异甘草酸镁注射液联合紫杉醇诱导乳腺癌细胞株凋亡及其机制

Magnesium isoglycyrrhizinate injection synergisitcally enhance paclitaxel-induced apoptosis in breast cancer cells and its mechanism

目的探讨异甘草酸镁注射液、紫杉醇以及两药联合诱导乳腺癌细胞MCF-7凋亡作用及分子机制。方法将不同浓度的异甘草酸镁注射液单药以及联合紫杉醇干预乳腺癌细胞MCF-7,采用细胞增殖毒性(Cell Counting Kit-8,CCK8)法检测细胞增殖情况; AnnexinV-FITC/PI流式细胞术检测细胞凋亡情况;蛋白质印迹法(Western blot)检测细胞凋亡相关蛋白磷酸化B淋巴细胞瘤-2基因(phosphate-B-cell lymphoma-2,pB cl-2)、半胱天冬酶(cysteinyl aspartate specific proteinase,Caspase)-3以及缺口蛋白(Notch)通路Notch2的表达情况。结果异甘草酸镁注射液处理48 h对乳腺癌细胞MCF-7的体外增殖具有抑制作用,并在100 mmol/L达到最大值27%;两药联合组对MCF-7细胞增殖的抑制作用较单药组明显增强尤其在100μmol/L(60. 89%比26. 98%,t=-0. 017,P <0. 05);两药联合处理MCF-7细胞早期凋亡率和晚期凋亡率均与空白对照组、异甘草酸镁注射液组和紫杉醇组差异有统计学意义(F=209. 839,P <0. 001; F=40. 742,P <0. 001);凋亡相关蛋白pB cl-2在空白对照组、异甘草酸镁注射液组、紫杉醇组和联合组差异有统计学意义,F=11. 064,P <0. 001,联合组的pB cl-2蛋白表达均高于其他三组(1. 94,0. 79,0. 83和1. 01,均P <0. 001)。与空白对照组相比,异甘草酸镁联合紫杉醇可上调细胞凋亡相关蛋白Caspase-3和Notch2表达(3. 17比2. 01,P=0. 027; 1. 17比0. 67,P=0. 018),联合组与单药紫杉醇的Casepase-3和Notch2表达差异无统计学意义(3. 17比2. 46,P=0. 160; 1. 17比0. 82,P=0. 121)。结论异甘草酸镁注射液与紫杉醇能协同诱导乳腺癌细胞MCF-7的细胞凋亡,其机制可能与部分促进紫杉醇通过Notch通路上调MCF-7细胞pB cl-2、Caspase-3凋亡蛋白表达有关。

Objective To evaluate the effect of magnesium isoglycyrrhizinate injection combined with paclitaxel on proliferation and apoptosis of breast cancer cells(MCF-7),and further investigate its potential mechanism.Methods MCF-7 cells were treated with various concentrations of magnesium isoglycyrrhizinate injection alone or combined with paclitaxel.Cell proliferation inhibition and appotosis rates were measured by Cell Counting Kit-8(CCK-8)assay and flow cytometry in MCF-7.Western blot was used to quantify the alternation of the apoptosis-related protein pBcl-2,caspase-3 and Notch2.Results Magnesium isoglycyrrhizinate injection could lightly induce apoptosis in breast cancer cell line MCF-7,at the peak in the 100 mmol/L concentration(60.89%vs.26.98%,t=-0.017,P<0.05),and proliferation inhibition was significantly higher in combined group than others isoglycyrrhizinate injection synergistically enhanced paclitaxel-induced early and late stage apoptosis rates in MCF-7(F=209.839,P<0.001;F=40.742,P<0.001).The expression level of the apoptosis-related factors pBcl-2 was upregulated in group of isoglycyrrhizinate injection combined with paclitaxel than other groups(1.94,0.79,0.83 and 1.01,all P<0.001),and combined groups showed upregulation of the expression of caspase-3 and Notch2 protein(3.17 vs.2.01,P=0.027;1.17 vs.0.67,P=0.018).No differences were found between paclitaxel and combined groups on expression of caspase-3 and Notch2 protein(3.17 vs.2.46,P=0.160;1.17 vs.0.82,P=0.121).Conclusions Magnesium isoglycyrrhizinate injection combined with paclitaxel can synergistically promote the apoptosis in breast cancer cell line(MCF-7).The potential mechanism may be magnesium isoglycyrrhizinate injection promoted paclitaxel-induced upregulation of pBcl-2 and caspase-3 via Notch pathway.