Objective: To observe the expression of Toll-like receptor 8 (TLR8) in human cervical cancer cell-line HeLa cells, and the effects of TLR8 agonist CL075 on the survival and proliferation of HeLa cells.
Methods: PCR and RT-PCR were used to detect the expression of TLR8 in 13 cancer cell lines, and the expression of COX-2, Bcl-2, VEGF mRNA in the HeLa cells stimulated by TLR8 agonist CL075 were also measured by RT-PCR. Immunofluorescence technique was used to determine the exact location of TLR8 in the cells. The percentage of viable cells was determined by trypan blue exclusion after the HeLa cells were stimulated with TLR8 agonist CL075 (0.1 µg/ml, 0.5 µg/ml, 1.0 µg/ml, 2.5 µg/ml), and cell cycle and apoptosis were analyzed by flow cytometry, and the proliferation was measured by MTT.
Results: Compared with the other cancer cell lines, the expression of TLR8 in HeLa cells was the highest (703.7 ± 20.6). After stimulation by CL075, the cells had a remarkable increase of the percentage of cells in G(2)/M + S phases. In the control group, the percentage of cells in G(2)/M +S phases was (39.02 ± 2.33)%, whereas after stimulated with 1.0 µg/ml CL075, the percentage of cells in G(2)/M + S phases reached the highest ratio (57.67 ± 1.73)%, and the percentage of cells in G(2)/M + S phases had a less decrease after 2.5 µg/ml CL075 stimulation and the percentage was (56.14 ± 3.73)%. After the CL075 treatment, there was no significant changes of apoptosis compared with that of the control cells (P > 0.05), but after DDP treatment the apoptosis had a significant change (P < 0.01). After stimulation by 1.0 µg/ml CL075 for 24 h, no significant difference (P > 0.05) was found by MTT test, but a significant difference was found at 48 h and 72 h (P < 0.01). An increased expression of COX-2, Bcl-2 and VEGF mRNA was observed in HeLa cells after stimulation by TLR8 agonist CL075 for 24 h and 48 h (P < 0.05).
Conclusions: Expression of TLR8 is significantly increased in HeLa cells. The proportion of cells at different phases has a significant change after CL075 stimulation, which may up-regulate the proliferation of HeLa cells. These data suggested that TLR8 agonist may influence the tumor development and TLR8 may become a potential target in the treatment for cervical cancer.