丙泊酚和瑞马唑仑对前列腺癌细胞增殖的影响

Effect of propofol and remimazolam on the proliferation of prostate cancer cells

中文关键词: 前列腺癌  丙泊酚  瑞马唑仑  细胞增殖  信号传导及转录激活蛋白3

英文关键词: Prostate cancer  Propofol  Remimazolam  Cell proliferation  Signal transducer and activator of transcription 3

中文摘要:

      
目的： 探讨丙泊酚和瑞马唑仑对前列腺癌细胞增殖的影响及相关机制。
方法： 选择人前列腺癌细胞系DU145和PC3，培养至对数生长期。将细胞随机分为四组：对照组（C组）、丙泊酚组（P组）、瑞马唑仑组（R组）和丙泊酚复合瑞马唑仑组（PR组）。C组用完全培养基培养，P组和R组分别在完全培养基中加入丙泊酚和瑞马唑仑的半数抑制浓度（IC₅₀）培养（DU145和PC3细胞中丙泊酚的IC₅₀分别为120 μg/ml和100 μg/ml，瑞马唑仑的IC₅₀分别为500 μmol/L和400 μmol/L），PR组DU145细胞加入低浓度丙泊酚100 μg/ml和瑞马唑仑400 μmol/L共培养，PC3细胞加入低浓度丙泊酚80 μg/ml和瑞马唑仑300 μmol/L共培养。于培养后0、6、12、24、36 h采用CCK-8法测定待检测孔的吸光度。于培养后14 d采用平板克隆形成实验计算集落数量。采用qPCR法和Western blot法检测c-Myc和cyclin D1 mRNA表达量和蛋白含量。通过网络药理学分析找出丙泊酚和瑞马唑仑作用于前列腺癌细胞的关键靶基因，并采用qPCR法和Western blot法检测该靶基因的mRNA表达量和蛋白含量。
结果： 与C组比较，DU145和PC3细胞P组、R组和PR组培养后36 h 吸光度均明显降低、集落数量明显减少、c-Myc和cyclin D1 mRNA表达量和蛋白含量明显降低（P<0.05）。与P组比较，DU145细胞R组和PR组培养后36 h吸光度、cyclin D1 mRNA表达量明显降低，R组c-Myc mRNA表达量明显升高，PR组集落数量明显减少、c-Myc蛋白含量明显降低（P<0.05）；PC3细胞R组cyclin D1 mRNA表达量和蛋白含量明显降低，PR组培养后36 h吸光度明显降低、c-Myc和cyclin D1 mRNA表达量和蛋白含量明显降低（P<0.05）。与R组比较，DU145和PC3细胞PR组集落数量明显减少，c-Myc mRNA表达量和蛋白含量明显降低（P<0.05）。网络药理学分析结果显示：丙泊酚、瑞马唑仑与前列腺癌的共同靶点为信号传导及转录激活蛋白3（STAT3）。与C组比较，DU145和PC3细胞P组、R组和PR组STAT3 mRNA表达量和蛋白含量明显降低（P<0.05）。与P组比较，DU145细胞R组STAT3 mRNA表达量和蛋白含量明显升高，PR组STAT3蛋白含量明显升高（P<0.05）；PC3细胞R组STAT3 mRNA表达量明显降低（P<0.05）。与R组比较，DU145细胞PR组STAT3蛋白含量明显降低（P<0.05）。
结论： 丙泊酚和瑞马唑仑可以单独或协同抑制前列腺癌细胞增殖，降低前列腺癌细胞c-Myc和cyclin D1 mRNA表达量和蛋白含量，其机制可能与抑制HIF-1α通路中的STAT3靶点有关。

英文摘要:

      
Objective: To investigate the effects of propofol and remimazolam on the proliferation of prostate cancer cells and related mechanisms.
Methods: Human prostate cancer cell lines DU145 and PC3 were selected and cultured to the logarithmic growth phase. Cells were randomly divided into four groups: the control group (group C), the propofol group (group P), the remimazolam group (group R), and the propofol combined with remimazolam group (group PR). Group C was cultured with complete medium, groups P and R were cultured with semi-inhibitory concentrations (IC₅₀) of propofol and remimazolam (IC₅₀ of propofol in DU145 and PC3 cells were 120 μg/ml and 100 μg/ml, IC₅₀ of remimazolam were 500 μmol/L and 400 μmol/L), and DU145 cells in group PR were co-cultured with low concentration propofol 100 μg/ml and remimazolam 400 μmol/L, PC3 cells were co-cultured with low concentrations of propofol 80 μg/ml and remimazolam 300 μmol/L. The absorbance of 0, 6, 12, 24, and 36 hours after incubation of the wells was determined by CCK-8. The number of colonies of 14 days after incubation was calculated by colony formation assay. qPCR and Western blot were used to detect the expression of mRNA and protein of c-Myc and cyclin D1. The key target genes of propofol and remimazolam on prostate cancer cells were identified by network pharmacological analysis, and the expression of mRNA and protein of the target gene were detected by qPCR and Western blot.
Results: Compared with group C, the absorbance at 36 hours, the number of colonies and the expression of mRNA and protein of c-Myc and cyclin D1 were decreased significantly in groups P, R, and PR of DU145 and PC3 cells (P < 0.05). Compared with group P, in DU145 cells, the absorbance at 36 hours and the expression of mRNA of cyclin D1 in groups R and PR were significantly reduced, the expression of mRNA of c-Myc in group R was increased significantly, the number of colonies and the expression of protein of c-Myc in group PR were significantly reduced (P < 0.05). In PC3 cells, the expression of mRNA and protein of cyclin D1 in the group R were significantly reduced, the absorbance at 36 hours and the expression of mRNA and protein of c-Myc and cyclin D1 in group PR were significantly reduced (P < 0.05). Compared with group R, the number of colonies and the expression of mRNA and protein of c-Myc in group PR of DU145 and PC3 cells were significantly reduced (P < 0.05). The results of network pharmacological analysis showed that the common target of propofol, remimazolam and prostate cancer was signal transducer and activator of transcription 3 (STAT3). Compared with group C, the expression of mRNA and protein of STAT3 were decreased significantly in groups P, R, and PR in DU145 and PC3 cells (P < 0.05). Compared with group P, in DU145 cells, the expression of mRNA and protein of STAT3 were increased significantly in group R, the expression of protein of STAT3 was increased significantly in group PR (P < 0.05). In PC3 cells, the expression of mRNA of STAT3 was decreased significantly in group R (P < 0.05). Compared with group R, the expression of protein of STAT3 was decreased significantly in group PR in DU145 cells(P < 0.05).
Conclusion: The proliferation of prostate cancer cells can be inhibited by propofol and remimazolam alone or synergistically and they can also reduce the expression of mRNA and protein of c-Myc and cyclin D1, the mechanism may be related to the inhibition of expression of STAT3 which is the member of the HIF-1α pathway.