Isolated dentinogenesis imperfecta: Novel DSPP variants and insights on genetic counselling.

OBJECTIVE: Dentinogenesis imperfecta (DI) is an inherited dentin defect and may be isolated or associated with disorders such as osteogenesis imperfecta, odontochondrodysplasia Ehler-Danlos and others. Isolated DI is caused mainly by pathogenic variants in DSPP gene and around 50 different variants have been described in this gene. Herein, we report on 19 patients from two unrelated Egyptian families with isolated DI. Additionally, we focused on genetic counselling of the two families. MATERIALS AND METHODS: The patients were examined clinically and dentally. Panoramic X-rays were done to some patients. Whole exome sequencing (WES) and Sanger sequencing were used. RESULTS: WES revealed two new nonsense variants in DSPP gene, c.288T > A (p.Tyr96Ter) and c.255G > A (p.Trp85Ter). Segregation analysis by Sanger sequencing confirmed the presence of the first variant in all affected members of Family 1 while the second variant was confirmed to be de novo in the patient of Family 2. CONCLUSIONS AND CLINICAL RELEVANCE: Our study extends the number of DSPP pathogenic variants and strengthens the fact that DSPP is the most common DI causative gene irrespective of patients' ethnicity. In addition, we provide insights on genetic counseling issues in patients with inherited DSPP variants taking into consideration the variable religion, culture and laws in our society.

The Role of DSPP in Dentine Formation and Hereditary Dentine Defects.

The dentine sialophosphoprotein (DSPP) gene is the only identified causative gene for dentinogenesis imperfecta type 2 (DGI-II), dentinogenesis imperfecta type 3 (DGI-III) and dentine dysplasia type 2 (DD-II). These three disorders may have similar molecular mechanisms involved in bridging the DSPP mutations and the resulting abnormal dentine mineralisation. The DSPP encoding proteins DSP (dentine sialoprotein) and DPP (dentine phosphoprotein) are positive regulators of dentine formation and perform a function during dentinogenesis. The present review focused on the recent findings and viewpoints regarding the relationship between DSPP and dentinogenesis as well as mineralisation from multiple perspectives, involving studies relating to spatial structure and tissue localisation of DSPP, DSP and DPP, the biochemical characteristics and biological function of these molecules, and the causative role of the proteins in phenotypes of the knockout mouse model and in hereditary dentine defects.

[Recognition on dentin dysplasia type â ¡].

Dentin dysplasia type Ⅱ (DD-Ⅱ) is a subtype of hereditary dentin disorders. The dentin sialophosphoprotein (DSPP) gene has been revealed to be the causative gene, whose mutations could affect the normal tooth development process. The lesions involve both deciduous and permanent dentition, mainly manifested as tooth discoloration, attrition and even the subsequent malocclusion. If not treated in time, it will significantly affect the physical and psychological health of patients. The disease is difficult to be diagnosed in clinic accurately as its low incidence and hidden manifestations. The present article aims to discuss the clinical and radiographic characteristics, diagnosis, treatment of DD-Ⅱ, in order to improve the overall understanding on DD-Ⅱ for clinicians.

Novel dentin sialophosphoprotein gene frameshift mutations affect dentin mineralization.

OBJECTIVE: This study aimed to identify candidate genes for inheritable dentin defects in three Chinese pedigrees and characterize the property of affected teeth. DESIGN: Clinical and radiological features were recorded for the affected individuals. Genomic DNA obtained from peripheral venous blood or saliva were analyzed by whole-exome sequencing. The density and microhardness of affected dentin was measured. Scanning electron microscopy (SEM) was also performed to obtain the microstructure phenotype. RESULTS: 1) General appearance: the affected dentitions shared yellowish-brown or milky color. Radiographs showed that the pulp cavity and root canals were obliterated in varying degrees or exhibited a pulp aspect in the 'thistle tube'. Some patients exhibited periapical infections without pulpal exposure, and some affected individuals showed shortened, abnormally thin roots accompanied by severe alveolar bone loss. 2) Genomic analysis: three new frameshift mutations (NM_014208.3: c.2833delA, c.2852delGand c.3239delA) were identified in exon 5 of dentin sialophosphoprotein (DSPP) gene, altering dentin phosphoprotein (DPP) as result. In vitro studies showed that the density and microhardness of affected dentin were decreased, the dentinal tubules were sparse and arranged disorderly, and the dentinal-enamel-junction (DEJ) was abnormal. CONCLUSIONS: In this study, we identified three novel frameshift mutations of dentin sialophosphoprotein gene related to inherited dentin defects. These mutations are speculated to cause abnormal coding of dentin phosphoprotein C-terminus, which affect dentin mineralization. These results expand the spectrum of dentin sialophosphoprotein gene mutations causing inheritable dentin defects and broaden our understanding of the biological mechanisms by which dentin forms.

Hereditary dentin defects with systemic diseases.

OBJECTIVE: This review aimed to summarize recent progress on syndromic dentin defects, promoting a better understanding of systemic diseases with dentin malformations, the molecules involved, and related mechanisms. SUBJECTS AND METHODS: References on genetic diseases with dentin malformations were obtained from various sources, including PubMed, OMIM, NCBI, and other websites. The clinical phenotypes and genetic backgrounds of these diseases were then summarized, analyzed, and compared. RESULTS: Over 10 systemic diseases, including osteogenesis imperfecta, hypophosphatemic rickets, vitamin D-dependent rickets, familial tumoral calcinosis, Ehlers-Danlos syndrome, Schimke immuno-osseous dysplasia, hypophosphatasia, Elsahy-Waters syndrome, Singleton-Merten syndrome, odontochondrodysplasia, and microcephalic osteodysplastic primordial dwarfism type II were examined. Most of these are bone disorders, and their pathogenic genes may regulate both dentin and bone development, involving extracellular matrix, cell differentiation, and metabolism of calcium, phosphorus, and vitamin D. The phenotypes of these syndromic dentin defects various with the involved genes, part of them are similar to dentinogenesis imperfecta or dentin dysplasia, while others only present one or two types of dentin abnormalities such as discoloration, irregular enlarged or obliterated pulp and canal, or root malformation. CONCLUSION: Some specific dentin defects associated with systemic diseases may serve as important phenotypes for dentists to diagnose. Furthermore, mechanistic studies on syndromic dentin defects may provide valuable insights into isolated dentin defects and general dentin development or mineralization.

Identification of DSPP novel variants and phenotype analysis in dentinogenesis dysplasia Shields type II patients.

OBJECTIVES: To investigate the genetic causes and teeth characteristics of dentin dysplasia Shields type II(DD-II) in three Chinese families. MATERIALS AND METHODS: Data from three Chinese families affected with DD-II were collected. Whole-exome sequencing (WES) and whole-genome sequencing (WGS) were conducted to screen for variations, and Sanger sequencing was used to verify mutation sites. The physical and chemical characteristics of the affected teeth including tooth structure, hardness, mineral content, and ultrastructure were investigated. RESULTS: A novel frameshift deletion mutation c.1871_1874del(p.Ser624fs) in DSPP was found in families A and B, while no pathogenic mutation was found in family C. The affected teeth's pulp cavities were obliterated, and the root canals were smaller than normal teeth and irregularly distributed comprising a network. The patients' teeth also had reduced dentin hardness and highly irregular dentinal tubules. The Mg content of the teeth was significantly lower than that of the controls, but the Na content was obviously higher than that of the controls. CONCLUSIONS: A novel frameshift deletion mutation, c.1871_1874del (p.Ser624fs), in the DPP region of the DSPP gene causes DD-II. The DD-II teeth demonstrated compromised mechanical properties and changed ultrastructure, suggesting an impaired function of DPP. Our findings expand the mutational spectrum of the DSPP gene and strengthen the understanding of clinical phenotypes related to the frameshift deletion in the DPP region of the DSPP gene. CLINICAL RELEVANCE: A DSPP mutation can alter the characteristics of the affected teeth, including tooth structure, hardness, mineral content, and ultrastructure.

AAV6-Mediated Gene Therapy Prevents Developmental Dentin Defects in a Dentinogenesis Imperfecta Type â ¢ Mouse Model.

Dentin is a major type of hard tissue of teeth and plays essential roles for normal tooth function. Odontoblasts are responsible for dentin formation. Mutations or deficiency in various genes affect the differentiation of odontoblasts, leading to irreversible dentin developmental defects in animals and humans. Whether such dentin defects can be reversed by gene therapy for odontoblasts remains unknown. In this study, we compare the infection efficiencies of six commonly used adeno-associated virus (AAV) serotypes (AAV1, AAV5, AAV6, AAV8, AAV9, and AAVDJ) in cultured mouse odontoblast-like cells (OLCs). We show that AAV6 serotype infects OLCs with the highest efficiency among the six AAVs. Two cellular receptors, which are able to recognize AAV6, AAV receptor (AAVR), and epidermal growth factor receptor (EGFR), are strongly expressed in the odontoblast layer of mouse teeth. After local administration to mouse molars, AAV6 infects the odontoblast layer with high efficiency. Furthermore, AAV6-Mdm2 was successfully delivered to teeth and prevents the defects in odontoblast differentiation and dentin formation in Mdm2 conditional knockout mice (a mouse model of dentinogenesis imperfecta type Ⅲ). These results suggest that AAV6 can serve as a reliable and efficient vehicle for gene delivery to odontoblasts through local injection. In addition, human OLCs were also successfully infected by AAV6 with high efficiency, and both AAVR and EGFR are strongly expressed in the odontoblast layer of extracted human developing teeth. These findings suggest that AAV6-mediated gene therapy through local injection may be a promising treatment approach for hereditary dentin disorders in humans.

Unequal Impact of COL1A1 and COL1A2 Variants on Dentinogenesis Imperfecta.

Dentinogenesis imperfecta (DI) is the main orodental manifestation of osteogenesis imperfecta (OI) caused by COL1A1 or COL1A2 heterozygous pathogenic variants. Its prevalence varies according to the studied population. Here, we report the molecular analysis of 81 patients with OI followed at reference centers in Brazil and France presenting COL1A1 or COL1A2 variants. Patients were submitted to clinical and radiographic dental examinations to diagnose the presence of DI. In addition, a systematic literature search and a descriptive statistical analysis were performed to investigate OI/DI phenotype-genotype correlation in a worldwide sample. In our cohort, 50 patients had COL1A1 pathogenic variants, and 31 patients had COL1A2 variants. A total of 25 novel variants were identified. Overall, data from a total of 906 individuals with OI were assessed. Results show that DI was more frequent in severe and moderate OI cases. DI prevalence was also more often associated with COL1A2 (67.6%) than with COL1A1 variants (45.4%) because COL1A2 variants mainly lead to qualitative defects that predispose to DI more than quantitative defects. For the first time, 4 DI hotspots were identified. In addition, we showed that 1) glycine substitution by branched and charged amino acids in the α2(I) chain and 2) substitutions occurring in major ligand binding regions-MLRB2 in α1(I) and MLBR 3 in α2(I)-could significantly predict DI (P < 0.05). The accumulated variant data analysis in this study provides a further basis for increasing our comprehension to better predict the occurrence and severity of DI and appropriate OI patient management.

[Mutation of dentin sialophosphoprotein and hereditary malformations of dentin].

The classification as well as the clinical manifestations of hereditary malformations of dentin are of great concern and have been deeply elucidated. The understanding of its genetic basis also increases progressively. Dentin sialophosphoprotein (DSPP) is the pathogenic gene of dentinogenesis imperfecta type Ⅱ, dentinogenesis imperfecta type Ⅲ and dentin dysplasia type Ⅱ. In this article, the classification of DSPP mutations as well as the resultant dysfunction of the mutant DSPP are summarized respectively and the corresponding clinical manifestations are analyzed. This work will provide a reference for the diagnosis and treatment of hereditary malformations of dentin.

A novel DSPP frameshift mutation causing dentin dysplasia type 2 and disease management strategies.

The present study aims to investigate the mutation in a Chinese family with dentin dysplasia type II (DD-II) and to summarize mutation hotspots, clinical manifestations, and disease management strategies. Phenotype analysis, clinical intervention, mutation screening, and cosegregation analysis within the enrolled family were performed. A summary of the reported mutations in the dentin phosphoprotein (DPP) region of dentin sialophosphoprotein (DSPP) was analyzed. Pathogenicity prediction analysis of the physical properties and function of DSPP variants was performed by bioinformatic processing. Clinical management strategies are discussed. A novel pathogenic mutation (c.2035delA) in the DPP region of DSPP was identified, which was cosegregated in the family. The immature permanent teeth of patients with DD-II presented with X-shaped root canal phenotypes. Most of the identified mutations for DD-II were clustered in the DPP region between nucleotides 1686-2134. Points of differential diagnosis, clinical interventions, and management strategies are proposed. This study revealed a novel DSPP frameshift mutation and presented new clinical features of DD-II. The locus involving nucleotides 1686-2134 of DSPP may represent a mutational hotspot for the disease. Appropriate management of DD-II at different stages is important to avoid the development of secondary dental lesions.

[Clinical and genetic analysis of a pedigree affected with hereditary dentinogenesis imperfecta type II].

OBJECTIVE: To explore the clinical and genetic characteristics of a Chinese pedigree affected with hereditary dentinogenesis imperfecta (DGI) type II. METHODS: Clinical data of the pedigree members were collected. Genomic DNA was extracted from peripheral blood samples and subjected to whole exome sequencing. RESULTS: Clinical characteristics of the affected family members have included amber teeth along with significant attrition, constricted roots and dentine hypertrophy leading to pulpal obliteration, which were suggestive of DGI type II. All of the affected members were found to have harbored a novel heterozygous c.2837delA (p.Asp946Valfs*368) variant of the DSPP gene which was predicted to be likely pathogenic. CONCLUSION: The c.2837delA variant of the DSPP gene probably underlay the disease in this pedigree. Above finding has expanded the variant spectrum of DSPP gene and provided a basis for molecular diagnosis and genetic counseling for this pedigree.

A Review of Dentinogenesis Imperfecta and Primary Dentin Disorders in Dogs.

This review describes the clinical, radiographic and histologic characteristics of dentinogenesis imperfecta diagnosed in two unrelated young dogs without evidence of concurrent osteogenesis imperfecta. The dentition was noted to have generalized coronal discoloration ranging from grey-blue to golden brown. Clinical pulp exposure, coronal wear and fractures were observed as was radiographic evidence of endodontic disease, thin dentin walls or dystrophic obliteration of the pulp canal. The enamel was severely affected by attrition and abrasion despite histologically normal areas; loss was most likely due to poor adherence or support by the underlying abnormal dentin. Histologically, permanent and deciduous teeth examined showed thin, amorphous dentin without organized dentin tubules and odontoblasts had dysplastic cell morphology. Primary dentin disorders, including dentinogenesis imperfecta and dentin dysplasia, have been extensively studied and genetically characterized in humans but infrequently reported in dogs. Treatment in human patients is aimed at early recognition and multi-disciplinary intervention to restore and maintain normal occlusion, aesthetics, mastication and speech. Treatment in both humans and canine patients is discussed as is the documented genetic heritability of primary dentin disorders in humans.

The Modified Shields Classification and 12 Families with Defined DSPP Mutations.

Mutations in Dentin Sialophosphoprotein (DSPP) are known to cause, in order of increasing severity, dentin dysplasia type-II (DD-II), dentinogenesis imperfecta type-II (DGI-II), and dentinogenesis imperfecta type-III (DGI-III). DSPP mutations fall into two groups: a 5'-group that affects protein targeting and a 3'-group that shifts translation into the −1 reading frame. Using whole-exome sequence (WES) analyses and Single Molecule Real-Time (SMRT) sequencing, we identified disease-causing DSPP mutations in 12 families. Three of the mutations are novel: c.53T>C/p.(Val18Ala); c.3461delG/p.(Ser1154Metfs*160); and c.3700delA/p.(Ser1234Alafs*80). We propose genetic analysis start with WES analysis of proband DNA to identify mutations in COL1A1 and COL1A2 causing dominant forms of osteogenesis imperfecta, 5'-DSPP mutations, and 3'-DSPP frameshifts near the margins of the DSPP repeat region, and SMRT sequencing when the disease-causing mutation is not identified. After reviewing the literature and incorporating new information showing distinct differences in the cell pathology observed between knockin mice with 5'-Dspp or 3'-Dspp mutations, we propose a modified Shields Classification based upon the causative mutation rather than phenotypic severity such that patients identified with 5'-DSPP defects be diagnosed as DGI-III, while those with 3'-DSPP defects be diagnosed as DGI-II.

Mouse Dspp frameshift model of human dentinogenesis imperfecta.

Non-syndromic inherited defects of tooth dentin are caused by two classes of dominant negative/gain-of-function mutations in dentin sialophosphoprotein (DSPP): 5' mutations affecting an N-terminal targeting sequence and 3' mutations that shift translation into the - 1 reading frame. DSPP defects cause an overlapping spectrum of phenotypes classified as dentin dysplasia type II and dentinogenesis imperfecta types II and III. Using CRISPR/Cas9, we generated a Dspp-1fs mouse model by introducing a FLAG-tag followed by a single nucleotide deletion that translated 493 extraneous amino acids before termination. Developing incisors and/or molars from this mouse and a DsppP19L mouse were characterized by morphological assessment, bSEM, nanohardness testing, histological analysis, in situ hybridization and immunohistochemistry. DsppP19L dentin contained dentinal tubules but grew slowly and was softer and less mineralized than the wild-type. DsppP19L incisor enamel was softer than normal, while molar enamel showed reduced rod/interrod definition. Dspp-1fs dentin formation was analogous to reparative dentin: it lacked dentinal tubules, contained cellular debris, and was significantly softer and thinner than Dspp+/+ and DsppP19L dentin. The Dspp-1fs incisor enamel appeared normal and was comparable to the wild-type in hardness. We conclude that 5' and 3' Dspp mutations cause dental malformations through different pathological mechanisms and can be regarded as distinct disorders.

Combined exome sequencing and deep phenotyping in highly selected fetuses with skeletal dysplasia during the first and second trimesters improves diagnostic yield.

OBJECTIVE: To investigate the genetic etiology of skeletal dysplasia in highly selected fetuses during the first and second trimesters using deep phenotyping and exome sequencing (ES). METHOD: Fetuses with short femurs were identified using the established prenatal diagnostic approach. A multidisciplinary team reviewed fetal phenotypic information (prenatal ultrasound findings, fetal postmortem, and radiographs) in a cohort of highly selected fetuses with skeletal dysplasia during the first and second trimesters. The affected families underwent multiplatform genetic tests. RESULTS: Of the 27 affected fetuses, 21 (77.8%) had pathogenic or potential pathogenic variations in the following genes: COL1A1, FGFR3, COL2A1, COL1A2, FLNB, DYNC2LI1, and TRIP11. Two fetuses had compound heterozygous mutations in DYNC2LI1 and TRIP11, respectively, and the other 19 carried de novo autosomal dominant variants. Novel variants were identified in COL1A1, COL2A1, COL1A2, DYNC2LI1, and TRIP11 in 11 fetuses. We also included the first description of the phenotype of odontochondrodysplasia in a prenatal setting. CONCLUSIONS: ES or panel sequencing offers a high diagnostic yield for fetal skeletal dysplasia during the first and second trimesters. Comprehensive and complete phenotypic information is indispensable for genetic analysis and the expansion of genotype-phenotype correlations in fetal skeletal abnormalities.

Phenotypic features of dentinogenesis imperfecta associated with osteogenesis imperfecta and COL1A2 mutations.

OBJECTIVE: Dentinogenesis imperfecta (DI) requires dental treatment. This study investigated the characteristics of DI teeth associated with osteogenesis imperfecta (OI) and COL1A2 mutations. STUDY DESIGN: Whole exome and Sanger sequencing were performed. Three primary teeth (called "OIDI teeth") obtained from 3 unrelated COL1A2 patients were investigated and compared with 9 control teeth from age-matched healthy individuals using colorimetry, micro-computed tomography, Knoop microhardness, energy dispersive X-ray spectroscopy, scanning electron microscopy, and histology. RESULTS: All patients were identified with heterozygous glycine substitutions in COL1A2. The COL1A2 mutations, c.1531G>T and c.2027G>T, were de novo, whereas c.3106G>C was inherited. OIDI1, 2, and 3 teeth had a substantial decrease in dentin microhardness and lightness. OIDI2 enamel microhardness was significantly reduced, whereas OIDI1 and 3 had enamel microhardness comparable to that of control individuals. The OIDI1 pulp cavity was large; OIDI2 was narrow; and OIDI3 was obliterated. OIDI1 and 3 had significantly higher carbon levels than those in control individuals. Numerous ectopic calcified masses, sparse and obstructed dentinal tubules, dentin holes, and collagen disorientation were observed. CONCLUSIONS: OIDI teeth had reduced lightness and variable pulp morphology. Weak dentin, mineral disproportion, and abnormal ultrastructure could contribute to the brittleness of OIDI teeth and adhesive restoration failure. Here, we expand the phenotypic spectrum of COL1A2 mutations and raise awareness among dentists seeing patients with OI.

Tooth ultrastructure of a novel COL1A2 mutation expanding its genotypic and phenotypic spectra.

OBJECTIVES: To investigate tooth ultrastructure and mutation of two patients in a family affected with osteogenesis imperfecta (OI) type IV and dentinogenesis imperfecta (DGI). METHODS: Mutations were detected by whole exome and Sanger sequencing. The permanent second molar obtained from the proband (DGI1) and the primary first molar from his affected son (DGI2) were studied for their color, roughness, mineral density, hardness, elastic modulus, mineral content, and ultrastructure, compared to the controls. RESULTS: Two novel missense COL1A2 variants, c.752C > T (p.Ser251Phe) and c.758G > T (p.Gly253Val), were identified in both patients. The c.758G > T was predicted to be the causative mutation. Pulp cavities of DGI1 (permanent teeth) were obliterated while those of DGI2 (primary teeth) were wide. The patients' teeth had darker and redder colors; reduced dentin hardness; decreased, disorganized, and scattered dentinal tubules and collagen fibers; and irregular dentinoenamel junction (DEJ), compared to controls. Lacunae-like structures were present in DGI2. CONCLUSIONS: We reported the novel causative mutation, c.758G > T (p.Gly253Val), in COL1A2 for OI type IV and DGI. The DGI dentin demonstrated inferior mechanical property and ultrastructure, suggesting severe disturbances of dentin formation. These could contribute to fragility and prone to infection of DGI teeth. This study expands phenotypic and genotypic spectra of COL1A2 mutations.

Human dentin characteristics of patients with osteogenesis imperfecta: insights into collagen-based biomaterials.

Osteogenesis imperfecta (OI), also known as "brittle bone disease", is a rare genetic disorder of the skeleton, whose most benign form I corresponds to autosomal dominant mutations in the genes encoding type I collagen (COLA1, COLA2). Several associated skeletal manifestations are often observed but, surprisingly, while dentin defects often reflect genetic bone disorders, about half of OI patients have no obvious oral manifestations. Here, we investigated the collagen, mineral and mechanical properties of dentin from deciduous teeth collected from patients with mild form of OI and displaying no obvious clinical signs of dentinogenesis imperfecta. For the first time, an increase in the hardness of OI dentin associated with an increase in mineral content compared to healthy patients was reported. In addition, OI altered the tissue characteristics of the dentin-enamel junction but the interfacial gradient was preserved. The impact of changes in molecular structure due to mutations in OI was assessed by Raman microspectroscopy. Our results highlighted a change in the hydroxyproline-proline ratio in direct association with collagen mineralization. Our findings suggest that the evaluation of teeth could be an important aid for mild types of OI that are often difficult to diagnose clinically and provide experimental evidence that hydroxyproline content should be considered in future studies on collagen-based biomaterials.

Physicochemical properties of dentinogenesis imperfecta with a known DSPP mutation.

AIM: To investigate the chemical and mechanical properties of teeth affected by a 1-bp deletion (c.2688delT) in the DSPP gene. METHODS AND MATERIALS: Maxillary first premolars were extracted from the affected individual at age 9 years due to the orthodontic reason for crowding. A sample was imbedded in epoxy resin and sectioned buccolingually, after micro-computerized tomography (µCT) images were taken. Scanning Electron Microscopy (SEM), Energy Dispersive Spectrometry (EDS) and Vickers microhardness testing were also performed. RESULTS: µCT reconstruction and analysis showed an irregularly obliterated pulp chamber and an extremely small pulpal volume in the DGI-II sample. The mineral density and microhardness scores were smaller in the dentin of the DGI-II sample compared to the wild-type. Mg content was lower in the dentin of the DGI-II sample compared to the wild-type. CONCLUSION: This study shows that dentin affected by a 1-bp deletion in DSPP has a reduced mineral density, diminished microhardness and reduced Mg content.