.
  
  
  METHODS
  We performed a genome-wide association study in a cohort of Caucasian subjects with known pathogenicJAG1mutations, comparing patients with mild vs severe liver disease, followed by functional characterization of a candidate locus.
  
  
  RESULTS
  We identified a locus that reached suggestive genome-level significance upstream of the thrombospondin 2 (THBS2) gene.THBS2codes for a secreted matricellular protein that regulates cell proliferation, apoptosis, and angiogenesis, and has been shown to affect Notch signaling. By using a reporter mouse line, we detected thrombospondin 2 expression in bile ducts and periportal regions of the mouse liver. Examination ofThbs2-null mouse livers showed increased microvessels in the portal regions of adult mice. We also showed that thrombospondin 2 interacts with NOTCH1 and NOTCH2 and can inhibit JAG1-NOTCH2 interactions.
  
  
  CONCLUSIONS
  Based on the genome-wide association study results, thrombospondin 2 localization within bile ducts, and demonstration of interactions of thrombospondin 2 with JAG1 and NOTCH2, we propose that changes in thrombospondin 2 expression may further perturb JAG1-NOTCH2 signaling in patients harboring aJAG1mutation and lead to a more severe liver phenotype. These results implicateTHBS2as a plausible candidate genetic modifier of liver disease severity in Alagille syndrome.
authors.affiliation: Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania; Genomics and Computational Biology Graduate Group, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Ellen A
authors.initials: EA
authors.lastname: Tsai
authors.affiliation: Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Melissa A
authors.initials: MA
authors.lastname: Gilbert
authors.affiliation: Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Grochowski
authors.affiliation: Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Lara A
authors.initials: LA
authors.lastname: Underkoffler
authors.affiliation: Department of Neurology, University of Michigan, Ann Arbor, Michigan.
authors.forename: He
authors.initials: H
authors.lastname: Meng
authors.affiliation: Department of Neurology, University of Michigan, Ann Arbor, Michigan.
authors.forename: Xiaojie
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Neurology, University of Michigan, Ann Arbor, Michigan; Department of Physiology, University of Michigan, Ann Arbor, Michigan; VA Ann Arbor Healthcare System, Ann Arbor, Michigan.
authors.forename: Michael M
authors.initials: MM
authors.lastname: Wang
authors.affiliation: Medical Scientist Training Program, University of Michigan, Ann Arbor, Michigan.
authors.forename: Hailu
authors.initials: H
authors.lastname: Shitaye
authors.affiliation: Department of Physiology, Department of Small Animal Clinical Sciences, Colleges of Natural Science, Osteopathic Medicine, and Veterinary Medicine, Michigan State University, East Lansing, Michigan; Department of Orthopaedic Surgery, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Kurt D
authors.initials: KD
authors.lastname: Hankenson
authors.affiliation: Division of Pediatric Gastroenterology, Hepatology, and Nutrition, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: David
authors.initials: D
authors.lastname: Piccoli
authors.affiliation: Division of Pediatric Gastroenterology, Hepatology, and Nutrition, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Henry
authors.initials: H
authors.lastname: Lin
authors.affiliation: Division of Gastroenterology, Hepatology, and Nutrition, Hospital for Sick Children, University of Toronto, Toronto, Canada.
authors.forename: Binita M
authors.initials: BM
authors.lastname: Kamath
authors.affiliation: Division of Genetics, Department of Pediatrics, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania; Department of Biostatistics and Epidemiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania; Department of Molecular Medicine, University La Sapienza, Rome, Italy.
authors.forename: Marcella
authors.initials: M
authors.lastname: Devoto
authors.affiliation: Department of Pathology and Laboratory Medicine, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Nancy B
authors.initials: NB
authors.lastname: Spinner
authors.affiliation: Division of Pediatric Gastroenterology, Hepatology, and Nutrition, Children's Hospital of Philadelphia, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Kathleen M
authors.initials: KM
authors.lastname: Loomes
keywords: ALGS, Alagille syndrome; BSA, bovine serum albumin; CK19, cytokeratin 19; ChiLDReN, Childhood Liver Disease Research Network; Cholestasis; GFP, green fluorescent protein; GWAS, genome-wide association study; Gene Modifier; Genome-Wide Association Study; JAG1; NOTCH2; PCR, polymerase chain reaction; SNP, single-nucleotide polymorphism; THBS2, thrombospondin 2; cDNA, complementary DNA; ddPCR, droplet digital polymerase chain reaction