Abstract
Traumas, infections, tumors, and some congenital malformations can lead to bone defects or even bone loss. The goal of the present study was to investigate whether inclusion of endothelial progenitor cells derived from peripheral blood (PB–EPCs) in cell-seeded partially deproteinized bone (PDPB) implants would stimulate recruitment of systemically injected bone marrow stromal cells (BMSCs) to the implant. Methods: BMSCs were injected intravenously with lentiviral expression vector expressing enhanced green fluorescent protein (eGFP) for tracing. Recruitment of eGFP-positive BMSCs was tested for the following implant configurations: 1) seeded with both BMSC and PB-EPC, 2) BMSC alone, 3) PB-EPC alone, and 4) unseeded PDPB. Protein and mRNA levels of endogenous stromal-derived factor-1 (SDF-1) and its receptor CXCR4, as well as monocyte chemotactic protein-1 (MCP-1) and its receptor CCR2, were evaluated on the 8th week. Immunohistochemical staining was performed to determine eGFP-positive areas at the defective sites. Masson’s trichrome staining was conducted to observe the distribution of collagen deposition and evaluate the extent of osteogenesis. Results: The mRNA and protein levels of SDF-1 and CXCR4 in the co-culture group were higher than those in other groups (p < 0.05) 8 weeks after the surgery. MCP-1 mRNA level in the co-culture group was also higher than that in the other groups (p < 0.05). Immunohistochemical assays revealed that the area covered by eGFP-positive cells was larger in the co-culture group than in the other groups (p < 0.05) after 4 weeks. Masson’s trichrome staining revealed better osteogenic potential of the co-culture group compared to the other groups (p < 0.05). Conclusion: These experiments demonstrate an association between PB-EPC and BMSC recruitment mediated by the SDF-1/CXCR4 axis that can enhance repair of bone defects.
Highlights
Bone defects and morphological abnormality caused by trauma and congenital malformation are common clinical problems
Binding of Stromal cell-derived factor-1 (SDF-1) and its receptor CXCR4 leads to activation of the SDF-1/CXCR4 axis, which is important in the recruitment of Bone marrow stromal cells (BMSCs) and directed migration
MSCM (Sciencell), EGM-2 BulletKit (Lonza), L-DMEM (Gibco), PBS (Life), EDTA (Cxbio), monoclonal antibodies specific for rabbit CD29, CD34, CD45 (BD Bioscience, San Diego, CA), enhanced green fluorescent protein (eGFP) expression plasmid (GuangZhou FuNeng), lentivirus kit (Gene Copoeia), low-density lipoprotein acetylated DiI complex (Invitrogen), FITC-labeled Ulex europaeus agglutinin I (Sigma), Matrigel (BD Biosciences), SDF-1/MCP-1 enzyme-linked immunosorbent assay (ELISA) kits (Cloud-clone Corp), DEPC (AMRESCO), Trizol (MRC, TR118), Revert Aid TM First Strand cDNA Synthesis Kit (Fermentas), SYBR Green Master Mix (Fermentas), CXCR4, CCR2 primary antibody (Santa Cruz), PVDF membranes (BioRad), eGFP primary antibody (Millipore), eGFP secondary antibody kit (ZhongShan JinQiao pv-9000), and Masson’s trichrome staining kit (Sigma) were the materials used in this experiment
Summary
Bone defects and morphological abnormality caused by trauma and congenital malformation are common clinical problems. Developing methods for combining cytology and materiology to construct tissue-engineered bone is an important goal in regenerative medicine. Bone marrow stromal cells (BMSCs) are popular stem cells in tissue engineering and regenerative medicine. Transplantation of BMSCs can increase the quantity of osteoblasts and accelerate bone repair. When implanting tissue-engineered bones into the body, attention should be paid to the transformation of exogenous stem cells and the potential of intrinsic mesenchymal stem cells to accelerate bone repair. Repair of tissue-engineered bones can be accelerated if intrinsic stem cells move toward the defects while promoting proliferation and osteogenesis of seed cells. Promotion of stem cell homing is a key challenge in the development of regenerative medicine, from construction of 3D structures to clinical applications
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