CRISPR/Cas9-Induced Fam83h Knock-out Leads to Impaired Wnt/β-Catenin Pathway and Altered Expression of Tooth Mineralization Genes in Mice.

Abstract

Background: Dental enamel formation is a complex process that is regulated by various genes. One such gene, Family With Sequence Similarity 83 Member H (Fam83h), has been identified as an essential factor for dental enamel formation. Additionally, Fam83h has been found to be potentially linked to the Wnt/β-catenin pathway.

Objectives: This study aimed to investigate the effects of the Fam83h knockout gene on mineralization and formation of teeth, along with mediators of the Wnt/β-catenin pathway as a development aspect in mice.

Materials and methods: To confirm the Fam83h-KnockOut mice, both Sanger sequencing and Western blot methods were used. then used qPCR to measure the expression levels of genes related to tooth mineralization and formation of dental root, including Fam20a, Dspp, Dmp1, Enam, Ambn, Sppl2a, Mmp20, and Wnt/β-catenin pathway mediators, in both the Fam83h-Knockout and wild-type mice at 5, 11 and 18 days of age. also the expression level of Fgf10 and mediators of the Wnt/β-catenin pathway was measured in the skin of both Knockout and wild-type mice using qPCR. A histological assessment was then performed to further investigate the results.

Results: A significant reduction in the expression levels of Ambn, Mmp20, Dspp, and Fgf10 in the dental root of Fam83h-Knockout mice compared to their wild-type counterparts was demonstrated by our results, indicating potential disruptions in tooth development. Significant down-regulation of CK1a, CK1e, and β-catenin in the dental root of Fam83h-Knockout mice was associated with a reduction in mineralization and formation-related gene. Additionally, the skin analysis of Fam83h-Knockout mice revealed reduced levels of Fgf10, CK1a, CK1e, and β-catenin. Further histological assessment confirmed that the concurrent reduction of Fgf10 expression level and Wnt/β-catenin genes were associated with alterations in hair follicle maturation.

Conclusions: The concurrent reduction in the expression level of both Wnt/β-catenin mediators and mineralization-related genes, resulting in the disruption of dental mineralization and formation, was caused by the deficiency of Fam83h. Our findings suggest a cumulative effect and multi-factorial interplay between Fam83h, Wnt/Β-Catenin signaling, and dental mineralization-related genes subsequently, during the dental formation process.