Bone Regeneration In Rats Research Articles

Programmable editing of primary MicroRNA switches stem cell differentiation and improves tissue regeneration

Programmable RNA editing is harnessed for modifying mRNA. Besides mRNA, miRNA also regulates numerous biological activities, but current RNA editors have yet to be exploited for miRNA manipulation. To engineer primary miRNA (pri-miRNA), the miRNA precursor, we present a customizable editor REPRESS (RNA Editing of Pri-miRNA for Efficient Suppression of miRNA) and characterize critical parameters. The optimized REPRESS is distinct from other mRNA editing tools in design rationale, hence enabling editing of pri-miRNAs that are not editable by other RNA editing systems. We edit various pri-miRNAs in different cells including adipose-derived stem cells (ASCs), hence attenuating mature miRNA levels without disturbing host gene expression. We further develop an improved REPRESS (iREPRESS) that enhances and prolongs pri-miR-21 editing for at least 10 days, with minimal perturbation of transcriptome and miRNAome. iREPRESS reprograms ASCs differentiation, promotes in vitro cartilage formation and augments calvarial bone regeneration in rats, thus implicating its potentials for engineering miRNA and applications such as stem cell reprogramming and tissue regeneration.

Bone Regeneration of Rat Critical-Sized Calvarial Defects by the Combination of Photobiomodulation and Adipose-Derived Mesenchymal Stem Cells.

Introduction: This study explored the synergistic effects of low-level laser therapy (LLLT) and adipose-derived stem cells (ADSCs) on cranial bone regeneration in rats, addressing the limitations of autogenous grafts and advancing bone tissue engineering with innovative photobiomodulation (PBM) applications. Methods: Sixty Wistar rats were allocated to 5 separate groups randomly; (1) natural bovine bone mineral (NBBM); (2) NBBM+LLLT; (3) NBBM+allogenic ADSCs; (4) NBBM+allogenic ADSCs+LLLT; (5) Only defects. 8-mm calvarial defects were made in each rat in the surgical procedure. A diode laser was applied with the following parameters (continuous mode, power of 100mW, wavelength of 808nm, and 4 J/cm2 energy density) immediately after the procedure and every other day. Bone samples were obtained and assessed histomorphometrically and histologically after staining with hematoxylin and eosin (H&E). Results: Different volumes of bony material were observed in two weeks; 2.94%±1.00 in group 1, 5.1%±1.92 in group 2, 7.11%±2.82 in group 3, 7.34%±2.31 in group 4, and 2.01%±0.83 in group 5 (P<0.05). On the other hand, foreign body residuals were up by 23% in the groups with scaffolding by the end of 2 weeks. Four weeks of observation led to 6.74 %±1.95, 13.24%±1.98, 15.76%±1.19, 15.92%±3.4, and 3.11%±1.00 bone formation in groups 1 to 5, respectively (P<0.05). Generally, the difference between groups 2-4 was not statistically significant based on different types of bone and the extent of inflammation. Conclusion: Bearing in mind the limitations of our research, it was demonstrated that ADSCs in combination with PBM have promising effects on bone tissue regeneration in sizeable bony defects. Furthermore, this study also showed that PBM usage improved the newly regenerated bone quality.

IMPROVEMENT OF REPARATIVE REGENERATION OF THE JAW BONE IN EXPERIMENT

We conducted an experimental study on the effectiveness of the combined use of osteoplastic material Collapan-L with adhered MSCs to improve the processes of jaw bone regeneration in rats. The animals were divided into two groups: experimental and control groups of 50 animals each. Bone damage was induced in rats by drilling a defect with a dental bur in the area of attachment of the masticatory muscles in the angle of the mandible. We used only Collapan-L in the control group, and Collapan-L with adherent MSCs in the experimental group. During the cultivation process, statistically significant changes in concentration, depending on the day of cultivation, were observed for all cytokines: IL-1β (p&lt;0.001 ANOVA), IL-2 (p=0.007 ANOVA), IL-4 (p=0.02 ANOVA), IL-6 (p=0.003 ANOVA), IL-8 (p=0.003 by Kruskal-Wallis test) and TNF-α (p&lt;0.001 ANOVA). The dynamics of changes in the cytokines had a wave-like character. By the day 30 of the experiment, in animals with injected Collapan-L, the total volume of bone tissue in the regeneration zone was 50.14%, and in the Collapan-L + MSCs group it was 75.4%. The use of osteoplastic material Collapan-L with adhered MSCs is an effective combination for improving regeneration processes and reducing the inflammatory response of bone tissue.

Cross-linking N-succinyl chitosan-oxidated hyaluronic acid-based hydrogel loaded with bone marrow mesenchymal stem cell-derived exosomes induce bone regeneration in cranial defects

BackgroundHydrogel carrying exosomes shows better prospects for bone regeneration. ObjectiveThis work investigated the effect of cross-linking N-succinyl chitosan-oxidated hyaluronic acid-based hydrogel loaded with bone marrow mesenchymal stem cell-derived exosomes on bone regeneration in cranial defects. MethodsCross-linking N-succinyl chitosan-oxidated hyaluronic acid-based hydrogel loaded with bone marrow mesenchymal stem cell-derived exosomes was prepared, and utilized to treat bone marrow mesenchymal stem cells, human umbilical vein endothelial cells, and rats with cranial defects. Logical experiments were implemented to research the effect of the prepared hydrogel on osteogenesis and angiogenesis of cells in vitro, and on bone regeneration in rats with cranial defects. ResultsCross-linking N-succinyl chitosan-oxidated hyaluronic acid-based hydrogel loaded with bone marrow mesenchymal stem cell-derived exosomes showed good biocompatibility and self-healing property. It promoted osteogenic differentiation of bone marrow mesenchymal stem cells and angiogenesis of human umbilical vein endothelial cells, and facilitated osteogenic differentiation, angiogenesis and bone regeneration in rats with cranial defects. ConclusionCross-linking N-succinyl chitosan-oxidated hyaluronic acid-based hydrogel loaded with bone marrow mesenchymal stem cell-derived exosomes facilitated osteogenesis, angiogenesis and bone regeneration in rats with cranial defects. It might be an effective strategy for inducing bone regeneration in individuals with cranial defects.

Tricarboxylic Acid Cycle Metabolite-Coordinated Biohydrogels Augment Cranial Bone Regeneration Through Neutrophil-Stimulated Mesenchymal Stem Cell Recruitment and Histone Acetylation-Mediated Osteogenesis.

Cranial bone defects remain a major clinical challenge, increasing patients' life burdens. Tricarboxylic acid (TCA) cycle metabolites play crucial roles in facilitating bone tissue regeneration. However, the development of TCA cycle metabolite-modified biomimetic grafts for skull bone regeneration still needs to be improved. The mechanism underlying the release of TCA cycle metabolites from biomaterials in regulating immune responses and mesenchymal stem cell (MSC) fate (migration and differentiation) remains unknown. Herein, this work constructs biomimetic hydrogels composed of gelatin and chitosan networks covalently cross-linked by genipin (CGG hydrogels). A series of TCA cycle metabolite-coordinated CGG hydrogels with strong mechanical and antiswelling performances are subsequently developed. Remarkably, the citrate (Na3Cit, Cit)-coordinated CGG hydrogels (CGG-Cit hydrogels) with the highest mechanical modulus and strength significantly promote skull bone regeneration in rat and murine cranial defects. Mechanistically, using a transgenic mouse model, bulk RNA sequencing, and single-cell RNA sequencing, this work demonstrates that CGG-Cit hydrogels promote Gli1+ MSC migration via neutrophil-secreted oncostatin M. Results also indicate that citrate improves osteogenesis via enhanced histone H3K9 acetylation on osteogenic master genes. Taken together, the immune microenvironment- and MSC fate-regulated CGG-Cit hydrogels represent a highly efficient and facile approach toward skull bone tissue regeneration with great potential for bench-to-bedside translation.

Effects of Photobiomodulation Using Low-Power Diode Laser Therapy and Nano-bone on Mandibular Bone Regeneration in Rats.

Introduction: Recently, the positive effects of photobiomodulation (PBM) and nano-bone on bone regeneration have garnered significant attention. The purpose of the research was to assess the impact of PBM and nano-bone on the process of mandibular bone repair in mice. Methods: A 4-mm diameter bone defect was created in the left mandibular angle of 24 mice separated into 4 equal groups: group I: control; group II: PBM by irradiation at 100 mW of a 980 nm diode laser for one minute (three sessions per week; day on and day off); group III: nano-bone; group IV: PBM with nano-bone. Every group was sectioned into 3 equal subgroups corresponding to the evaluation method period: (A) one week, (B) two weeks, and (C) four weeks. Histological examination was done with hematoxylin, eosin, and Masson's Trichrome after one, two and four weeks for inflammation, bone defect coverage, vascularization within the newly formed bone, and new bone formation. Statistical analysis of the data was done and presented as percentage values using chi-square. The significance level was set at P value≤0.05 within all tests. Results: In general, by histological examination of the mandibular bone defect of the rats, the intensity of inflammation was the least in group IV when compared with groups II and III and the control group at all evaluation periods (P<0.001). Also, group IV showed a high significant rise in the percentage of new bone formation following four weeks when compared with the control (P≤ 0.001) and groups II and III (P<0.001). Conclusion: The present research results confirmed that the combination of PBM and nano-bone can aid in the repair of mandibular bone abnormalities. This animal study suggests that the use of PBM and nano-bone should be investigated further in clinical studies.

Biomimetic composite hydrogel promotes new bone formation in rat bone defects through regulation of miR-19b-3p/WWP1 axis by loaded extracellular vesicles

ObjectiveThis study aims to investigate the mechanism by which biomimetic composite hydrogels loaded with bone marrow mesenchymal stem cells (BMSCs) derived microRNA-19b-3p/WWP1 axis through extracellular vesicles (EVs) affect the new bone formation in rat bone defects.MethodsFirst, synthesize the bionic composite hydrogel Gel-OCS/MBGN. Characterize it through field-emission scanning electron microscopy (FE-SEM), X-ray diffraction (XRD), and FTIR. Then, conduct performance tests such as rheology, dynamic mechanical analysis, in vitro mineralization, and degradation. Rat BMSCs were selected for in vitro cell experiments, and EVs derived from BMSCs were obtained by differential centrifugation. The EVs were loaded onto Gel-OCS/MBGN to obtain Gel-OCS/MBGN@EVs hydrogel. Cell viability and proliferation were detected by live/dead cell staining and CCK-8 assay, respectively. ALP and ARS staining was used to evaluate the osteogenic differentiation of BMSCs. Differential gene expression analysis of osteogenic differentiation was performed using high-throughput sequencing. TargetScan database predicted the binding site between miR-19b-3p and WWP1, and a dual-luciferase reporter assay was performed to confirm the targeting binding site. A rat bone defect model was established, and new bone formation was evaluated by Micro-CT, H&E staining, and Masson's trichrome staining. Immunofluorescence staining and immunohistochemistry were used to detect the expression levels of osteogenic-related factors in rat BMSCs. RT-qPCR and Western blot were used to detect the expression levels of genes and proteins in tissues and cells.ResultGel-OCS/MBGN was successfully constructed and loaded with EVs, resulting in Gel-OCS/MBGN@EVs. The in vitro drug release experiment results show that Gel-OCS/MBGN could sustainably release EVs. Further experiments have shown that Gel-OCS/MBGN@EVs could significantly promote the differentiation of BMSCs into osteoblasts. Experiments have shown that WWP1 is a key factor in osteogenic differentiation and is regulated by miR-19b-3p. EVs promote osteogenic differentiation by suppressing WWP1 expression through the transmission of miR-19b-3p. In vivo animal experiments have demonstrated that Gel-OCS/MBGN@EVs significantly promote bone repair in rats with bone defects by regulating the miR-19b-3p/WWP1 signaling axis.ConclusionFunctional Gel-OCS/MBGN@EVs were obtained by constructing Gel-OCS/MBGN and loading EVs onto it. EVs could deliver miR-19b-3p to BMSCs, inhibit the expression of WWP1, and promote the osteogenic differentiation of BMSCs, ultimately promoting bone regeneration in rats with bone defects.Graphical

Exosomal mRNAs for Angiogenic-Osteogenic Coupled Bone Repair.

Regenerative medicine in tissue engineering often relies on stem cells and specific growth factors at a supraphysiological dose. These approaches are costly and may cause severe side effects. Herein, therapeutic small extracellular vesicles (t-sEVs) endogenously loaded with a cocktail of human vascular endothelial growth factor A (VEGF-A) and human bone morphogenetic protein 2 (BMP-2) mRNAs within a customized injectable PEGylated poly (glycerol sebacate) acrylate (PEGS-A) hydrogel for bone regeneration in rats with challenging femur critical-size defects are introduced. Abundant t-sEVs are produced by a facile cellular nanoelectroporation system based on a commercially available track-etched membrane (TM-nanoEP) to deliver plasmid DNAs to human adipose-derived mesenchymal stem cells (hAdMSCs). Upregulated microRNAs associated with the therapeutic mRNAs are enriched in t-sEVs for enhanced angiogenic-osteogenic regeneration. Localized and controlled release of t-sEVs within the PEGS-A hydrogel leads to the retention of therapeutics in the defect site for highly efficient bone regeneration with minimal low accumulation in other organs.

Reduced Bone Regeneration in Rats With Type 2 Diabetes Mellitus as a Result of Impaired Stromal Cell and Osteoblast Function-A Computer Modeling Study.

Bone has the fascinating ability to self-regenerate. However, under certain conditions, such as type 2 diabetes mellitus (T2DM), this ability is impaired. T2DM is a chronic metabolic disease known by the presence of elevated blood glucose levels that is associated with reduced bone regeneration capability, high fracture risk, and eventual non-union risk after a fracture. Several mechanical and biological factors relevant to bone regeneration have been shown to be affected in a diabetic environment. However, whether impaired bone regeneration in T2DM can be explained due to mechanical or biological alterations remains unknown. To elucidate the relevance of either one, the aim of this study was to investigate the relative contribution of T2DM-related alterations on either cellular activity or mechanical stimuli driving bone regeneration. A previously validated in silico computer modeling approach that was capable of explaining bone regeneration in uneventful conditions of healing was further developed to investigate bone regeneration in T2DM. Aspects analyzed included the presence of mesenchymal stromal cells (MSCs), cellular migration, proliferation, differentiation, apoptosis, and cellular mechanosensitivity. To further verify the computer model findings against in vivo data, an experimental setup was replicated, in which regeneration was compared in healthy and diabetic after a rat femur bone osteotomy stabilized with plate fixation. We found that mechanical alterations had little effect on the reduced bone regeneration in T2DM and that alterations in MSC proliferation, MSC migration, and osteoblast differentiation had the highest effect. In silico predictions of regenerated bone in T2DM matched qualitatively and quantitatively those from ex vivo μCT at 12 weeks post-surgery when reduced cellular activities reported in previous in vitro and in vivo studies were included in the model. The presented findings here could have clinical implications in the treatment of bone fractures in patients with T2DM. © 2023 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Injectable periodontal ligament stem cell-metformin-calcium phosphate scaffold for bone regeneration and vascularization in rats.

Injectable and self-setting calcium phosphate cement scaffold (CPC) capable of encapsulating and delivering stem cells and bioactive agents would be highly beneficial for dental and craniofacial repairs. The objectives of this study were to: (1) develop a novel injectable CPC scaffold encapsulating human periodontal ligament stem cells (hPDLSCs) and metformin (Met) for bone engineering; (2) test bone regeneration efficacy in vitro and in vivo. hPDLSCs were encapsulated in degradable alginate fibers, which were then mixed into CPC paste. Five groups were tested: (1) CPC control; (2) CPC +hPDLSC-fibers +0% Met (CPC + hPDLSCs + 0%Met); (3) CPC +hPDLSC-fibers +0.1% Met (CPC + hPDLSCs + 0.1%Met); (4) CPC +hPDLSC-fibers +0.2% Met (CPC + hPDLSCs + 0.2%Met); (5) CPC +hPDLSC-fibers +0.4% Met (CPC + hPDLSCs + 0.4%Met). The injectability, mechanical properties, metformin release, and hPDLSC osteogenic differentiation and bone mineral were determined in vitro. A rat cranial defect model was used to evaluate new bone formation. The novel construct had good injectability and physical properties. Alginate fibers degraded in 7 days and released hPDLSCs, with 5-fold increase of proliferation (p＜0.05). The ALP activity and mineral synthesis of hPDLSCs were increased by Met delivery (p＜0.05). Among all groups, CPC+hPDLSCs+0.1%Met showed the greatest cell mineralization and osteogenesis, which were 1.5-10folds those without Met (p＜0.05). Compared to CPC control, CPC+hPDLSCs+0.1%Met enhanced bone regeneration in rats by 9 folds, and increased vascularization by 3 folds (p＜0.05). The novel injectable construct with hPDLSC and Met encapsulation demonstrated excellent efficacy for bone regeneration and vascularization in vivo in an animal model. CPC+hPDLSCs+0.1%Met is highly promising for dental and craniofacial applications.

The effect of green synthesis of TiO2 nanoparticles/collagen/HA scaffold in bone regeneration: As an animal study.

The bone defects cannot heal by themselves when their range exceeds the critical size defect (CSD). In clinical treatment, significant bone defects are often caused by trauma, developmental deformity, tumour resection and infection. The purpose of this study was to investigate the effect of green synthesis of TiO2 from propolis extract/collagen/HA (Hydroxyapatite) scaffolds on bone regeneration in rats. Water uptake, biodegradability, porosity and biodegradation of the scaffolds were evaluated after they were synthesised using freeze-dry method. Cell viability by MTT assay was then evaluated. During the 4, 8 and 12 weeks following the scaffold implantation, the bone regeneration was evaluated using macroscopic and microscopic tests to determine the effectiveness of green synthesis of TiO2 from propolis extract/collagen/HA scaffolds. Compared to the HA/Coll scaffold, ProTiO2 /HA/Coll scaffold was reduced porosity, water absorption and degradability porosity. Based on in vitro tests, both synthetic scaffolds induced cell growth and were less toxic and stimulated cell growth. Based on histopathological testing, the ProTiO2 /HA/Coll scaffolds formed high levels of bone during 12 weeks in comparison with HA/Coll and control group. ProTiO2 /HA/Coll composite can be used in regenerative medicine, bone fillers and scaffolds. As a result, this research suggests that ProTiO2 /HA/Coll composites could be promising candidates for bone regeneration.

Melatonin Engineering M2 Macrophage-Derived Exosomes Mediate Endoplasmic Reticulum Stress and Immune Reprogramming for Periodontitis Therapy.

Periodontitis is a chronic infectious disease caused by bacterial irritation. As an essential component of the host immunity, macrophages are highly plastic and play a crucial role in inflammatory response. An appropriate and timely transition from proinflammatory (M1) to anti-inflammatory (M2) macrophages is indispensable for treating periodontitis. As M2 macrophage-derived exosomes (M2-exos) can actively target inflammatory sites and modulate immune microenvironments, M2-exos can effectively treat periodontitis. Excessive endoplasmic reticulum stress (ER stress) and unfolded protein response (UPR) are highly destructive pathological characteristics during inflammatory periodontal bone loss. Although melatonin has antioxidant and anti-inflammatory effects, studies focusing on melatonin ER stress modulation remain limited. This study fabricates engineered M2-exos loading with melatonin (Mel@M2-exos) for treating periodontitis. As a result, M2-exos drive an appropriate and timely macrophage reprogramming from M1 to M2 type, which resolves chronic inflammation and accelerated periodontal healing. Melatonin released from Mel@M2-exos rescues the osteogenic and cementogenic differentiation capacity in inflammatory human periodontal ligament cells (hPDLCs) by reducing excessive ER stress and UPR. Injectable gelatin methacryloyl (GelMA) hydrogels with sustained-release Mel@M2-exos accelerate periodontal bone regeneration in rats with ligation-induced periodontitis. Taken together, melatonin engineering M2 macrophage-derived exosomes are promising candidates for inflammatory periodontal tissue regeneration.

PF127 Hydrogel-Based Delivery of Exosomal CTNNB1 from Mesenchymal Stem Cells Induces Osteogenic Differentiation during the Repair of Alveolar Bone Defects.

Pluronic F127 (PF127) hydrogel has been highlighted as a promising biomaterial for bone regeneration, but the specific molecular mechanism remains largely unknown. Herein, we addressed this issue in a temperature-responsive PF127 hydrogel loaded with bone marrow mesenchymal stem cells (BMSCs)-derived exosomes (Exos) (PF127 hydrogel@BMSC-Exos) during alveolar bone regeneration. Genes enriched in BMSC-Exos and upregulated during the osteogenic differentiation of BMSCs and their downstream regulators were predicted by bioinformatics analyses. CTNNB1 was predicted to be the key gene of BMSC-Exos in the osteogenic differentiation of BMSCs, during which miR-146a-5p, IRAK1, and TRAF6 might be the downstream factors. Osteogenic differentiation was induced in BMSCs, in which ectopic expression of CTNNB1 was introduced and from which Exos were isolated. The CTNNB1-enriched PF127 hydrogel@BMSC-Exos were constructed and implanted into in vivo rat models of alveolar bone defects. In vitro experiment data showed that PF127 hydrogel@BMSC-Exos efficiently delivered CTNNB1 to BMSCs, which subsequently promoted the osteogenic differentiation of BMSCs, as evidenced by enhanced ALP staining intensity and activity, extracellular matrix mineralization (p < 0.05), and upregulated RUNX2 and OCN expression (p < 0.05). Functional experiments were conducted to examine the relationships among CTNNB1, microRNA (miR)-146a-5p, and IRAK1 and TRAF6. Mechanistically, CTNNB1 activated miR-146a-5p transcription to downregulate IRAK1 and TRAF6 (p < 0.05), which induced the osteogenic differentiation of BMSCs and facilitated alveolar bone regeneration in rats (increased new bone formation and elevated BV/TV ratio and BMD, all with p < 0.05). Collectively, CTNNB1-containing PF127 hydrogel@BMSC-Exos promote the osteogenic differentiation of BMSCs by regulating the miR-146a-5p/IRAK1/TRAF6 axis, thus inducing the repair of alveolar bone defects in rats.

Different Species of Marine Sponges Diverge in Osteogenic Potential When Therapeutically Applied as Natural Scaffolds for Bone Regeneration in Rats.

A highly porous structure, and an inorganic (biosilica) and collagen-like organic content (spongin) makes marine sponges potential candidates to be used as natural scaffolds in bone tissue engineering. The aim of this study was to characterize (through SEM, FTIR, EDS, XRD, pH, mass degradation and porosity tests) scaffolds produced from two species of marine sponges, Dragmacidon reticulatum (DR) and Amphimedon viridis (AV), and to evaluate the osteogenic potential of these scaffolds by using a bone defect model in rats. First, it was shown that the same chemical composition and porosity (84 ± 5% for DR and 90 ± 2% for AV) occurs among scaffolds from the two species. Higher material degradation was observed in the scaffolds of the DR group, with a greater loss of organic matter after incubation. Later, scaffolds from both species were surgically introduced in rat tibial defects, and histopathological analysis after 15 days showed the presence of neo-formed bone and osteoid tissue within the bone defect in DR, always around the silica spicules. In turn, AV exhibited a fibrous capsule around the lesion (19.9 ± 17.1%), no formation of bone tissue and only a small amount of osteoid tissue. The results showed that scaffolds manufactured from Dragmacidon reticulatum presented a more suitable structure for stimulation of osteoid tissue formation when compared to Amphimedon viridis marine sponge species.

The Promoting Effect of a Local Pulsatile Parathyroid Hormone Delivery on Healing of the Mandibular Fracture in Rats.

Parathyroid hormone (PTH) can promote bone formation and mineralization in mandibular fractures, and is systemically administered through daily injections. In this study, the local delivery of PTH using carboxymethyl chitosan/polyvinyl alcohol and alginate was investigated. Bovine serum albumin was used as a drug substitute, and the delivery system was verified to release drugs in a pulsed rhythm. After the delivery system was subcutaneously implanted in Sprague-Dawley (SD) rats, no rejection reaction was detected, indicating that it has good biocompatibility and biodegradability in vivo. Then, an SD rat model of mandibular fracture was established, and 24 rats were randomly divided into two groups. The control group was reduced and fixed with screws and a microplate, and the experimental group received pulsatile PTH release system (14 μg PTH) + screws and microplate fixation. The animals were euthanized on postoperative weeks 1-4. Observation of gross specimens, digital radiography, and hematoxylin and eosin showed that the local PTH pulsatile release system promoted osteogenesis and accelerated fracture healing. In summary, PTH can be loaded by biomaterials to locally target the fracture and stimulate bone formation. Moreover, the pulsatile PTH release system provides a potential therapeutic protocol for mandibular fracture. Impact statement Our study prepares a drug release system that could impulsively release parathyroid hormone. The system could enhance bone regeneration in rats with mandibular fracture. These data provide a foundation for future studies aimed to understand and optimize the use of bioactive molecule pulsatile delivery for bone regeneration and tissue engineering applications.

Angiogenetic and anti-inflammatory effects of photobiomodulation on bone regeneration in rat: A histopathological, immunohistochemical, and molecular analysis.

Post-surgical bone defects require new alternative approaches for a better healing process. For this matter, photobiomodulation therapy (PBMT) has been used in order to improve the process of healing, pain, and inflammation reduction and tissue rejuvenation. This study is set to evaluate the effect of PBMT on angiogenic and inflammatory factors for bone regeneration in rat post-surgical cranial defects. Thirty male Wistar rats were distributed accidentally into two groups (Subdivided into 3 groups according to their follow-up durations). During operation, an 8-mm critical-sized calvarial defect was made in each rat. A continuous diode laser was used (power density 100mW/cm2, wavelength 810nm, the energy density of 4J/cm2). Bone samples were assessed histomorphometrically and histologically after hematoxylin and eosin (H&E) staining. ALP, PTGIR, OCN, and IL-1 levels were measured by RT-PCR. VEGF expression was studied by immunohistochemistry analysis. The level of IL-1 expression decreased significantly in the PBMT group compared to the control after 7days (p<0.05), while, the PTGIR level was improved significantly compared to the control group after 7days. Furthermore, levels of OCN and ALP improved after PBM use; however, the alterations were not statistically meaningful (p>0.05). Evaluation with IHC displayed a significant rise in VEGF expression after 3days in the PBMT group compared to the control (p>0.05). In this study's conditions, the results showed a meaningful alteration in osteogenic, inflammatory, and angiogenic mediators in post-surgical calvarial defect following PBMT. It appears that PBM can accelerate angiogenesis in the bone healing procedure which can be helpful in bone tissue engineering.

Performance of Polydioxanone-Based Membrane in Association with 3D-Printed Bioceramic Scaffolds in Bone Regeneration

This study evaluated the bioactivity of 3D-printed β-tricalcium phosphate (β-TCP) scaffolds or hydroxyapatite (HA) scaffolds associated with polydioxanone (PDO) membrane (Plenum® Guide) for guided bone regeneration in rats. Fifty-four rats were divided into three groups (n = 18 animals): autogenous bone + PDO membrane (Auto/PG); 3D-printed β-TCP + PDO membrane (TCP/PG); and 3D-printed HA + PDO membrane (HA/PG). A surgical defect in the parietal bone was made and filled with the respective scaffolds and PDO membrane. The animals were euthanized 7, 30, and 60 days after the surgical procedure for micro-CT, histomorphometric, and immunolabeling analyses. Micro-CT showed an increase in trabecular thickness and a decrease in trabecular separation, even with similar bone volume percentages between TCP/PG and HA/PG vs. Auto/PG. Histometric analysis showed increased bone formation at 30 days in the groups compared to 7 days postoperatively. Immunolabeling analysis showed an increase in proteins related to bone formation at 30 days, and both groups showed a similar immunolabeling pattern. This study concludes that 3D-printed scaffolds associated with PDO membrane (Plenum® Guide) present similar results to autogenous bone for bone regeneration.

An altered oral microbiota induced by injections of superparamagnetic iron oxide nanoparticle-labeled periodontal ligament stem cells helps periodontal bone regeneration in rats.

Stem cell injection is good for periodontal regeneration due to the capacity of stem cells to differentiate toward osteogenic direction and to regulate the production of pro- and anti-inflammatory cytokines. However, injected cells are difficult to track in vivo. And there is microbiota in oral cavity, the dysbiosis of which leads to the damage and loss of periodontal tissue. Here, we demonstrated an enhanced periodontal repair was due to an altered oral microbiota. Periodontal defects were surgically prepared in rats, and periodontal ligament stem cells (PDLSCs) labeled by superparamagnetic iron oxide (SPIO) nanoparticles (PC-SPIO) were injected, with PDLSCs and saline treatments as controls. Detected by magnetic resonance imaging (MRI) and histological staining, PC-SPIO was major at limited areas in regenerated periodontal tissues. PC-SPIO-treated rats achieved better periodontal regeneration than the other two groups. Concurrently, the oral microbiota of PC-SPIO-treated rats was changed, presenting SPIO-Lac as a biomarker. SPIO-Lac assisted periodontal repair in vivo, inhibited the inflammation of macrophages induced by lipopolysaccharide (LPS) and antibacterial in vitro. Therefore, our study proved that SPIO-labeled cells can be tracked in periodontal defect and highlighted a potential positive role of an oral microbiota in periodontal regeneration, suggesting the possibility of periodontal repair promotion by manipulating oral microbiota.

Tracking of Stem Cells from Human Exfoliated Deciduous Teeth Labeled with Molday ION Rhodamine-B during Periodontal Bone Regeneration in Rats.

Chronic periodontitis can lead to alveolar bone resorption and eventually tooth loss. Stem cells from exfoliated deciduous teeth (SHED) are appropriate bone regeneration seed cells. To track the survival, migration, and differentiation of the transplanted SHED, we used super paramagnetic iron oxide particles (SPIO) Molday ION Rhodamine-B (MIRB) to label and monitor the transplanted cells while repairing periodontal bone defects. We determined an appropriate dose of MIRB for labeling SHED by examining the growth and osteogenic differentiation of labeled SHED. Finally, SHED was labeled with 25 μg Fe/ml MIRB before being transplanted into rats. Magnetic resonance imaging was used to track SHED survival and migration in vivo due to a low-intensity signal artifact caused by MIRB. HE and immunohistochemical analyses revealed that both MIRB-labeled and unlabeled SHED could promote periodontal bone regeneration. The colocalization of hNUC and MIRB demonstrated that SHED transplanted into rats could survive in vivo. Furthermore, some MIRB-positive cells expressed the osteoblast and osteocyte markers OCN and DMP1, respectively. Enzyme-linked immunosorbent assay revealed that SHED could secrete protein factors, such as IGF-1, OCN, ALP, IL-4, VEGF, and bFGF, which promote bone regeneration. Immunofluorescence staining revealed that the transplanted SHED was surrounded by a large number of host-derived Runx2- and Col II-positive cells that played important roles in the bone healing process. SHED could promote periodontal bone regeneration in rats, and the survival of SHED could be tracked in vivo by labeling them with MIRB. SHED are likely to promote bone healing through both direct differentiation and paracrine mechanisms.

Evaluation of the effects of Ankaferd haemostat application on bone regeneration in rats with calvarial defects: histochemical, immunohistochemical and scintigraphic study.

Bone wax, a haemostatic agent, is widely used in craniospinal surgical procedures for a long time, in spite of controversial results regarding its negative influence upon bone regeneration. In this experimental study, the effects of Ankaferd Blood Stopper (ABS), as an alternative haemostatic agent, were evaluated through histochemical, immunohistochemical and scintigraphic studies. The total of 30 adult female Wistar albino rats was randomly divided into three groups: intact control group (n = 10), bone wax group (n = 10), and ABS group (n = 10). Surgically, a 3.0 mm hole in diameter was drilled on the right side of calvarium of the rats using a Class Mini Grinder set in all three groups, as described previously. At the end of 8 weeks, bone healing and connective tissue alterations surrounding drilled calvarial defect areas of the rats were determined via haematoxylin and eosin and the Mallory's trichrome staining and anti-bone sialoprotein immunohistochemistry. Image Pro Express 4.5 programme was used for histomorphometric calculation of new bone and fibrotic tissue areas. All statistical analyses were made with SPSS 25.0 and analysis of variance (one-way ANOVA) followed by Bonferroni post hoc test was performed, p < 0.001 was considered as significance level. Histomorphometrically, it was found that he had the largest hole diameter and the least fibrotic scar area in the bone-wax group. In the bone wax group, it was observed that the material closed the hole and there was only a fibrotic scar tissue in the area between the bone tissue at the edge of the hole and bone wax, and a fibrotic tissue was formed in the bone wax area. During the histological procedure, this bone-wax material was poured and the sections were seen as a gap in this area. In the ABS haemostat group, the smallest hole diameter and the least fibrotic scar tissue were observed. Fibrotic scar tissue close to each other was found in the ABS haemostat and bone wax groups. Histological analysis of samples also showed a statistical significance for fibrotic connective tissue area between groups (p < 0.05). Scintigraphically, osteoblastic activity related to blood flow in the animal taken from the group with application of ABS haemostat was more pronounced compared to the other two groups. In our study, it has been concluded that the ABS yields affirmative effects on the bone healing, while bone wax leads to negative impact on the bone regeneration. Scintigraphic, histochemical and immunohistochemical data support the affirmative impact of the ABS haemostat application upon the bone regeneration apart from the quick stop of haemorrhage.