[Effects of anti-sense Smad4 gene on the biological characteristics of the fat-storing cell line CFSC].
OBJECTIVE
To investigate the effects of antisense Smad(4) on the biological characteristics of the fat-storing cell line CFSC.


METHODS
Fat-storing cells of line CFSC from rat with liver fibrosis were cultured and transfected with 50 MOI of recombinant adenoviral vector carrying antisense Smad(4) (AdvATSmad(4)) or the control empty adenovirus (Adv0), both produced by 293 packaging cells, respectively. Two, four, and six days after the transfection the cultured cells were collected to undergo trypan blue staining and cell counting. The growth curves were drawn. The presence of antisense Smad(4) was detected by RT-PCR and Western blotting. (3)H-TdR was added into the culture media to be co-cultured for 6 hours. Then the cells were collected to examine the (3)H-TdR incorporation rate. RT-PCR and immunohistochemistry were used to examine the expression of COL1A1 and type I collagen, kinds of extracellular matrix (ECM).


RESULTS
Compared with the control CFSC and the Adv0-transfected CFSC cells, the cell growth curve, (3)H-TdR incorporation rate, proline incorporation rate, expression of Smad(4), and expression of extracellular matrix were markedly decreased in the AdvATSmad(4)-transfected CFSCs.


CONCLUSIONS
The antisense Smad(4) gene inhibits the expression of Smad(4) mRNA and protein, proline incorporation and cell growth, thus down-regulating the production of ECM. Antisense Smad(4) gene may be used as a choice of gene therapy for liver fibrosis.
Xu
Xin-bao
XB
Department of Hepatobiliary Surgery, People's Hospital, Peking University, Beijing 100044, China.
Leng
Xi-sheng
XS
He
Zhen-ping
ZP
Liang
Zhi-qing
ZQ
Lin
Kai
K
Yu
Xin
X
Wei
Yu-hua
YH
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D015153:Blotting, Western; D002455:Cell Division; D002460:Cell Line; D024042:Collagen Type I / Q000378:metabolism; D016373:DNA, Antisense / Q000235:genetics / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D008099:Liver / Q000378:metabolism / Q000473:pathology; D011392:Proline / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051901:Smad4 Protein; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D024042:Collagen Type I; D016373:DNA, Antisense; D004268:DNA-Binding Proteins; D051901:Smad4 Protein; C490811:Smad4 protein, rat; D015534:Trans-Activators; D011392:Proline