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Ann Thorac Surg 2007;83:1501-1507
© 2007 The Society of Thoracic Surgeons

Original Articles: Cardiovascular

Novel Method of Decellularization of Porcine Valves Using Polyethylene Glycol and Gamma Irradiation

Takeyoshi Ota, MD, PhD*,a, Satoshi Taketani, MDb, Shigemitsu Iwai, MD, PhDb, Shuji Miyagawa, MD, PhDc, Masakazu Furuta, PhDd, Masayuki Hara, PhDd, Eiichiro Uchimura, PhDe, Yutaka Okita, MD, PhDa, Yoshiki Sawa, MD, PhDb

a Department of Cardiovascular Surgery, Kobe University Graduate School of Medicine, Kobe, Japan
b Department of Cardiovascular Surgery, Osaka University Graduate School of Medicine, Osaka, Japan
c Department of Regenerative Medicine, Osaka University Graduate School of Medicine, Osaka, Japan
d Department of Biological Science, Graduate School of Science, Osaka Prefecture University, Osaka, Japan
e Cardio, Inc, Kobe, Japan

Accepted for publication November 28, 2006.

* Address correspondence to Dr Ota, Division of Cardiovascular Surgery, Kobe University Graduate School of Medicine, 7-5-2 Kusunoki-cho, Chuo-ku, Kobe 650-0017, Japan (Email: toota-cvs{at}umin.ac.jp).

Background: Recent tissue-engineered valves are in need of a breakthrough to overcome several limitations against clinical applications. We have developed a new method of decellularization using polyethylene glycol and gamma irradiation.

Methods: Fresh porcine aortic valves were decellularized using polyethylene glycol and gamma irradiation. These were evaluated by histologic, biochemical (DNA, solubilized protein and collagen content), mechanical (strength test, transmission electron microscopy) and immunologic (porcine endogenous retrovirus and the {alpha}-1.3 galactosyl epitope) analyses. Implantations into the subcutaneous tissue of rats (1 week, n = 10; 2 months, n = 10) and into the descending aorta of dogs (2 months, n = 6; 6 months, n = 3) were used as in vivo studies.

Results: Complete decellularization was confirmed by histologic examination and by determining the DNA and solubilized protein content. The decellularized valve showed no significant differences in its mechanical strength or collagen content compared with native porcine tissues. The ultrastructure was well preserved in transmission electron microscope images. The DNA sequence of a porcine endogenous retrovirus and the {alpha}-1.3 galactosyl epitope were eliminated after the decellularizing process. No acute rejection and little calcification was noted in the rat model. In the dog model at 2 months, the surface of the graft was completely covered with a monolayer of endothelial cells. In addition, several layers of vimentin-positive cells lay under the endothelial cells. At 6 months after implantation, many smooth muscle cells, monolayer endothelial cells, and some vasculogenesis were seen.

Conclusions: The decellularizing method provided low immunogenicity, low risk of unknown infections, and was little subject to calcification. The decellularized tissues showed acceptable durability and recellularization.




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