The Promise of Regenerative Medicine in the Reconstruction of Auricular Cartilage Deformities.

There are many physiologic and psychologic challenges associated with ear cartilage deformities which are incredibly distasteful to patients, particularly children. The development of regenerative medicine (RM) sciences has opened up a new window for the reconstruction of auricular cartilage because it allows the creation of a structure similar to the auricular in appearance and function. As part of this review, we discuss the role that each RM tool, including tissue engineering, cells, and biomolecules, plays in developing engineered auricular tissue. In previous studies, it was shown that the simultaneous use of natural and synthetic biomaterials as well as three-dimensional printing techniques could improve the biological and mechanical properties of this tissue. Another critical issue is using stem cells and differentiated cartilage cells to produce tissue-specific cellular structures and extracellular matrix. Also, the importance of choosing a suitable animal model in terms of handling and care facilities, physiologic similarities to humans, and breed uniformity in the preclinical assessments have been highlighted. Then, the clinical trials registered on the clinicaltrials.gov website, and the commercialized product, called AuriNovo, have been comprehensively explained. Overall, it is important to provide engineered auricular cartilage structures with acceptable safety and efficacy compared with standard methods, autologous cartilage transplantation, and prosthetic reconstruction in RM.

Tissue buccal fat pad-stromal vascular fraction as a safe source in maxillofacial bone regeneration: A clinical pilot study.

AIM: The purpose of this study was to examine the biological properties of the buccal fat pad (BFP)-derived tissue stromal vascular fraction (tSVF) in vitro and compare them with BFP-derived cellular SVF (cSVF). Furthermore, a clinical pilot study assessed the safety of using BFP-derived tSVF for maxillofacial bone regeneration. MATERIALS AND METHODS: This study was performed in two sections: 1) experimental section: BFP tissue was harvested from three healthy donors, and then cSVF and tSVF were isolated by enzymatic and mechanical methods to assess their biological properties and 2) clinical section: Ten patients with maxillofacial bone defects were enrolled according to eligibility criteria and offered two options for surgery, including autologous BFP-tSVF (n = 5) and autologous bone grafting (n = 5), to evaluate safety after a year of follow-up. RESULTS: The BFP-tSVF exhibited high cell viability and various cell surface markers, including CD45, CD31, and CD34. There was no population-doubling time and multilineage differentiation capacity compared with BFP-cSVF. BFP-tSVF is safe because of the lack of intervention-related adverse events reported in donor and surgery sites during a one-year period. In addition, cell therapy was feasible because it can be performed during surgery and requires little preparation time. Patients in the ABG group experienced pain and tenderness in the iliac crest, leading to dissatisfaction and complications. CONCLUSION: The experimental results confirmed that the cells isolated from BFP-tSVF have stemness properties similar to BFP-cSVF. Clinical evaluation also indicated that this cellular product could be used safely to regenerate maxillofacial bone defects.

Surface Coating of Polyurethane Films with Gelatin, Aspirin and Heparin to Increase the Hemocompatibility of Artificial Vascular Grafts.

Purpose: A hemocompatible substrate can offer a wonderful facility for nitric oxide (NO) production by vascular endothelial cells in reaction to the inflammation following injuries. NO inhibits platelet aggregation this is especially critical in small-diameter vessels. Methods: The substrate films were made of polyurethane (PU) in a casting process and after plasma treatments, their surface was chemically decorated with polyethylene glycol (PEG) 2000, gelatin, gelatin-aspirin, gelatin-heparin and gelatin-aspirin-heparin. The concentrations of these ingredients were optimized in order to achieve the biocompatible values and the resulting modifications were characterized by water contact angle and Fourier transform infra-red (FTIR) assays. The values of NO production and platelet adhesion were then examined. Results: The water contact angle of the modified surface was reduced to 26±4∘ and the newly developed hydrophilic chemical groups were confirmed by FTIR. The respective concentrations of 0.05 mg/ml and 100 mg/mL were found to be the IC50 values for aspirin and heparin. However, after the surface modification with aspirin, the bioactivity of the substrate increased in compared to the other experimental groups. In addition, there was a synergistic effect between these reagents for NO synthesis. While, heparin inhibited platelet adhesion more than aspirin. Conclusion: Because of the highly hydrophilic nature of heparin, this reagent was hydrolyzed faster than aspirin and therefore its influence on platelet aggregation and cell growth was greater. Taken together, the results give the biocompatible concentrations of both biomolecules that are required for endothelial cell proliferation, NO synthesis and platelet adhesion.

Comparative evaluation of the effects of three hydraulic calcium silicate cements on odontoblastic differentiation of human dental pulp stem cells: an in vitro study.

OBJECTIVE: The study aimed to compare the response of human dental pulp stem cells (hDPSCs) towards three hydraulic calcium silicate cements (HCSCs) by measuring cytotoxicity and expression of dentinogenic genes. METHODOLOGY: Dental pulps of five impacted mandibular third molars were extirpated as a source for hDPSCs. Next to culturing, hDPSCs were subjected to fluorescence-activated cell sorting after the third passage to validate stemness of the cells. Human DPSCs were exposed to diluted supernatants of OrthoMTA (OMTA), Biodentine (BD) and Calcium-Enriched Mixture (CEM) at concentrations 10, 25, 50 and 100% at the first, third and fifth day of culture. Then, cells were exposed to 10% concentrations supernatant of HCSCs to determine DSPP and DMP1 gene expression, using a quantitative polymerase-chain reaction. Data were analyzed using one-way and three-way ANOVA, followed by Tukey post hoc statistical tests. RESULTS: Optimal cell proliferation was observed in all groups, regardless of concentration and time-point. HCSC supernatants were non-cytotoxic to hDPSCs at all three time-points, except for 100% Biodentine on day five. On day seven, OMTA group significantly upregulated the expression of DSPP and DMP1 genes. On day 14, expression of DMP1 and DSPP genes were significantly higher in BD and OMTA groups, respectively. CONCLUSION: Biodentine significantly upregulated DMP1 gene expression over 14 days, whereas CEM was associated with only minimal expression of DSPP and DMP1 .

Prefabrication technique by preserving a muscular pedicle from masseter muscle as an in vivo bioreactor for reconstruction of mandibular critical-sized bone defects in canine models.

In vivo bioreactors serve as regenerative niches that improve vascularization and regeneration of bone grafts. This study has evaluated the masseter muscle as a natural bioreactor for ßTCP or PCL/ßTCP scaffolds, in terms of bone regeneration. The effect of pedicle preservation, along with sole, or MSC- or rhBMP2-combined application of scaffolds, has also been studied. Twenty-four mongrel dogs were randomly placed in six groups, including ßTCP, ßTCP/rhBMP2, ßTCP/MSCs, PCL/ßTCP, PCL/ßTCP/rhBMP2, and PCL/ßTCP/MSCs. During the first surgery, the scaffolds were implanted into the masseter muscle for being prefabricated. After 2 months, each group was divided into two subgroups prior to mandibular bone defect reconstruction; one with a preserved vascularized pedicle and one without. After 12 weeks, animals were euthanized, and new bone formation was evaluated using histological analysis. Histological analysis showed that all ß-TCP scaffold groups had resulted in significantly greater rates of new bone formation, either with a pedicle surgical approach or non-pedicle surgical approach, comparing to their parallel groups of ßTCP/PCL scaffolds (p ≤ .05). Pedicled ß-TCP scaffold groups that were treated with either rhBMP2 (48.443% ± 0.250%) or MSCs (46.577% ± 0.601%) demonstrated the highest rates of new bone formation (p ≤ .05). Therefore, masseter muscle can be used as a local in vivo bioreactor with potential clinical advantages in reconstruction of human mandibular defects. In addition, scaffold composition, pedicle preservation, and treatment with MSCs or rhBMP2, influence new bone formation and scaffold degradation rates in the prefabrication technique.

Comparative evaluation of the effects of three hydraulic calcium silicate cements on odontoblastic differentiation of human dental pulp stem cells: an in vitro study

Abstract Objective The study aimed to compare the response of human dental pulp stem cells (hDPSCs) towards three hydraulic calcium silicate cements (HCSCs) by measuring cytotoxicity and expression of dentinogenic genes. Methodology Dental pulps of five impacted mandibular third molars were extirpated as a source for hDPSCs. Next to culturing, hDPSCs were subjected to fluorescence-activated cell sorting after the third passage to validate stemness of the cells. Human DPSCs were exposed to diluted supernatants of OrthoMTA (OMTA), Biodentine (BD) and Calcium-Enriched Mixture (CEM) at concentrations 10, 25, 50 and 100% at the first, third and fifth day of culture. Then, cells were exposed to 10% concentrations supernatant of HCSCs to determine DSPP and DMP1 gene expression, using a quantitative polymerase-chain reaction. Data were analyzed using one-way and three-way ANOVA, followed by Tukey post hoc statistical tests. Results Optimal cell proliferation was observed in all groups, regardless of concentration and time-point. HCSC supernatants were non-cytotoxic to hDPSCs at all three time-points, except for 100% Biodentine on day five. On day seven, OMTA group significantly upregulated the expression of DSPP and DMP1 genes. On day 14, expression of DMP1 and DSPP genes were significantly higher in BD and OMTA groups, respectively. Conclusion Biodentine significantly upregulated DMP1 gene expression over 14 days, whereas CEM was associated with only minimal expression of DSPP and DMP1 .

Reconstruction of bilateral ramus-condyle unit defect using custom titanium prosthesis with preservation of both condyles.

BACKGROUND: Novel technologies for management and reconstruction of complex bony defects regarding both function and facial appearance are interestingly used in maxillofacial surgery. In the current study, we demonstrated reconstruction of a bilateral ramus-condyle unit (RCU) defect while preserving both condyles by a novel designed titanium prosthesis using virtual surgical planning (VSP), computer-aided design and manufacturing (CAD/CAM), and Selective Laser Melting (SLM) technologies. MATERIALS AND METHODS: A 3D customized titanium prosthesis was designed for a 49 -year-old patient with bilateral mandibular aggressive central giant cell granuloma (CGCG) according to mandibular normal anatomy and structure while preserving bilateral intact condyles. Finite element study was performed to investigate the effects of new design strength and the stress shielding phenomenon. The design of macro-pores inside the body of prosthesis allowed it to act as a scaffold for bone tissue engineering under load bearing conditions. RESULTS: Analysis of the strength and stress shielding phenomenon demonstrated favorable outcomes regarding the novel design. For instance, there was no stress shielding in any of the preserved condyles with regard to the size and distribution of stresses. Also, the stress distribution around the pores showed that these pores had no effect on the strength of the prosthesis. Thirty month follow-ups after reconstruction of bilateral RCU defect showed normal jaw function with a favorable facial appearance and mandibular contour. CONCLUSION: We design a novel patient-specific prosthesis with desirable biomechanical features for reconstruction of bilateral RCU defect after resection of the benign tumor with preservation of bilateral intact condyles.

Regenerative benefits of using growth factors in treatment of periodontal defects: A systematic review and meta-analysis with Trial Sequential Analysis on preclinical studies.

The ultimate goal in periodontal treatments is to achieve a functional and anatomical regeneration of the lost tissues. Numerous studies have in some way illustrated the beneficial effects of biologic modifiers in this process, yet they are subject to a rather large degree of diversity in their results. Thanks to the promising outcomes of bioengineering techniques in the field of periodontal regeneration, this systematic review aims to evaluate the effect of various biologic modifiers used in periodontal defects of animal models. Electronic databases (Medline, Scopus, Embase, Web of Science, and Google Scholar) were searched (March 2010-December 2020) for every study that used biomolecules for regeneration of periodontal osseous defects in animal models. Regenerated bone height or area, new cementum, new connective tissues, new regenerated periodontal ligament and the dimensions of epithelial attachment (either in mm/mm2 or percentage) were the investigated outcomes. The risk of bias of the included studies was assessed using the SYRCLE tool. In closing, there was a meta-analysis carried out on the outcomes of interest. Trial Sequential Analysis was also carried out to figure out the power of meta-analytic outcomes. From 1995 studies which were found in the initial search, 34 studies were included in this review, and 20 of them were selected for the meta-analysis. The eligible studies were categorized according to the morphology of the experimental periodontal defects as one-, two-, and three-wall intrabony defects; furcation defects, and recession-type defects. The most studied biomolecules were rhFGF-2, rhGDF-5, platelet-derived growth factor, bone morphogenetic protein-2, and enamel matrix derivative (EMD). Based on the meta-analysis findings, combined application of biomolecules with regenerative treatments could improve new bone and cementum formation near 1 mm when compared to the control groups in one, two and three-wall intrabony defect models (p < 0.001). In furcation grade II defect, the addition of biomolecules was observed to enhance bone area gain and cementum height regeneration up to almost 2 mm (p < 0.001). Trial Sequential Analysis results confirmed the significant effect in the aforementioned meta-analyses. In cases of the buccal recession model, the application of rhFGF-2 and rhGDF-5 decreased the dimension of epithelial attachments besides regenerative advantages on bone and cementum formation, but EMD deposition exerted no inhibitory effect on epithelial down-growth. Application of biologic modifiers especially FGF-2 and GDF-5, could positively improve the regeneration of periodontal tissues, particularly cementum and bone in animal models. Trial Sequential Analysis confirmed the results but the power of the evidences was high just in some subgroup meta-analyses, like bone and cementum regeneration in furcation grade II model and cementum regeneration in one-wall intrabony defects. The outcomes of this study can potentially endow clinicians with guidelines for the appropriate application of growth factors in periodontal regenerative therapies.

Fabrication of Decellularized Engineered Extracellular Matrix through Bioreactor-Based Environment for Bone Tissue Engineering.

Extracellular matrix (ECM)-contained grafts can be achieved by decellularization of native bones or synthetic scaffolds. Limitations associated with harvesting the native bone has raised interest in preparing in vitro ECM bioscaffold for bone tissue engineering. Here, we intend to develop an ECM-contained construct via decellularizing an engineered gelatin-coated ß-tricalcium phosphate (gTCP) scaffold. In order to find an optimal protocol for decellularization of cell-loaded gTCP scaffolds, they were seeded with buccal fat pad-derived stem cells. Then, four decellularization protocols including sodium dodecyl sulfate, trypsin, Triton X-100, and combined solution methods were compared for the amounts of residual cells and remnant collagen and alteration of scaffold structure. Then, the efficacy of the selected protocol in removing cells from gTCP scaffolds incubated in a rotating and perfusion bioreactor for 24 days was evaluated and compared with static condition using histological analysis. Finally, decellularized scaffolds, reloaded with cells, and their cytotoxicity and osteoinductive capability were evaluated. Complete removal of cells from gTCP scaffolds was achieved from all protocols. However, treatment with Triton X-100 showed significantly higher amount of remnant ECM. Bioreactor-incubated scaffolds possessed greater magnitude of ECM proteins including collagen and glycosaminoglycans. Reseeding the decellularized scaffolds also represented higher osteoinductivity of bioreactor-based scaffolds. Application of Triton X-100 as decellularization protocol and usage of bioreactors are suggested as a suitable technique for designing ECM-contained grafts for bone tissue engineering.

Computational modeling of media flow through perfusion-based bioreactors for bone tissue engineering.

Bioreactor system has been used in bone tissue engineering in order to simulate dynamic nature of bone tissue environments. Perfusion bioreactors have been reported as the most efficient types of shear-loading bioreactor. Also, combination of forces, such as rotation plus perfusion, has been reported to enhance cell growth and osteogenic differentiation. Mathematical modeling using sophisticated infrastructure processes could be helpful and streamline the development of functional grafts by estimating and defining an effective range of bioreactor settings for better augmentation of tissue engineering. This study is aimed to conduct computational modeling for newly designed bioreactors in order to alleviate the time and material consuming for evaluating bioreactor parameters and effect of fluid flow hydrodynamics (various amounts of shear stress) on osteogenesis. Also, biological assessments were performed in order to validate similar parameters under implementing the perfusion or rotating and perfusion fluid motions in bioreactors' prototype. Finite element method was used to investigate the effect of hydrodynamic of fluid flow inside the bioreactors. The equations used in the simulation to calculate the velocity values and consequently the shear stress values include Navier-Stokes and Brinkman equations. It has been shown that rotational fluid motion in rotating and perfusion bioreactor produces more velocity and shear stress compared with perfusion bioreactor. Moreover, implementing the perfusion together with rotational force in rotating and perfusion bioreactors has been shown to have more cell proliferation and higher activity of alkaline phosphatase enzyme as well as formation of extra cellular matrix sheet, as an indicator of bone-like tissue formation.

Bioreactor cultivation condition for engineered bone tissue: Effect of various bioreactor designs on extra cellular matrix synthesis.

Dynamic-based systems are bio-designed in order to mimic the micro-environments of the bone tissue. There is limited direct comparison between perfusion and perfusion-rotation forces in designing a bioreactor. Hence, in current study, we aimed to compare given bioreactors for bone regeneration. Two types of bioreactors including rotating & perfusion and perfusion bioreactors were designed. Mesenchymal stem cells derived from buccal fat pad were loaded on a gelatin/ß-Tricalcium phosphate scaffold. Cell-scaffold constructs were subjected to different treatment condition and place in either of the bioreactors. Effect of different dynamic conditions on cellular behavior including cell proliferation, cell adhesion, and osteogenic differentiation were assessed. Osteogenic assessment of scaffolds after 24 days revealed that rotating & perfusion bioreactor led to significantly higher expression of OCN and RUNX2 genes and also greater amount of ALP and collagen I protein production compared to static groups and perfusion bioreactor. Observation of cellular sheets which filled the scaffold porosities in SEM images, approved the better cell responses to rotating & perfusion forces of the bioreactor. The outcomes demonstrated that rotating & perfusion bioreactor action on bone regeneration is much preferable than perfusion bioreactor. Therefore, it seems that exertion of multi-stimuli is more effective for bone engineering.