Tissue-Engineered Bone for Treatment of Combat Related Limb Injuries

Abstract

Aim: Based on our preliminary positive clinical results with use of cultured bone marrow-derived multipotent mesenchymal stem/stromal cells in traumatology, our aim was to develop living three-dimensional tissue-engineered bone equivalent transplantation technology for restoration of critical sized bone defects caused by combat related high energy trauma. Materials and Methods: To fabricate bone equivalent we used devitalized allogeneic bone scaffolds (blocks and chips) seeded with cultured autologous cells: bone marrow-derived multipotent mesenchymal stem/stromal cells in mix with periosteal progenitor cells and endothelial progenitor cells. Quality/identity of cell cultures was assured by donor and cell culture infection screening (immunofluorescence assay, polymerase chain reaction), flow cytometry (cell phenotype), karyotyping (GTG banding), functional assays (colony forming units analysis, multilineage differentiation assay). Bone defect treatment with bone equivalent application was fully completed in 39 combat-injured with 42 defects. New bone formation was assessed by the radiographic examination. Results: Casualties were included in a treatment program an average of 10.1 months after injury, provided the ineffectiveness of conventional surgery methods. All cell type cultures had a normal karyotype and appropriate phenotype, differentiation potential and functional properties, ~30% colony forming units frequency and hadn’t any signs of cell senescence. The fluorescein diacetate/propidium iodide combined staining and histology analysis of graft samples before transplantation showed their regular seeding with viable cells. Pathomorphological analysis of bone equivalent specimens 3–6 months post-op revealed the active remodeling processes and immature bone tissue formation. Bone defect restoration was observed 5–6 months post-op. Conclusion: The developed biotechnology of living three-dimensional tissue-engineered bone equivalent transplantation with overall effectiveness 90.4% allows restoring the bone integrity, forming new bone tissue in a site of bone defect, and significantly reducing the rehabilitation period of a patient.

Recent Advances in Endothelial Progenitor Cells Toward Their Use in Clinical Translation

“…This lack of standardization makes it difficult to compare the results of different clinical trials and will in the future hinder the translation of these therapies into medical practice. A similar lack of consistency is again observed with the methods to culture EPCs with some reports administering EPCs without prior culture (31, 32), some culturing on fibronectin before re-injection into the patient (27, 28, 33, 34), and Vaysilev et al culturing cells on uncoated flasks with EGM-2 medium (30). Despite the positive results from the above papers, EPC therapy is still limited by the immunogenicity of allogeneic EPCs, along with its poor definition, isolation and expansion standardization, as outlined above.…”

“…Two papers were published beyond the date range specified in our search (23, 26). Finally, two other papers from Table 1 met our classification criteria did not appear in our literature search (27, 30). Granton et al used 7–50 × 10 6 peripheral blood eNOS modified EPCs for pulmonary arterial hypertension and showed an improvement in pulmonary resistance after treatment (27), whereas Vasyliev et al used 20–60 × 10 6 autologous peripheral blood derived EPCs in a scaffold of allogeneic bone and fibrin gel for bone fracture repair and demonstrated an improvement in bone regeneration (30).…”

“…From the published EPC human clinical trials, there is a marked inconsistency in terms of the culture conditions and characterization of EPCs with D'Avola et al and Vasyliev et al using multiple different cell surface markers to identify the EPCs they used (28, 30), Lara-Hernandez et al and Granton et al using two cell surface markers (27, 31), Tanaka et al only utilizing one cell surface marker (32), and Zhu et al and Zhu et al did not mention the marker used for defining the EPCs in their studies (33, 34). This lack of standardization makes it difficult to compare the results of different clinical trials and will in the future hinder the translation of these therapies into medical practice.…”

The Few Who Made It: Commercially and Clinically Successful Innovative Bone Grafts

“…Furthermore, a clinical study performed in 2017 utilized a cocktail of expanded autologous BMSCs, periosteal progenitor cells and endothelial progenitor cells on a fibrin hydrogel-DBM composite, to restore critical-sized bone defects of 47 casualties with complicated gunshot bone wound. X-ray examination determined that within 4–6 months post-operatory, 90.4% of the treated defects regained native integrity ( Vasyliev et al, 2017 ).…”

Recent Advances in Endothelial Progenitor Cells Toward Their Use in Clinical Translation

“…Finally, two other papers from Table 1 met our classification criteria did not appear in our literature search (27,30). Granton et al used 7-50 × 10 6 peripheral blood eNOS modified EPCs for pulmonary arterial hypertension and showed an improvement in pulmonary resistance after treatment (27), whereas Vasyliev et al used 20-60 × 10 6 autologous peripheral blood derived EPCs in a scaffold of allogeneic bone and fibrin gel for bone fracture repair and demonstrated an improvement in bone regeneration (30). Of note, neither study had a control group to compare against.…”