Effects from combined exposure to arsenic and dichloromethane: in vitro expression of genotoxicity, transformation and metastatic potential for cholangiocarcinogenesis.

Cholangiocarcinoma (CCA) is a common and devastating cancer in Thailand and in certain Asian countries. Among risk factors for CCA, exposure to environmental carcinogens such as arsenic and dichloromethane (DCM) has gained attention, especially because such exposure is preventable. Using a human cholangiocyte cell line (MMNK-1) and a CCA cell line (HuCCA-1), our study aimed to identify effects from exposure to arsenite alone and in combination with DCM on pathways associated with CCA development. MMNK-1 cells treated with 0.5 μM arsenite showed significantly increased formation of 8-OHdG and 8-nitroguanine; altered expression of genes and proteins involved in carcinogenesis, as well as increased cell transformation. Treatment with arsenite combined with 25 μM DCM (24 h) showed additive effects regarding the formation of 8-OHdG (1.88-fold), expression of CXCL8 and MMP9, as well as increased cell transformation. The synergistic effects observed in 8-nitroguanine (3.66-fold) highlighted the significant impact of combined-treatment on DNA damage formation. Transformed cells were subsequently cultured to form spheroids, which exhibited increased levels of CD133 and ECT2 in all treated cultures. Furthermore, the migration and invasion capabilities of HuCCA-1 cells increased with arsenite treatment compared to the control, with more pronounced increase from combined-treatment. This change was consistent with the increases in CXCL8 and MMP9 levels. Our data indicates that arsenite and DCM are relevant environmental chemicals which induced cellular changes that are consistent with cholangiocarcinogenesis. Our in vitro approach, like others, may also be useful as a valuable compliment to in vivo carcinogenesis investigations.