Figure 6.

Cucurbitacin I inhibits M2 polarization of macrophages by decreasing the expression of Hmox1 and augments the tumoricidal activity, which can be reversed by hemin. (A–D) Cell viability (%) of HCT116 cells and MCF7 cells was assayed, including those cocultured with macrophages alone, with macrophages after being treated with cucurbitacin I, or in a triple coculture system including macrophages and cucurbitacin I under normoxic or hypoxic conditions. (E) Hemin reversed the mRNA level of HMOX1 downregulated by cucurbitacin I cocultured with MCF7 cells, cocultured with HCT116 cells, or under hypoxia. (F–H) Hemin restored the protein levels of HMOX1 downregulated by cucurbitacin I, either in a coculture system with HCT116 cells (F), in a coculture system with MCF7 cells (G), or under hypoxia (H). (I,J) Cell migration and invasion capacity of HCT116 alone or cocultured with macrophages (alone, cucurbitacin I-treated macrophages, or both cucurbitacin I and hemin-treated macrophages), as determined by a wound-healing assay and Transwell coculture system, respectively. The concentration of hemin is 1 μM, and the concentration of cucurbitacin I is 500 nM. Scale bar represents 100 μm (I) and 50 μm (J). The values presented are expressed as the mean ± standard deviation (SD) and are based on data obtained from three biologically independent samples. No significance: ns, p < 0.05: *, p < 0.01: **, p < 0.001: ***.