Y. Torrente et al., Extracorporeal circulation as a new experimental pathway for myoblast implantation in mdx mice, CELL TRANSP, 8(3), 1999, pp. 247-258
The deficiency of dystrophin, a sarcolemmal associated protein, is responsi
ble for Duchenne muscular dystrophy (DMD). Gene replacement is attractive a
s a potential therapy. In this article, we describe a new method for myobla
st transplantation and expression of dystrophin in skeletal muscle tissue o
f dystrophin-deficient mdx mouse through iliac vessels extracorporeal circu
lation. We evaluated the extracorporeal circulation as an alternative route
of delivering myoblasts to the target tissue. Two series of experiments we
re performed with the extracorporeal circulation. In a first series, L6 rat
myoblasts, transfected with LacZ reporter gene, were perfused in limbs of
15 rats. In the second series, the muscle limbs of three 6-8-week-old mdx w
ere perfused with myoblasts of donor C57BL10J mice. Before these perfusions
, the right tibialis anterior (TA) muscle of the rats and mdx was injected
three times at several sites with bupivacaine (BPVC) and hyaluronidase. The
ability of injected cells to migrate in the host tissue was assessed in ra
ts by technetium-99m cell labeling. No radioactivity was detected in the lu
ngs, bowels, and liver of animals treated with extracorporeal circulation.
The tissue integration and viability of the myoblasts were ultimately confi
rmed by genetic and histochemical analysis of LacZ reporter gene. Following
a single extracorporeal perfusion of myoblasts from donor C57BL10J, sarcol
emmal expression of dystrophin was observed in clusters of myofibers in tib
ialis anterior muscles previously treated with BPVC and hyaluronjdase. Furt
hermore, large clusters of dystrophin-positive fibers were observed in musc
les up to 21 days after repeated treatments. These clusters represented an
average of 4.2% of the total muscle fibers. These results demonstrate that
the extracorporeal circulation allows selective myoblast-mediated gene tran
sfer to muscles, opening new perspectives in muscular dystrophy gene therap
y.