Effect Of Total Saponins Of Panaxginseng On JAK2,STAT Of K562 Cells

Leukemia is a malignant haematopoietic tumor which threatens the health of humanity. It's pathogeny is due to abnormal proliferation and disdifferentiation of bone marrow haematopoietic stem cells.So far, traditional chemotherapy and radiotherapy are still the main methods, but they cause many side effects and high recurrence rate, and greatly damage the normal immunity and haematopoiesis system too. More and more researches have shown that the traditional Chinese medicine may play an important role in the cancer therapy for it's little side affection on human body and many other advantages. Therefore, finding natural traditional Chinese medicine which has the effect of promoting normal haematopoiesis and inhibiting the proliferation of leukemia cells becomes much attractive issue to treat leukemia.Objective:Our previous study suggested that TSPG can not only improve the proliferation and differentaition of haematopoietic stem/progenitor cell, but also inhibit the proliferation of leukemia cells, and induce them apoptosis and differentiation toward the mature cell. But these mechanism remain to be studied. Some researches show that JAK2 and STAT play an important role in the proliferation and differentiation of cells. So We study on the signal transduction way and chose JAKs/STATs pathway to clarify the mechanism of the regulation of TSPG on haematogenesis in order to establish a foundation for pursuit the molecule target of TSPG acting on proliferation , differentiation and apoptosis of leukemic cell, and provided a guidance for clinic treatment of haematologic disease.Methods:1,Techniques of cell culture were used in this experiment. The effect of TSPG on proliferation of K562 was examined by MTT and Flow Cytometry ; 2,The change of Hemoglobin of K562 treated with TSPG was examined by UV spectrophotometer; 3,we adopted Immuocytochemistry to detect the effect of TSPG on expression of JAK2 in K562 cell; 4,The distribution of STAT5 of in K562 treated with TSPG was examined by laser scanning confocal microscope(LSCM); 5,We chose Western blotting was performed To detect the expression of JAK2,STAT5,STAT3,STAT1,P-JAK2,P-STAT5 and P-STAT3 of K562 cells treated at different times.Results: 1,TSPG could significantly inhibit the proliferation of K562 cells ex vivo in time and concentration-dependent manner; 2,After treated with TSPG, G0/G1% cells increased and more cells were detained at S cell cycle stage, which suggested that TSPG significantly inhibit the proliferation of K562 cells ex vivo through regulating cell cycle;3,TSPG promoted the synthesis of Hb of K562 cells ; 4,By means of Immunocytochemical stain for JAK2, The cytoplasma of K562 cells in both of the two groups is stained,and the expression of JAK2 in both groups have no statistical difference; 5,Under LSCM, fluorescence intensity in nucleus of K562 cells stimulated by 200 mg/L TSPG for 24 h was weaker than that of control group;6,As determined by Western blotting,the expression of STAT5 in nucleus of K562 cells decreased after TSPG(200 mg/L) treatment for 24h , while the expression of STAT5 in cytoplasm of K562 cell increased compared with control group; after treated for 24 h,48 h and 72 h, the expression of JAK2,STAT5,STAT1,STAT3 almost had no change compared with the control group;after treated for 24h the expression of phosphor-STAT5 decreased and Phosphor-JAK2 increased, for 48h the expression of phosphor-STAT5 notably decreased and Phospho-STAT3 greatly increased, and for 72h phosphor-STAT5 was almost the same with that of the control group, for 24h and 72h the expression of phosphor-STAT3 notably decreased; we treated K562 cells with TSPG and AG490 together, for 24 h,48 h and 72 h the expression of JAK2,STAT5,STAT1,STAT3 almost did not change; for 24h and 72h the expression of phosphor-STAT5 notably decreased , for 72h phosphor-STAT5 decreased compared with the control group; for 48h phosphor-STAT3 had apparently changed. Conclusion: TSPG can significantly inhibit the proliferation and induce the differentiation towards erythroid cell of K562 ex vivo;TSPG may play an important role on regulating the phosphor-JAk2,phosphor-STAT1,phosphor-STAT3 and phosphor-STAT5 to inhibit the proliferation and differentiation to the mature cells of K562 cells.