Evaluating osteogenic potential of a 3D-printed bioceramic-based scaffold for critical-sized defect treatment: an in vivo and in vitro investigation.

The integration of precision medicine principles into bone tissue engineering has ignited a wave of research focused on customizing intricate scaffolds through advanced 3D printing techniques. Bioceramics, known for their exceptional biocompatibility and osteoconductivity, have emerged as a promising material in this field. This article aims to evaluate the regenerative capabilities of a composite scaffold composed of 3D-printed gelatin combined with hydroxyapatite/tricalcium phosphate bioceramics (G/HA/TCP), incorporating human dental pulp-derived stem cells (hDPSCs). Using 3D powder printing, we created cross-shaped biphasic calcium phosphate scaffolds with a gelatin layer. The bone-regenerating potential of these scaffolds, along with hDPSCs, was assessed through in vitro analyses and in vivo studies with 60 rats and critical-sized calvarial defects. The assessment included analyzing cellular proliferation, differentiation, and alkaline phosphatase activity (ALP), and concluded with a detailed histological evaluation of bone regeneration. Our study revealed a highly favorable scenario, displaying not only desirable cellular attachment and proliferation on the scaffolds but also a notable enhancement in the ALP activity of hDPSCs, underscoring their pivotal role in bone regeneration. However, the histological examination of calvarial defects at the 12-wk mark yielded a rather modest level of bone regeneration across all experimental groups. The test and cell group exhibited significant bone formation compared to all other groups except the control and cell group. This underscores the complexity of the regenerative process and paves the way for further in-depth investigations aimed at improving the potential of the composite scaffolds.

Effect of simultaneous and sequential use of TGF-ß1 and TGF-ß3 with FGF-2 on teno/ligamentogenic differentiation of periodontal ligament stem cells.

OBJECTIVE: The periodontal ligament is a crucial part of the periodontium, and its regeneration is challenging. This study compares the effect of simultaneous and sequential use of FGF-2 and TGF-ß1 with FGF-2 and TGF-ß3 on the periodontal ligament stem cells (PDLSCs) teno/ligamentogenic differentiation. DESIGN: This study comprises ten different groups. A control group with only PDLSCs; FGF-2 group containing PDLSCs with a medium culture supplemented with FGF-2 (50 ng/mL). In other experimental groups, different concentrations (5 ng/mL or 10 ng/mL) of TGF-ß1&-ß3 simultaneously or sequentially were combined with FGF-2 on the cultured PDLSCs. TGF-ß was added to the medium after day 3 in the sequential groups. Methyl Thiazolyl Tetrazolium (MTT) assay on days 3, 5, and 7 and Quantitative Real-time Polymerase Chain Reaction (RT-qPCR) analysis after day 7 were conducted to investigate PLAP1, SCX, and COL3A1, RUNX2 genes. All experiments were conducted in a triplicate. The One-way and Two-way ANOVA with Tukey post hoc were utilized to analyze the results of the MTT and RT-qPCR tests, respectively. A p-value less than 0.05 is considered significant. RESULTS: The proliferation of cells on days 3, 5, and 7 was not significantly different among different experimental groups (P > 0.05). A higher expression of the PLAP1, SCX, and COL3A1 have been seen in groups with sequential use of growth factors; among these groups, the group using 5 ng/mL of TGF-ß3 led other groups with the most amount of significant upregulation in PLAP1(17.69 ± 1.11 fold; P < 0.0001), SCX (5.71 ± 0.38 fold; P < 0.0001), and COL1A3 (6.35 ± 0.39 fold; P < 0.0001) expression, compared to the control group. The expression of the RUNX2 decreased in all groups compared to the control group; this reduction was more in groups with sequential use of growth factors. CONCLUSION: The sequential use of growth factors can be more effective than simultaneous use in teno/ligamentogenic differentiation of PDLSCs. Moreover, treatment with 5 ng/mL TGF-ß3 after FGF-2 was more effective than TGF-ß1.

Application of biodegradable Patient-specific scaffolds for maxillofacial bone regeneration: a scoping review of clinical studies.

This study aimed to systematically review clinical studies in which biodegradable patient-specific scaffolds were used for bone regeneration. Studies in which biodegradable scaffolds were fabricated through computer-assisted design and computer-assisted manufacturing (CAD-CAM) procedures were included. Those that applied non-biodegradable materials or used biodegradable materials in a condensable powder or block form were excluded. Among a total of 26 included studies, 11 used customised allogeneic bone blocks, five used polycaprolactone (PCL)-containing scaffolds, four used hydroxyapatite (HA) scaffolds, and four biphasic calcium phosphate (BCP). The majority of the studies applied scaffolds for minor intraoral defects. All the large defects were reconstructed with polymer-containing scaffolds. Results of the included studies showed partial to complete filling of the defect following the application of biodegradable scaffolds. However, limited graft exposure was reported when using PCL, BCP, and HA scaffolds. Tissue engineering can be considered a potential method for the treatment of maxillofacial bone defects. However, more evidence is required, especially for the application of biodegradable scaffolds in large defects.

Management of recurrent aphthous ulcers with therapeutic Nd:YAG laser, using two different methods.

BACKGROUND: Low-level laser therapy (LLLT) has been applied for the management of craniomaxillofacial disorders, including intraoral wounds, as well as recurrent aphthous stomatitis (RAS) lesions. However, the proper combination of laser features and tissue characteristics remains the major challenge in the realm of photobiomodulation (PBM). OBJECTIVES: The aim of the present study was to assess the feasibility of neodymium-doped yttrium aluminum garnet (Nd:YAG) laser therapy in treating RAS lesions, and to compare 2 techniques, different with regard to the distance between the fiber tip and the ulcer. MATERIAL AND METHODS: A total of 138 patients (94 males and 44 females) with untreated RAS were divided into 3 groups: focused laser (energy density: 48 J/cm2; power density: 0.797 W/cm2; spot size: 0.1256 cm2); defocused laser (energy density: 21 J/cm2; power density: 0.354 W/cm2; spot size: 0.2826 cm2); and placebo. In the focused group, laser irradiation was performed with the laser tip kept 1 mm away from the lesion. Acrylic cylinders were prepared to precisely fit the handpiece tip and hold it in the proper position. In the defocused group, acrylic cylinders were prepared to set the laser tip 6 mm away from the lesion to obtain defocused irradiation. Finally, in the placebo group, a routine laser therapy procedure was carried out with a helium-neon (He-Ne) red light laser. The lesion size, and pain intensity and duration were recorded. RESULTS: Photobiomodulation showed a significantly more efficient pain relief as compared to the placebo group (p < 0.001) and also significantly better results in decreasing pain duration (p < 0.001). Besides, the diameter of the lesions in the exposed cases decreased during the 3 consecutive days of the study, while an increase in the diameter of the lesions was noticed in the placebo group. CONCLUSIONS: The Nd:YAG laser therapy, with the conditions and adjustments of the present study, may be successfully applied to manage RAS lesions, using either focused and defocused scanning techniques.

Comparative Biocompatibility and Odonto-/Osteogenesis Effects of Hydraulic Calcium Silicate-Based Cements in Simulated Direct and Indirect Approaches for Regenerative Endodontic Treatments: A Systematic Review.

BACKGROUND: Regenerative dentistry is the operation of restoring dental, oral and maxillofacial tissues. Currently, there are no guidelines for the ideal cement/material in regenerative endodontic treatments (RET). Hydraulic calcium silicate-based cements (hCSCs) are currently the material of choice for RET. OBJECTIVES: This systematic review was conducted to gather all of the different direct and indirect approaches of using hCSCs in RET in vitro and in vivo, and to ascertain if there are any superiorities to indirect approaches. METHODS AND MATERIALS: This systematic review was conducted according to the 2020 PRISMA guidelines. The study question according to the PICO format was as follows: Comparison of the biological behavior (O) of stem cells (P) exposed to hCSCs through direct and indirect methods (I) with untreated stem cells (C). An electronic search was executed in Scopus, Google Scholar, and PubMed. RESULTS: A total of 78 studies were included. Studies were published between 2010 and 2022. Twenty-eight commercially available and eighteen modified hCSCs were used. Seven exposure methods (four direct and three indirect contacts) were assessed. ProRoot MTA and Biodentine were the most used hCSCs and had the most desirable results. hCSCs were either freshly mixed or set before application. Most studies allowed hCSCs to set in incubation for 24 h before application, which resulted in the most desirable biological outcomes. Freshly mixed hCSCs had the worst outcomes. Indirect methods had significantly better viability/proliferation and odonto-/osteogenesis outcomes. CONCLUSION: Biodentine and ProRoot MTA used in indirect exposure methods result in desirable biological outcomes.

The effect of antibacterial photodynamic therapy with diode laser on chromogenic bacteria associated with dental black staining: An in-vitro study.

BACKGROUND: Aggregatibacter actinomycetemcomitan (A.a) and Actinomyces naeslundii (A.n) are two gram-negative chromogenic bacteria involved in the formation of dental black stainings. Our study aimed to investigate the antibacterial effect of photodynamic therapy (aPDT) using two photosensitizers, Methylene Blue (MB) and Indocyanine Green (ICG). MATERIALS AND METHODS: In this in-vitro study, two isolates of each selected bacterium were cultured and treated as follows; Negative control with no treatment; CHX as a positive control; ICG; MB; ICG with 808 nm laser activation; and MB with 660 nm laser activation. The number of colonies (CFU/mL) was determined to compare the groups. The qualitative evaluation of biofilm formation was done by scanning electron microscopy of treated enamel pieces. The logarithmic values of the colony counts were compared using One-way ANOVA and the Welch test Tukey HSD and Games-Howell tests were used for multiple comparisons. P-values of less than 0.05 were considered statistically significant. RESULTS: The use of ICG alone or along with laser irradiation at the wavelength of 808 nm significantly reduced the number of colonies of A.a and A.n bacteria. Comparing the colony counts in the MB group with the positive control showed no significant decrease in bacterial load. On the contrary, activation of MB with 660 nm radiation of diode laser showed a significant antibacterial effect. The density of bacterial biofilm was significantly lower in the groups treated with MB and ICG without laser activation than in the control group; however, the reduction in bacteria biofilm density was more robust using photodynamic therapy with ICG. CONCLUSION: aPDT using MB with 660 nm laser and ICG with 808 nm laser significantly reduced the number of chromogenic A.a and A.n bacteria, and photodynamic therapy with ICG was proven to be significantly more effective than MB with or without laser radiation.

Bilayer scaffolds/membranes for bone tissue engineering applications: A systematic review.

OBJECTIVE: This systematic review evaluates the purpose, materials, physio-mechanical, and biological effects of bilayer scaffolds/membranes used for bone tissue engineering applications. METHODS: A comprehensive electronic search of English-language literature from 2012 to October 2022 was conducted in PubMed, Scopus, ScienceDirect, and Google Scholar online databases according to the PRISMA 2020 guidelines. The quality of animal studies was evaluated through the SYRCLE's risk of bias tool. RESULTS: A total of 77 studies were sought for retrieval, and 39 studies met the inclusion criteria. According to the synthesis results, most bilayers had a dense barrier layer that prevented connective tissue penetration and a loose osteogenic layer that supported cell migration and osteogenesis. PLGA, PCL, and chitosan were the most common polymers in the barrier layers, while the most utilized polymers in osteogenic layers were PLGA and gelatin. Electrospinning and solvent casting were the most common fabrication methods to design the bilayer structures. Many studies reported higher biological results for bilayers compared to their single layers. Also, fabricated bilayers' in vitro osteogenesis and in vivo new bone formation were significantly superior or at least comparable to the frequently used commercial membranes. CONCLUSION: 1) Bilayers with two distinct layers and different materials, porosities, mechanical properties, and biological behavior can significantly improve heterogeneous bone regeneration; 2) the addition of ceramics and/or drugs to the osteogenic layer enhances the osteogenic properties of the bilayers; 3) fabrication method and pore size of the layers play an important role in determining the mechanical and biological behavior of them.

3D-printed MgO nanoparticle loaded polycaprolactone ß-tricalcium phosphate composite scaffold for bone tissue engineering applications: In-vitro and in-vivo evaluation.

Magnesium (Mg) plays an important role in controlling bone apatite structure and density and is a potential bioactive material in repairing critical-sized bone defects. In this study, we aimed to evaluate the effect of adding NanoMgO to polycaprolactone/beta-tricalcium phosphate (PCL/ß-TCP) scaffolds on bone regeneration. Novel 3D-printed porous PCL/ß-TCP composite scaffolds containing 10% nanoMgO were fabricated by fused deposition modeling (FDM) and compared with PCL/ß-TCP (1:1) scaffolds (control). The morphology and physicochemical properties of the scaffolds were characterized by ATR-FTIR, XRD, scanning electron microscope-energy dispersive X-ray analysis (SEM-EDX), transmission-electron-microscopy (TEM), water contact angle, and compressive strength tests and correlated to its cytocompatibility and osteogenic capacity in-vitro. To evaluate in-vivo osteogenic capacity, bone-marrow-derived stem cell (BMSC)-loaded scaffolds were implanted into 8 mm rat critical-sized calvarial defects for 12 weeks. The hydrophilic scaffolds showed 50% porosity (pore size = 504 µm). MgO nanoparticles (91.5 ± 27.6 nm) were homogenously dispersed and did not adversely affect BMSCs' viability and differentiation. Magnesium significantly increased elastic modulus, pH, and degradation. New bone formation (NBF) in Micro-CT was 30.16 ± 0.31% and 23.56 ± 1.76% in PCL/ß-TCP/nanoMgO scaffolds with and without BMSCs respectively, and 19.38 ± 2.15% and 15.75 ± 2.24% in PCL/ß-TCP scaffolds with and without BMSCs respectively. Angiogenesis was least remarkable in PCL/ß-TCP compared with other groups (p < .05). Our results suggest that the PCL/ß-TCP/nanoMgO scaffold is a more suitable bone substitute compared to PCL/ß-TCP in critical-sized calvarial defects.

Periosteum as a covering vascular flap in posterior mandibular augmentation: A retrospective cohort study.

OBJECTIVE: To analyze the impact of creating periosteal vascular flaps on the amount of bone augmentation following inlay bone grafting (IBG) and cortical autogenous tenting (CAT). MATERIALS AND METHODS: This was a retrospective cohort study enrolling a sample cohort of patients presented to a private clinic in 2015 and 2019 for posterior mandibular ridge augmentation before dental implant placement. The predictor variables were surgical methods: CAT vs. CAT in conjunction with periosteal flap (CATP) vs. IBG vs. IBG in conjunction with periosteal flap (IBGP). The primary outcome variables were supra bundle bone (SBB) superior to the inferior alveolar canal (ΔH) and crestal width difference (ΔW) at a 4-month follow-up. Appropriate statistics were computed at 0.05 significance level. RESULTS: A total of 29 cases (10 males and 19 females) with a mean age of 57.96±7.14 years were included. A total of 33 sites were augmented through CATP, 16 sites through IBGP, 33 sites through CAT, and 11 sites through IBG techniques. All patients healed uneventfully without permanent neurosensory changes, and adequate horizontal (ΔW:3.33±0.71 mm) and vertical (ΔH:5.10±2.04 mm) bone dimensions were restored that allowed implant placement. Using periosteal vascular flaps significantly increased bone augmentation in both vertical and horizontal dimensions (P < 0.01). CONCLUSION: Periosteal vascular flaps can increase the efficacy of mandibular augmentation techniques and decrease post-surgical complications.

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 .

Current Trends, Advances, and Challenges of Tissue Engineering-Based Approaches of Tooth Regeneration: A Review of The Literature.

INTRODUCTION: Tooth loss is a significant health issue. Currently, this situation is often treated with the use of synthetic materials such as implants and prostheses. However, these treatment modalities do not fully meet patients' biological and mechanical needs and have limited longevity. Regenerative medicine focuses on the restoration of patients' natural tissues via tissue engineering techniques instead of rehabilitating with artificial appliances. Therefore, a tissue-engineered tooth regeneration strategy seems like a promising option to treat tooth loss. OBJECTIVE: This review aims to demonstrate recent advances in tooth regeneration strategies and discoveries about underlying mechanisms and pathways of tooth formation. RESULTS AND DISCUSSION: Whole tooth regeneration, tooth root formation, and dentin-pulp organoid generation have been achieved by using different seed cells and various materials for scaffold production. Bioactive agents are critical elements for the induction of cells into odontoblast or ameloblast lineage. Some substantial pathways enrolled in tooth development have been figured out, helping researchers design their experiments more effectively and aligned with the natural process of tooth formation. CONCLUSION: According to current knowledge, tooth regeneration is possible in case of proper selection of stem cells, appropriate design and manufacturing of a biocompatible scaffold, and meticulous application of bioactive agents for odontogenic induction. Understanding innate odontogenesis pathways play a crucial role in accurately planning regenerative therapeutic interventions in order to reproduce teeth.

Evaluating the Preemptive Analgesic Effect of Photo-biomodulation Therapy on Pain Perception During Local Anesthesia Injection in Children: A Split-mouth Triple-blind Randomized Controlled Clinical Trial.

To evaluate the impact of photobiomodulation therapy (PBMT) on injection pain perception and compare it with a topical oral anesthetic gel. A total of 30 patients of 6 to 9 years-old seeking pulpotomy treatment of maxillary secondary primary molars of both sides were considered for this split-mouth triple-blind randomized clinical trial. On one side of the maxilla, the low-level laser (diode laser, 808 nm, 250 mW; 16.25 J; 32.5 J cm-2 ) was irradiated upon the buccal gingiva of the tooth, while a Benzocaine 20% topical anesthetic gel was applied on the other side. A gel with the same taste (strawberry) was applied for the placebo. The Wong-Baker Faces Pain Rating Scale was used to evaluate the injection pain and postoperation pain at two timestamps, 1 h and 24 h after treatment. Patients' heart rate was also evaluated. Paired t, Wilcoxon signed-rank test, McNemar and Friedman tests were used for statistical analyses. Results demonstrated that PBMT could significantly decrease the injection pain perception and heart rate alternations compared to the topical anesthetic gels (P = 0.000). However, no significant differences were documented between the two methods concerning the 1-h (P = 0.26) and 24-h (P = 1.00) postoperation pain. PBMT can be an effective nonpharmacological technique for controlling injection pain.

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.

Identification Osteogenic Signaling Pathways Following Mechanical Stimulation: A Systematic Review.

INTRODUCTION: It has been shown that mechanical forces can induce or promote osteogenic differentiation as well as remodeling of the new created bone tissues. To apply this characteristic in bone tissue engineering, it is important to know which mechanical stimuli through which signaling pathway has a more significant impact on osteogenesis. METHODS: In this systematic study, an electronic search was conducted using PubMed and Google Scholar databases. This study has been prepared and organized according to the preferred reporting items for systematic reviews and meta-analyses (PRISMA) guidelines. Included studies were first categorized according to the in vivo and in vitro studies. RESULTS: Six types of mechanical stresses were used in these articles and the most commonly used mechanical force and cell source were tension and bone marrow-derived mesenchymal stem cells (BMMSCs), respectively. These forces were able to trigger twelve signaling pathways in which Wnt pathway was so prominent. CONCLUSION: 1) Although specific signaling pathways are induced through specific mechanical forces, Wnt signaling pathways are predominantly activated by almost all types of force/stimulation, 2) All signaling pathways regulate expression of RUNX2, which is known as a master regulator of osteogenesis, 3) In Tension force, the mode of force administration, i.e, continuous or noncontinuous tension is more important than the percentage of elongation.

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 .

Cartilage regeneration with dual-drug-releasing injectable hydrogel/microparticle system: In vitro and in vivo study.

In this study, we developed an injectable in situ forming hydrogel/microparticle system consisting of two drugs, melatonin and methylprednisolone, to investigate the capability of the system for chondrogenesis in vitro and in vivo. The chemical, mechanical, and rheological properties of the hydrogel/microparticle were investigated. For in vitro evaluation, the adipose-derived stem cells might be mixed with hydrogel/microparticles, then cellular viability was analyzed by acridine orange/propidium iodide and 4',6-diamidino-2-phenylindole staining and also dimethylmethylene blue assay were conducted to find the amount of proteoglycan. The real-time polymerase chain reaction for aggrecan, sex-determining region Y-Box 9, collagen I (COL1), and COL2 gene expression was performed after 14 and 21 days. For evaluation of cartilage regeneration, the samples were implanted in rabbit knees with cartilaginous experimental defects. Defects were created in both knees of three groups of rabbits. Group 1 was the control with no injection, and Groups 2 and 3 were loaded with hydrogel/cell and hydrogel/microparticle/cell; respectively. Then, after 3 and 6 months, histological evaluations of the defected sites were carried out. The amount of glycosaminoglycans after 14 and 21 days increased significantly in hydrogels/microparticles loaded with cells. The expression of marker genes was also significant in hydrogels/microparticles loaded with cells. According to histology analysis, the hydrogels/microparticles loaded with cells showed the best cartilage regeneration. Overall, our study revealed that the developed injectable hydrogel/microparticle can be used for cartilage regeneration.

Effect of Photobiomodulation on Relapse in an Experimental Rapid Maxillary Expansion Model in Rat.

Rapid maxillary expansion (RME) is performed on transversely deficient maxilla. As all orthodontic treatments, retention is important in maintaining therapeutic outcomes. Fixed /removable retainers are used post-RME causing hygiene and compliance problems. Given photobiomodulation's positive effects on the quantity and quality of bone regeneration, its effect on post-RME relapse was studied. Thirty Sprague Dawley rats were randomly divided into group R, non-irradiated RME-treated (n = 12), group P, irradiated RME-treated (n = 12) and group C, non-RME non-irradiated (n = 6). A 1.5 mm metal ring inserted between maxillary incisors at days 0 and 15 was expanded until 1.5 mm space was obtained at day 30. In group P, Ga-Al-As diode laser (810 nm, 100 mW, 4J/cm2 , 30 secs) was applied on days 0, 2, 4, 6, 8, 10, 12 and 14 as predictor variable. The relapse was measured as the space lost between incisors for 30 days after appliance removal (primary outcome variable) and compared with t-test. In week 2, space loss in group P was significantly lower (P < 0.05) than all other groups. The relapse during weeks 2 and 3 was significantly lower in group P than group R. However, no significant difference in relapse amount was found between groups during first and fourth week. There was a significant difference (P < 0.05) between groups in relapse rates (secondary outcome variable) but not in total relapse after 4 weeks. Photobiomodulation proved beneficial in resisting relapse in our study, and it is suggested to be continued until the end of expansion.

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.