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Modulation of human melanoma cell proliferation and apoptosis by hydatid cyst fluid of Echinococcus granulosus
Abstract
Objective
The objective of this paper was to assess the effects of hydatid cyst fluid (HCF) of Echinococcus granulosus on melanoma A375 cell proliferation and apoptosis.
Methods
A375 cells were classified into five groups by in vitro culture: normal group, control group, 10% HCF group, 20% HCF group and 30% HCF group. Trypan blue staining method was employed to detect the toxicity of HCF. Effects of different concentrations of HCF on melanoma A375 cell proliferation at different time points were evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Flow cytometry and propidium iodide (PI) staining were used to detect cell cycle, and Annexin-V/PI double staining method was used to determine A375 cell apoptotic rate. Western blotting was applied to detect the expression of phosphorylated extracellular regulated protein kinases, proliferating cell nuclear antigen (PCNA), cell-cycle-related proteins (cyclin A, cyclin B1, cyclin D1 and cyclin E) and apoptosis-related proteins (Bcl-2, Bax and caspase-3).
Results
HCF with a high concentration was considered as atoxic to A375 cells. HCF promoted A375 cell proliferation, and the effects got stronger with an increase in concentrations but was retarded after reaching a certain range of concentrations. HCF increased phosphorylation level and expression of extracellular regulated protein kinase, as well as PCNA expression. HCF also promoted the transferring progression of A375 cells from the G0/G1 phase to the S phase to increase the cell number in S phase and increased the expression of cyclin A, cyclin D1 and cyclin E. HCF increased the expression of procaspase-3 (the precursor of apoptosis-related protein caspase-3) and antiapoptotic protein-Bcl-2, and decreased the expression of proapoptotic factor Bax, thereby inhibiting cell apoptosis.
