Assessment of the Bone Healing Process Mediated by Periosteum-Derived Mesenchymal Stem Cells' Secretome and a Xenogenic Bioceramic-An In Vivo Study in the Rabbit Critical Size Calvarial Defect Model.
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ABSTRACT:
SUBMITTER: Pranskunas M
PROVIDER: S-EPMC8269548 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Hedgehog (Hh) signaling positively regulates both endochondral and intramembranous ossification. Use of small molecules for tissue engineering applications poses several advantages. In this study, we examined whether use of an acellular scaffold treated with the small molecule Smoothened agonist (SAG) could aid in critical-size mouse calvarial defect repair. First, we verified the pro-osteogenic effect of SAG in vitro, using primary neonatal mouse calvarial cells (NMCCs). Next, a 4?mm nonhealing defect was created in the mid-parietal bone of 10-week-old CD-1 mice. The scaffold consisted of a custom-fabricated poly(lactic-co-glycolic acid) disc with hydroxyapatite coating (measuring 4?mm diameter?×?0.5?mm thickness). Treatment groups included dimethylsulfoxide control (n?=?6), 0.5?mM SAG (n?=?7) or 1.0?mM SAG (n?=?7). Evaluation was performed at 4 and 8 weeks postoperative, by a combination of high-resolution microcomputed tomography, histology (H & E, Masson's Trichrome), histomorphometry, and immunohistochemistry (BSP, OCN, VEGF). In vivo results showed that SAG treatment induced a significant and dose-dependent increase in calvarial bone healing by all radiographic parameters. Histomorphometric analysis showed an increase in all parameters of bone formation with SAG treatment, but also an increase in blood vessel number and density. In summary, SAG is a pro-osteogenic, provasculogenic stimulus when applied locally in a bone defect environment.
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Project description:Injectable hydrogels have long been gaining attention in the bone tissue engineering field owing to their ability to mix homogeneously with cells and therapeutic agents, minimally invasive administration, and seamless defect filling. Despite the advantages, the use of injectable hydrogels as cell delivery carriers is currently limited by the challenge of mimicking the natural microenvironment of the loaded cells, promoting cell proliferation, and enhancing bone regeneration. To overcome these problems, we aimed to develop an injectable and in situ-forming nanocomposite hydrogel composed of gelatin, alginate, and LAPONITE® to mimic the architecture and composition of the extracellular matrix. The encapsulated rat bone marrow mesenchymal stem cells (rBMSCs) survived in the nanocomposite hydrogel, and the gel promoted cell proliferation in vitro. Systematic in vivo research of the biomimetic hydrogel with or without cells was conducted in a critical-size (8 mm) rat bone defect model. The in vivo results proved that the hydrogel loaded with rBMSCs significantly promoted bone healing in rat calvarial defects, compared to the hydrogel without cells, and that the hydrogel did not provoked side effects on the recipients. Given these advantageous properties, the developed cell-loaded injectable nanocomposite hydrogel can greatly accelerate the bone healing in critical bone defects, thus providing a clinical potential candidate for orthopedic applications.
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Project description:Peripheral blood mesenchymal stem cells (PBMSCs) may be easily harvested from patients, permitting autologous grafts for bone tissue engineering in the future. However, the PBMSC's capabilities of survival, osteogenesis and production of new bone matrix in the defect area are still unclear. Herein, PBMSCs were seeded into a nanofiber scaffold of self-assembling peptide (SAP) and cultured in osteogenic medium. The results indicated SAP can serve as a promising scaffold for PBMSCs survival and osteogenic differentiation in 3D conditions. Furthermore, the SAP seeded with the induced PBMSCs was splinted by two membranes of poly(lactic)-glycolic acid (PLGA) to fabricate a composited scaffold which was then used to repair a critical-size calvarial bone defect model in rat. Twelve weeks later the defect healing and mineralization were assessed by H&E staining and microcomputerized tomography (micro-CT). The osteogenesis and new bone formation of grafted cells in the scaffold were evaluated by immunohistochemistry. To our knowledge this is the first report with solid evidence demonstrating PBMSCs can survive in the bone defect area and directly contribute to new bone formation. Moreover, the present data also indicated the tissue engineering with PBMSCs/SAP/PLGA scaffold can serve as a novel prospective strategy for healing large size cranial defects.
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Project description:Reconstruction of bone due to surgical removal or disease-related bony defects is a clinical challenge. It is known that the immune system exerts positive immunomodulatory effects on tissue repair and regeneration. In this study, we evaluated the in vivo efficacy of autologous neutrophils on bone regeneration using a rabbit calvarial defect model. Methods: Twelve rabbits, each with two surgically created calvarial bone defects (10 mm diameter), were randomly divided into two groups; (i) single application of neutrophils (SA-NP) vs. SA-NP control, and (ii) repetitive application of neutrophils (RA-NP) vs. RA-NP control. The animals were euthanized at 4 and 8 weeks post-operatively and the treatment outcomes were evaluated by micro-computed tomography, histology, and histomorphometric analyses. Results: The micro-CT analysis showed a significantly higher bone volume fraction (bone volume/total volume) in the neutrophil-treated groups, i.e., median interquartile range (IQR) SA-NP (18) and RA-NP (24), compared with the untreated controls, i.e., SA-NP (7) and RA-NP (14) at 4 weeks (p < 0.05). Similarly, new bone area fraction (bone area/total area) was significantly higher in neutrophil-treated groups at 4 weeks (p < 0.05). Both SA-NP and RA-NP had a considerably higher bone volume and bone area at 8 weeks, although the difference was not statistically significant. In addition, immunohistochemical analysis at 8 weeks revealed a higher expression of osteocalcin in both SA-NP and RA-NP groups. Conclusions: The present study provides first hand evidence that autologous neutrophils may have a positive effect on promoting new bone formation. Future studies should be performed with a larger sample size in non-human primate models. If proven feasible, this new promising strategy could bring clinical benefits for bone defects to the field of oral and maxillofacial surgery.
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Project description:Background/objectives:Accelerating the process of bone regeneration is of great interest for surgeons and basic scientists alike. Recently, umbilical cord mesenchymal stem cells (UCMSCs) are considered clinically applicable for tissue regeneration due to their noninvasive harvesting and better viability. Nonetheless, the bone regenerative ability of human UCMSCs (HUCMSCs) is largely unknown. This study aimed to investigate whether Wnt10b-overexpressing HUCMSCs have enhanced bone regeneration ability in a rat model. Method:A rat calvarial defect was performed on 8-week old male Sprague Dawley rats. Commercially purchased HUCMSCsEmp in hydrogel, HUCMSCsWnt10b in hydrogel and HUCMSCsWnt10b with IWR-1 were placed in the calvarial bone defect right after surgery on rats (N = 8 rats for each group). Calvaria were harvested for micro-CT analysis and histology four weeks after surgery. CFU-F and multi-differentiation assay by oil red staining, alizarin red staining and RT-PCR (real-time polymerase chain reaction) were performed on HUCMSCsEmp and HUCMSCsWnt10b in vitro. Conditioned media from HUCMSCsEmp and HUCMSCsWnt10b were collected and used to treat human umbilical cord vein endothelial cells in Matrigel to access vessel formation capacity by tube formation assay. Results:Alizarin red staining, oil red staining and RT-PCR results showed robust osteogenic differentiation but poor adipogenic differentiation ability of HUCMSCsWnt10b. Furthermore, HUCMSCsWnt10b could accelerate bone defect healing, which was likely due to enhanced angiogenesis after the HUCMSCsWnt10b treatment, because more CD31+ vessels and increased vascular endothelial growth factor-A (VEGF-A) expression were observed, compared with the HUCMSCsEmp treatment. Conditioned media from HUCMSCsWnt10b also induced endothelial cells to form vessel tubes in a tube formation assay, which could be abolished by SU5416, an angiogenesis inhibitor. Conclusion:To our knowledge, this is the first study providing empirical evidence that HUCMSCsWnt10b can enhance their ability to heal calvarial bone defects via VEGF-mediated angiogenesis. The translational potential of this article:HUCMSCsWnt10b can accelerate critical size calvaria and are a new promising therapeutic cell source for fracture nonunion healing.
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Project description:Mesenchymal stem cells sheets have been verified as a promising non-scaffold strategy for bone regeneration. Alveolar bone marrow mesenchymal stem cells, derived from neural crest, have the character of easily obtained and strong multi-differential potential. However, the bone regenerative features of alveolar bone marrow mesenchymal stem cells sheets in the craniofacial region remain unclear. The purpose of the present study was to compare the osteogenic differentiation and bone defect repairment characteristics of bone marrow mesenchymal stem cells sheets derived from alveolar bone (alveolar bone marrow mesenchymal stem cells) and iliac bone (Lon-bone marrow mesenchymal stem cells) in vitro and in vivo. Histology character, osteogenic differentiation, and osteogenic gene expression of human alveolar bone marrow mesenchymal stem cells and Lon-bone marrow mesenchymal stem cells were compared in vitro. The cell sheets were implanted in rabbit calvarial defects to evaluate tissue regeneration characteristics. Integrated bioinformatics analysis was used to reveal the specific gene and pathways expression profile of alveolar bone marrow mesenchymal stem cells. Our results showed that alveolar bone marrow mesenchymal stem cells had higher osteogenic differentiation than Lon-bone marrow mesenchymal stem cells. Although no obvious differences were found in the histological structure, fibronectin and integrin ?1 expression between them, alveolar-bone marrow mesenchymal stem cells sheet exhibited higher mineral deposition and expression levels of osteogenic marker genes. After being transplanted in the rabbit calvarial defects area, the results showed that greater bone volume and trabecular thickness regeneration were found in bone marrow mesenchymal stem cells sheet group compared to Lon-bone marrow mesenchymal stem cells group at both 4?weeks and 8?weeks. Finally, datasets of bone marrow mesenchymal stem cells versus Lon-bone marrow mesenchymal stem cells, and periodontal ligament mesenchymal stem cells (another neural crest derived mesenchymal stem cells) versus umbilical cord mesenchymal stem cells were analyzed. Total 71 differential genes were identified by overlap between the 2 datasets. Homeobox genes, such as LHX8, MKX, PAX9, MSX, and HOX, were identified as the most significantly changed and would be potential specific genes in neural crest mesenchymal stem cells. In conclusion, the Al-bone marrow mesenchymal stem cells sheet-based tissue regeneration appears to be a promising strategy for craniofacial defect repair in future clinical applications.
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