Spatial Multi-omics Reveals the Role of the Wnt Modulator, Dkk2, in Palatogenesis'.

Multiple genetic and environmental etiologies contribute to the pathogenesis of cleft palate, which is the most common of the inherited disorders of the craniofacial complex. Insights into the molecular mechanisms regulating osteogenic differentiation and patterning in the palate during embryogenesis are limited and needed for the development of innovative diagnostics and cures. This study used the Pax9-/- mouse model with a consistent phenotype of cleft secondary palate to investigate the role of Pax9 in the process of palatal osteogenesis. Although prior research has identified the upregulation of Wnt pathway modulators Dkk1 and Dkk2 in Pax9-/- palate mesenchyme, limitations of spatial resolution and technology restricted a more robust analysis. Here, data from single-nucleus transcriptomics and chromatin accessibility assays validated by in situ highly multiplex targeted single-cell spatial profiling technology suggest a distinct relationship between Pax9+ and osteogenic populations. Loss of Pax9 results in spatially restricted osteogenic domains bounded by Dkk2, which normally interfaces with Pax9 in the mesenchyme. Moreover, the loss of Pax9 leads to a disruption in the normal osteodifferentiaion of palatal osteogenic mesenchymal cells. These results suggest that Pax9-dependent Wnt signaling modulators influence osteogenic programming during palate formation, potentially contributing to the observed cleft palate phenotype.

Fracture Toughness, Flexural Strength, and Flexural Modulus of New CAD/CAM Resin Composite Blocks.

PURPOSE: To determine and compare the fracture toughness, flexural strength and flexural modulus of four new, commercially available CAD/CAM resin composite blocks and one new CAD/CAM lithium disilicate glass-ceramic block, tested under dry and aged conditions. MATERIALS AND METHODS: Three dispersed-fillers resin composite blocks, CERASMART, KZR-CAD-HR2, and CAMouflage NOW, one polymer-infiltrated ceramic network resin composite block, Enamic, along with Obsidian, a lithium disilicate glass-ceramic block, were characterized. Fracture toughness was determined through the notchless triangular prism specimen test, while flexural strength and flexural modulus were determined by three-point bend testing. Blocks were cut and ground to obtain (6 × 6 × 6 × 12) mm prisms and 10:1 span-to-thickness ratio bars (n = 25/group); half of the resin composite specimens were aged in 37°C distilled water for 30 days before testing. Fractured surfaces were characterized using a scanning electron microscope. Results were analyzed using Weibull statistics and two-way ANOVA, followed by Scheffé multiple means comparisons (α = 0.05). RESULTS: With regards to fracture toughness, KZR stood out among resin composites with a dry value of 1.37 MPa·m1/2 ; this was significantly affected by ageing, while the fracture toughness of the other dispersed-fillers resin composite blocks was not. Obsidian had the highest fracture toughness at 1.47 MPa·m1/2 . With regards to flexural strength, Obsidian > CERASMART = KZR > CAMouflage > Enamic. The flexural strength of the resin composites was lowered by ageing. Enamic was found to have the highest flexural modulus among the resin composites (33.02 GPa), but its value was significantly lower than that of Obsidian (76.46 GPa); flexural modulus was not affected by ageing. CONCLUSION: There was a significant difference in flexural strength between the materials, but not unanimously in flexural modulus and fracture toughness. The tested resin composite block materials had inferior flexural strength, flexural modulus and fracture toughness compared with the tested lithium disilicate glass-ceramic block (Obsidian). Enamic, the polymer infiltrated ceramic network material, had a significantly higher flexural modulus than the dispersed-fillers materials. Ageing had a deleterious impact on the flexural strength of all RCB, while its effect on the flexural modulus was insignificant. The selection of any restorative material requires a thorough analysis of its advantages and limitations to inform the clinical decision in a case-by-case approach.

Determination of the allelic frequency in Smith-Lemli-Opitz syndrome by analysis of massively parallel sequencing data sets.

Data from massively parallel sequencing or 'Next Generation Sequencing' of the human exome has reached a critical mass in both public and private databases, in that these collections now allow researchers to critically evaluate population genetics in a manner that was not feasible a decade ago. The ability to determine pathogenic allele frequencies by evaluation of the full coding sequences and not merely a single nucleotide polymorphism (SNP) or series of SNPs will lead to more accurate estimations of incidence. For demonstrative purposes, we analyzed the causative gene for the disorder Smith-Lemli-Opitz Syndrome (SLOS), the 7-dehydrocholesterol reductase (DHCR7) gene and determined both the carrier frequency for DHCR7 mutations, and predicted an expected incidence of the disorder. Estimations of the incidence of SLOS have ranged widely from 1:10,000 to 1:70,000 while the carrier frequency has been reported as high as 1 in 30. Using four exome data sets with a total of 17,836 chromosomes, we ascertained a carrier frequency of pathogenic DHRC7 mutations of 1.01%, and predict a SLOS disease incidence of 1/39,215 conceptions. This approach highlights yet another valuable aspect of the exome sequencing databases, to inform clinical and health policy decisions related to genetic counseling, prenatal testing and newborn screening.