[Tissue-engineered graft constructed by bone marrow mononuclear cells and heterogeneous acellularized tissue matrix: an animal experiment].
OBJECTIVE
To construct a tissue-engineered graft by using bone marrow cells as seeding cells and heterogeneous acellularized matrix as scaffold.


METHODS
Mononuclear cells were isolated from the bone marrow from piglets and cultured in different mediums including either vessel endothelial growth factor (VEGF) or platelet derived growth factor BB (PDGF-BB) to observe the differentiation of cells. Immunoassay was used to detect the expression of specific markers of endothelial cells or specific markers of smooth muscle cells. Adult dogs were killed and their thoracic or abdominal aortas were taken out and processed by multi-step decellularizing technique to remove the original cells while the elastic and collagen fibers were preserved. The undifferentiated bone marrow mononuclear cells were seeded onto the acellularized matrix and incubated in vitro. The cell-seeded grafts were then transplanted to the bone marrow donating piglets to substitute part of the native pulmonary artery. Three weeks later right ventriculography was performed. 100 days later the piglets were sacrificed. The transplanted vessels and the nearby tissues of native pulmonary vessels were excised for inspection.


RESULTS
The mononuclear cells cultured in the medium including VEGF showed the morphological features of endothelial cells and were positive of the specific markers of endothelial cells: platelet-endothelial cell adhesion molecule-l, vascular endothelial growth factor receptor Flk-1, VE-cadherin, and plasma factor VIII. The cells cultured in the medium including PDGF-BB showed morphological feature of smooth muscle cells and were positive of he specific marks of smooth muscle cells: alpha-SMA and calponin. One hundred days after transplantation, the inner surfaces of the grafts were smooth without thrombosis, calcification, and aneurysm. The maximal load was 2.76 N and the maximal elongation was 20.31 mm. Under the microscopy a great number of growing endothelial cells and smooth muscle cells could be seen and elastic and collagen fibers were abundant.


CONCLUSIONS
The mononuclear cells from bone marrow and acellularized matrix may be used to construct tissue-engineered graft.
Huang
Hui-min
HM
Shanghai Children's Medical Center, Medical College, Shanghai Jiaotong University, Shanghai 200127, China. wenfeik@online.sh.cn
Ma
Liang-long
LL
Ren
Hong
H
Wu
Shao-feng
SF
Jiang
Zu-ming
ZM
D000199:Actins / Q000032:analysis; D000818:Animals; D015703:Antigens, CD / Q000032:analysis; D001807:Blood Vessel Prosthesis; D001854:Bone Marrow Cells / Q000166:cytology*; D015820:Cadherins / Q000032:analysis; D002135:Calcium-Binding Proteins / Q000032:analysis; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D004285:Dogs; D004547:Elastic Tissue / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D005169:Factor VIII / Q000032:analysis; D007963:Leukocytes, Mononuclear / Q000166:cytology*; D008840:Microfilament Proteins / Q000032:analysis; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000032:analysis; D013552:Swine; D013997:Time Factors; D023822:Tissue Engineering / Q000379:methods*; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000032:analysis; D000096985:Calponins; D000099152:Cadherin 5
D000199:Actins; D015703:Antigens, CD; D015820:Cadherins; D002135:Calcium-Binding Proteins; D003094:Collagen; D005169:Factor VIII; D008840:Microfilament Proteins; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D040301:Vascular Endothelial Growth Factor Receptor-2; D000096985:Calponins; D000099152:Cadherin 5