DELIVERY OF HUMAN FIBROBLAST GROWTH-FACTOR-I GENE TO BRAIN BY MODIFIED RAT-BRAIN ENDOTHELIAL-CELLS

Fibroblast growth factor (FGF) is an endothelial cell mitogen and serv es as a mitogen and/or differentiating factor that can be neuroprotect ive for other cell types within the CNS. We established brain microvas cular endothelial cell lines that secrete FGF-1 with the ultimate goal of examining their usefulness as a cellular platform for FGF gene del ivery to brain. A chimeric gene consisting of the secretory sequence o f FGF-4 linked at the 5' end of human FGF-1 (sp-hst/KS3:FGF-1) was tra nsfected into rat microvascular endothelial cells previously altered t o express the lacZ reporter gene (RBEZ), and numerous clones were foun d to secrete FGF-1 (RBEZ-FGF). Immunoblotting of conditioned medium de monstrated an 18-kDa protein corresponding to FGF-1. Conditioned mediu m from RBEZ-FGF cells enhanced [H-3]thymidine incorporation in BALB/c3 T3 fibroblasts by up to sevenfold when compared with conditioned mediu m of control cell lines, corresponding to as much as 110 ng of active FGF-1/mg of cell protein/24 h. RBEZ-FGF cell lines remained contact-in hibited and proliferated independent of exogenous endothelial mitogens , in contrast to control lines that are mitogen-dependent. Incubation of PC12 cells with RBEZ-FGF cells or their conditioned medium induced neurite outgrowth by PC12 cells. RBEZ-FGF cells survived following imp lantation to neonatal and adult rat caudate-putamen for at least 21 da ys based on 5-bromo-4-chloro-3-indolyl beta-D-galactopyranoside (X-gal ) histochemistry, and FGF-1 gene expression by these cells in vivo was demonstrated by in situ hybridization and reverse transcriptase-PCR. These findings suggest that endothelial cells may be useful for FGF ge ne delivery to the CNS.