Experimental Study of Arbutin Regulating the Expression of TLR4 and Affecting the Proliferation, Cell Cycle and Apoptosis of Multiple Myeloma Cells RPMI-8226
TAO Jian1, WANG Ling1, ZHANG Liyu1, GU Zheyun1, ZHOU Xiaodan1, ZHANG Dongmei2*
The study aimed to investigate whether arbutin regulated the activation of TLR4 (Toll-like receptor 4)/JAK2/STAT3 (signal transducers and activators of transcription 3) signaling pathway to affect the proliferation, cell cycle and apoptosis of multiple myeloma cells RPMI-8226. RPMI-8226 cells were divided into control group (NC group), low-, medium- and high- dose groups (50, 100 and 200 µg/mL arbutin), si-NC group (transfected with si-NC), si-TLR4 group (transfected with si-TLR4), pcDNA-NC+high-dose group (transfected with pcDNA-NC and 200 µg/mL arbutin), pcDNA-TLR4+high-dose group (transfected with pcDNA-TLR4 and 200 µg/mL arbutin). The thiazole blue tetrazolium bromide (MTT) method was used to monitor cell proliferation. Flow cytometry was implemented to assay cell cycle and apoptosis. The protein expression levels of CyclinD1, Cleaved-caspase-3, TLR4, p-JAK2, and p-STAT3 were detected by Western blot. The qRT-PCR was used to measure TLR4 mRNA expression. Compared with the NC group, arbutin treatment dose-dependently decreased CyclinD1 and TLR4 expression, cell proliferation, and the cell proportion in S phase but increased the cell proportion in G0/G1 phase, Cleaved-caspase-3 expression and apoptotic rate in RPMI-8226 cells (P<0.05). Moreover, 200 µg/mL arbutin reduced the protein expression levels of p-JAK2 and p-STAT3 in RPMI-8226 cells (P<0.05). The CyclinD1 protein expression level, proliferation activity, and S-phase cell ratio of RPMI-8226 cells in the siTLR4 group were lower than those in the si-NC group, and the G0/G1 phase cell ratio, Cleaved-caspase-3 protein expression level and apoptosis rate were higher than those in the si-NC group (all P<0.05). pcDNA-TLR4 could reverse the effects of arbutin on the proliferation, cell cycle, apoptosis and protein expression of p-JAK2 and pSTAT3 in RPMI-8226 cells. Above all, arbutin inhibits the proliferation, arrests the cell cycle and promotes apoptosis of multiple myeloma cells RPMI-8226 by blocking the activation of TLR4/JAK2/STAT3 signaling pathways.
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