P1.11 GENE TRANSFER INTO THE DEVELOPING FOLLICULAR TISSUES BY ADMINISTRATING RETROVIRUS VECTORS IN UTERO

Atsuko Wada, Hisako Kayama, Setsunosuke Ihara and Takashi Matsuzaki

Course of Biological Science, Graduate School of Life and Environmental Science, Shimane University, Matsue, Shimane, Japan

Objectives: Specific gene transfer techniques are required to investigate functions of interesting genes in the follicle tissues, which allow us to modify those expression patterns and install structurally modified genes. For this purpose, we searched a preferable condition for gene transfer into embryonic skin using retroviral vectors.

Methods: Retrovirus vectors carrying a lac-Z gene were microinjected into the amniotic fluid of pregnant mice at 8.0-13.5 dps and were infected to embryonic keratinocytes in utero. The vectors were administrated with Polybrene, poly-L-lysine, poly-L-ornithine or Retronectin. Gene transfer efficiency was estimated by X-gal staining of infected embryos. 

Results: Distribution of the lac-Z-positive cells varied, depending on the age of embryos at inoculation, virus number, and additives. Transfection to E9.5-10.5 embryos worked well and addition of poly-L-lysine increased the efficiency. Patches of positive cells were found in the skin where the hair follicles were stained almost uniformly or in variegated patterns, suggesting that the hair follicles were developed from a small number of ancestors. Positive cells were detected even at 11 weeks after birth.

Conclusions: We obtained a preferable condition for gene transfer into hair follicles with retrovirus vectors as follows: injection of 1x 10⁵ cfu of retrovirus in 1 microlitter with 30 microgram/ml of poly-L-lysine into the amniotic fluid at 9.5-10.5 dps.