Novel Method of Decellularization of Porcine Valves Using Polyethylene Glycol and Gamma Irradiation Takeyoshi Ota, MD, PhD, Satoshi Taketani, MD, Shigemitsu.

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Presentation transcript:

Novel Method of Decellularization of Porcine Valves Using Polyethylene Glycol and Gamma Irradiation Takeyoshi Ota, MD, PhD, Satoshi Taketani, MD, Shigemitsu Iwai, MD, PhD, Shuji Miyagawa, MD, PhD, Masakazu Furuta, PhD, Masayuki Hara, PhD, Eiichiro Uchimura, PhD, Yutaka Okita, MD, PhD, Yoshiki Sawa, MD, PhD The Annals of Thoracic Surgery Volume 83, Issue 4, Pages 1501-1507 (April 2007) DOI: 10.1016/j.athoracsur.2006.11.083 Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 1 Histologic preparations (A–D) and transmission electron microscopy (E, F) of the decellularized valve. (A, B) Decellularized wall. (C) Decellularized cusp. (D) Multilayer structure of the elastic lamina and collagen. (E) Decellularized wall. (F) Decellularized cusp, elastin (e) and collagen (co) ultrastructure was well preserved. (A–C, hematoxylin and eosin stain; D, elastica van Gieson; Original magnification: A and C, ×40; B and D, ×100; E and F, ×10,000.) The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 2 (A) Mechanical strength. (B) Collagen content. (C) DNA content. (D) Protein content. (DV = decellularized valve tissue; NS = not significant; PN = porcine native tissue. The results are presented as mean and standard deviation; *p < 0.05.) The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 3 Polymerase chain reaction analysis for porcine endogenous retrovirus (PERV). PERV sequences were not detected in decellularized valve tissues (#1–#5). (DV = decellularized valve tissue; GAPDH = glyceraldehyde-3-phosphate dehydrogenase; PC = positive control.) The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 4 Histologic preparations of decellularized walls. (A, B) α-1.3 Gal was detected in the porcine native tissue and the tissue treated with polyethylene glycol and Dnase. (C, D) It was not detectable in the native tissue treated with 100 kGy gamma irradiation or the decellularized tissue. (Griffonia simplicifolia lectin I-B4 isolectin stain, all at original magnifications ×100. DV = decellularized valve tissue; PN = porcine native tissue.) The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 5 Acute rejection analysis in rats is shown in A and B. The arrows (→) show the margin between the vascular walls and the subcutaneous tissue. (A) The invasion of inflammatory cells was not seen in the decellularized tissue, (B) but invading inflammatory cells (}) were seen in the porcine native tissue (hematoxylin and eosin stain, original magnification ×100). Calcification analysis is shown in C and D. (C) A small amount of calcification was observed in the decellularized tissue, but (D) much stronger calcification was seen in the glutaraldehyde-treated porcine tissue (von Kossa stain, original magnification ×40). The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions

Fig 6 Histologic preparations of decellularized walls. (A–C) Two months after implantation into dogs. (A) Several layers of cells were observed, consisting of (B) several layers of vimentin-positive cells (}), and (C) a monolayer of factor VIII–positive cells (arrow). (D–G) Six months after implantation. (D) Good repopulation of cells was seen along with (E) some vasculogenesis (arrow). (F) Many cells positive for α-smooth muscle actin (α-SMA) had spread over the tissue (}), and (G) the monolayer cells on the surface were factor VIII–positive (arrow). (A, D and E, hematoxylin and eosin stain; B, vimentin immunoreactivity; C and G, factor VIII immunoreactivity; F, α-SMA immunoreactivity. All at original magnification ×100.) The Annals of Thoracic Surgery 2007 83, 1501-1507DOI: (10.1016/j.athoracsur.2006.11.083) Copyright © 2007 The Society of Thoracic Surgeons Terms and Conditions