Volume 115 Number 1 January 1998 The Journal of THORACIC AND CARDIOVASCULAR SURGERY CARDIAC AND PULMONARY REPLACEMENT EFFICIENCY OF A HIGH-TITER RETROVIRAL VECTOR FOR GENE TRANSFER INTO SKELETAL MYOBLASTS Reida M. El Oakley, FRCS, MD a Nigel J. Brand, PhD a Paul B. J. Burton, BSc, MBBS a Madeleine C. McMullen b Gregor B. Adams, BSc (Hons) b Mark C. Poznansky, MRCP, PhD c Paul J. R. Barton, PhD a Magdi H. Yacoub, FRCS, DSc a Background: Genetic transformation of skeletal myoblasts for myocardial repair is dependent on an efficient gene transfer system that integrates the genes of interest into the genome of the target cell and its progeny. The aim of this investigation was to evaluate the use of a new retrovirally based gene transfer system for this purpose. Methods: MFGnlslacZ retroviral vector, packaged in high-titer, split-genome packaging cell line (FLYA4) was used to transduce the skeletal myoblast cell line L6. L6 cells, cultured in 10% fetal calf serum, were transduced with the MFGnlslacZ vector by means of filtered supernatant from FLYA4 cells. Transduced L6 cells were divided into four groups. Group I cells were fixed as myoblasts 3 days after transduction. Group II cells were allowed to differentiate into myotubes. Group III cells were split every 3 days for 4 months. Group IV cells were split as in group III but then allowed to differentiate into myotubes. All samples were fixed and stained for -galactosidase activity. The effects on gene transfer of transforming growth factor–, insulin-like growth factor–I, and platelet-derived growth factor were determined by spectrophotometric assay of -galactosidase activity in cells transduced in the presence or absence of serum with 0 to 200 ng/ml of each growth factor. Results: Morphometric analysis showed that 66.3%  3% to 69.6%  6% of cells in From the Departments of Cardiothoracic Surgery, Imperial College School of Medicine at The National Heart and Lung Institute, a London, Department of Genitourinary Medicine, Imperial College School of Medicine at St. Mary’s Hospital, b London, United Kingdom, and the Department of Infectious Diseases Medicine, The Dana Farber Cancer Institute, Har- vard Medical School, c Boston, Mass. Supported by The British Heart Foundation (PG/96085). Read at the Seventy-seventh Annual Meeting of The American Asso- ciation for Thoracic Surgery, Washington, D.C., May 4-7, 1997. Received for publication May 7, 1997; revisions requested July 11, 1997; revisions received August 7, 1997; accepted for publication August 8, 1997. Address for reprints: Reida El Oakley, FRCS, MD, Imperial College School of Medicine at The National Heart and Lung Institute, Dovehouse St., London SW3 6LY, United Kingdom. J Thorac Cardiovasc Surg 1998;115:1-8 Copyright © 1998 by Mosby, Inc. 0022-5223/98 $5.00 + 0 12/6/85372 1