Low concentrations of tumor necrosis factor-alpha promote human periodontal ligament stem cells osteogenic differentiation by activation of autophagy via inhibition of AKT/mTOR pathway.

BACKGROUND: Tumor necrosis factor-alpha (TNF-α) is one of the crucial inflammatory factors in alveolar bone metabolism during the process of periodontitis. Autophagy is indispensable for proper osteoblast function. However, the effects of autophagy on osteogenic differentiation of human periodontal ligament stem cells (hPDLSCs) in inflammatory microenvironment and the underlying mechanisms remain to be clarified. The aim of the present study was to investigate whether autophagy participates in hPDLSCs differentiation after treated with TNF-α and explore the underlying mechanisms. METHODS AND RESULTS: Characterizations of hPDLSCs were evaluated by Alizarin-red S staining, Oil red staining and flow cytometry. hPDLSCs were treated with various concentrations of TNF-α. Rapamycin or 3MA was used to achieve or inhibit autophagy activation. AKT signaling was inhibited using ARQ092. Cell proliferation was evaluated using Cell Counting Kit-8 (CCK8) assay. Real-time reverse transcriptase-polymerase chain reaction assay (RT-PCR), western blot, alkaline phosphatase (ALP) staining and Alizarin Red S staining were applied to evaluate levels of osteogenic differentiation and autophagy. CCK8 showed that low concentrations of TNF-α had no influence on cell proliferation, while high concentrations of TNF-α inhibited proliferation. Low concentrations of TNF-α promoted osteogenic differentiation and autophagy, while high concentrations of TNF-α inhibited osteogenic differentiation and autophagy in hPDLSCs. The levels of osteogenic differentiation in hPDLSCs were partly effected after co-incubation with 0.1 ng/mL TNF-α with 3MA or Rapamycin. ARQ092 enhanced 0.1 ng/mL TNF-α-induced ALP expression and mineral nodule formation. CONCLUSION: Low concentrations of TNF-α promote hPDLSCs osteogenic differentiation by activation of autophagy via inhibition of AKT/mTOR signaling.

[Influence of different designs and porcelain/glaze firing on the marginal and internal fit of implant-supported zirconia crowns].

OBJECTIVE: This study investigated the influence of different designs and porcelain/glaze firing on the marginal and internal fit of three kinds of computer aided design/computer aided manufacturing (CAD/CAM) zirconia ceramic implant-supported crowns. METHODS: Three groups of zirconia ceramic implant-supported crowns with different designs were produced from copings by using a Cercon CAD/CAM system (n = 8). The first two groups comprised double-layer crowns (zirconia coping + veneer) with regular (Group A) and full circumferential zirconia-collar marginal designs (Group B). The third group was composed of anatomic single-layer zirconia crowns without cores (Group C). Initially, the marginal and internal gaps of the copings and crowns were individually replicated by light-body silicon and then measured by micro-computed tomography scanning before and after porcelain/glaze firing. Five measurements were employed: vertical marginal gap (MG); horizontal marginal discrepancy (HMD); chamfer area (CA); axial wall (AW); and axial-occlusal transition area (AOT). Statistical analyses were performed by SPSS 17.0. RESULTS: HMD measurements in Group A were statistically higher than those in Groups B and C (P < 0.05), regardless of whether the values were obtained before or after porcelain/glaze firing. By contrast, the HIMD measurements in Groups B and C showed no significant difference (P > 0.05). Moreover, no differences were noted in MG, CA, AW, and AOT among the three groups (P > 0.05). All the measurements in the three groups showed no significant change after porcelain/glaze firing (P > 0.05), except for MG in Group A, which significantly decreased (P < 0.05). CONCLUSION: The marginal fits of the double-layer crowns with full circumferential zirconia-collar and the anatomic single-layer zirconia crowns were superior to that of the double-layer crowns with regular margins. The MG of the crowns with regular margins was obviously influenced by porcelain firing.

Influence of different designs and porcelain/glaze firing on the marginal and internal fit of implant-supported zirconia crowns / 华西口腔医学杂志

<p><b>OBJECTIVE</b>This study investigated the influence of different designs and porcelain/glaze firing on the marginal and internal fit of three kinds of computer aided design/computer aided manufacturing (CAD/CAM) zirconia ceramic implant-supported crowns.</p><p><b>METHODS</b>Three groups of zirconia ceramic implant-supported crowns with different designs were produced from copings by using a Cercon CAD/CAM system (n = 8). The first two groups comprised double-layer crowns (zirconia coping + veneer) with regular (Group A) and full circumferential zirconia-collar marginal designs (Group B). The third group was composed of anatomic single-layer zirconia crowns without cores (Group C). Initially, the marginal and internal gaps of the copings and crowns were individually replicated by light-body silicon and then measured by micro-computed tomography scanning before and after porcelain/glaze firing. Five measurements were employed: vertical marginal gap (MG); horizontal marginal discrepancy (HMD); chamfer area (CA); axial wall (AW); and axial-occlusal transition area (AOT). Statistical analyses were performed by SPSS 17.0.</p><p><b>RESULTS</b>HMD measurements in Group A were statistically higher than those in Groups B and C (P < 0.05), regardless of whether the values were obtained before or after porcelain/glaze firing. By contrast, the HIMD measurements in Groups B and C showed no significant difference (P > 0.05). Moreover, no differences were noted in MG, CA, AW, and AOT among the three groups (P > 0.05). All the measurements in the three groups showed no significant change after porcelain/glaze firing (P > 0.05), except for MG in Group A, which significantly decreased (P < 0.05).</p><p><b>CONCLUSION</b>The marginal fits of the double-layer crowns with full circumferential zirconia-collar and the anatomic single-layer zirconia crowns were superior to that of the double-layer crowns with regular margins. The MG of the crowns with regular margins was obviously influenced by porcelain firing.</p>

High-level transcription of human mitochondrial tRNA(Trp) based on ribozyme and characterization on its aminoacylation activity / 生物工程学报

In vitro transcription systems with T7 RNA polymerase (T7 RNAP) were widely used in preparation of RNA because of their simplicity and high efficiency. The transcripts would have additional 5' sequence since T7 promoter spans the transcription start site, while deletion of the transcription start site would severely reduce the T7 RNAP transcriptional activity. We successfully developed an in vitro transcription by combining of T7 RNAP high efficient transcription system and highly specific self-splicing technology of ribozymes, in this system, ribozyme self-splices at the designed specific site and releases the aim RNA without affecting transcription efficiency of T7 RNAP, the aminoacylation activity of human mitochondrial tRNA(Trp) (HmtRNA(Trp) (UCA)) is 113.6 pmol/microg. This method with its high efficiency on transcription and good repeatability is very suitable for preparation of accurate RNA in large scale.