Efficiently engineered cell sheet using a complex of polyethylenimine–alginate nanocomposites plus bone morphogenetic protein 2 gene to promote new bone formation

Abstract

Regeneration of large bone defects is a common clinical problem. Recently, stem cell sheet has been an emerging strategy in bone tissue engineering. To enhance the osteogenic potential of stem cell sheet, we fabricated bone morphogenetic protein 2 ( BMP-2 ) gene-engineered cell sheet using a complex of polyethylenimine–alginate (PEI–al) nanocomposites plus human BMP-2 complementary(c)DNA plasmid, and studied its osteogenesis in vitro and in vivo. PEI–al nanocomposites carrying BMP-2 gene could efficiently transfect bone marrow mesenchymal stem cells. The cell sheet was made by culturing the cells in medium containing vitamin C for 10 days. Assays on the cell culture showed that the genetically engineered cells released the BMP-2 for at least 14 days. The expression of osteogenesis-related gene was increased, which demonstrated that released BMP-2 could effectively induce the cell sheet osteogenic differentiation in vitro. To further test the osteogenic potential of the cell sheet in vivo, enhanced green fluorescent protein or BMP-2-producing cell sheets were treated on the cranial bone defects. The results indicated that the BMP-2-producing cell sheet group was more efficient than other groups in promoting bone formation in the defect area. Our results suggested that PEI–al nanocomposites efficiently deliver the BMP-2 gene to bone marrow mesenchymal stem cells and that BMP-2 gene-engineered cell sheet is an effective way for promoting bone regeneration.

A Concise Review on the Use of Mesenchymal Stem Cells in Cell Sheet-Based Tissue Engineering with Special Emphasis on Bone Tissue Regeneration

“…There are ongoing investigations to obtain the best results in bone regeneration with MSC sheets. Some of the biomaterials and/or biological agents reported to be combined with MSC sheets to enhance osteogenesis include coral particles [ 47 ], ceramics [ 48 ], surface-modified titanium and zirconia [ 82 ], simvastatin [ 83 ], β -tricalcium phosphate [ 84 ], coumarin-like derivative osthole [ 85 ], CD34+ peripheral blood cells [ 86 ], a complex of polyethylenimine-alginate nanocomposites plus BMP2 gene [ 87 ], nonviral oligonucleotide antimiR-138 delivery to MSC sheets [ 88 ], platelet-rich fibrin [ 89 , 90 ], vitamin C [ 91 ], poly(dimethylsiloxane) surface silanization [ 92 ], stromal cell-derived factor-1 [ 93 ], microRNA-21-loaded chitosan/hyaluronic acid nanoparticles [ 94 ], hydroxyapatite particles [ 90 , 95 ], and Notch activation by Jagged1 in MSC sheet cultures [ 96 ].…”

Hybrid composites of mesenchymal stem cell sheets, hydroxyapatite, and platelet-rich fibrin granules for bone regeneration in a rabbit calvarial critical-size defect model

“…The cell sheet technique was used to harvest cells and produce tissue-engineered bone. The application of cell sheets, as a scaffold-free cell delivery system, has previously been reported in the transplantation of MSCs to enhance the bone formation ( 16 , 19 ). The use of cell sheets has several advantages compared with current methods; firstly, as no invasive enzymatic treatment is used, the intact cell sheet is harvested along with its ECM, which is important for cell differentiation and bone formation ( 51 ).…”

“…Cell sheet engineering, as a novel tissue engineering technique, recreates a biological microenvironment similar to that of a regenerative milieu (16). Without typical proteolytic enzyme or EDTA treatments, cultured cells are able to be harvested as a single contiguous cell sheet, retaining cell-cell junctions, as well as deposited extracellular matrix (ECM) on the basal sheet surface (17).…”

“…Scaffolds used for tissue engineering applications are typically biocompatible and biodegradable, support cell proliferation and differentiation, and are able to provide appropriate mechanical support ( 15 ). Cell sheet engineering, as a novel tissue engineering technique, recreates a biological microenvironment similar to that of a regenerative milieu ( 16 ). Without typical proteolytic enzyme or EDTA treatments, cultured cells are able to be harvested as a single contiguous cell sheet, retaining cell-cell junctions, as well as deposited extracellular matrix (ECM) on the basal sheet surface ( 17 ).…”