Mouse Model of Alagille Syndrome and Mechanisms of Jagged1 Missense Mutations.

BACKGROUND & AIMS: Alagille syndrome is a genetic disorder characterized by cholestasis, ocular abnormalities, characteristic facial features, heart defects, and vertebral malformations. Most cases are associated with mutations in JAGGED1 (JAG1), which encodes a Notch ligand, although it is not clear how these contribute to disease development. We aimed to develop a mouse model of Alagille syndrome to elucidate these mechanisms. METHODS: Mice with a missense mutation (H268Q) in Jag1 (Jag1+/Ndr mice) were outbred to a C3H/C57bl6 background to generate a mouse model for Alagille syndrome (Jag1Ndr/Ndr mice). Liver tissues were collected at different timepoints during development, analyzed by histology, and liver organoids were cultured and analyzed. We performed transcriptome analysis of Jag1Ndr/Ndr livers and livers from patients with Alagille syndrome, cross-referenced to the Human Protein Atlas, to identify commonly dysregulated pathways and biliary markers. We used species-specific transcriptome separation and ligand-receptor interaction assays to measure Notch signaling and the ability of JAG1Ndr to bind or activate Notch receptors. We studied signaling of JAG1 and JAG1Ndr via NOTCH 1, NOTCH2, and NOTCH3 and resulting gene expression patterns in parental and NOTCH1-expressing C2C12 cell lines. RESULTS: Jag1Ndr/Ndr mice had many features of Alagille syndrome, including eye, heart, and liver defects. Bile duct differentiation, morphogenesis, and function were dysregulated in newborn Jag1Ndr/Ndr mice, with aberrations in cholangiocyte polarity, but these defects improved in adult mice. Jag1Ndr/Ndr liver organoids collapsed in culture, indicating structural instability. Whole-transcriptome sequence analyses of liver tissues from mice and patients with Alagille syndrome identified dysregulated genes encoding proteins enriched at the apical side of cholangiocytes, including CFTR and SLC5A1, as well as reduced expression of IGF1. Exposure of Notch-expressing cells to JAG1Ndr, compared with JAG1, led to hypomorphic Notch signaling, based on transcriptome analysis. JAG1-expressing cells, but not JAG1Ndr-expressing cells, bound soluble Notch1 extracellular domain, quantified by flow cytometry. However, JAG1 and JAG1Ndr cells each bound NOTCH2, and signaling from NOTCH2 signaling was reduced but not completely inhibited, in response to JAG1Ndr compared with JAG1. CONCLUSIONS: In mice, expression of a missense mutant of Jag1 (Jag1Ndr) disrupts bile duct development and recapitulates Alagille syndrome phenotypes in heart, eye, and craniofacial dysmorphology. JAG1Ndr does not bind NOTCH1, but binds NOTCH2, and elicits hypomorphic signaling. This mouse model can be used to study other features of Alagille syndrome and organ development.

Lollipop containing Glycyrrhiza uralensis extract reduces Streptococcus mutans colonization and maintains oral microbial diversity in Chinese preschool children.

The anticariogenic activity of the extract of Glycyrrhiza uralensis (licorice) has been well documented. We recently developed an herbal lollipop containing licorice extracts with Glycyrrhizol A, the compound displaying strong antimicrobial activity against Streptococcus mutans. Preliminary testing showed that the herbal lollipop reduced salivary S. mutans counts in vivo. In this study, we aimed to further test the efficacy of this herbal lollipop for reducing salivary S. mutans levels, and investigate its impact on salivary microbiome. Using a well-established in vitro oral microbiome model, we showed that licorice extract displays targeted killing against S. mutans without affecting the biodiversity of the community. In vivo study corroborated in vitro findings, showing for high caries-risk children aged 3-6 with salivary S. mutans levels >5x105 cells/ml, daily use of 2 licorice-containing lollipops for 3 weeks significantly reduced salivary S. mutans levels compared to the control group. Salivary microbiome analysis showed either no change or even increase in phylogenetic diversity of the oral community following herbal lollipop usage. Although further study with longer term observation is needed, these results suggest that use of licorice extract-containing lollipops can be as a simple and effective way to reduce the risk of dental caries in children.