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. 2021 Jun 14;9:660005. doi: 10.3389/fcell.2021.660005

FIGURE 6.

FIGURE 6

Combination of sorafenib and TMZ-induced autophagy is associated with the inhibition of STAT3. (A) Western blot analysis showed that the combination of sorafenib and TMZ upregulated the level of LC3II and downregulated the expression of p62. Meanwhile, WP-1006 enhanced sorafenib and TMZ treatment-induced autophagy of U251 cells. (B) After the same treatment as (A), representative images under a fluorescence microscope revealed that the combination of sorafenib and TMZ induced the colocalization of p62 and LC3 at 12 h and WP-1006 enhanced the colocalization induced by the combination of sorafenib and TMZ in U251 cells (120×) (bar, 10 μm). (C) After the same treatment as (A), Western blot assay showed that the combination of sorafenib and TMZ increased the expression of autophagy-related protein Beclin 1 and WP-1006 promoted this upregulation of Beclin 1 in U251 cells. (D) After U251 cells were treated with 3-MA or CQ and the combination of sorafenib and TMZ, flow cytometry analysis with Annexin V and PI double staining showed that 3-MA or CQ prevented cell apoptosis induced by the combination of sorafenib and TMZ (*P < 0.05 and **P < 0.01 compared to the control group, #P < 0.05 versus the combination of sorafenib and TMZ).