| Objective: By observing the the effection of Marsdenia tenacissima Injection, which combined with specific cell cycle of drugs, acting on cell cycle, apoptosis and growth of lung A549 cancer cell, and the related gene expression of Cyclin D1,Cyclin E and P53 m RNA, exploring its possible working mechanism.Methods: The inhibitory effect of was lung A549 cancer cell detected by MTT.The impaction of cell cycle and apoptosis were tested by flow cytometry. The expression of cyclin D1, cyclin E and P53 m RNA were detected by RT-PCR method.Results:1 By using MTT method, it showed that, in a certain range of concentration, the inhibition effect of Marsdenia tenacissima, fluorouracil and mitomycin, was positively related to the dosage and time. The24h、 48h、 72 h IC50 of Marsdenia tenacissima, fluorouracil and mitomycin, were 72.40 ul/ml 17.93 ul/ml 7.98ul/ml, 57.56ug/ml 15.02ug/ml 2.09 ug/ml, 1195.23 ug/ml 182.26 ug/ml 24.99 ug/ml respectively. In different period of time, the value of OD of Marsdenia tenacissima(2ul/ml, 8ul/ml, 32ul/ml) combined with mitomycin C(2 ug/ml) was higher than single drug group,(P<0.05), its OD value increased with the increase of concentration(P<0.05). Similarly, the value of OD Marsdenia tenacissima(2ul/ml, 8ul/ml, 32ul/ml) combined with fluorouracil were high than the single drug group,(P<0.05), its OD value increases with the increase of concentration of Marsdenia tenacissima(P<0.05).2 About to effection of cell cycle and apoptosis, the results showed Marsdenia tenacissima group and mitomycin group, compared with control group, the lung A549 cancer cell percentage of G0/G1 phase cells were significantly higher, 48.43% VS 34.99%, 50.69% VS 34.99%, P<0.05, respectively. Compared with fluorouracil in control group, the percentage of S phase cells in the lung A549 cancer cell were significantly higher than those in the control group, 50.40% VS 40.86%, P<0.05, with significant difference. And for apoptosis, Marsdenia tenacissima group, fluorouracil group and mitomycin group compared with the control group, the apoptosis rate of the lung A549 cancer cell was obviously higher than that of the control group, respectively 1.99%, 4.68%, 4.56%, 0.45%, respectively(P<0.05). Combination of Marsdenia tenacissima with mitomycin and fluorouracil, the apoptosis rate of lung A549 cancer cell was increased, from 4.68%, 4.56% to 9.56%, 13.45%, respectively, with significant difference(P<0.05).3 Through the examination of RT-PCR test, it was showed that Marsdenia tenacissima, fluorouracil and mitomycin could increase p53 m RNA expression, compared to control group,the relative expression quantity is 0.25. 0.45. 0.53. 0.09, respectively,(P<0.05). Marsdenia tenacissima combined with fluorouracil and mitomycin could enhance the expression of p53 m RNA, which had a significantly differentce with Marsdenia tenacissima group, fluorouracil group and mitomycin group, 0.80 VS 0.45 、0.83 VS 0.53,respectively,(P<0.05). About to the expression of Cyclin D1 gene, the results showed Marsdenia tenacissima, mitomycin and fluorouracil can reduce the expression of Cyclin D1 m RNA, 0.51. 0.46. 0.59. 0.88, respectively(P<0.05), the expression of Cyclin D1 m RNA can significantly reduce by the combination of Marsdenia tenacissima with mitomycin,(0.26 VS 0.46,P=0.015<0.05),but the effect of Marsdenia tenacissima combined with fluorouracil on the expression of Cyclin D1 gene was closed to Marsdenia tenacissima group( 0.53 VS 0.59, P=0.314>0.05),with non-significant difference. For the expression of Cyclin E gene, the results showed Marsdenia tenacissima group, mitomycin group, Marsdenia tenacissima with mitomycin in combination group compared with the control group, were not abled to reduce the expression of Cyclin E gene(0.81. 0.79. 0.88. 0.85,P>0.05). Fluorouracil compared with the control group, can reduce the expression of Cyclin E gene in lung A549 cancer cells(0.49 VS 0.85 P<0.05).But Marsdenia tenacissima combined with fluorouracil did not deline the expression of Cyclin E m RNA in lung compared to fluorouracil group( 0.56 VS 0.49 P=0.136>0.05), with no significant difference.Conclusions:1 Marsdenia tenacissima, fluorouracil and mitomycin have thefunction of inhibition effect on on lung A549 cancer cell, with time and dose dependence in each drug. In the certain scope, Marsdenia tenacissima combined with fluorouracil and mitomycin can enhance the inhibition effect on lung A549 cancer cell.2 Marsdenia tenacissima, fluorouracil, mitomycin can induce apoptosis of the lung A549 cancer cell, Marsdenia tenacissima combined with fluorouracil and mitomycin can improve the effection of apoptosis on lung A549 cancer cell, its mechanism may be related to the up expression of P53 m RNA.3 Marsdenia tenacissima, fluorouracil, mitomycin blocked the cell cycle of lung cancer A549 cells in G0/G1, S, G0/G1 phase, respectively. Marsdenia tenacissima combined with mitomycin can improve the effect of blocking of lung A549 cancer in G0/G1, the mechanism of action may be related with down-regulation of Cyclin D1 m RNA. Marsdenia tenacissima combined with fluorouracil can enhance the the lung A549 cancer blocked in S phase of cell cycle, but,it still not clear the mechanism relate to the down-regulation of Cyclin D1, Cyclin E m RNA expression. |
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