Research On The Antiproliferative And Apoptosic Action Of Marsdenia Tenacissima Injection Alone And When Combined With Oxaliplation In Human Hepatoma Cell

Objective:Observe the effects of Marsdenia tenacissima injection alone and the effects when it is combined with oxaliplatin on human hepatoma cell lines HepG-2 in vitro, and investigate the mechanism, to provide the theoretical foundation for the combination of chemotherapy with traditional medicine to cure hepatocarcinoma.Methods:Use various methods to detect the growth condition of human hepatoma cell lines HepG-2 in vitro when they are affected by different concentration of Marsdenia tenacissima injection, oxaliplatin alone and by the combinative use of the drugs above: (1)Estimating the inhibition of proliferation by MTT-test; (2) Observing the morphologic changes by light microscope; (3) Observing the dyed nucleolus of apoptosis cell by TUNEL method; (4) Observing the morphologic characteristics of apoptosis by fluorescence microscope; (5) Analyzing the distribution of cell cycle and early apoptosis by flow cytometry; (6) Detecting the expressions of apoptosis-associated gene proteins:Survivin, Bcl-2 and Caspase-3 by immunohistochemical technique.Results:(1) Marsdenia tenacissima injection could inhibit the proliferation of HepG-2 cell lines, and the inhibition depends on the exposure time and dose; there were additive effects when combine Marsdenia tenacissima injection of 17.5μl/ml with oxaliplatin of 0.5μg/ml; (2) Observed by light microscope, the Cells shrank and turned round, and the contacts among cells became loose; (3) The TUNEL method detected that the nucleuses of apoptotic cells were dyed deep brown, the apoptosis index (AI) increased when combined with oxaliplatin, and the difference is significant statistically; (4)Observed by fluorescence microscope, the apoptosis took place in cells, the phenomenon become more obvious when Marsdenia tenacissima injection combined oxaliplatin(P<0.001); (5)The analysis of cell cycle showed that Marsdenia tenacissima injection blocked cells at G0/G1 phase while the combination of Marsdenia tenacissima injection and oxaliplatin blocked cells at S phase; The analysis of cell cycle showed that Marsdenia tenacissima injection can make cells'apoptosis take place, the phenomenon become more obvious when Marsdenia tenacissima injection combined oxaliplatin (P<0.05); (6) The expressions of Caspase-3 were down-regulated (P<0.01), while the expressions of Survivin and Bcl-2 were up-regulated (P<0.01); The expressions of Caspase-3 was decreased while Survivin and Bcl-2 was increase when combined with oxaliplatin (P<0.01)Conclusion:Marsdenia tenacissima injection could inhibit the proliferation of HepG-2 cell lines, the effects were achieved by apoptosis related to the blocking of cell cycle at G0/G1 phase, down-regulation of the expressions of Caspase-3 and up-regulation of the expressions of Survi-vin and Bcl-2. When combined with oxaliplatin, it strengthened the inhibition of proliferation and inducement of apoptosis. This research provides the experimental foundation for the combination of TCM with western medicine to cure hepatocarcinoma and it has a degree of significance.