蛋白激酶A活性对膀胱癌细胞株T24增殖的影响

Protein Kinase A activity suppresses the proliferation of T24 cell line

目的探讨蛋白激酶A（PKA）活性增高对膀胱癌细胞株T24增殖的影响及其机制。方法用PKA信号传导通路的激动剂forskolin诱导T24细胞后，用放免法检测forskolin作用下T24细胞中PKA活性的变化；用噻唑蓝（MTr）比色法检测forskolin作用下T24细胞活力的变化；用流式细胞仪检测forskolin作用下细胞周期分布的变化；用Western blot检测forskolin作用下T24细胞中Cyclin D1表达的变化。结果forskolin能够使124细胞中的PKA活性增高，并显著降低细胞活力。在20μmol／L forskolin作用24h后，流式细胞仪检测可见T24细胞增殖受到抑制，G1期细胞74．46％，显著高于对照组50．21％（P〈0．01），S期和G2／M期细胞分别占7．63％和14．13％，显著低于对照组的22，25％和24．37％（S期P〈0．01，G2／M期P〈0．05）。用20μmol／L forskolln诱导T24细胞后，Cyclin D1表达呈下降趋势。结论forskolin能够显著上调T24细胞中PKA的活性，PKA活性增高使T24细胞处于G1期静止，抑制其增殖，并显著降低细胞活力。PKA活性增高所致的G1期静止可能与CyclinD1的表达下降有关。

Objective To investigate the effects of increased protein kinase A （PKA） activity on the proliferation of T24 cell line and the possible mechanism. Methods T24 cells were cultured with PKA activator-forskdin with different concentrations, and the changes in PKA activity, cell viability, cell cycle and the expression of Cyclin D1 were measured at different time points. Results （ 1 ） Forskolin could significantly upregulate PKA activity in T24 ceils （P 〈 0.01 ） ; （2） After treated with 20 μmol/L forskolin for 24 h,the proliferation of T24 cells was suppressed and cell viability was decreased. The percentage of T24 ceils in G1 phase was 74.46% ,higher than that in control group （50.21% ,P 〈0.01 ） ,and the percentage in S phase and G2/M phase was 7.63% and 14. 13% respectively, lower than that in control group （22.25% and 24.37% respectively,for S phase P 〈 0.01, for G2/M phase, P 〈 0.05 ） ; （3） After cultured with 20 p.mol/L forskolin,the expression of Cyclin D1 was downregulated significantly. Conclusion Upregulating PKA activity can significantly decrease cell viability of 124 cells, and can lead to reduction of cyclin D1 expression,resulting in G1 arrest and proliferation inhibition.