Bone alteration and esthetics associated with implant-supported prostheses in the anterior maxilla under different implant placement timing: A retrospective clinical study of 1 to 3 years.

STATEMENT OF PROBLEM: Different factors influence alterations in facial bone thickness and esthetic outcomes after implant placement. Whether the timing of implant placement influences alterations in the bone dimensional and esthetic outcomes is unclear. PURPOSE: The purpose of this retrospective clinical study was to assess the influence of the timing of implant placement on alveolar bone alterations and esthetic outcome. MATERIAL AND METHODS: Data were collected from 40 patients who had received guided bone regeneration (GBR) performed simultaneously with immediate, early, or delayed single-tooth implant placement in the anterior maxilla. Facial and palatal horizontal bone thicknesses (FHBT, PHBT) and vertical bone level (FVBL, PVBL) immediately after surgery (T0), at 6 months after implant placement (T1), and at 1 to 3 years follow-up (T2) were measured, and the changes calculated. The pink esthetic score (PES) and white esthetic score (WES) were evaluated at the 1- to 3-year follow-up. The Kruskal-Wallis followed by the Dunn t test was applied to evaluate bone alteration among groups, and the Bonferroni method was used for adjusting multiple comparisons. The 1-way ANOVA test was used to determine any significance in the esthetic outcome in the 3 groups (α=.05). RESULTS: The reduction in the FHBT0 of the immediate, early, and delayed implant placement group (T2-T0) was -1.17 (-1.70, -0.61) mm, -1.53 (-1.69, -0.49) mm, and -1.47 (-2.30, -0.20) mm, respectively. The FHBT around the implant apices remained basically stable. No obvious changes in the PHBT around the implants of the immediate and delayed implant placement group were noted. The FVBL significantly decreased in each group during the follow-up period (-1.34 (01.88, -0.56) mm, immediate; -2.88 (-3.79, -1.07) mm, early; -1.26 (-2.52, -0.48) mm, delayed). The PVBL change in the early implant placement group (-2.18 (-3.26, -0.86) mm) was more significant than that in the immediate (-0.55 (-2.10, -0.17) mm) and delayed (-0.51 (-1.29, 0.02) mm) implantation groups (P =.013). The mean ±standard deviation PES/WES score of the immediate (15.6 ±1.84) and early (15.00 ±1.13) implant placement groups was higher than that of the delayed implant placement group (13.92 ±2.10) without significant difference. CONCLUSIONS: Similar bone changes and esthetic outcomes were found around implants of the immediate, early, and delayed implant placement groups.

Effect of mitophagy in the formation of osteomorphs derived from osteoclasts.

Osteoclasts are specialized multinucleated giant cells with unique bone-destroying capacities. A recent study revealed that osteoclasts undergo an alternative cell fate by dividing into daughter cells called osteomorphs. To date, no studies have focused on the mechanisms of osteoclast fission. In this study, we analyzed the alternative cell fate process in vitro and, herein, reported the high expression of mitophagy-related proteins during osteoclast fission. Mitophagy was further confirmed by the colocalization of mitochondria with lysosomes, as observed in fluorescence images and transmission electron microscopy. We investigated the role played by mitophagy in osteoclast fission via drug stimulation experiments. The results showed that mitophagy promoted osteoclast division, and inhibition of mitophagy induced osteoclast apoptosis. In summary, this study reveals the role played by mitophagy as the decisive link in osteoclasts' fate, providing a new therapeutic target and perspective for the clinical treatment of osteoclast-related diseases.

Porous composite hydrogels with improved MSC survival for robust epithelial sealing around implants and M2 macrophage polarization.

The application of mesenchymal stem cell (MSC)-based therapy is expected to make a significant contribution to the improvement of epithelial sealing around implants. However, there is currently no optimal MSC delivery biomaterial for clinical application in peri-implant epithelium (PIE) integration. In this study, we show that injectable photo-cross-linkable porous gelatin methacryloyl (GelMA)/silk fibroin glycidyl methacrylate (SilMA) hydrogels encapsulating gingival tissue-derived MSCs (GMSCs) are a simple and practical approach for re-epithelization applications. The hydrogels played a prominent role in supporting the proliferation, survival, and spread of GMSCs. Moreover, it was found that GMSCs-laden Porous GelMA/SilMA hydrogels could significantly upregulate the hemidesmosomes (HDs)-related genes and proteins expression and promote M2 polarization while inhibiting M1 polarization in vitro. Based on a rat model of early implant placement, application of the MSC-loaded hydrogels could enhance the protein expression of LAMA3 and BP180 (COL17A1) at the implant-PIE interface and reduce horseradish peroxidase (HRP) penetration between the implants and PIE. Noticeably, hydrogel-based MSC therapy contributed to augmenting M2 macrophage infiltration at two time points in the gingival connective tissue around implants. These findings demonstrated that GMSCs-laden Porous GelMA/SilMA hydrogels could facilitate epithelial sealing around implants and M2-polarized macrophages and may be a novel and facile therapeutic strategy for implant-PIE integration. STATEMENT OF SIGNIFICANCE: In the case of poor integration between the implant and gingival epithelium, peri-implantitis can develop, which is one of the main causes of implant failure. While stem cell therapy has tremendous potential for addressing this issue, poor cell survival and engraftment compromise the effectiveness of the therapy. Due to the excellent modifiable and tunable properties of gelatin and silk fibroin, injectable photo-cross-linkable porous hydrogels were developed using gelatin methacryloyl (GelMA) and silk fibroin glycidyl methacrylate (SilMA) as delivery vehicles for gingiva-derived MSCs (GMSCs). Porous GelMA/SilMA not only enhanced the proliferation and viability of GMSCs but also promoted their immunomodulatory capability for favorable epithelial sealing around implants. Overall, GMSCs-seeded porous hydrogels could be promising strategies for re-epithelization treatment.

circRNA422 enhanced osteogenic differentiation of bone marrow mesenchymal stem cells during early osseointegration through the SP7/LRP5 axis.

Circular RNAs (circRNAs) play an important role in biological activities, especially in regulating osteogenic differentiation of stem cells. However, no studies have reported the role of circRNAs in early osseointegration. Here we identified a new circRNA, circRNA422, from rat bone marrow mesenchymal stem cells (BMSCs) cultured on sandblasted, large-grit, acid-etched titanium surfaces. The results showed that circRNA422 significantly enhanced osteogenic differentiation of BMSCs with increased expression levels of alkaline phosphatase, the SP7 transcription factor (SP7/osterix), and lipoprotein receptor-related protein 5 (LRP5). Silencing of circRNA422 had opposite effects. There were two SP7 binding sites on the LRP5 promoter, indicating a direct regulatory relationship between SP7 and LRP5. circRNA422 could regulate early osseointegration in in vivo experiments. These findings revealed an important function of circRNA422 during early osseointegration. Therefore, circRNA422 may be a potential therapeutic target for enhancing implant osseointegration.

Activation of Nell-1 in BMSC Sheet Promotes Implant Osseointegration Through Regulating Runx2/Osterix Axis.

Neural epidermal growth factor-like 1 protein (Nell-1) is first studied because of its association with human craniosynostosis. Nell-1 has been used to accelerate the process of fracture healing because of the osteoinductive ability in recent years. However, the role of Nell-1 during the process of osteointegration is unknown. Here we show that activation of Nell-1 in the BMSC sheet promotes osseointegration in vivo and in vitro. We found that overexpression of Nell-1 improved osteogenic differentiation and enhanced matrix mineralization of BMSCs through increasing expression of Runx2 and Osterix. Activation of Nell-1 up-regulated the expression ratio of OPG/RANKL, which might have a negative influence on osteoclast differentiation. Furthermore, we obtained BMSC sheet-implant complexes transfected with lentivirus overexpressing and interfering Nell-1 in in vivo study, and confirmed that overexpression of Nell-1 promoted new bone formation around the implant and increased the bone-implant contacting area percentage. Our results demonstrate that activation of Nell-1 improves implant osteointegration by regulating Runx2/Osterix axis and shows the potential of BMSC sheet-implant complexes in gene therapy.

Adenovirus-Mediated LAMA3 Transduction Enhances Hemidesmosome Formation and Periodontal Reattachment during Wound Healing.

A robust dento-epithelial junction prevents external pathogenic factors from entering connective tissue and could be crucial for periodontal reattachment after periodontal surgery. The junctional epithelium (JE) is attached to the tooth surface through the hemidesmosome (HD) and internal basal lamina, where the primary component is laminin-332. Destruction of the JE leads to the loss of periodontal attachment. Traditional treatments are effective in eliminating local inflammation of the gingiva; however, few directly promote periodontal reattachment and HD formation. Here, we designed a gene-therapy strategy using the adenovirus-mediated human laminin-332 α3 chain (LAMA3) gene (Ad-LAMA3) transduced into a human-immortalized epidermal cell line (HaCaT) to study the formation of HD in vitro. Ad-LAMA3 promoted early adhesion and fast migration of HaCaT cells and increased expression of LAMA3 and type XVII collagen (BP180) significantly. Furthermore, HaCaT cells could facilitate formation of mature HDs after LAMA3 overexpression. In vivo experiments demonstrated that the JE transduced with Ad-LAMA3 could increase expression of LAMA3 and BP180 and "biological sealing" between the tooth and gingival epithelium. These results suggested that adenovirus-mediated LAMA3 transduction is a novel therapeutic strategy that promotes the stability and integration of the JE around the tooth during wound healing.

BMP2-mimicking peptide modified with E7 coupling to calcined bovine bone enhanced bone regeneration associating with activation of the Runx2/SP7 signaling axis.

Commercial bone substitute, such as calcined bovine bone (CBB), is currently extensively used as an alternative to autogenous bone. However, CBB lacks osteoinductivity and merely serves as a scaffold for native bone formation. To address this issue, we designed and prepared a heptaglutamate (E7)-modified BMP2-mimicking peptide (7E) and carried out a series of comprehensive physical characterizations and in vivo and in vitro studies to evaluate its role in the repair of cranial defects. The data elucidated that the amount of peptide anchoring to the bone graft materials was remarkably increased after modified with E7. Of note, 7E had a relatively stable and durable release, which promoted the osteogenic differentiation of rat derived bone marrow mesenchymal stem cells (BMSCs) and enhanced the bone regeneration of a rabbit calvarial defect by regulating the expression of the Runx2/SP7 axis. In summary, the composite biomaterials incorporating the E7-modified BMP2-mimicking peptide and CBB prepared in this study is a novel bone augmentation material with the merits of non-immunotoxicity, convenience, and low cost. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 108B:80-93, 2020.

Light-Controlled BMSC Sheet-Implant Complexes with Improved Osteogenesis via an LRP5/ß-Catenin/Runx2 Regulatory Loop.

The combination of bone marrow mesenchymal stem cell (BMSC) sheets and titanium implants (BMSC sheet-implant complexes) can accelerate osseointegration. However, methods of fabricating BMSC sheet-implant complexes are quite limited, and the survival of BMSC sheet-implant complexes is one of the key barriers. Here, we show that a light-controlled fabricating system can generate less injured BMSC sheet-implant complexes with improved viability and osteogenesis and that noninvasive monitoring of the viability of BMSC sheet-implant complexes using a lentiviral delivery system is feasible. Enhanced green fluorescent protein- and luciferase-expressing BMSC sheets were used to track the viability of BMSC sheet-implant complexes in vivo. The experiments of micro-computed tomography analysis and hard tissue slices were performed to evaluate the osteogenic ability of BMSC sheet-implant complexes in vivo. The results showed that BMSC sheet-implant complexes survived for almost 1 month after implantation. Notably, BMSC sheet-implant complexes fabricated by the light-controlled fabricating system had upregulating expression levels of low-density lipoprotein-receptor-related protein 5 (LRP5), ß-catenin, and runt-related transcription factor 2 (Runx2) compared to the complexes fabricated by mechanical scraping. Furthermore, we found that Runx2 directly bound to the rat LRP5 promoter and the LRP5/ß-catenin/Runx2 regulatory loop contributed to the enhancement of the osseointegrating potentials. In this study, we successfully fabricated BMSC sheet-implant complexes with improved viability and osteogenesis and established a feasible, noninvasive, and continuous method for tracking BMSC sheet-implant complexes in vivo. Our findings lay the foundation for the application of BMSC sheet-implant complexes in vivo and open new avenues for engineered BMSC sheet-implant complexes.

Laminin-521 Promotes Rat Bone Marrow Mesenchymal Stem Cell Sheet Formation on Light-Induced Cell Sheet Technology.

Rat bone marrow mesenchymal stem cell sheets (rBMSC sheets) are attractive for cell-based tissue engineering. However, methods of culturing rBMSC sheets are critically limited. In order to obtain intact rBMSC sheets, a light-induced cell sheet method was used in this study. TiO2 nanodot films were coated with (TL) or without (TN) laminin-521. We investigated the effects of laminin-521 on rBMSCs during cell sheet culturing. The fabricated rBMSC sheets were subsequently assessed to study cell sheet viability, reattachment ability, cell sheet thickness, collagen type I deposition, and multilineage potential. The results showed that laminin-521 could promote the formation of rBMSC sheets with good viability under hyperconfluent conditions. Cell sheet thickness increased from an initial 26.7 ± 1.5 µm (day 5) up to 47.7 ± 3.0 µm (day 10). Moreover, rBMSC sheets maintained their potential of osteogenic, adipogenic, and chondrogenic differentiation. This study provides a new strategy to obtain rBMSC sheets using light-induced cell sheet technology.

Combination of simvastatin, calcium silicate/gypsum, and gelatin and bone regeneration in rabbit calvarial defects.

The present study was performed to determine whether simvastatin improves bone regeneration when combined with calcium silicate/gypsum and gelatin (CS-GEL). The surface morphology was determined using field-emission scanning electron microscopy (FSEM). Degradation in vitro was evaluated by monitoring the weight change of the composites soaked in phosphate buffered saline (PBS). Drug release was evaluated using high-performance liquid chromatography (HPLC). Cytotoxicity testing was performed to assess the biocompatibility of composites. Four 5 mm-diameter bone defects were created in rabbit calvaria. Three sites were filled with CS-GEL, 0.5 mg simvastatin-loaded CS-GEL (SIM-0.5) and 1.0 mg simvastatin-loaded CS-GEL (SIM-1.0), respectively, and the fourth was left empty as the control group. Micro-computed tomography (micro-CT) and histological analysis were carried out at 4 and 12 weeks postoperatively. The composites all exhibited three-dimensional structures and showed the residue with nearly 80% after 4 weeks of immersion. Drug release was explosive on the first day and then the release rate remained stable. The composites did not induce any cytotoxicity. The results in vivo demonstrated that the new bone formation and the expressions of BMP-2, OC and type I collagen were improved in the simvastatin-loaded CS-GEL group. It was concluded that the simvastatin-loaded CS-GEL may improve bone regeneration.