Mahmoodzadeh Hosseini Hamideh, Imani Fooladi Abbas Ali, Soleimanirad Jafar, Nourani Mohammad Reza, Davaran Soodabeh, Mahdavi Mehdi
Department of Pharmaceutical Biotechnology, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran.
Tumour Biol. 2014 Apr;35(4):3699-707. doi: 10.1007/s13277-013-1489-1. Epub 2014 Jan 8.
Exosomes (EXO) are acellular vehicles used for cancer immunotherapy due to their immune inducing properties. To identify whether designed structure based on tumoral EXO have a cytotoxic effect together with a potent immunological property, we synthesized a novel structure based on EXO and staphylococcal entrotoxin B (SEB), two immune inducer substances, and surveyed its cytostatic effect on the breast cancer cell line. EXO were purified from tumor cells and SEB was anchored on it by protein transfer method. To determine the cytotoxic and apoptosis inducing effect of this structure, treated cells with different concentrations of EXO/SEB were examined by MTT assay and Hoechst staining method. In addition, the expression rate of bcl-2, bax, bak, fas, bcl-xl and the activity of caspase-3 and caspase-9 were assessed. We observed that EXO/SEB significantly decreased the cell proliferation and stimulated apoptosis (P < 0.001) at all concentration after 24 h (P < 0.001). Furthermore, EXO/SEB raised the expression rate of bax and bak (P < 0.001) but no impact on fas and bcl-xl after 48 h. We observed reducing effect of EXO/SEB on the mRNA expression of bcl-2. After 24 h of exposing the cell with the EXO/SEB, a significant increase was found in the activity of caspase at the concentration of 2.5, 5 and 10 μg/100 μl for caspase-9 and at all concentrations for caspase-3 (P < 0.001). Our designed structure, the EXO/SEB, is a novel model for apopto-immunotherapy being able to induce apoptosis in ER(-) breast cancer cells.
外泌体(EXO)因其免疫诱导特性而被用作癌症免疫治疗的无细胞载体。为了确定基于肿瘤外泌体设计的结构是否具有细胞毒性作用以及强大的免疫特性,我们合成了一种基于外泌体和葡萄球菌肠毒素B(SEB)这两种免疫诱导物质的新型结构,并研究了其对乳腺癌细胞系的细胞生长抑制作用。从肿瘤细胞中纯化出外泌体,并通过蛋白质转移方法将SEB锚定在其上。为了确定这种结构的细胞毒性和诱导凋亡作用,采用MTT法和Hoechst染色法检测用不同浓度的EXO/SEB处理后的细胞。此外,评估了bcl-2、bax、bak、fas、bcl-xl的表达率以及caspase-3和caspase-9的活性。我们观察到,24小时后,EXO/SEB在所有浓度下均显著降低细胞增殖并刺激细胞凋亡(P<0.001)。此外,48小时后,EXO/SEB提高了bax和bak的表达率(P<0.001),但对fas和bcl-xl没有影响。我们观察到EXO/SEB对bcl-2的mRNA表达有降低作用。在用EXO/SEB处理细胞24小时后,发现caspase-9在浓度为2.5、5和10μg/100μl时活性显著增加,caspase-3在所有浓度下活性均显著增加(P<0.001)。我们设计的结构EXO/SEB是一种新型的凋亡免疫治疗模型,能够诱导雌激素受体阴性(ER(-))乳腺癌细胞凋亡。