Inhibition of COL1A1 and COL3A1 expression by RNA interference in human skin fibroblasts.
- Author:
Qiong WANG
1
;
Zhen-Hui PENG
;
Wan-Juan WANG
Author Information
- Publication Type:Journal Article
- MeSH: Blotting, Western; Cells, Cultured; Collagen Type I; biosynthesis; genetics; Collagen Type III; biosynthesis; genetics; Fibroblasts; cytology; metabolism; Humans; RNA Interference; RNA, Small Interfering; genetics; Reverse Transcriptase Polymerase Chain Reaction; Skin; cytology; Transfection; methods
- From: Journal of Southern Medical University 2008;28(1):1-6
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo suppress COL1A1 and COL3A1 gene expressions in human skin fibroblasts (HSFs) by means of RNA interference (RNAi).
METHODSThree small interfering RNA (siRNA) expression cassette (SEC) sequences were designed for each of the COL1A1 and COL3A1 gene sequences available in GenBank. The synthesized SECs capable of effective gene suppression were transfected into cultured HSFs, either after cloning into the expression vector or mediated by Lipofectamine 2000, and the suppression of the target genes at both mRNA and protein levels was determined by quantitative fluorescence RT-PCR and Western blotting, respectively.
RESULTSTransfection of the SECs into HSFs resulted in specific depression of COL1A1 and COL3A1 expressions (down to 5.00% and 6.48%, respectively). The expression vector-mediated RNAi established a HSF cell line with persistent gene knockdown for over 30 days (to 25.21% and 22.12%, respectively).
CONCLUSIONCOL1A1 and COL3A1 gene expressions can be specifically and efficiently inhibited in HSFs by either liposome- or vector-mediated SEC transfection.
