In vivo transfer of small interfering RNA or small hairpin RNA targeting glomeruli.
Small synthetic interfering RNA duplexes (siRNAs) can selectively suppress gene expression in somatic mammalian cells without the nonselective toxic effects associated with double-stranded RNA (dsRNA). However, in vivo delivery of siRNA targeting the kidney has been described in only a few reports. We have found that injection of synthetic siRNAs via the renal artery, followed by electroporation, can be therapeutically effective in silencing the expression of specific genes in the glomerulus. Here we provide details of an experimental protocol showing that 1) delivery of siRNA targeting enhanced green fluorescent protein (EGFP) to the kidney in the transgenic "green" rat reduces endogenous EGFP expression, mainly in the glomerular mesangial cells, and that 2) delivery of siRNA targeting transforming growth factor (TGF)-beta1 to the kidney significantly suppresses messenger RNA (mRNA) and protein expression of TGF-beta1, thereby ameliorating the progression of matrix expansion in experimental glomerulonephritis. In addition, we describe the application of vector-based RNA interference (RNAi) (small hairpin RNA [shRNA]), which also inhibits TGF-beta1 expression in vivo.
Takabatake
Yoshitsugu
Y
Department of Nephrology, Osaka University Graduate School of Medicine, Osaka, Japan.
Isaka
Yoshitaka
Y
Imai
Enyu
E
D000818:Animals; D005456:Fluorescent Dyes / Q000378:metabolism; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007678:Kidney Glomerulus / Q000166:cytology / Q000502:physiology*; D009393:Nephritis / Q000139:chemically induced / Q000503:physiopathology; D009690:Nucleic Acid Conformation; D034741:RNA, Small Interfering / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism
D005456:Fluorescent Dyes; D034741:RNA, Small Interfering; D053773:Transforming Growth Factor beta1; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins