Abstract
A replication-deficient adenovirus was used for ex vivo gene transfer into rat heart grafts under conditions simulating clinical transplantation. The adenoviral vector, AdHCMVsp1LacZ, containing an expression cassette of Escherichiae coli lacZ, was used to perfuse heart grafts during cold ischemia before transplantation. Heart grafts were perfused with University of Wisconsin (UW) solution containing either 0 pfu, 5×1010 pfu, or 1×1011 pfu of viral vector, and were preserved for either 2 or 4 h and then transplanted into syngeneic recipients. The animals were killed at 1, 7, and 14 days after transplantation. The infection rate was assessed by histochemical staining for β-galactosidase. Using polymerase chain reaction (PCR), viral DNA presence was confirmed in every graft perfused with viral vectors. The protein production from the transfected gene was confirmed by a functional protein assay. An efficient gene transfer was achieved with an infection rate of 1%–1.5% for all cardiac myocytes, as assessed by 5-bromo-4-chloro-indolyl-β-d-galactopyranoside (X-gal) staining. All studies were negative in the control grafts. Gene expression persisted for at least 10 days after transplantation. We thus conclude that an efficient adenovirus-mediated gene transfection and expression of gene products can be achieved in ex vivo perfusion of the heart graft during cold preservation.
Similar content being viewed by others
References
Graham FL, Prevec L (1991) Manipulation of adenovirus vector. In: Murray EJ (eds) Mehtods in molecular biology, vol 7: Gene transfer and expression protocols. Humana, New Jersey, pp 109–128
Shaked A, Csete ME, Shiraishi M, Miller AR, Moen RC, Busuttil RW, Economou JS (1994) Retroviral-mediated gene transfer into rat experimental liver transplant. Transplantation 57:32–34
Kass EA, Falck PE, Alvira M, Rivera J, Buttrick PM, Wittenberg BA, Cipriani L, Leinwand LA (1993) Quantitative determination of adenovirus-mediated gene delivery to rat cardiac myocytes in vitro and in vivo. Proc Natl Acad Sci USA 90:11498–11502
Guzman RJ, Lemarchand P, Crystal RG, Epstein SE, Finkel T (1993) Efficient gene transfer into myocardium by direct injection of adenovirus vectors. Circ Res 73:1202–1207
Ono K, Lindsey ES (1969) Improved technique of heart transplantation in rats. J Thorac Cardiovasc Surg 57:225–229
Csete ME, Benhamou PY, Drazan KE, Wu L, Mclntee DF, Afra R, Mullen Y, Busuttil RW, Shaked A (1995) Efficient gene transfer to pancreatic islets mediated by adenoviral vectors. Transplantation 59:263–268
Csete ME, Drazan KE, Van Bree M, Mclntee DF, McBride WH, Bett A, Graham FL, Busuttil RW, Berk AJ, Shaked A (1993) Adenovirus-mediated gene transfer in the transplant setting. Part I: Conditions for expression of transferred genes in cold-preserved hepatocytes. Transplantation 57:1502–1507
Shaked A, Csete ME, Drazan KE, Bullington D, Wu L, Busuttil RW, Berk AJ (1994) Adenovirus-mediated gene transfer in the transplant setting. Part II: Successful expression of transferred cDNA in syngeneic liver grafts. Transplantation 57:1508–1511
Qin L, Chavin KD, Ding Y, Woodward JE, Favaro JP, Lin J, Bromberg JS (1994) Gene transfer for transplantation. Prolongation of allograft survival with transforming growth factor-beta 1. Ann Surg 220:508–519
Ojamaa K, Petrie JF, Balkman C, Hong C, Klein I (1994) Post-transcriptional modification of myosin heavy-chain gene expression in the hypertrophied rat myocardium. Proc Natl Acad Sci USA 91:3468–3472
Geissler EK, Wang J, Fechner JJ, Burlingham WJ, Knechtle SJ (1994) Immunity to MHC class I antigen after direct DNA transfer into skeletal muscle. J Immunol 152:413–421
Drazan KE, Wu L, Shen XD, Bullington D, Jurim O, Busuttil RW, Shaked A (1995) Adenovirus-mediated gene transfer in the transplant setting. Part III: Variables affecting gene transfer in liver grafts. Transplantation 59:670–673
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Shiraishi, M., Kusano, T., Hara, J. et al. Adenovirus-mediated gene transfer using ex vivo perfusion of the heart graft. Surg Today 26, 624–628 (1996). https://doi.org/10.1007/BF00311668
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00311668