[Construction of bicistronic eukaryotic vector containing basic fibroblast growth factor and study of their functions in gene therapy for hearing impairment].
OBJECTIVE
To construct basic fibroblast growth factor(bFGR) and enhance green fluorescence protein(EGFP) fusion gene eukaryotic expression vector internal ribosome entry site (pIRES)-bFGF-GFP and to evaluate the effect of transduction bFGF gene on noise induced hearing-loss in inner ear hair cells of guinea pigs.


METHODS
Human bFGF cDNA was inserted into mammalian expressed plasmid pIRES-EGFP. The recombinant expression plasmid pIRES-bFGF-EGFP was transfected into inner ear of guinea pigs, using lipofectin method. The transduced bFGF gene was mediated by SA lipidsome. IRES- bFGF-GFP was administered into the round window as rescue agent at the same time of noise exposure or as a protective agent 7 days before.


RESULTS
SA liposome-mediated bFGF expressed at a high level in the cochlea of guinea pigs, and in the rescue group, a significant lower hearing thresholds was displayed. pIRES- bFGF-EGFP could protect hair cells. It demonstrated that pIRES- bFGF-GFP could protect the inner ear both structurally and functionally. bFGF/EGFP gene could be transcripted and translated into inner ear hair cells of guinea pig. bFGF/EGFP gene could express a specific protein. The recombinant bFGF/EGFP had significant protective effect as well as manifestation of autonomous fluorescence.


CONCLUSIONS
bFGF/EGFP fusion protein not only expressed in hair cells of guinea pig but also showed significant bFGF activity and autonomous fluorescence. IRES induced exogenous gene could enter the hair cells.
Shi
Li
L
Institute of Otorhinolaryngology, PLA General Hospital, Beijing 100853, China. shient@sina.com
Dong
Ming-min
MM
Shi
Wen-jie
WJ
Hu
Yin-yan
YY
Guo
Wei
W
Zhai
Suo-qiang
SQ
Yang
Wei-yan
WY
D000818:Animals; D005260:Female; D016222:Fibroblast Growth Factor 2 / Q000235:genetics* / Q000502:physiology; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D006168:Guinea Pigs; D006198:Hair Cells, Auditory; D006317:Hearing Loss, Noise-Induced / Q000628:therapy*; D006801:Humans; D008297:Male
D016222:Fibroblast Growth Factor 2