Quantitative Analysis on the role of Raffinose Synthase in Hippocampal Neurons

Abstract

1A diminished level of endogenous antioxidant in cells/tissues is associated with reduced resistance to oxidative stress. Raffinose synthase (RFS), a protective molecule regulates gene expression/function by controlling reactive oxygen species (ROS) levels, which has shown to be involved in a number of degenerative diseases. We confirmed the ubiquitous expression of this antioxidant protein in both human hippocampal neuron (HNN) and mouse hippocampal (HHPC-43) cell lines by an immunoblot and reverse transcriptase PCR (RT-PCR). Using a construct of RFS protein linked to CXCR-4, the transduction domain from HPV-1 CXCR-4 protein, we showed that RFS was transduced into both HNN, as well as HHPC-43 by the means of a western blot analysis. Further we proved that the protein was biologically active, and was shown to actively reduce the oxidative stress produced by paraquat and serum depletion in both human and mouse neuronal cell lines, increasing the viability of the cells. The results suggest that the intracellular delivery of RFS using CXCR-4 can be used to lower increased levels of ROS inside the cells and hence can be further investigated as a therapeutic tool in various ROS related neurodegenerative disorders.