层粘连蛋白 A 表达对卵巢癌细胞迁移和核稳定性的影响。
Effect of lamin-A expression on migration and nuclear stability of ovarian cancer cells.
机构信息
Department of Pathology, Xiangya Hospital, Central South University, Changsha, Hunan, China; Department of Pathology, School of Basic Medical Science, Central South University, Changsha, Hunan, China.
Department of Pathology, School of Basic Medical Science, Central South University, Changsha, Hunan, China.
出版信息
Gynecol Oncol. 2019 Jan;152(1):166-176. doi: 10.1016/j.ygyno.2018.10.030. Epub 2018 Oct 29.
OBJECTIVE
Nuclear lamina plays important roles in nuclear shape and mechanical stability. Many studies demonstrated that defects of lamin-A were associated with several diseases, but little research was found on its potential roles in ovarian cancer.
METHODS
GEPIA and GEO database were used to analyze lamin-A in ovarian tissues, followed by assessing lamin-A and prognosis of ovarian cancer patients with Kaplan-Meier plotter. Then, transient transfected HO-8910 cells with shRNA to knockdown lamin-A. Knockdown efficiency was determined by western blot, qRT-PCR and immunofluorescence. Meanwhile, lamin-A was overexpressed in HO-8910 PM cells. Then, 2D migration, 3D migration through 3 μm and 8 μm pores were carried out, followed by immunofluorescence and TEM observation.
RESULTS
Lamin-A tended to be lower in ovarian cancer, and higher expression of lamin-A was associated with better survival. After lamin-A knockdown, 2D and 3D migration (3 μm, 8 μm) abilities of HO-8910 cells were significantly increased (p < 0.001), while overexpression of lamin-A in HO-8910PM impeded migration. Meanwhile, when HO-8910 cells migrated through 3 μm pores, nuclei became strikingly elongated, and down-regulation of lamin-A promoted nuclear plasticity, making the circularity of nucleus increased. Besides, further knockdown group had the highest proportion of γ-H2AX, with micronuclei forming. Furthermore, western blot showed that the expression of BRCA1, Ku80 and Rad50 decreased significantly after further knockdown, suggesting impairment of DNA damage repair.
CONCLUSIONS
Lamin-A was down-regulated in ovarian cancer, and higher lamin-A was associated with better prognosis. Nuclei with high lamin-A were severely deformed through constricted pores. Moderate lamin-A enhanced nuclear plasticity, so as to strengthen migration ability. When lamin-A was further knockdown, ovarian cancer cells that migrated through restricted pores decreased, with DNA damage, genomic instability and impairment of DNA damage repair.
目的
核层在核形状和机械稳定性中发挥重要作用。许多研究表明，核层 A 的缺陷与几种疾病有关，但关于其在卵巢癌中的潜在作用的研究甚少。
方法
使用 GEPIA 和 GEO 数据库分析卵巢组织中的核层 A，然后使用 Kaplan-Meier plotter 评估核层 A 与卵巢癌患者预后的关系。然后，使用 shRNA 瞬时转染 HO-8910 细胞以敲低核层 A。通过 Western blot、qRT-PCR 和免疫荧光测定敲低效率。同时，在 HO-8910 PM 细胞中过表达核层 A。然后进行 2D 迁移、通过 3μm 和 8μm 孔的 3D 迁移，随后进行免疫荧光和 TEM 观察。
结果
核层 A 在卵巢癌中呈下降趋势，核层 A 的高表达与生存改善相关。敲低核层 A 后，HO-8910 细胞的 2D 和 3D 迁移（3μm、8μm）能力显著增强（p<0.001），而在 HO-8910PM 中过表达核层 A 则阻碍了迁移。同时，当 HO-8910 细胞穿过 3μm 孔时，细胞核变得明显拉长，核层 A 的下调促进了核可塑性，使细胞核的圆形度增加。此外，进一步敲低组中γ-H2AX 的比例最高，形成微核。此外，Western blot 显示，进一步敲低后 BRCA1、Ku80 和 Rad50 的表达显著下降，表明 DNA 损伤修复受损。
结论
核层 A 在卵巢癌中下调，核层 A 水平较高与预后较好相关。通过狭窄的孔，高核层 A 的细胞核严重变形。适度的核层 A 增强了核可塑性，从而增强了迁移能力。当核层 A 进一步敲低时，穿过受限孔的卵巢癌细胞减少，出现 DNA 损伤、基因组不稳定和 DNA 损伤修复受损。
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