Abstract
Introduction and aims: To identify the genetic cause of dentinogenesis imperfecta type II (DGI-II) in a family, analyse tooth microstructure and evaluate functional defects in patient-derived stem cells (SHED) to explore genotype-phenotype links. Methods: A 13-member family with DGI-II underwent clinical evaluation. Whole-exome sequencing (WES) and Sanger sequencing identified pathogenic variants. Exfoliated tooth was assessed via micro-CT, microhardness testing and SEM. SHEDs from patients and controls were isolated, validated for stemness (flow cytometry, adipogenic induction) and tested for proliferation (CCK-8), migration (scratch assay) and mineralisation (osteogenic induction). Expression of DSPP, RUNX2, OCN and ALP was analysed by qRT-PCR and Western blot. Green florescence protein (GFP)-fused wild-type/mutant DSPP localisation was studied via immunocytochemistry. Results: A novel heterozygous DSPP variant (c.2470_2479del:p.S824Vfs*487) co-segregated with DGI-II: 8 affected members carried the variant. Micro-CT showed irregular dentinal tubules and reduced dentin density; patient tooth exhibited lower microhardness and localised enamel defects. SHEDs from patients displayed impaired proliferation, migration and mineralisation, with downregulated DSPP, RUNX2, OCN and ALP. Mutant DSPP accumulated in the endoplasmic reticulum (ER). Conclusion: The novel DSPP frameshift mutation (c.2470_2479del) underlies DGI-II, driving dentin defects and SHEDs dysfunction. The aberrant endoplasmic reticulum retention of mutant DSPP protein, implicates ER stress in mediating the genotype-phenotype correlation. Clinical relevance: This study expands DGI-II's genetic profile and highlights DSPP's role in dentinogenesis. Gene sequencing is essential to identify the DSPP variant site, which relate closely to the clinical phenotype and treatment of DGI.