EXPRESSION OF BETA-GALACTOSIDASE IN MOUSE-BRAIN - UTILIZATION OF A NOVEL NONREPLICATIVE SINDBIS VIRUS VECTOR AS A NEURONAL GENE DELIVERY SYSTEM

Sindbis virus expression has been used for in vitro investigations of antigen processing, presentation and epitope mapping. The recent devel opment of a replication-deficient recombinant Sindbis virus expression vector has made in vivo expression possible with minimal pathogenic r isk. Advantages of Sindbis virus over other available viral systems in clude a comparatively smaller genome size making it possible to clone larger inserts, the ability to infect a wide range of host cell types with reduced pathogenicity for humans. These features suggest the poss ible utility of Sindbis virus for the in vivo delivery of genes to neu ral cells. We used the recombinant Sindbis viral expression system to target delivery of the lacZ gene to neuronal cells of mice by stereota ctic surgery. Sindbis viral mRNA obtained by in vitro transcription wa s used to transfect baby hamster kidney (BHK) cells. As shown by histo chemistry, beta-galactosidase (beta-gal) was expressed in approximatel y 50% of transfected cells. Cells were then cotransfected I with DH-BB helper sequences that enabled the recombinant Sindbis virus RNA packa ging. Nonreplicative Sindbis viral stock was collected 24 h after tran sfection. BHK cells were then infected with viral stock and histochemi stry analysis was performed. Again, approximately 50% of the cells exp ressed beta-gal. The same viral stock was infused into the nucleus cau datus/putamen and nucleus accumbens septi and histochemical analysis o n frozen sections from the relevant brain areas confirmed that beta-ga l was expressed in neurons in a time-dependent manner. beta-Gal was de tected at 24 h after inoculation and was present for at least 14 days, with maximum expression at 48 h. These results suggest that a nonrepl icative Sindbis virus expression system may be useful for delivery of foreign genes into the central nervous system (CNS).