DNAzyme for TGF-beta suppressed extracellular matrix accumulation in experimental glomerulonephritis.
BACKGROUND
We developed an electroporation-mediated gene transfer method targeting glomerular mesangial cells. Injecting DNA solution via renal artery followed by electric pulses using tweezers-type electrodes could result in efficient transfection in mesangial cells. Therefore, this gene transfer system opened a feasible strategy to manipulate the function of several cytokines and growth factors in mesangial cells. Recently, a new generation of catalytic nucleic acid composed of DNA, named DNA enzyme (DNAzyme), has been developed.


METHODS
We generated a DNAzyme (TGFDE) targeting transforming growth factor-beta1 (TGF-beta1), and examined the therapeutic effect of TGFDE in vitro and in vivo.


RESULTS
In cultured rat mesangial cells, treatment with TGFDE blocked TGF-beta1 mRNA expression, and thereby suppressed type I collagen mRNA expression. Next, we introduced TGFDE or scrambled DNAzyme (TGFSCR) into anti-Thy-1 model of nephritic rats by electroporation 3 days after disease induction. Northern blot analysis and immunohistochemical staining demonstrated that glomerular message and protein expression of TGF-beta1, alpha-smooth muscle actin (alpha-SMA), and type I collagen were suppressed in TGFDE-transfected nephritic rats compared with untreated nephritic rats and TGFSCR-transfected rats on day 7. Consequently, we observed significant reduction in glomerular matrix score in TGFDE-transfected nephritic rats.


CONCLUSIONS
Inhibition of TGF-beta1 expression by electroporation-mediated DNAzyme transfer might be useful for the therapy of glomerulonephritis.
Isaka
Yoshitaka
Y
Department of Internal Medicine and Therapeutics, Osaka University Graduate School of Medicine, Osaka, Japan. isaka@medone.med.osaka-u.ac.jp
Nakamura
Hiroyuki
H
Mizui
Masayuki
M
Takabatake
Yoshitsugu
Y
Horio
Masaru
M
Kawachi
Hiroshi
H
Shimizu
Fujio
F
Imai
Enyu
E
Hori
Masatsugu
M
D000199:Actins / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics; D021881:DNA, Catalytic / Q000235:genetics* / Q000378:metabolism; D018274:Electroporation / Q000379:methods*; D005109:Extracellular Matrix / Q000378:metabolism; D015870:Gene Expression; D005920:Glomerular Mesangium / Q000166:cytology / Q000378:metabolism*; D005921:Glomerulonephritis / Q000378:metabolism* / Q000503:physiopathology / Q000628:therapy; D051381:Rats; D016212:Transforming Growth Factor beta / Q000235:genetics* / Q000378:metabolism; D053773:Transforming Growth Factor beta1
D000199:Actins; D024042:Collagen Type I; D021881:DNA, Catalytic; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1