Spheroid model study comparing the biocompatibility of Biodentine and MTA.

The primary objective of this study was to assess the biological effects of a new dentine substitute based on Ca3SiO5 (Biodentine™) for use in pulp-capping treatment, on pseudo-odontoblastic (MDPC-23) and pulp (Od-21) cells. The secondary objective was to evaluate the effects of Biodentine and mineral trioxide aggregate (MTA) on gene expression in cultured spheroids. We used the acid phosphatase assay to compare the biocompatibility of Biodentine and MTA. Cell differentiation was investigated by RT-qPCR. We investigated the expression of genes involved in odontogenic differentiation (Runx2), matrix secretion (Col1a1, Spp1) and mineralisation (Alp). ANOVA and PLSD tests were used for data analysis. MDPC-23 cells cultured in the presence of MTA had higher levels of viability than those cultured in the presence of Biodentine and control cells on day 7 (P = 0.0065 and P = 0.0126, respectively). For Od-21 cells, proliferation rates on day 7 were significantly lower in the presence of Biodentine or MTA than for control (P < 0.0001). Col1a1 expression levels were slightly lower in cells cultured in the presence of MTA than in those cultured in the presence of Biodentine and in control cells. Biodentine and MTA may modify the proliferation of pulp cell lines. Their effects may fluctuate over time, depending on the cell line considered. The observed similarity between Biodentine and MTA validates the indication for direct pulp-capping claimed by the manufacturers.

Transcriptional regulation of MSX1 natural antisense transcript.

Msx homeogenes play an important role in the epithelial-mesenchymal interactions leading development. Msx1 is relevant for dental and craniofacial morphogenesis, as suggested by phenotypes of Msx1 mutations in human and Msx1 KO mice. Our group showed that Msx1 gene expression can be regulated by a bidirectional transcription generating long noncoding antisense (AS) RNA the expression which is linked to the Msx1 sense (S) RNA level. Thus, the aim of the present study was to analyze the synthesis of Msx1 (AS) RNA. In vivo Msx1 AS expression analysis showed that (i) the putative promoter sequence but not the transcribed sequence was important and necessary for its expression, (ii) 2 different areas of alveolar bone can be distinguished depending on Msx1 S and AS expression, and (iii) Msx1 presence was necessary for Msx1 AS RNA full expression. In silico analysis of the Msx1 AS putative promoter region showed the presence of 4 Msx response elements possibly involved in Msx1 regulation. Msx1 constitutes an example of a bidirectionally transcribed gene giving rise to an S/AS RNA pair included in the big and growing family of AS noncoding RNAs. Our results contribute to defining a link between Msx1 S and AS RNAs resulting in a fine-tuned regulatory loop of Msx1 expression. The significance of this finding is that disturbance of the balance between Msx1 S and AS RNA status may be associated with tooth agenesis and bone loss.

Autoregulatory loop of Msx1 expression involving its antisense transcripts.

The Msx1 homeogene plays an important role in epithelial-mesenchymal interactions leading organogenesis. Msx1 gene is submitted to bidirectional transcription generating a long non-coding antisense (AS) RNA potentially involved in Msx1 expression regulation. RT-Q-PCR and RNA-FISH studies indicated that transient overexpression of the Msx1 AS transcript in 705IC5 mouse odontoblasts decreased the abundance of endogenous Msx1 S mRNA at the post-transcriptional level. Conversely, Msx1 overexpression increased the AS RNA level probably by activating AS transcription. In vivo mapping by RT-PCR evidenced both Msx1 RNAs in all adult mouse tissues tested raising the issue of Msx1 function during adulthood. The expression patterns of the two RNAs were similar, confirming the tight S/AS relationship. In particular, both Msx1 mRNAs and Msx1 protein were similarly distributed in eyes, and were found in regions with a common ectodermic origin and in cells potentially involved in regeneration. In conclusion, we report that Msx1 S RNA is negatively controlled by its AS RNA at a post-transcriptional level, and that the AS RNA is retrocontrolled positively by Msx1. The tight link between Msx1 S and AS RNAs constitutes a regulatory loop resulting in a fine-tuned expression of Msx1 which appears to be significant for adult homeostasis.

Msx1 expression regulation by its own antisense RNA: consequence on tooth development and bone regeneration.

Msx homeogenes play an important role in epithelial-mesenchymal interactions leading development. Msx1 is relevant for dental and craniofacial morphogenesis, as suggested by phenotypes of Msx1 mutations in human and Msx1 KO mice. During adulthood, Msx1 is still expressed in the skeleton where its role is largely unknown. Our group showed that the Msx1 gene is submitted to bidirectional transcription generating a long noncoding antisense (AS) RNA. During tooth development, Msx1 sense (S) and AS RNAs showed specific patterns of expression. Thus, the aim of the present study was to analyze the relation between Msx1 S and AS RNAs. In vivo mapping in adult mice showed that both Msx1 RNAs were detected in tested tissues such as bone. In vitro, Msx1 AS RNA decreased endogenous Msx1 S expression and modified Msx1 protein cell distribution. Regulations of Dlx5 and Bmp4 expression involving Msx1 S and AS RNAs showed that Msx1 AS RNA could modulate Msx1 function. The study of Msx1 S and AS RNA status is interesting in the case of tooth agenesis and bone loss to see if a disturbance of this balance could be associated with a disturbance of bone homeostasis. In that sense, our current results suggest a clear involvement of Msx1 in alveolar bone.

[Relations between extraction of wisdom teeth and temporomandibular disorders: a case/control study]. / Relations entre les extractions de dents de sagesse et les dysfonctionnements de l'appareil manducateur : étude cas / témoins.

The aim of this study was to assess the impact of extraction of third molars on the occurrence of temporo-mandibular disorders (TMD). A review of the literature and a case-control study have been conducted. The case-control study compares the frequency of extraction of third molars between the sample with TMD (case) and the sample without TMD (control). The proportion of patients who had undergone extractions of wisdom teeth was higher in the case group than in the control group. The difference was statistically significant when patients had undergone extraction of all four wisdom teeth or when the extraction of four wisdom teeth underwent in one sitting or under general anesthesia. The study of patients in case sample shows that all signs of TMD were more common in patients who had undergone extractions in several sessions and under local anesthesia. The temporomandibular joint sounds are significantly more frequent with local anesthesia. In the case group, 85 to 92% of patients have parafunctions and 5 to 11% have malocclusion. This demonstrates the multifactorial etiology of temporomandibular disorders.

Msx1 role in craniofacial bone morphogenesis.

The homeobox gene Msx1 encodes a transcription factor that is highly expressed during embryogenesis and postnatal development in bone. Mutations of the MSX1 gene in humans are associated with cleft palate and (or) tooth agenesis. A similar phenotype is observed in newborn mice invalidated for the Msx1 gene. However, little is known about Msx1 function in osteoblast differentiation and bone mineralization in vivo. In the present study, we aimed to explore the variations of individualized bone shape in a subtle way avoiding the often severe consequences associated with gene mutations. We established transgenic mice that specifically express Msx1 in mineral-matrix-secreting cells under the control of the mouse 2.3kb collagen 1 alpha 1 (Col1α1) promoter, which enabled us to investigate Msx1 function in bone in vivo. Adult transgenic mice (Msx1-Tg) presented altered skull shape and mineralization resulting from increased Msx1 expression during bone development. Serial section analysis of the mandibles showed a high amount of bone matrix in these mice. In addition, osteoblast number, cell proliferation and apoptosis were higher in Msx1-Tg mice than in controls with regional differences that could account for alterations of bone shape. However, Von Kossa staining and µCT analysis showed that bone mineralization was lower in Msx1-Tg mice than in controls due to alteration of osteoblastic differentiation. Msx1 appears to act as a modeling factor for membranous bone; it stimulates trabecular bone metabolism but limits cortical bone growth by promoting apoptosis, and concomitantly controls the collagen-based mineralization process.