The Study On The Effects Of Giesenoside Rh₂ On SP2/O Myeloma Cells And Its Mechanism

Multiple myeloma(MM) is the prototype of a clonal B-cell tumor of differentiated and usually proliferating plasma.This disorder is frequently accompanied by monoclonal protein production and diffuse osteoporosis or lytic bone lesions. It is one of the malignant diseases for human being.The occurrence of myeloma has relationship with many cytokines, such as interleukin-6 (IL-6),Tumor necrosis factor-alpha (TNF-α),Insulin growth factor-binding protein-4 and macrophage inflammatory protein-1α(MIP-1α). The previous studies showed that the cytokines mentioned above play great important role in the growth and development of myeloma;They can stimulate the development of the osteoclasts, and has close relationship with the development and prognosis of myeloma, the higher concentration, the worse prognosis of cases. The symptom of cases could be lessened, the prognosis could be better and the life quality improved if these cytokines were inhibited. Ginsenoside (G) is the main active ingredient in Ginsen, Araliaceae. It has biological effects on anti-aging, anti-fatigue, anti-aggregation of platelets and radioprotection. Recently, more and more studies showed that G plays an important role on cancer prevention and treatment. G has many ways to anti-tumor by Directly retaining the growth of tumor cells, Inducing the cell apoptosis and differentiate, Inhibiting the metastasis of tumor through many channels, Enhancing the sensitivity of chemotherapy though reversing the resistance to chemical medicine and Adjusting the activity of many enzymes in cell or enhancing the immunity so that indirectly inhibit the growth of tumor cells. It was found that G has effects on many kinds of tumor cells in the literature, but without information on myeloma cells yet. Objective: This study was try to explore the effects of Gin-senoside Rh2 on mouse myeloma cell line SP2/O; And themechanism of Rh2 effects on myeloma. Methods: The G-Rh2 were added in the culture of SP2/O cell line when the exponential proliferation phase was showed. The check on cell proliferation was studied with MTT method. The apoptosis cell ratio and changes of cell cycle was analyzed with flow cytometry. The morphology of apoptosis cells were observed with conventional Giemsa staining and DAPI counterstained. DNA ladder of apoptosis were checked on regular agarose gel. ELISA method was used to measure the TNFα,MIP-1αon the culture supernatant of SP2/O. Results 1 The proliferation of SP2/O cell line was inhibited by G-Rh2, and the inhibition was dose and time-dependent in certain dose range of Rh2:The proliferation of SP2/O was significantlyinhibited in dose range of 10μg/ml to 120μg/ml (P<0.05) according to the 0μg/ml group.And the inhibition was enhanced with the G-Rh2 dose level and time. 2 G-Rh2 did not change the level of TNF-αin SPS2/0:The levels of TNF-αwere not changed in dose range of 20μg/ml to 100μg/ml for 24 hrs. or 48 hrs.,compared with the 0μg/ml group (P>0.05). 3 G-Rh2 did change the level of MIP-1αin SPS2/0: The levels of MIP-1αwere significantly decreased in dose range of 20μg/ ml to 100μg/ml for 24 hrs. or 48 hrs.,compared with the 0μg/ ml group (P<0.05). 4 G-Rh2 could induce the apoptosis of SP2/O: The cells were collected after G-Rh2 treatment for certain hours,and the flow cytometry was used to analyze the apoptosis cell rate. The ratio of apoptosis in 40μg/ml to 100μg/ml for 24 hrs. was significantly higher than that of 0μg/ml (P<0.05), but the ratio of apoptosis in 20μg/ml was not significantly higher than that of 0μg/ml (P>0.05). The ratio of apoptosis in 20μg/ml to 100 μg/ml for 48 hrs. was significantly higher than that of 0μg/ml (P<0.05). 5 G-Rh2 could be increased the cells in S phase in all dose levels of 20μg/ml to 100μg/ml,compared with that of 0μg/ml (P<0.05). 6 The morphology of SP2/O showed the characteristic apoptosis change after G-Rh2 treatment,such as condensation of chromatin,bubble,marginal chromatin and apoptosis body.7 The characteristic DNA ladder was showed in SP2/O cells after G-Rh2 treatment. Conc...