[TNF-α regulates the proliferation of human breast cancer cells via regulation of ceramide content].

Objective To determine whether tumor necrosis factor α (TNF-α) regulates the proliferation of MCF-7 and MDA-MB231 cells by modifying ceramide (Cer) production. Methods The optimum dosage and time of TNF-α treatment was determined at first. Immunocytochemistry combined with confocal microcopy was adopted to measure Cer content. MCF-7 and MDA-MB231 cells were treated with TNF-α (50 ng/mL) alone or combined with the interference of ASM inhibitor, desipramine (Des) or sphingosine kinase 1 (SPHK1) inhibitor, dimethylsphingosine (DMS) to investigate the role of Cer in the proliferative regulation of TNF-α on breast cancer cells. MTT assay was used to determine the proliferation of the cancer cells and Western blotting was performed to measure the protein expressions of acid sphingomyelinase (ASM), SPHK1 and apoptosis or proliferation relevant factors including Bcl2, Bax and proliferating cell nuclear antigen (PCNA). Results TNF-α reduced the proliferation of MCF-7 cells, but enhanced the growth of MDA-MB-231 cells in a time- and dose-dependent manner. After TNF-α treatment, the protein expression of ASM, but not SPHK1 increased in MCF-7 cells, but both of them were elevated in MDA-MB-231 cells. Accordingly, Cer content was increase in MCF-7 cells treated by TNF-α, which was blocked by the pretreatment of ASM inhibitor Des. Cer content in MDA-MB231 cells was reduced by TNF-α, which was prevented by the pretreatment of SPHK1 inhibitor DMS. Des and DMS could respectively reverse the TNF-α-induced decrease and increase of proliferation in MCF-7 and MDA-MB-231 cells. Moreover, the protein expression of Bcl2 decreased and Bax increased in MCF-7 cells, and the protein expression of PCNA increased in MDA-MB231 cells after TNF-α treatment. Conclusion TNF-α can alter Cer production by regulating the expressions of ASM and SPHK1 to control the proliferation of MCF-7 and MDA- MB231 cells.