Background Increasing evidence has revealed that long non-coding RNAs (lncRNAs) exert critical roles in biological mineralization. As a critical process for dentin formation, odontoblastic differentiation is regulated by complex signaling networks. The present study aimed to investigate the biological role and regulatory mechanisms of lncRNA-H19 (H19) in regulating the odontoblastic differentiation of human dental pulp stem cells (hDPSCs). Methods We performed lncRNA microarray assay to reveal the expression patterns of lncRNAs involved in odontoblastic differentiation. H19 was identified and verified as a critical factor by qRT-PCR. The gain- and loss-of-function studies were performed to investigate the biological role of H19 in regulating odontoblastic differentiation of hDPSCs in vitro and in vivo. Odontoblastic differentiation was evaluated through qRT-PCR, Western blot, and Alizarin Red S staining. Bioinformatics analysis identified that H19 could directly interact with miR-140-5p, which was further verified by luciferase reporter assay. After overexpression of miR-140-5p in hDPSCs, odontoblastic differentiation was determined. Moreover, the potential target genes of miR-140-5p were investigated and the biological functions of BMP-2 and FGF9 in hDPSCs were verified. Co-transfection experiments were conducted to validate miR-140-5p was involved in H19-mediated odontoblastic differentiation in hDPSCs. Results The expression of H19 was significantly upregulated in hDPSCs undergoing odontoblastic differentiation. Overexpression of H19 stimulated odontoblastic differentiation in vitro and in vivo, whereas downregulation of H19 revealed the opposite effect. H19 binds directly to miR-140-5p and overexpression of miR-140-5p inhibited odontoblastic differentiation of hDPSCs. H19 acted as a miR-140-5p sponge, resulting in regulated the expression of BMP-2 and FGF9. Overexpression of H19 abrogated the inhibitory effect of miR-140-5p on odontoblastic differentiation. Conclusion Our data revealed that H19 plays a positive regulatory role in odontoblastic differentiation of hDPSCs through miR-140-5p/BMP-2/FGF9 axis, suggesting that H19 may be a stimulatory regulator of odontogenesis.
To compare the efficacy of various irrigants (citric acid, ethylenediaminetetraacetic acid (EDTA) and NaOCl) and techniques in removing Ca(OH)2 in two types of curved root canal systems, simulated root canals with specific curvatures were used to investigate the effects of different irrigants and instruments on Ca(OH)2 removal. The optimal methods were verified on extracted human teeth. Simulated root canals were assigned to one of two groups based on the irrigation solution: 10% citric acid or 2.5% NaOCl. Each group was divided into four subgroups according to the technique used to remove Ca(OH)2. The percentage of Ca(OH)2 removal in different sections of root canals was calculated. On the basis of the results obtained for the simulated canals, 10% citric acid and 17% EDTA were applied to remove Ca(OH)2 from the extracted human teeth with curved root canal systems. The teeth were scanned by micro computed tomography to calculate the percentage of Ca(OH)2 removal in the canals. In simulated root canals, we found that 10% citric acid removed more Ca(OH)2 than 2.5% NaOCl in the 0–1 mm group from the apex level (P<0.05). Ultrasonic and EndoActivator activation significantly removed more Ca(OH)2 than a size 30 K file in the apical third (P<0.05). However, there were no significant differences in any sections of the canals for 10% citric acid or 17% EDTA in removing Ca(OH)2 in extracted human teeth. We concluded that it was effective to remove residual Ca(OH)2 using the decalcifying solution with EndoActivator or Passive Ultrasonic Irrigation in a curved root canal system. A protocol for Ca(OH)2 removal was provided based on the conclusions of this study and the methods recommended in previous studies.
Dental pulp cells were sensitive to compressive stress, especially after 12 and 24 h of applied force. Proliferation and odontogenic differentiation were significantly promoted in this in vitro model.
Root canal calcification is considered a great challenge during root canal treatment. Although the application of ultrasonic instruments and dental operating microscope (DOM) has advantages, dealing with calcified root canals still suffers a great risk of failure because of limited information about the location, length, and direction of obliteration on periapical radiographs. In this work, a cone-beam computed tomography- (CBCT-) aided method aimed at solving complicated calcified root canals in which conventional approaches could not work was proposed. Thirteen teeth with sixteen calcified canals (12 calcified in the upper third, 4 calcified in the middle third), which cannot be negotiated with conventional methods, were treated with the aid of CBCT. The location of calcification and depth of instrumentation and operating direction were calculated and assessed in three dimensions with ultrasonic instruments under DOM. In all thirteen teeth, canals with upper and middle thirds calcification were treated successfully. Finally, a guideline was proposed to help achieve consistent apical patency in calcified canals.
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