Inhibition of high-mobility group box 1 expression by siRNA in rat hepatic stellate cells.
OBJECTIVE
To explore the role of high-mobility group box 1 (HMGB1) protein during liver fibrogenesis and investigate the functional effects of HMGB1 gene silencing in hepatic stellate cells (HSCs) using siRNA.


METHODS
Hepatic fibrosis in rats was induced throu-gh serial subcutaneous injections of dimethylnitrosamine, and expression of HMGB1 was detected by immunohistochemistry. HMGB1 siRNAs were developed and transiently transfected into HSC-T6 cells using Lipofectamine 2000. HMGB1 expression was evaluated by real-time polymerase chain reaction (PCR) and Western blotting analysis. Expression of α-smooth muscle actin (α-SMA) and collagen types I and III was evaluated by real-time PCR. Cell proliferation and the cell cycle were determined using the methyl thiazolyl tetrazolium method. Finally, collagen content in HSC supernatant was evaluated by an enzyme-linked immunosorbent assay.


RESULTS
The results showed that HMGB1 was upregulated during liver fibrosis and that its expression was closely correlated with the deposition of collagen. siRNA molecules were successfully transfected into HSCs and induced inhibition of HMGB1 expression in a time-dependent manner. Moreover, HMGB1 siRNA treatment inhibited synthesis of α-SMA and collagen types I and III in transfected HSCs.


CONCLUSIONS
This study suggests a significant fun-ctional role for HMGB1 in the development of liver fibrosis. It also demonstrates that downregulation of HMGB1 expression might be a potential strategy to treat liver fibrosis.
Department of Gastroenterology, Shanghai Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai 200092, China.
Wen-Song
WS
Ge
Jian-Xin
JX
Wu
Jian-Gao
JG
Fan
Yao-Jun
YJ
Wang
Ying-Wei
YW
Chen
D000818:Animals; D002453:Cell Cycle / Q000235:genetics; D002460:Cell Line; D049109:Cell Proliferation; D004128:Dimethylnitrosamine / Q000494:pharmacology; D020868:Gene Silencing; D024243:HMGB1 Protein / Q000235:genetics / Q000378:metabolism*; D055166:Hepatic Stellate Cells / Q000502:physiology*; D008099:Liver / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008103:Liver Cirrhosis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008297:Male; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D024243:HMGB1 Protein; D034741:RNA, Small Interfering; D004128:Dimethylnitrosamine