Fig. 4

Regulation of hGH expression in transduced C2C12 myoblasts in vitro and in vivo

(A) In vitro regulation of hGH expression. C2C12 myoblasts were infected with different retroviral vectors for constitutive or regulated hGH expression, as indicated on the y axis, as described in Materials and Methods. Two days after infection, 1 × 10⁶ cells were plated into replica plates, one of which contained 1 µg/ml doxycyline in the growth medium. The following day the growth medium was replaced and 24 hours later the supernatant was harvested and levels of secreted hGH in the off- (solid bar) or the on state (shaded bar) were determined by ELISA. (B) In vivo regulation of hGH expression. Groups of 5 to 6 C3H/HeJ mice received i.m. injections of transduced C2C12 myoblast populations as described in Materials and Methods. Mice were left untreated up until day 16 after injection when half of the mice of each group were put on doxycycline treatment (dark symbols), while the other half continued to be untreated (light symbols). The treatment of both subgroups was then changed to the opposite on day 46 after injection (e.g. mice treated with doxycycline were taken off the drug and mice not previously treated were given doxycycline). Serum samples of individual mice were taken at the time points indicated in the graph and hGH serum levels were determined by RIA as described in Materials and Methods. Individual values in the graph are the mean of the hGH serum levels of 2 to 6 mice.