LIPOSOME-MEDIATED TRANSFER OF GENES CONTAINING HLA-CLASS-II ALPHA-CHAIN INTO CULTURED EMBRYONIC CHICK CARDIAC MYOCYTES AND COS7 CELLS

Rejection remains a major problem after cardiac transplantation. One h ypothesis is that transfer of recipient HLA genes could lead to expres sion of the antigens on the surface of donor cells and so facilitate g raft acceptance. This paper describes a pilot study for relevant gene- transfer (transfection) experiments on adult cardiac myocytes, investi gating the feasibility of transfection using cationic liposomes. The p lasmid pcDV1-pL2 vector containing HLA-DR-alpha chain gene was incubat ed with Lipofectin(R), a DOTMA and DOPE lipid mixture, for 10 minutes. Embryonic chide cardiac myocytes (ECCM) and COS7 monkey cells were in cubated with DNA: Lipofectin(R) ratios of 1:1, 1:2, 1:4, and 1 :10 for 16 hours (hrs). Using a fixed ratio of 1 :4, incubation periods of 4, 8, and 16 hrs were compared. Finally, cells were incubated for 16 hrs and consecutively cultured for 6 days. Expression of the HLA-DR-alpha chain antigen was detected by indirect immunohistochemical staining. Highest transfection rates were achieved with a DNA: Lipofectin ratio of 1:4 for ECCM and COS7 cells (2.7% +/- 0.6% and 2.4% +/- 0.3% after 16 hrs incubation) and a transfection time of 4 hrs (5.8% +/- 0.6% and 5.3% +/- 1.7%). Immunohistochemically positive cells were present aft er 6 days (2.0% +/- 1.2% and 2.1% +/- 0.3%). We found a low level of e xpression of HLA-DR-alpha chain gene, influenced by transfection time and DNA: Lipofectin ratio. To increase the efficiency of liposome-medi ated gene transfer and assess potential applications in cardiac transp lantation, further investigation of cell properties promoting transfec tion is necessary.