Cancer-Associated Fibroblasts Autophagy Enhances Progression of Triple-Negative Breast Cancer Cells.

BACKGROUND Cancer-associated fibroblasts (CAFs) are key factors in malignant tumor initiation, progression, and metastasis. However, the effect of CAFs autophagy on triple-negative breast cancer (TNBC) cells is not clear. In this study, the growth effect of TNBC cells regulated by CAFs autophagy was evaluated. MATERIAL AND METHODS CAFs were obtained from invasive TNBC tumors and identified by Western blot and immunofluorescence staining assay. CAFs were co-cultured with TNBC cells, and migration and invasion were evaluated by Matrigel-coated Transwell and Transwell inserts. TNBC cells growth was detected by MTT assay, and epithelial-mesenchymal transition (EMT) regulated by CAFs was evaluated by Western blot assay. RESULTS CAFs were identified by the high expression of α-smooth muscle actin (α-SMA) protein. Autophagy-relevant Beclin 1 and LC3-II/I protein conversion levels in CAFs were higher than those in NFs (P<0.05). TNBC cells migration, invasion, and proliferation levels were significantly improved in the CAFs-conditioned medium (CAFs-CM) group, compared with the other 3 groups (P<0.05). TNBC cells vimentin and N-cadherin protein levels were upregulated and E-cadherin protein level was downregulated in the CAFs-CM group compared with the control group (P<0.05). Further study indicated b-catenin and P-GSK-3ß protein levels, which are the key proteins in the Wnt/ß-catenin pathway, were upregulated in the CAFs-CM group compared with the control group (P<0.05). CONCLUSIONS Our data demonstrated CAFs autophagy can enhance TNBC cell migration, invasion, and proliferation, and CAFs autophagy can induce TNBC cells to engage in the EMT process through the Wnt/ß-catenin pathway.

Genotoxicity of 1-methylpyrene and 1-hydroxymethylpyrene in Chinese hamster V79-derived cells expressing both human CYP2E1 and SULT1A1.

1-Methylpyrene (1-MP) is a widespread pollutant that is carcinogenic in animals following metabolic activation. Previous studies have shown that benzylic hydroxylation of 1-MP, catalyzed by multiple CYP isoforms, gives rise to 1-hydroxymethylpyrene (1-HMP), which becomes bioreactive following further metabolism by various sulfotransferase (SULT) isoforms. However, the mutagenic and chromosome damaging effects of 1-MP and 1-HMP in mammalian cells have not been investigated. In this study a Chinese hamster V79-derived cell line expressing both human CYP2E1 and human SULT1A1 was used to investigate the ability of 1-MP and 1-HMP to induce cytotoxicity (using the CCK-8 assay), micronuclei and Hprt gene mutations. The role of each enzyme was investigated through co-exposure in the presence of an enzyme inhibitor. We found that at concentrations of 0.5-4 µM and 5-20 µM, under conditions where no reduction in cell viability/growth occurred, 1-HMP and 1-MP induced micronuclei in V79-hCYP2E1-hSULT1A1 cells in a concentration-dependent manner; however, both compounds were inactive in V79 cells. Similarly, they both caused an increase in Hprt mutant frequency in V79-hCYP2E1-hSULT1A1 cells in these concentration ranges, with 1-MP impairing cell viability/growth at 10 µM and above in the mutagenicity assay. The compounds were again both inactive in V79 cells. The effects of 1-HMP in V79-hCYP2E1-hSULT1A1 cells were blocked or reduced by addition of pentachlorophenol (PCP), a SULT1 inhibitor; the genotoxicity of 1-MP was significantly reduced by either 1-aminobenotrazole, a CYP2E1 inhibitor, or PCP. The results suggest that human CYP2E1 and SULT1A1 cooperate to activate 1-MP and cause genotoxicity in mammalian cells.