TRANSIENT INTRODUCTION OF A FOREIGN GENE INTO HEALING RAT PATELLAR LIGAMENT

We investigated the in vivo introduction of a reporter gene into heali ng rat patellar ligaments using the hemagglutinating virus of Japan (H VJ)-liposome-mediated gene transfer method. The mid-portion of the med ial half of the patellar ligament was cut transversely with a scalpel in 14-wk-old male Wistar rats, A HVJ-liposome suspension containing be ta-galactosidase (beta-gal) cDNA was injected directly into the injure d site and pooled in the fascial pocket covering the injured site 3 d postoperatively. Thereafter, beta-gal-labeled cells were observed in t he wound site accounting for 3% of the wound cells on the first day, 2 % on the third, 7% on the seventh, 6% on the 14th, 2% on the 28th, and 0.2% on the 56th day after injection, The beta-gal-labeled cells were initially localized in and adjacent to the wound site, but they were observed spreading into the ligament substance away from the wound on the seventh day after injection, On day 28, beta-gal-labeled cells wer e observed throughout the length of the ligament substance, With doubl e-labeling for marker antigens for monocyte/macrophage (ED-1) and for collagen I aminopropeptide (pN collagen I), it was revealed that fibro blastic (pN collagen I-positive) cells accounted for 63% and monocyte/ macrophage lineage cells for 32% of the beta-gal-labeled cells in the day 7 wound. On day 28, they formed 58 and 35% of the beta-gal-labeled cells in the wound, respectively. Thus, we succeeded in introducing t he beta-gal gene into healing rat patellar ligament. Moreover, labelin g of the transfected cells made it possible to identify a biological e vent, namely that the cells in and around the wound site infiltrate in to the uninjured ligament substance and come to populate the whole len gth of the ligament substance as repair progresses, These results sugg est that ligament healing may involve not only the repair of the wound site itself but also extensive cellular infiltration of ligament subs tance adjacent to the wound.