苦参碱联合DC-CIK细胞对乳腺癌细胞的杀伤作用

Killing effect of matrine combined with DC-CIK cells on breast cancer cells

目的研究苦参碱联合树突状细胞(DC)-细胞因子诱导的杀伤(CIK)细胞对乳腺癌细胞的杀伤作用及其可能的作用机制。方法收集本院体检的健康志愿者的外周血单核细胞,用粒细胞-巨噬细胞刺激因子(GM-CSF)及白细胞介素-4(IL-4)联合培养体系诱导DC细胞,用γ干扰素(IFN-γ)、白细胞介素-2(IL-2)、白细胞介素-1α(IL-1α)等细胞因子诱导CIK细胞,并将DC细胞与CIK细胞共培养。将成功诱导DC-CIK细胞联合苦参碱作用于乳腺癌MCF-7细胞。将MCF-7细胞分为4组:Control组(正常培养)、Matrine组(1.0 mg·mL^(-1)苦参碱)、DC-CIK组(DC-CIK细胞处理)、Combine组(1.0 mg·mL^(-1)苦参碱+DC-CIK细胞处理)。用噻唑蓝(MTT)实验检测不同效靶比的DC-CIK细胞联合苦参碱对MCF-7细胞增殖的抑制作用;以蛋白质印迹法检测各组细胞蛋白表达情况。结果Control组、Matrine组、DC-CIK组、Combine组MCF-7细胞核相关抗原Ki67(Ki67)蛋白表达水平分别为0.91±0.03、0.68±0.07、0.63±0.04、0.33±0.03;E-cadherin蛋白表达水平分别为0.34±0.03、0.51±0.10、0.62±0.08、0.84±0.07;N-cadherin蛋白表达水平分别为0.91±0.08、0.72±0.05、0.67±0.02、0.45±0.03;磷酸化JAK2(p-JAK2)蛋白表达水平分别为1.07±0.08、0.83±0.07、0.65±0.04、0.50±0.05;磷酸化STAT3(p-STAT3)蛋白表达水平分别为0.89±0.04、0.79±0.05、0.72±0.04、0.39±0.05。以上指标,Matrine组、DC-CIK组、Combine组与Control组比较,Combine组与Matrine组、DC-CIK组比较,差异均有统计学意义(均P<0.05)。结论DC-CIK细胞可增强苦参碱对乳腺癌细胞的体外杀伤作用,这可能与抑制JAK2/STAT3通路有关。

Objective To explore the killing effect of matrine combined with dendritic cell(DC)-cytokine induced killer(CIK)cells on breast cancer cells and its possible mechanism.Methods Peripheral blood mononuclear cells were collected from healthy volunteers in our hospital.DC cells were induced by the combined culture system of granulocyte-macrophage stimulating factor(GM-CSF)and interleukin(IL)-4,and CIK cells were induced by interferon-γ(IFN-γ),recombinant IL-2,IL-1αand other cytokines.DC cells were co-cultured with CIK cells.DC-CIK cells were successfully induced and used as MCF-7 cells for breast cancer.The MCF-7 cells were divided into four groups:control group(normal culture),matrine group(1.0 mg·mL^(-1) matrine),DC-CIK group(DC-CIK cell treatment),and combine group(1.0 mg·mL^(-1) matrine+DC-CIK cell treatment).MTT assay was used to detect the inhibitory effect of DC-CIK cells combined with matrine on the proliferation of MCF-7 cells;protein expression level in each group was detected by Western blotting.Results The expression levels of Ki67 protein in the control group,the matrine group,the DC-CIK group and the combine group were 0.91±0.03,0.68±0.07,0.63±0.04,0.33±0.03,respectively;the expression levels of E-cadherin protein were 0.34±0.03,0.51±0.10,0.62±0.08,0.84±0.07,respectively;the expression levels of N-cadherin protein were 0.91±0.08,0.72±0.05,0.67±0.02,0.45±0.03,respectively;the protein expression levels of p-JAK2 were 1.07±0.08,0.83±0.07,0.65±0.04,0.50±0.05,respectively;p-STAT3 protein expression levels were 0.89±0.04,0.79±0.05,0.72±0.04,0.39±0.05,respectively.Compared control group with combine group,matrine group,DC-CIK group,and compared combine group with matrine group,DC-CIK group,the differences of the above indicators were statistically significant(all P<0.05).Conclusion DC-CIK cells can enhance the killing effect of matrine on breast cancer cells in vitro,which may be related to the inhibition of JAK2/STAT3 pathway.