左玲, 乔淦, 郭铭悦, 林秀坤, 刘明华. 负载Bcl-2 siRNA外泌体的制备及其对胰腺癌细胞生长抑制作用J. 药学学报, 2020,55(12): 2918-2923. doi: 10.16438/j.0513-4870.2020-1011
引用本文: 左玲, 乔淦, 郭铭悦, 林秀坤, 刘明华. 负载Bcl-2 siRNA外泌体的制备及其对胰腺癌细胞生长抑制作用J. 药学学报, 2020,55(12): 2918-2923. doi: 10.16438/j.0513-4870.2020-1011
ZUO Ling, QIAO Gan, GUO Ming-yue, LIN Xiu-kun, LIU Ming-hua. Preparation of exosome-coated Bcl-2 siRNA and its inhibitory effect on the growth of pancreatic cancer cellsJ. Acta Pharmaceutica Sinica, 2020,55(12): 2918-2923. doi: 10.16438/j.0513-4870.2020-1011
Citation: ZUO Ling, QIAO Gan, GUO Ming-yue, LIN Xiu-kun, LIU Ming-hua. Preparation of exosome-coated Bcl-2 siRNA and its inhibitory effect on the growth of pancreatic cancer cellsJ. Acta Pharmaceutica Sinica, 2020,55(12): 2918-2923. doi: 10.16438/j.0513-4870.2020-1011

负载Bcl-2 siRNA外泌体的制备及其对胰腺癌细胞生长抑制作用

Preparation of exosome-coated Bcl-2 siRNA and its inhibitory effect on the growth of pancreatic cancer cells

  • 摘要: Bcl-2高表达与胰腺癌的发生、发展密切相关。本研究采用超速离心法从人胚肾(HEK293)细胞上清培养液中分离外泌体,利用电转技术制备了负载Bcl-2 siRNA的外泌体(exosiBcl-2),透射电镜观察表明,外泌体呈现典型内凹的杯状结构,Western blot分析表明外泌体特征蛋白CD9、CD81、CD63和TSG101呈现高表达,采用共聚焦显微镜、流式细胞术分析证实,exosiBcl-2可以高效转染人胰腺癌Miapaca-2细胞,转染效率为77.2%。MTS分析表明,exosiBcl-2可以显著抑制Miapaca-2细胞增殖,抑制率达到63%。本研究证实,exosiBcl-2可高效穿透细胞膜,通过抑制靶基因Bcl-2表达发挥抑制肿瘤细胞生长的作用。

     

    Abstract: High expression of Bcl-2 is associated with the development of pancreatic cancer, and downregulation of Bcl-2 is an effective approach for the treatment of pancreatic malignancy. In the present study exosomes were isolated from the cultured medium of human embryonic kidney cells (HEK293) by ultracentrifugation and exosome-coated Bcl-2 siRNA (exosiBcl-2) was synthesized using electroporation. The results showed that the particle size of exosiBcl-2 was 67.3±9.7 nm and the morphology of exosomes displayed a concave ring structure as determined by transmission electron microscopy (TEM). Western blot analysis indicated that exosomal proteins including CD9, CD81, CD63 and TSG101 were highly expressed. Confocal microscopy revealed that exosiBcl-2 was widely distributed in Miapaca-2 cells, and the transfection efficiency of exosiBcl-2 in Miapaca-2 was 77.2% as determined by flow cytometry. Treatment with exosiBcl-2 at a concentration of 100 nmol·L-1 resulted in an inhibitory effect on the growth of Miapaca-2 cells with an inhibition rate of 63%. ExosiBcl-2 treatment can downregulate Bcl-2 and upregulate Bax protein. This study provides evidence that exosiBcl-2 is able to inhibit the growth of pancreatic cancer cells and the nanoparticles have potential to be developed as a novel anticancer agent.

     

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