Digitally printed custom trays assembled with occlusion rims and gothic arch tracing devices attached with tenon-and-mortise joints for biofunctional complete dentures: A dental technique.

A novel technique of digitally printed custom trays assembled with occlusion rims and gothic arch tracing devices attached with tenon-and-mortise joints for biofunctional complete dentures that could be delivered in 2 visits is presented. This technique takes advantage of closed-mouth impressions and objective jaw relation records by following the biofunctional prosthetic system concept with high efficiency and reduced labor.

Stage-Specific Role of Amelx Activation in Stepwise Ameloblast Induction from Mouse Induced Pluripotent Stem Cells.

Amelogenin comprises ~90% of enamel proteins; however, the involvement of Amelx transcriptional activation in regulating ameloblast differentiation from induced pluripotent stem cells (iPSCs) remains unknown. In this study, we generated doxycycline-inducible Amelx-expressing mouse iPSCs (Amelx-iPSCs). We then established a three-stage ameloblast induction strategy from Amelx-iPSCs, including induction of surface ectoderm (stage 1), dental epithelial cells (DECs; stage 2), and ameloblast lineage (stage 3) in sequence, by manipulating several signaling molecules. We found that adjunctive use of lithium chloride (LiCl) in addition to bone morphogenetic protein 4 and retinoic acid promoted concentration-dependent differentiation of DECs. The resulting cells had a cobblestone appearance and keratin14 positivity. Attenuation of LiCl at stage 3 together with transforming growth factor ß1 and epidermal growth factor resulted in an ameloblast lineage with elongated cell morphology, positivity for ameloblast markers, and calcium deposition. Although stage-specific activation of Amelx did not produce noticeable phenotypic changes in ameloblast differentiation, Amelx activation at stage 3 significantly enhanced cell adhesion as well as decreased proliferation and migration. These results suggest that the combination of inducible Amelx transcription and stage-specific ameloblast induction for iPSCs represents a powerful tool to highlight underlying mechanisms in ameloblast differentiation and function in association with Amelx expression.

Promoted Abutment-Soft Tissue Integration Around Self-Glazed Zirconia Surfaces with Nanotopography Fabricated by Additive 3D Gel Deposition.

Introduction: Improving the biological sealing around dental abutments could promote the long-term success of implants. Although titanium abutments have a wide range of clinical applications, they incur esthetic risks due to their color, especially in the esthetic zone. Currently, zirconia has been applied as an esthetic alternative material for implant abutments; however, zirconia is purported to be an inert biomaterial. How to improve the biological activities of zirconia has thus become a popular research topic. In this study, we presented a novel self-glazed zirconia (SZ) surface with nanotopography fabricated by additive 3D gel deposition and investigated its soft tissue integration capability compared to that of clinically used titanium and polished conventional zirconia surfaces. Materials and Methods: Three groups of disc samples were prepared for in vitro study and the three groups of abutment samples were prepared for in vivo study. The surface topography, roughness, wettability and chemical composition of the samples were examined. Moreover, we analyzed the effect of the three groups of samples on protein adsorption and on the biological behavior of human gingival keratinocytes (HGKs) and human gingival fibroblasts (HGFs). Furthermore, we conducted an in vivo study in which the bilateral mandibular anterior teeth of rabbits were extracted and replaced with implants and corresponding abutments. Results: The surface of SZ showed a unique nanotopography with nm range roughness and a greater ability to absorb protein. The promoted expression of adhesion molecules in both HGKs and HGFs was observed on the SZ surface compared to the surfaces of Ti and PCZ, while the cell viability and proliferation of HGKs and the number of HGFs adhesion were not significant among all groups. In vivo results showed that the SZ abutment formed strong biological sealing at the abutment-soft tissue interface and exhibited markedly more hemidesmosomes when observed with a transmission electron microscope. Conclusion: These results demonstrated that the novel SZ surface with nanotopography promoted soft tissue integration, suggesting its promising application as a zirconia surface for the dental abutment.

Epiprofin Transcriptional Activation Promotes Ameloblast Induction From Mouse Induced Pluripotent Stem Cells via the BMP-Smad Signaling Axis.

The transcriptional regulation of induced pluripotent stem cells (iPSCs) holds promise for their directed differentiation into ameloblasts, which are usually lost after tooth eruption. Ameloblast differentiation is regulated by multiple signaling molecules, including bone morphogenetic proteins (BMPs). Epiprofin (Epfn), a transcription factor, is expressed in the dental epithelium, and epithelial Epfn overexpression results in ectopic ameloblast differentiation and enamel formation in mouse incisor, a striking phenotype resembling that of mice with deletion of follistatin (a BMP inhibitor). However, it remains unknown whether and how Epfn transcriptional activation promotes ameloblast induction from mouse iPSCs. Here, we generated doxycycline-inducible Epfn-expressing mouse iPSCs (Epfn-iPSCs). Ameloblasts, which are characterized by positive staining for keratin 14 and amelogenin and alizarin red S staining, were successfully derived from Epfn-iPSCs based on a stage-specific induction protocol, which involved the induction of the surface ectoderm, dental epithelial cells, and ameloblasts at stages 1, 2, and 3, respectively. Epfn activation by doxycycline at stages 2 and/or 3 decreased cell proliferation and promoted ameloblast differentiation, along with the upregulation of p-Smad1/5/8, a key regulator of the BMP-Smad signaling pathway. Gene analysis of the BMP-Smad signaling pathway-associated molecules revealed that Epfn activation decreased follistatin expression at stage 2, but increased BMP2/4/7 expression at stage 3. Perturbations in the ameloblast differentiation process were observed when the BMP-Smad signaling pathway was inhibited by a BMP receptor inhibitor (LDN-193189). Simultaneous LDN-193189 treatment and Epfn activation largely reversed the perturbations in ameloblast induction, with partial recovery of p-Smad1/5/8 expression, suggesting that Epfn activation promotes ameloblast induction from mouse iPSCs partially by upregulating BMP-Smad activity. These results reveal the potential regulatory networks between Epfn and the BMP-Smad pathway and suggest that Epfn is a promising target for inducing the differentiation of ameloblasts, which can be used in enamel and tooth regeneration.

Novel Mesenchymal Stem Cell Spheroids with Enhanced Stem Cell Characteristics and Bone Regeneration Ability.

Mesenchymal stem cells (MSCs) exhibit self-renewal, multi-lineage differentiation potential and immunomodulatory properties, and are promising candidates for cellular therapy of various tissues. Despite the effective function of MSCs, the gradual loss of stem cell characteristics that occurs with repeated passages may significantly limit their therapeutic potential. A novel 3D shaking method was previously established to generate MSC spheroids in growth medium (GM-spheroids) and successfully maintain the multipotency of expanded MSCs, yet the expression of MSC-related genes was still low. In this study, we used a neurosphere culture technique to optimize the shaking culture method using human bone marrow-derived MSCs (BM-MSCs). MSC spheroids generated in neurosphere medium (NM-spheroids) maintained high expression of MSC-related genes during 3 weeks of prolonged shaking culture. Moreover, NM-spheroids generated from expanded MSCs showed high viability, upregulation of MSC-related and immune-related genes, and recovery of differentiation potential in vitro. Expanded adherent MSCs, GM-spheroids, and NM-spheroids were transplanted into a rat femur bone defect model to investigate their therapeutic potential in bone repair. Adherent MSCs and GM-spheroids showed delayed bone healing. In contrast, NM-spheroids showed high transplantation efficiency and enhanced bone regeneration. These data suggest that NM-spheroids generated using modified neurosphere culture conditions under continuous shaking recovered their stem cell characteristics in vitro and enhanced bone regeneration in vivo. Therefore, NM-spheroids should have great clinical potential for bone and tissue regenerative therapies as a stem cell-based biomaterial therapy.

Rapid and efficient generation of cartilage pellets from mouse induced pluripotent stem cells by transcriptional activation of BMP-4 with shaking culture.

Induced pluripotent stem cells (iPSCs) offer an unlimited source for cartilage regeneration as they can generate a wide spectrum of cell types. Here, we established a tetracycline (tet) controlled bone morphogenetic protein-4 (BMP-4) expressing iPSC (iPSC-Tet/BMP-4) line in which transcriptional activation of BMP-4 was associated with enhanced chondrogenesis. Moreover, we developed an efficient and simple approach for directly guiding iPSC-Tet/BMP-4 differentiation into chondrocytes in scaffold-free cartilaginous pellets using a combination of transcriptional activation of BMP-4 and a 3D shaking suspension culture system. In chondrogenic induction medium, shaking culture alone significantly upregulated the chondrogenic markers Sox9, Col2a1, and Aggrecan in iPSCs-Tet/BMP-4 by day 21. Of note, transcriptional activation of BMP-4 by addition of tet (doxycycline) greatly enhanced the expression of these genes. The cartilaginous pellets derived from iPSCs-Tet/BMP-4 showed an oval morphology and white smooth appearance by day 21. After day 21, the cells presented a typical round morphology and the extracellular matrix was stained intensively with Safranin O, alcian blue, and type II collagen. In addition, the homogenous cartilaginous pellets derived from iPSCs-Tet/BMP-4 with 28 days of induction repaired joint osteochondral defects in immunosuppressed rats and integrated well with the adjacent host cartilage. The regenerated cartilage expressed the neomycin resistance gene, indicating that the newly formed cartilage was generated by the transplanted iPSCs-Tet/BMP-4. Thus, our culture system could be a useful tool for further investigation of the mechanism of BMP-4 in regulating iPSC differentiation toward the chondrogenic lineage, and should facilitate research in cartilage development, repair, and osteoarthritis.

Modulatory effect of simultaneously released magnesium, strontium, and silicon ions on injectable silk hydrogels for bone regeneration.

Injectable silk hydrogels are ideal carriers of therapeutic agents due to their biocompatibility and low immunogenicity. Injectable silk hydrogels for bone regeneration have been previously developed but often utilize expensive biologics. In this study, we have developed an injectable silk composite incorporated with a triphasic ceramic called MSM-10 (54 Mg2SiO4, 36 Si3Sr5 and 10 MgO (wt%)) capable of simultaneously releasing magnesium, silicon, and strontium ions into its environment. These ions have been previously reported to possess therapeutic effects for bone regeneration. MSM-10 particles were incorporated into the silk hydrogels at various weight percentages [0.1 (SMH-0.1), 0.6 (SMH-0.6), 1 (SMH-1) and 2 (SMH-2)]. The effects of the released ions on the physicochemical and biological properties of the silk hydrogel were comprehensively evaluated. Increased MSM-10 loading was found to hinder the gelation kinetics of the silk hydrogel through the reduction of beta-sheet phase formation, which in turn affected the required sonication time for gelation, compressive strength, force of injection, microstructure and in vitro degradation rate. Primary human osteoblasts seeded on SMH-0.6 demonstrated increased proliferation and early alkaline phosphatase activity, as well as enhanced osteogenic gene expression compared to pure silk hydrogel and SMH-0.1. In vivo results in subcutaneous mouse models showed both decreased fibrous capsule formation and increased number of new blood vessels around the injected SMH-0.1 and SMH-0.6 implants compared to pure silk hydrogels. The results in this study indicate that the ions released from MSM-10 is able to influence the physicochemical and biological properties of silk hydrogels, and SMH-0.6 in particular shows promising properties for bone regeneration.

The osteoimmunomodulatory properties of MBG scaffold coated with amino functional groups.

Mesoporous bioactive glass (MBG) is a good scaffold for bone regeneration. In this study, amino functionalized MBG (N-MBG) was used as a model scaffold to examine the effect of the scaffold to bone marrow stromal cells (BMSCs) and macrophages. The MTT results revealed that the proliferation of BMSCs from ovariectomized rabbits was enhanced by N-MBG. Compared to the control group, the expression of osteogenic genes was significantly enhanced by N-MBG, which was related to CaSR pathway. Meanwhile, the anti-inflammatory cytokines (interleukin-10 and arginase-1) were also upregulated by N-MBG stimulation compared with MBG. Furthermore, the amino functionalization of MBG resulted in an increase in the pH value of the material extract. Interestingly, the formation of TRAP+ multinuclear cells was inhibited by the slightly alkaline extract to a certain extent, which reasonably explained the increase in TRAP+ multinuclear cells after adjusting the pH value of N-MBG extract. In vivo, the areas of new bone formation in the maxillary sinus floor elevation were increased in the N-MBG/BMSCs group with less TRAP+ multinuclear cells compared with the MBG/BMSCs group. These findings provided valuable insight that the osteogenic ability of MBG scaffold could be enhanced by amino functionalization due to coordinate BMSCs and macrophages differentiation.

The response of human osteoblasts, epithelial cells, fibroblasts, macrophages and oral bacteria to nanostructured titanium surfaces: a systematic study.

Nanotopography modification is a major focus of interest in current titanium surface design; however, the influence of the nanostructured surface on human cell/bacterium behavior has rarely been systematically evaluated. In this study, a homogeneous nanofiber structure was prepared on a titanium surface (Nano) by alkali-hydrothermal treatment, and the effects of this Nano surface on the behaviors of human MG-63 osteoblasts, human gingival epithelial cells (HGECs) and human gingival fibroblasts (HGFs) were evaluated in comparison with a smooth titanium surface (Smooth) by polishing and a micro-rough titanium surface (Micro) by sandblasting and acid etching. In addition, the impacts of these different surface morphologies on human THP-1 macrophage polarization and Streptococcus mutans attachment were also assessed. Our findings showed that the nanostructured surface enhanced the osteogenic activity of MG-63 cells (Nano=Micro>Smooth) at the same time that it improved the attachment of HGECs (Nano>Smooth>Micro) and HGFs (Nano=Micro>Smooth). Furthermore, the surface with nanotexture did not affect macrophage polarization (Nano=Micro=Smooth), but did reduce initial bacterial adhesion (Nano

M2 macrophages contribute to osteogenesis and angiogenesis on nanotubular TiO2 surfaces.

The monocyte/macrophage system plays an essential role in the host response and the fate of endosseous implanted materials. Macrophage behavior was thought to be regulated by nanostructured titanium which has been considered as a very promising candidate for dental implants. However, there is little known for subsequent effects of these activated macrophages on osteogenesis and angiogenesis which were essential for bone integration. Here we presented two different dimensions of titanium nanotubes generated by anodic oxidation at 10 V (NT 10) and 20 V (NT 20), respectively. The behavior of macrophages on the surfaces was evaluated, and their conditioned medium (CM) was collected to stimulate MC3T3 and HUVECs, with commercially pure titanium (cp Ti) as control. We found that NT 20 induced macrophage activation similar to the anti-inflammatory M2 macrophage state with the enhanced expression of IL-10 and ARG, while NT 10 was associated with M1 macrophage phenotype characterized by high levels of IL-1ß, iNOS and TNF-α. Furthermore, the osteogenic capacity of MC3T3 in CM from NT 20 was enhanced (NT 20 > NT 10 ≈ cp Ti) and the tube formation capacity of HUVECs was promoted in CM from nanotubular surfaces with increasing tube dimensions (NT 20 > NT 10 > cp Ti). Our data suggest that dental implants with the large nanotube dimension surface could result in a favorable osteoimmunomodulatory microenvironment for the establishment of osseointegration.

Tumor size predicts prognosis of head and neck synovial cell sarcoma.

Head and neck synoviosarcoma (HNSS) is uncommon. To the best of our knowledge, the specific clinicopathological characteristics, treatment outcome and prognostic factors of HNSS were uninvestigated at the time of writing, so a meta-analysis was performed. An online data collection was carried out using PubMed and Google Scholar. Studies that reported primary HNSS and the treatment, follow-up time and outcome were chosen for the present study. In total, 93 cases from 26 studies were included for analysis. The study sample consisted of 55 males and 38 females and the median age was 32.1 years (range, 4-76 years). The median follow-up period was 62.1 months (range, 1-373 months). The tumor size was correlated with local recurrence and metastasis of HNSS, as well as with mortality (P=0.001, P<0.0001 and P<0.0001, respectively). The three-year, five-year and 10-year survival rates were 82.1, 80.4 and 78.2% for treatment with surgery alone, and 88.5, 85.5 and 82% for treatment with surgery plus radiotherapy, respectively. A significant tumor size-dependent difference was found between the overall survival (OS) rates (P<0.0001), as tumors that were >5.0 cm in diameter were associated with a worse OS rate (hazard ratio, 6.460; 95% confidence interval, 206-18.917; P=0.001). The tumor size was found to be an independent adverse prognostic factor for the OS of HNSS patients. In conclusion, surgical excision is a mainstream treatment of HNSS and post-operative adjuvant radiotherapy improves the OS rate of HNSS patients.

Treatment outcome and prognostic factors of head and neck hemangiopericytoma: meta-analysis.

BACKGROUND: Head and neck hemangiopericytoma (HNHPC) is rare. Treatment outcome and specific prognostic factors were unexplored. METHODS: A case meta-analysis, in which treatment and outcome data were available, was performed. RESULTS: A total of 116 primary HNHPC cases were analyzed. Poor pathologic differentiation was associated with increased risk of tumor recurrence (odds ratio [OR] = 2.378; p = .005), metastasis (OR = 3.634; p = .011), and mortality (OR = 4.563; p = .002), whereas surgery was associated with decreased risk of tumor recurrence (OR = 0.109; p = .004). The tumor size >5.0 cm in diameter (hazard ratio [HR] = 6.391; p = .002), nonsurgical treatment (HR = 7.648; p = .000), and poor pathologic differentiation (HR = 1.705; p = .012) were the independent unfavorable prognostic factors for disease-free survival. Moreover, nonsurgical treatment (HR = 8.097; p = .002) and deep tumor location (HR = 4.074; p = .013) were independent adverse prognostic factors for overall survival (OS). CONCLUSION: These results suggest a management emphasizing the surgical removal of the tumor as first-line treatment. Tumor size >5.0 cm, poor pathologic differentiation, deep tumor location, and nonsurgical treatment were independent adverse prognostic factors.

Postoperative adjuvant radiotherapy improves loco-regional recurrence of head and neck mucosal melanoma.

Primary head and neck mucosal melanoma (HNMM) is a rare tumor with a poor prognosis. Controversy remains as to whether postoperative adjuvant radiotherapy (PORT) achieves a significant benefit in HNMM treatment. Because of the lack of available conclusive prospective data, we performed a systematic review and meta-analysis of all relevant available studies to clarify the benefits of PORT. A comprehensive literature search of PubMed and Google Scholar electronic databases was conducted to collect relevant studies until April 30, 2014. Studies published in the English language comparing surgery alone and surgery plus PORT for HNMM were included, with more than 15 study populations. All statistical analyses were performed using STATA version 12.0. A total of 423 patients were available from eight studies and the median sample size was 53 cases. The median follow-up time was 38.2 months (range 18.3-65.2 months). There was a positive association between PORT and loco-regional recurrence of HNMM (odds ratio [OR] = 0.36, 95% confidence interval [CI] = 0.22-0.60, P = 0.000). No associations were found between the PORT and 3-year and 5-year overall survival (OS) (OR = 1.41, 95% CI = 0.94-2.09, P = 0.093 and OR = 1.06, 95% CI = 0.70-1.61, P = 0.161, respectively). PORT had no impact on 3-year and 5-year OS (hazard ratio [HR] = 1.14, 95% CI = 0.80-1.61, P = 0.472 and HR = 1.34, 95% CI = 0.97-1.85, P = 0.227, respectively). PORT improved loco-regional recurrence of HNMM independent of OS.

[Treatment outcome and prognosis of head and neck hemangiopericytoma].

OBJECTIVE: Aim of the study is to report the unique clinicopathologic feature, treatment outcome and prognostic factors of head and neck hemangiopericytoma (HNHPC). METHOD: A retrospective data collection of reported HNHPC cases, in which therapy, follow-up and outcome data were available, was performed from the electronic database of PubMed, Embase, Google scholar, China National Knowledge Infrastructure, Wan Fang and Wei Pu until on December 31, 2013. RESULT: A total of 213 HNHPC cases were identified from 122 peer-reviewed articles. The recurrence rate was 24.4% (51/209). The positive surgical margin (OR= 3. 977, P<0. 01) and poor pathologic differentiation (OR=l. 890, P<0. 01) were associated with increased local recurrence. The metastasis rate was 15.8% (22/139). The positive surgical margin (OR=13. 833, P<0. 01), poor pathologic differentiation (OR=4. 661, P<0. 01) and non-surgical treatment (OR=2. 000, P<0. 01) were associated with increased distant metastasis. The mortality rate was 15. 0% (32/213). The tumor size >5. 0 cm in diameter (OR= 2. 860, P<0. 05), positive surgical margin (OR=9. 833, P<0. 01), poor pathologic differentiation (OR=4. 061, P<0. 01) and non-surgical treatment (OR=2. 032, P<0. 01) were associated with worse mortality. The treatment included surgery alone 139 cases, multiple treatments 64 cases and non-surgical treatment 10 cases. The overall survival (OS) of the 213 cases was 85%, and the 3-year, 5-year and 10-year OS were 86%, 78% and 74%, respectively. The 3-year, 5-year and 10-year OS for surgery alone were 95%, 88% and 84%, respectively. The 3- year, 5-year and 10-year OS for surgery plus radiotherapy were 90%, 80% and 80%, respectively. The 3-year, 5- year and 10-year OS for surgery plus chemotherapy were 75%, 25% and 25%, respectively. The 3-year, 5-year and 10-year OS for surgery plus radio-chemotherapy were 67%, 58% and 46%, respectively. There were signifi- cant survival difference in recurrence-free survival (RFS), metastasis free survival (MFS) and OS depending on surgical margins (P<0. 01). RFS, MFS and OS difference were identified depending on pathologic differentiation (P<0. 01). MFS and OS differences were observed on the different treatment modality (P<0. 01). OS differences was observed on the different tumor sizes (P<0. 05). Positive surgical margins was correlated with disease recurrence (HR= 3. 680, P<0.01), while poor pathologic differentiation was correlated with metastasis and death (HR=2. 619, P<0. 05 and HR=3. 188, P<0. 05). The tumor size >5. 0 cm in diameter and non-surgical treatment was correlated with death (HR= 5. 461, P<0. 01 and HR= 8. 563, P<0. 01, respectively). CONCLUSION: The surgical resection was the mainstream treatment and it was superior to multiple treatments. The tumor size, surgical margins, pathological differentiation and non-surgical treatment were independent prognostic factors.