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Published on: July 23, 2014
[Application of transforming growth factor-beta(1) on construction of tissue engineering heart valves: experimental
Nian-Guo Dong1, Yu-Ming Qiu, Jia-Wei Shi
1Department of Cardiovascular Surgery, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.
Objective:
To investigate the effects of transforming growth factor-beta1 (TGF-beta1) on construction of tissue engineering heart valves (TEHV).
Methods:
Fresh porcine aortic valves were decellularized with trypsinase and detergent Triton X-100. Myofibroblasts were obtained from rat thoracic aorta, cultured, transfected with the vector containing TGF-beta1 gene-plasmid pcDNA3.0/TGF-beta1 mediated by lipofectamine 2,000 after 48 hours, screened by G418 for for 3 weeks. Decellularized valves were divided into 3 groups: Group A, seeded with the transfected myofibroblasts and cultured in medium without TGF-beta1, Group B, seeded with the transfected myofibroblasts and cultured in medium with TGF-beta1 10 ng/ml, and Group C, seeded with non-transfected myofibroblasts and cultured in medium without TGF-beta1. Hematoxylin-eosin staining and transmission electron microscopy were performed to observe the cell proliferation. DNA contents were measured. Hydroxyproline content was measured so as to indirectly test the collagen production. AGS-J mechanical testing instrument was used to test the mechanical properties of the strips of valves.
Results:
Immunohistological investigation showed instant TGF-beta1 expression in the myofibroblasts 48 h after the transfection and stable TGF-beta1 expression 4 weeks later. Morphological examination showed that the myofibroblasts in Groups A and B were connected to one another closely with abundant extracellular matrix in the valves. The DNA contents of Groups A and B were (0.126 +/- 0.013) per thousand, and (0.109 +/- 0.004) per thousand, both significantly higher than that of Group [(0.089 +/- 0.011) per thousand, both P < 0.011], with a significant difference between Groups A and B (P < 0.05). The hydroxyproline content of Groups A and B were (5.83 +/- 0.67) per thousand and (5.02 +/- 0.40) per thousand, both significantly higher than that of Group C [(4.34 +/- 0.47) per thousand, both P < 0.05], with a significant difference between Groups A and B (P < 0.05). The maximum load of Groups A and B were (13.4 +/- 1.0) N and (11.7 +/- 1.4) N respectively, both significantly higher than that of Group C [(10.0 +/- 1.1) N, both P < 0.05], with a significant difference between Groups A and B (P < 0.05).
Conclusion:
TGF-beta1 is an important and effective bioactive factor for cell proliferation and extracellular matrix growth of heart valve. It is of great value for constructing TEHV in vitro.
