Data from: Loss of miR-143 and miR-145 in condyloma acuminatum promotes cellular proliferation and inhibits apoptosis by targeting NRAS

The expression profile of miRNAs and their function in condyloma acuminatum (CA) remains unknown. In this study, we aimed to detect the effects of miR-143 and miR-145, the most downregulated in CA samples using high-throughput sequencing, on cell proliferation and apoptosis, to determine a novel therapeutic target for CA recurrence. RT-qPCR was used to validate the lower expression of miR-143 and miR-145 in a larger size of CA samples, and the expression of NRAS in CA samples was significantly higher than self-controls as determined western blotting assay. Luciferase assay was performed to confirm that miR-143 or miR-145 targeted NRAS directly. Transduction of LV-pre-miR-143 or LV-pre-miR-145 to human papilloma virus (HPV)-infected SiHa cells led to reduced proliferation, greater apoptosis, and inhibition of expression of NRAS, PI3Kp110α, and pAKT. However, knockout of miR-143 or miR-145 in human epidermal keratinocytes (HEKs) by delivery of CRISPR/CAS9-gRNA for target miRNAs protected cells from apoptosis and upregulated expression of target genes as described above. MiR-143 and miR-145 sensitized cells to nutlin-3a, a p53 activator and MDM2 antagonist, while their loss protected cells from the stress of nutlin-3a. Furthermore, siRNA targeting NRAS showed similar effects on proliferation and apoptosis as miR-143 or miR-145. Taken together, our results suggest that loss of miR-143 or miR-145 in CA protects HPV-infected cells from apoptosis induced by environmental stress, in addition to promoting cellular proliferation and inhibiting apoptosis by targeting NRAS/PI3K/ATK. Restoration of miR-143 or miR-145 might provide an applicable and novel approach to block the recurrence and progression of CA.微小RNA（microRNAs, miRNAs）在尖锐湿疣（condyloma acuminatum, CA）中的表达特征与功能，目前仍未明确。本研究旨在通过高通量测序筛选出尖锐湿疣样本中表达下调最显著的miR-143与miR-145，并探究其对细胞增殖与凋亡的调控作用，以期为尖锐湿疣复发寻找全新的治疗靶点。我们采用实时荧光定量聚合酶链反应（real-time quantitative polymerase chain reaction, RT-qPCR）在更大样本量的尖锐湿疣组织中验证了miR-143与miR-145的低表达水平；同时通过蛋白质印迹（western blotting）实验发现，尖锐湿疣样本中NRAS的表达水平显著高于自身对照组织。我们通过荧光素酶报告基因实验（luciferase assay）证实，miR-143与miR-145可直接靶向结合NRAS基因。将携带pre-miR-143或pre-miR-145的慢病毒（lentivirus, LV）转染至人类乳头瘤病毒（human papilloma virus, HPV）感染的SiHa细胞后，细胞增殖能力显著降低、凋亡水平明显升高，且NRAS、PI3Kp110α以及pAKT的表达均受到抑制。然而，通过靶向miRNA的CRISPR/CAS9-gRNA系统敲除人表皮角质形成细胞（human epidermal keratinocytes, HEKs）中的miR-143或miR-145后，细胞凋亡受到抑制，且上述靶基因的表达水平显著上调。miR-143与miR-145可使细胞对nutlin-3a（一种p53激活剂与MDM2拮抗剂）敏感；而敲除这两种miRNA则可使细胞免受nutlin-3a诱导的应激损伤。此外，靶向NRAS的小干扰RNA（small interfering RNA, siRNA）对细胞增殖与凋亡的调控效果与miR-143、miR-145相似。综上，本研究结果显示，尖锐湿疣病灶中miR-143与miR-145的缺失，可通过靶向NRAS/PI3K/AKT通路促进细胞增殖并抑制细胞凋亡，同时还能使人乳头瘤病毒感染的细胞免受环境应激诱导的凋亡。恢复miR-143与miR-145的表达，或可为阻断尖锐湿疣的复发与进展提供一种全新且可行的治疗策略。