Construction and evaluation of an efficient C-Jun siRNA to downregulate matrix metalloproteinase in human keratinocytes and fibroblasts under UV exposure

Abstract

Background: C-Jun and EGFR have not been explored as targets via the mechanism of RNA silencing. Hence, this study designed an efficient C-Jun-h-825 small interfering RNA (siRNA) and investigated its effect on matrix metalloproteinase (MMP) and collagen expression in human keratinocytes exposed to UV radiation. Methods: Five C-Jun siRNAs were designed and screened for their ability to downregulate C-Jun expression in human fibroblasts. These constructs were used to study changes in skin cancer-related protein expression. HaCaT cells were grouped into 5-carboxyfluorescien (FAM-labeled) C-Jun-h-825 siRNA + 2 hr prior irradiation; mock transfected + 2 hr prior irradiation; normal control; irradiation only for 2 hr; and Blank. Twenty-four hours posttransfection, mRNA and protein levels of MMP-I, MMP-III, collagen-I and collagen-III were determined using standard RT-PCR and ELISA kits. Results: FAM-labeled C-Jun siRNA showed 80%–90% transfection efficiency. There was a significant increase in MMP-I and MMP-III and decrease in Col-I and III mRNA levels when the cells were exposed to UV irradiation without siRNA transfection compared to blank (p