The Effects of Somatic Neurotrophic Factor Gene Transfection for Retinal Pigment Epithelium

Abstract

Abstract: : Purpose:To examine the effect of somatic neurotrophic factor gene transfection for retinal pigment epithelium (RPE). Methods:Somatic transfection of bFGF or BDNF cDNA was performed in the human RPE cell line ARPE19 or D407RPE. The differences of the gene expression between bFGF-transfected (ARPE-19) and non-transfected cells were examined using DNA microarray system. We also performed real time-PCR of the genes from the results of the DNA microarray between bFGF or BDNF-transfected and non-transfected cells (D407RPE). Cell attachment was assayed using toluidine blue and SDS method. MTS assay for cell viability was also applied after hypoxia induction to these cells. Results:Somatic transfection of these cDNAs was performed successfully using appropriate antibiotics and confirmed by RT-PCR and enzyme-linked immunosorbent assay. The results of DNA microarray revealed that half of the genes showing higher expression in bFGF transfected ARPE-19 than those of non transfected cells were several neuron survival factors and neurogenesis factors. One of the genes that showed less expression than those of non transfected cells was lysyl oxidase like-1 gene, which play an important role for the maturation of the extracellular collagen and for cell attachment. It was also confirmed in somatic bFGF gene transfected D407RPE by real -time PCR. A statistically significant less cell attachment was observed in bFGF transfected D407RPE at 4 hour after starting the culture. These gene expressions were not always true in the cells of somatic BDNF gene transfection. When we cultured the cells in the condition of hypoxia, the cells with somatic bFGF transfection showed higher MTS activity than those of non-transfected cells. Conclusions:Somatic transfection of some neurotrophic factor genes may have its factor specific synexpression cluster of some genes and function of the transfected cells.

Keywords: gene transfer/gene therapy • transplantation • gene microarray