ABSTRACT

Cervical cancer is the fourth-most prevalent malignancy in women. For advanced cervical cancer, radiotherapy is a major treatment. Micro RNAs (miRNAs) are small, noncoding RNAs that negatively regulate the target gene expression posttranscriptionally. miR-22 is frequently downregulated in various cancers including cervical cancer, and is associated with a poor prognosis in cervical cancer. Exosomes are small endosomally secreted vesicles that carry components such as proteins, messenger RNA (mRNA), DNA and miRNA. We investigated whether or not exosomes can efficiently deliver miR-22 to recipient cervical cancer cells and affect the gene expression in the cells, as well as assessed the role of exosomal miR-22 in radiosensitivity. Exosomes containing high levels of miR-22 were extracted by ultracentrifugation and then characterized by Western blotting, a nanoparticle tracking analysis and electron microscopy. The high presence of miR-22 in the exosome was confirmed by real-time polymerase chain reaction. After the administration of the collected exosomal miR-22 to SKG-II and C4-I cervical cancer cells, the level of miR-22 in the cells was significantly increased, indicating the absorption of the exosomal miR-22. When miR-22 encapsulated in exosomes was administered to the SKG-II cells, the level of c-Myc binding protein (MYCBP) and human telomerase reverse transcriptase (hTERT) was significantly decreased in correlation with increased radiosensitivity determined by a clonogenic assay. Taken together, these results suggest that the administration of exosomal miR-22 may be a novel drug delivery system for cervical cancer radiotherapy.

摘要

宫颈癌是女性第四大常见恶性肿瘤。对于晚期宫颈癌，放疗是主要的治疗方法。微小 RNA (miRNA) 是小的非编码 RNA，可在转录后负调控靶基因的表达。miR-22 在包括宫颈癌在内的各种癌症中经常被下调，并且与宫颈癌的不良预后有关。外泌体是内体分泌的小囊泡，携带蛋白质、信使 RNA (mRNA)、DNA 和 miRNA 等成分。我们研究了外泌体是否可以有效地将 miR-22 递送至受体宫颈癌细胞并影响细胞中的基因表达，并评估外泌体 miR-22 在放射敏感性中的作用。通过超速离心提取含有高水平 miR-22 的外泌体，然后通过蛋白质印迹、纳米颗粒跟踪分析和电子显微镜进行表征。实时聚合酶链反应证实了外泌体中 miR-22 的高存在。将采集的外泌体miR-22给予SKG-II和C4-I宫颈癌细胞后，细胞内miR-22水平显着升高，表明外泌体miR-22被吸收。当包裹在外泌体中的 miR-22 被施用于 SKG-II 细胞时，c-Myc 结合蛋白 (MYCBP) 和人端粒酶逆转录酶 (hTERT) 的水平显着降低，与通过克隆测定确定的放射敏感性增加相关。综合起来，