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We employed primary septo-hippocampal cell cultures to determine the ability of liposome-mediated BDNF gene transfection to facilitate recovery of neurofilament loss caused by depolarization injury. After BDNF gene transfection in uninjured cultures, RT-PCR and immunohistochemical staining confirmed increases in BDNF mRNA and protein in transfected cells. Three days after depolarization injury, Western blot and immunohistochemical analyses detected significant loss of neurofilament proteins in non-transfected cultures, while BDNF transfection produced marked increases in neurofilament proteins following either pre-injury transfection or transfection 24 h following injury. Immunohistochemical studies also detected enhanced immunolabeling of BDNF and total neurofilament protein (phosphorylated and non-phosphorylated) in injured neurons following BDNF transfection or administration of exogenous BDNF protein, compared to untransfected, injured controls.

Although liposomes have been widely employed to transfect DNA into a variety of cell types, no previous studies have systematically examined conditions producing optimal liposomal-mediated transfection of DNA into central nervous system (CNS) cells. Thus, we used the beta-galactosidase (beta-gal) reporter gene to examine factors influencing the efficiency of liposome-mediated gene transfection in CNS cell cultures. Our results indicate that without increasing the amounts of DNA, increased liposome concentrations within certain limits enhanced transfection efficiency. However, higher liposome levels could produce cell lysis. Without increasing liposome concentrations, increased amounts of DNA did not improve transfection efficiency. Employing the optimal concentration (1 microgram DNA/3 microliters liposomes/well), beta-gal gene expression was sustained for at least two weeks after transfection in primary septo-hippocampal cultures.

We examined liposome-mediated gene transfection of nerve growth factor (NGF) in primary central nervous system cultures. RT-PCR analyses detected increased expression of NGF mRNA one day after liposome-mediated NGF gene transfection. ELISA studies detected large increases in NGF protein in cells and in culture medium after NGF gene transfection. Cells continued to secrete NGF into the medium for at least 2 weeks. NGF bioassays confirmed that the NGF secreted after gene transfection was biologically active.