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. 2020 Jun 24;10:694. doi: 10.3389/fonc.2020.00694

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Copyright © 2020 Shi, Zhang, Ma, Yan, Zhang, Cao, Wang, Wu and Jiang.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Effects of exogenous S1P on the sensitivity of HCC cells to regorafenib. (A) The regorafenib sensitivity of HCC cells stimulated with 1 μM S1P was examined by the CCK-8 assay. We further used the (B) annexin V–FITC/propidium iodide double-staining assay, (C) cell cycle analysis, and (D) colony formation assay to measure regorafenib effects on HCC cells in different groups. (E) The expression levels of Bcl2, cleaved PARP, cyclin D1, CDK2, and CDK4 were examined by Western blot analysis. The dose of regorafenib treatment in all assays was 10 μM for 48 h, with the exception of the colony formation assay (5 μM, 14 days for SMCC-7721 and 10 days for MHCC-97H, respectively). The result is representative for three independent experiments. The error bars represent mean ± SD from a representative experiment. *p < 0.05, **p < 0.01, ***p < 0.001.