DSPP -1 Frameshift Mouse Model of Dentinogenesis Imperfecta

Objectives: Dentin dysplasia type II (DD-II) and dentinogenesis imperfecta types II and III (DGI-II and DGI-III) are autosomal dominant inherited conditions caused by mutations in one allele of the dentin sialophosphoprotein (DSPP) gene. Over half of the DSPP mutations that cause DD-II, DGI-II and DGI-III are -1 frameshifts in the repetitive region of DSPP. The objective was to understanding the mechanism inherited dentin malformations by generating and characterizing a Dspp -1 frameshift mouse model.
Methods: We generated Dspp^+/-1fs mouse using CRISPRcas9 gene editing approach that inserts a short Flag-tag insert (DYKDDDDK) for antibody detection followed by a single nucleotide deletion that translates into a 493 amino acid frameshift sequence similar to the one that causes dentin malformations in humans. Dentin formation 7-week-old in mandibular incisors was characterized by focused ion beam scanning electron microscopy (FIB-SEM), micro-computed tomography (µCT), and backscatter scanning electron microscopy (bSEM).
Results: The entire Dspp coding sequence was characterized in founder mice. Dspp^+/-1fs mandibular incisors showed visible dentin defects and a susceptibility to incisor fractures. Enamel appeared normal in thickness and degree of mineralization, but delaminated from dentin at the incisal edge. Dentin was much thinner than normal in both molars and incisors. FIB-SEM of longitudinal sections showed that the onset of dentin mineralization occurred in a thicker layer of predentin, suggesting it was delayed relative to the wild-type. Significant odontoblast pathology and macrophages in the odontoblast layer were observed. bSEM images taken at 1 mm increments along the mandibular incisor showed only a relatively thin layer of reparative dentin with cell inclusions that lacked dentinal tubules.
Conclusions: DSPP -1 frameshifts are toxic to odontoblasts reflecting a dominant negative (rather than haploinsufficiency) pathological mechanism. Our next experimental objectives are to localize mutant DSPP accumulation and determine if dimerization reduces secretion of wild-type DSPP. This study is supported by NIH/NIDCR R01DE015846.