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Enhanced anti-fibrogenic effects of novel oridonin derivative CYD0692 in hepatic stellate cells

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Abstract

Oridonin, isolated from Rabdosia rubescens, has been proven to possess various anti-neoplastic and anti-inflammatory properties. Previously, we reported the anti-fibrogenic effects of oridonin for liver in vitro. In the present study, we investigated the effects of a newly designed analog CYD0692 in vitro. Cell viability was measured by Alamar Blue assay. Cell apoptosis was assessed by Cell Death ELISA and Yo-Pro-1 staining. Western blots were performed for cellular proteins. Flow cytometry was used to measure cell cycle regulation. CYD0692 significantly inhibited LX-2 cells proliferation in a dose- and time-dependent manner with an IC50 value of ~0.7 μM for 48 h, ~tenfold greater potency than oridonin. Similar results were observed in HSC-T6 cells. In contrast, on the human hepatocyte cell line C3A, only 12 % of the cell growth was inhibited with 5 μM of CYD0692 treatment for 48 h, while 30 % inhibited at 10 μM. After CYD0692 treatment on LX-2 cells, apoptosis and S-phase cell cycle arrest were induced; cleaved-PARP, p21, and p53 were activated while cyclin-B1 levels declined. In addition, α-smooth muscle actin, type I Collagen, and fibronectin (FN) were markedly down regulated. Transforming growth factor β1 (TGF β1) has been identified as a dominant stimulator for ECM production in HSC. Our results indicated that pretreatment with CYD0692 blocked TGF β1-induced FN expression, thereby decreasing the downstream factors of TGF β1 signaling, such as Phospho-Smad2/3 and phospho-ERK. In comparison with oridonin, its novel derivative CYD0692 has demonstrated to be a more potent and potentially safer anti-fibrogenic agent for the treatment of hepatic fibrosis.

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Acknowledgments

This work was supported by Grants P50 CA097007, P30 DA028821, R21 MH093844 (JZ), and T32-GM8256 (FJB) from the National Institutes of Health, Cancer Prevention Research Institute of Texas (CPRIT) award, R. A. Welch Foundation Chemistry and Biology Collaborative Grant (JZ) from the Gulf Coast Consortia, BP Fellows Endowment (AP), Shriner’s Hospital for Children (SHC 80100) (AP), Anderson Foundation (AP), and John Sealy Memorial Endowment Fund, and the Center for Addiction Research (JZ) from the University of Texas Medical Branch. We would also like to thank Karen Martin for her generous help in preparing our data for publication. We have no conflicts of interest to disclose.

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Correspondence to Jia Zhou or Ravi S. Radhakrishnan.

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Bohanon, F.J., Wang, X., Graham, B.M. et al. Enhanced anti-fibrogenic effects of novel oridonin derivative CYD0692 in hepatic stellate cells. Mol Cell Biochem 410, 293–300 (2015). https://doi.org/10.1007/s11010-015-2562-4

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