Published online Jan 21, 2015. doi: 10.3748/wjg.v21.i3.878
Peer-review started: April 24, 2014
First decision: May 29, 2014
Revised: June 18, 2014
Accepted: July 15, 2014
Article in press: July 16, 2014
Published online: January 21, 2015
Processing time: 272 Days and 0 Hours
AIM: To determine the role of Notch1 and Hes1 in regulating the activation of hepatic stellate cells (HSCs) and whether Hes1 is regulated by transforming growth factor (TGF)/bone morphogenetic protein (BMP) signaling.
METHODS: Immunofluorescence staining was used to detect the expression of desmin, glial fibrillary acidic protein and the myofibroblastic marker α-smooth muscle actin (α-SMA) after freshly isolated, normal rat HSCs had been activated in culture for different numbers of days (0, 1, 3, 7 and 10 d). The expression of α-SMA, collagen1α2 (COL1α2), Notch receptors (Notch1-4), and the Notch target genes Hes1 and Hey1 were analyzed by reverse transcriptase-polymerase chain reaction. Luciferase reporter assays and Western blot were used to study the regulation of α-SMA, COL1α1, COL1α2 and Hes1 by NICD1, Hes1, CA-ALK3, and CA-ALK5 in HSC-T6 cells. Moreover, the effects of inhibiting Hes1 function in HSC-T6 cells using a Hes1 decoy were also investigated.
RESULTS: The expression of Notch1 and Hes1 mRNAs was significantly down-regulated during the culture of freshly isolated HSCs. In HSC-T6 cells, Notch1 inhibited the promoter activities of α-SMA, COL1α1 and COL1α2. On the other hand, Hes1 enhanced the promoter activities of α-SMA and COL1α2, and this effect could be blocked by inhibiting Hes1 function with a Hes1 decoy. Furthermore, co-transfection of pcDNA3-CA-ALK3 (BMP signaling activin receptor-like kinase 3) and pcDNA3.1-NICD1 further increased the expression of Hes1 compared with transfection of either vector alone in HSC-T6 cells, while pcDNA3-CA-ALK5 (TGF-β signaling activin receptor-like kinase 5) reduced the effect of NICD1 on Hes1 expression.
CONCLUSION: Selective interruption of Hes1 or maintenance of Hes1 at a reasonable level decreases the promoter activities of α-SMA and COL1α2, and these conditions may provide an anti-fibrotic strategy against hepatic fibrosis.
Core tip: In our paper, we found that Notch signaling appears to crosstalk with TGF-β/BMP signaling and co-regulates the expression of Hes1 in hepatic stellate cells. Notch1 may keep Hes1 protein in a reasonable range due to Hes1 auto-negative feedback, allowing Notch1 to repress the expression of α-SMA and collagen1α2 (COL1α2). Over-expression of Hes1 via transfection, thus lacking the negative feedback, enhances the promoter activities of α-smooth muscle actin (α-SMA) and COL1α2, consistent with the finding when Hes1 is captured by a Hes1-decoy oligodeoxynucleotide. This may provide an anti-fibrotic strategy against hepatic fibrosis. Further investigations in the future on the molecular mechanism by which Hes1 regulates the expression of α-SMA and COL1α2 should provide additional support for such an approach.