[Regulatory Effect of MiR-543 on Proliferation and Apoptosis of Human Leukemia Cell Line K562 and Its Mechanism].

OBJECTIVE: To investigate the regulatory effect of miR-543 on proliferation and apoptosis of human leukemia cell line K562 and its mechanism. METHODS: 46 CML patients treated in our hospital from 2018.2-2018.9 was selected and enrolled in CML group, and 30 healthy persons were selected and enrolled in control group at the same time. After Lipofectamine 2000 liposome was used to transfer the miR-543 mimic and negative control (Scramble mimic) to K562 cells, CCK8 assay was used to detect the effect of miR-543 on proliferation of K562 cells; Luciferase reporter assay was used to report the targeting relationship of miR-543 to Wnt gene. Flow cytometry was used to detect the effect of miR-543 on apoptosis of K562 cells; Western blot method was used to detect the Wnt, ß-catenin, BCL-2, c-MYC and BAX expression level. RESULTS: The level of miRNA-543 in CML patients was significantly lower than that in healthy controls (P＜0.05). The expression level of miR-543 in mimic group was significantly higher than that in blank control group (P＜0.05). The Wnt gene expression level in mimic group was significantly lower than that in blank control group (P＜0.05). The expression of luciferase of Wnt wild plasmid was decreased by miR-543 mimic (P＜0.05), and the luciferase activity of Wnt mutant plasmid was not inhibited by miR-543 mimic after mutation of binding site (P＜0.05). There was no significant difference in the proliferation ability of K562 cells between the blank control group and the negative control (Scramble mimic) group (P＞0.05). The proliferation level of K562 cells in mimic group was significantly lower than that in blank control group and negative control (Scramble mimic) group (P＜0.05). Apoptotic level of K562 cells in miR-543 mimic group was significantly higher than that in blank control group and negative control (Scramble mimic) group (P＜0.05). Western blot assay showed that Wnt, ß-catenin, BCL-2 and c-MYC protein levels in miR-543 mimic group were significantly lower than those in blank control group and negative control (Scramble mimic) group (P＜0.05); BAX protein level in miR-543 mimic group was higher than that in blank control group and negative control (Scramble mimic) group (P＜0.05). CONCLUSION: miR-543 can target Wnt protein to inhibit the activity of Wnt signaling pathway, thereby inhibiting the proliferation of leukemia cells and increasing the level of apoptosis.

Down regulation of G protein-coupled receptor 137 expression inhibits proliferation and promotes apoptosis in leukemia cells.

BACKGROUND: G protein-coupled receptors (GPR) are involved in a wide range of physiological processes, some of which, however, can be hijacked by tumor cells. Over-expression of G protein-coupled receptors 137 (GPR137) are associated with the growth of tumor cells, but under-expression of GPR137 has shown to inhibit cell proliferation in several different types of cancers. Currently, the role of GPR137 in leukemia is still unclear. In this study, the effect of under-expression of GPR137 on inhibiting the proliferation of leukemia cells is explored, to identify a novel target for leukemia treatment. MATERIALS AND METHODS: In this study, lentivirus-mediated RNA interference (RNAi) was employed to investigate the role of GPR137 in two leukemia cell lines K562 and HL60. The gene expression of GPR137 was analyzed by RT-PCR and its protein expression was determined by Western blot. Flow cytometry and Annexin V/7-AAD Apoptosis Detection Kit was used respectively in cell cycle and apoptosis analysis. The protein expression of CyclinD1, CDK4, BCL-2 and caspase-3 were also determined. RESULTS: There was high level of constitutive expression of GPR137 in leukemia cancer cell lines K562 and HL60. Lentivirus-mediated RNAi could significantly down-regulate gene and protein expression of GPR137 in both cell lines. Down regulation of GPR137 was associated with the reduction in proliferation rate and colony forming capacity. In addition, down regulation of GPR137 arrested cells in the G0/G1 phase of cell cycle and induced apoptosis in both leukemia cell lines K562 and HL60. CONCLUSIONS: The expression of GPR137 is associated with the proliferation of leukemia cell lines. Down regulation of GPR137 could inhibit proliferation and promote apoptosis in leukemia cells, which makes it a promising bio-marker and therapeutic target to treat patients with leukemia.