自噬调控对JAK2 V617F阳性HEL细胞及真性红细胞增多症患者造血细胞增殖的影响

OBJECTIVE: To detect the activity of autophagy and explore the impact on survival and proliferation of HEL cells and hematopoietic cells of polycythemia vera (PV) patients with JAK2 V617F mutation. METHODS: Flow cytometry, AO staining and Western blot methods were used to detect the autophagy activity and the expression of LC3-Ⅱ protein of JAK2 V617F(+) HEL cells and hematopoietic cells of 12 newly diagnosed PV patients with JAK2 V617F mutation. HEL cells and bone marrow cells of 3 PV patients were treated with rapamycin or 3-MA to induce and inhibit autophagy, respectively. CellTiter Glo(®)method was used to detect the proliferation activity of cells. RESULTS: There was higher level of mean LC3-Ⅱ fluorescence intensity in HEL cells (159 389±29 001) than that in K562 cells (96 047±24 134) (P=0.044). The formation of autophagosome in HEL cells is more than that in K562 cells detected by microscope. What's more, the level of mean LC3-Ⅱ fluorescence intensity in 12 PV patients' myeloid cells (92 842±4 250) was higher than that of 15 healthy volunteers (86 633±2 504) (P=0.001). The expression of LC3-Ⅱ protein was higher in PV patients' peripheral blood cells than that in healthy volunteers detected by Western blot. After treated with rapamycin 12, 24, 48 h, the activity of autophagy in HEL cells and bone marrow cells of 3 PV patients were increased and the proliferation activity was higher than the control group, the proliferation activity at 48 h were (101 413±3 720), (18 744±1 015), respectively. However, after treated with 3-MA 12, 24, 48 h, the activity of autophagy was decreased and the proliferation activity was lower than the control group, the proliferation activity at 48 h were (5 732±166), (5 371±56), respectively. CONCLUSION: There is high basical activity of autophagy in JAK2 V617F(+) HEL cells and hematopoietic cells of PV patients with JAK2 V617F mutation. Up-regulated autophagy promotes proliferation of JAK2 V617F(+) HEL cells and bone marrow cells of PV patients with JAK2 V617F mutation. Decreased autophagy inhibits proliferation of JAK2 V617F(+) HEL cells and bone marrow cells of PV patients with JAK2 V617F mutation.