[摘 要] 目的：探究长链非编码RNA（lncRNA）内源性博尔纳病毒样核蛋白3假基因（EBLN3P）调控miR-369-3p/ CCND1轴对甲状腺癌B-CPAP细胞增殖、迁移和EMT进程的影响。方法：收集2020年5月至2021年5月间在海南省肿瘤医院 手术切除的20例甲状腺癌及相应癌旁组织标本，以及甲状腺癌B-CPAP细胞，qPCR法和WB法检测癌组织和细胞中 EBLN3P、miR-369-3p、CCND1 mRNA水平，双萤光素酶报告基因实验验证lncRNA EBLN3P、CCND1与miR-369-3p之间的靶向 关系。随机将B-CPAP细胞分为对照组、sh-NC组、sh-EBLN3P组、sh-EBLN3P + anti-NC组、sh-EBLN3P + anti-miR-369-3p组，通 过克隆形成实验、划痕愈合实验、Transwell实验分别检测各组细胞的增殖和迁移能力，WB法检测各组细胞中EMT相关蛋白 的表达。构建B-CPAP细胞裸鼠移植瘤模型，观察沉默EBLN3P对移植瘤生长的影响。结果：在甲状腺癌组织和B-CPAP细胞中 EBLN3P、CCND1 mRNA表达上调，miR-369-3p表达下调（均P < 0.05）；EBLN3P与miR-369-3p、CCND1与miR-369-3p之间有 结合位点，存在靶向关系。与sh-NC组比较，sh-EBLN3P组克隆形成数、划痕愈合率、细胞迁移数降低（均P < 0.05），EBLN3P、 CCND1、Ki67、MMP-2、N-cadherin、vimentin表达下调，miR-369-3p、E-cadherin表达上调（均P < 0.05）；与 sh-EBLN3P + anti-NC 组比 较，sh-EBLN3P + anti-miR-369-3p组miR-369-3p表达下调，克隆形成数、划痕愈合率、细胞迁移数均升高（均P < 0.05），CCND1、 Ki67、MMP-2、N-cadherin、vimentin表达均上调，E-cadherin表达下调（均P < 0.05）。与sh-NC组比较，sh-EBLN3P组B-CPAP细胞 裸鼠移植瘤的体积和质量均显著降低（均P < 0.05）。结论：在甲状腺癌组织和B-CPAP细胞中lncRNA EBLN3P表达上调，沉默 EBLN3P可靶向调控miR-369-3p/CCND1轴抑制甲状腺癌B-CPAP细胞的增殖、迁移、EMT进程。

[Abstract] Objective: To investigate the effects of long non-coding RNA (lncRNA) endogenous Bornavirus-like nucleoprotein 3 pseudogene (EBLN3P) on the proliferation, migration and epithelial-mesenchymal transition (EMT) of thyroid cancer B-CPAP cells by regulating the miR-369-3p/CCND1 axis. Methods: 20 samples of thyroid cancer and corresponding adjacent tissue specimens surgically removed at Hainan Cancer Hospital between May 2020 and May 2021, as well as thyroid cancer B-CPAP cells were collected. The levels of EBLN3P, miR-369-3p and CCND1 mRNA in cancer tissues and cells were detected using qPCR and Western blot (WB) methods. The dual-luciferase reporter gene assay was used to validate the targeting relationship among lncRNA EBLN3P, CCND1 and miR-369-3p. B-CPAP cells were randomly divided into the control group, sh-NC group, sh-EBLN3P group, sh-EBLN3P + anti-NC group and sh-EBLN3P + anti-miR-369-3p group. The colony formation assay was used to detect the number of colony formation in each group. The scratch wound healing assay and Transwell assay were performed to evaluate cell migration ability. WB was used to detect the expression of EMT-related proteins. The nude mouse xenograft model of B-CPAP cells was constructed to observe the effect of silencing EBLN3P on the growth of xenograft tumors. Results: The expressions of lncRNA EBLN3P and CCND1 mRNA were up-regulated, and the expression of miR-369-3p was down-regulated in thyroid cancer tissues and B-CPAP cells (all P < 0.05). lncRNA EBLN3P had binding sites with miR-369-3p, and CCND1 had binding sites with miR-369-3p, and there was a targeting relationship. Compared with those in the sh-NC group, the number of clone formation, the scratch healing rate and the number of cell migration in the sh-EBLN3P group decreased (all P < 0.05); the expressions of EBLN3P, CCND1, Ki67, MMP-2, N-cadherin and vimentin was down-regulated; the expressions of miR-369-3p and E-cadherin was up-regulated (all P < 0.05). Compared with those in the sh-EBLN3P + anti-NC group, the expression of miR-369-3p in the sh-EBLN3P + anti-miR-369-3p group was down-regulated; the number of clone formation, scratch healing rate and the number of cell migration increased (all P < 0.05); the expressions of CCND1, Ki67, MMP-2, N-cadherin and vimentin was up-regulated; the expression of E-cadherin was down-regulated (all P < 0.05). Compared with those in the sh-NC group, the volume and weight of B-CPAP cell nude mouse xenograft tumors in the sh-EBLN3P group were significantly reduced (both P < 0.05). Conclusion: The expression of lncRNA EBLN3P is up-regulated in thyroid cancer cells and tissues. Silencing EBLN3P can inhibit the proliferation, migration and EMT of thyroid cancer B-CPAP cells by targeting and regulating the miR-369-3p/CCND1 axis.

海南省自然科学基金高层次人才项目（No. 821RC723）