Dataset Information

Differential dynamics of bone graft transplantation and mesenchymal stem cell therapy during bone defect healing in a murine critical size defect.

ABSTRACT:

Background

A critical size bone defect is a clinical scenario in which bone is lost or excised due to trauma, infection, tumor, or other causes, and cannot completely heal spontaneously. The most common treatment for this condition is autologous bone grafting to the defect site. However, autologous bone graft is often insufficient in quantity or quality for transplantation to these large defects. Recently, tissue engineering methods using mesenchymal stem cells (MSCs) have been proposed as an alternative treatment. However, the underlying biological principles and optimal techniques for tissue regeneration of bone using stem cell therapy have not been completely elucidated.

Methods

In this study, we compare the early cellular dynamics of healing between bone graft transplantation and MSC therapy in a murine chronic femoral critical-size bone defect. We employ high-dimensional mass cytometry to provide a comprehensive view of the differences in cell composition, stem cell functionality, and immunomodulatory activity between these two treatment methods one week after transplantation.

Results

We reveal distinct cell compositions among tissues from bone defect sites compared with original bone graft, show active recruitment of MSCs to the bone defect sites, and demonstrate the phenotypic diversity of macrophages and T cells in each group that may affect the clinical outcome.

Conclusion

Our results provide critical data and future directions on the use of MSCs for treating critical size defects to regenerate bone.Translational Potential of this article: This study showed systematic comparisons of the cellular and immunomodulatory profiles among different interventions to improve the healing of the critical-size bone defect. The results provided potential strategies for designing robust therapeutic interventions for the unmet clinical need of treating critical-size bone defects.

SUBMITTER: Huang EE

PROVIDER: S-EPMC9357712 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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Publications

Differential dynamics of bone graft transplantation and mesenchymal stem cell therapy during bone defect healing in a murine critical size defect.

Huang Elijah Ejun EE Zhang Ning N Ganio Edward A EA Shen Huaishuang H Li Xueping X Ueno Masaya M Utsunomiya Takeshi T Maruyama Masahiro M Gao Qi Q Su Ni N Yao Zhenyu Z Yang Fan F Gaudillière Brice B Goodman Stuart B SB

Journal of orthopaedic translation 20220804

<h4>Background</h4>A critical size bone defect is a clinical scenario in which bone is lost or excised due to trauma, infection, tumor, or other causes, and cannot completely heal spontaneously. The most common treatment for this condition is autologous bone grafting to the defect site. However, autologous bone graft is often insufficient in quantity or quality for transplantation to these large defects. Recently, tissue engineering methods using mesenchymal stem cells (MSCs) have been proposed ...[more]

PMID: 35979174

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Project description:Bone regeneration poses a significant challenge in the field of tissue engineering, prompting ongoing research to explore innovative strategies for effective bone healing. The integration of stem cells and nanomaterial scaffolds has emerged as a promising approach, offering the potential to enhance regenerative outcomes. This study focuses on the application of a stem cell-laden nanomaterial scaffold designed for bone regeneration in rabbits. The in vivo study was conducted on thirty-six healthy skeletally mature New Zealand white rabbits that were randomly allocated into six groups. Group A was considered the control, wherein a 15 mm critical-sized defect was created and left as such without any treatment. In group B, this defect was filled with a polycaprolactone-hydroxyapatite (PCL + HAP) scaffold, whereas in group C, a PCL + HAP-carboxylated multiwalled carbon nanotube (PCL + HAP + MWCNT-COOH) scaffold was used. In group D, a PCL + HAP + MWCNT-COOH scaffold was used with local injection of bone morphogenetic protein-2 (BMP-2) on postoperative days 30, 45, and 60. The rabbit bone marrow-derived mesenchymal stem cells (rBMSCs) were seeded onto the PCL + HAP + MWCNT-COOH scaffold by the centrifugal method. In group E, an rBMSC-seeded PCL + HAP + MWCNT-COOH scaffold was used along with the local injection of rBMSC on postoperative days 7, 14, and 21. For group F, in addition to the treatment given to group E, BMP-2 was administered locally on postoperative days 30, 45, and 60. Gross observations, radiological observation, scanning electron microscopic assessment, and histological evaluation study showed that group F displayed the best healing properties, followed by group E, group D, group C, and B. Group A showed no healing with ends blunting minimal fibrous tissue. Incorporating growth factor BMP-2 in tissue-engineered rBMSC-loaded nanocomposite PCL + HAP + MWCNT-COOH construct can augment the osteoinductive and osteoconductive properties, thereby enhancing the healing in a critical-sized bone defect. This novel stem cell composite could prove worthy in the treatment of non-union and delayed union fractures in the near future.

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.

Project description:Bisphosphonates (BP) are anti-resorptive drugs that are widely used to prevent bone loss in osteoporosis. Since inhibition of bone resorption will cause a decrease in bone formation through a process called coupling, it is hypothesized that extended treatment protocols may impair bone healing. In this study, β-tri‑calcium-phosphate (βTCP) ceramics were inserted into critical-size long bone defects in estrogen-deficient mice under BP therapy. The study assessed the benefits of coating the ceramics with Bone Morphogenetic Protein-2 (BMP2) and an engineered BMP2 analogue (L51P) that inactivates BMP antagonists on the healing process, implant resorption, and bone formation. Female NMRI mice (11-12 weeks of age) were ovariectomized (OVX) or sham operated. Eight weeks later, after the manifestation of ovariectomy-induced osteoporotic bone changes, BP therapy with Alendronate (ALN) was commenced. After another five weeks, a femoral critical-size defect was generated, rigidly fixed, and βTCP-cylinders loaded with 0.25 μg or 2.5 μg BMP2, 2.5 μg L51P, and 0.25 μg BMP2/2.5 μg L51P, respectively, were inserted. Unloaded βTCP-cylinders were used as controls. Femora were collected six and twelve weeks post-implantation. Histological and micro-computer tomography (MicroCT) evaluation revealed that insertion of cylinders coated with 2.5 μg BMP2 accelerated fracture repair and induced significant bone formation compared to controls (unloaded cylinders or coated with 2.5 μg L51P, 0.25 μg BMP2) already six weeks post-implantation, independent of estrogen-deficiency and BP therapy. The simultaneous administration of BMP2 and L51P (0.25 μg BMP2/2.5 μg L51P) did not promote fracture healing six and twelve weeks post-implantation. Moreover, new bone formation within the critical-size defect was directly linked to the removal of the βTCP-implant in all experimental groups. No evidence was found that long-term therapy with ALN impaired the resorption of the implanted graft. However, osteoclast transcriptome signature was elevated in sham and OVX animals upon treatment with BP, with transcript levels being higher at six weeks than at twelve weeks post-surgery. Furthermore, the transcriptome profile of the developing repair tissue confirmed an accelerated repair process in animals treated with 2.5 μg BMP2 implants. L51P did not increase the bioefficacy of BMP2 in the applied defect model. The present study provides evidence that continuous administration of BP does not inhibit implant resorption and does not alter the kinetics of the healing process of critical-size long bone defects. Furthermore, the BMP2 variant L51P did not enhance the bioefficacy of BMP2 when applied simultaneously to the femoral critical-size defect in sham and OVX mice.

Project description:Osteochondral defects of the knee are common in orthopaedic patients. They are challenging to treat, especially in young, highly demanding patients who do not qualify for arthroplasty. Among the many possibilities to treat osteochondral lesions presented so far, none is ideal. Because of the poor healing potential of cartilage, treatment outcomes significantly worsen with larger lesions. The treatment of large defects usually requires expensive solutions, sometimes including second-stage surgery. Using mesenchymal stem cell transplantation and cancellous bone autografts, the technique presented here for osteochondral lesion reconstruction can be effectively used to treat large osteochondral lesions in a single-stage procedure. Technique Video Video 1 Osteochondral lesion reconstruction using mesenchymal stem cell transplantation and cancellous bone autografts; right knee. Patients are considered candidates after careful selection. To promote mesenchymal stem cell growth, 4 days before surgery, patients are administered granulocyte colony-stimulating factor. On the day of surgery, the mesenchymal stem cell suspension is obtained, using an MSC blood separator, under the control of the blood bank. A cytometric test is performed for the quantitative evaluation of cell lines, as well as a histopathological evaluation of stained preparations. The cell suspension is then transferred to the operating theater. The diagnosis is confirmed arthroscopically through a standard anterolateral portal, and concomitant lesions are excluded. Mini-open access is created next to the site of the lesion. After the lesion is exposed, delaminated cartilage or bony-cartilage sequestrum is removed. Using a bone spoon, the bony edges of the cavity are resected to healthy cartilaginous tissue. The next step is decortication and debridement of the bottom. To ensure the integration of bone grafts, it is necessary to remove the sclerotic subchondral bone layer until bleeding from the bone occurs. An additional factor improving the blood supply is the microfracturing the bottom of the cavity. From a separate site above the iliac crest, autologous cancellous bone grafts are harvested. The bottom of the lesion is filled with the debris of the cancellous bone. Restoration with a bone graft is performed up to the height of the surrounding healthy bone margin. During this time, the previously sized collagen sponge Tachosil is soaked in MSC suspension. The sponge is placed on the lesion and adjusted to the edges of the lesion using tweezers. The soaked implant should not cover surrounding healthy cartilage edges, but only fill the cavity. The entirety of the damaged fragment is covered with a 2-component fibrin glue. After binding, a suspension of mesenchymal cells, is administered under the fibrin layer using a no. 12 needle, and the application site is sealed with the rest of the glue. The wound is closed with layered sutures and sealed with sterile dressing. The operated joint is protected by cocoon dressing to limit mobility in the early postoperative period.

Dataset Information

Differential dynamics of bone graft transplantation and mesenchymal stem cell therapy during bone defect healing in a murine critical size defect.

Background

Methods

Results

Conclusion

Publications

Differential dynamics of bone graft transplantation and mesenchymal stem cell therapy during bone defect healing in a murine critical size defect.

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