KIF4A在卵巢癌中的预后、生物学功能及富集分析

KIF4A in Prognosis and Biological Function of Ovarian Cancer and its En-richment Analysis

目的:探讨驱动蛋白超家族成员4A(kinesin family member 4A,KIF4A)与卵巢癌患者预后的关系,对卵巢癌细胞生物学功能的影响,以及功能富集分析。方法:利用TCGA(the cancer genome atlas, TCGA)数据库中372例卵巢癌患者数据进行KIF4A基因与相关共表达基因的富集分析及Kaplan-Meier法预后分析。采用实时荧光定量PCR法(reverse transcription quantitative real-time PCR,RT-qPCR)及Western blot检测多种卵巢癌细胞系中KIF4A的表达量。将三组KIF4A-shRNA分别转染至KIF4A相对表达量最高的细胞系。采用RT-qPCR及Western blot检测转染效果,选取沉默效率最高的建立稳定沉默细胞。采用MTT实验、克隆形成实验检测细胞增殖情况,划痕实验和Transwell实验检测细胞的迁移能力,流式细胞仪检测细胞凋亡和周期分布。结果:KIF4A基因在卵巢癌临床样本中显著高表达(P<0.05),且KIF4A高表达组总生存期显著低于低表达组(P<0.01)。不同卵巢癌细胞株中的表达水平均高于正常人卵巢上皮细胞株IOSE80(P<0.05),其中在SKOV3中表达最高。沉默KIF4A的表达能有效抑制卵巢癌细胞的增殖和迁移能力(P<0.05),并促进卵巢癌细胞凋亡(P<0.05),并主要影响G0/G1期。基因富集分析显示KIF4A与MYH11、ACTG2、HSPB6等基因成负相关,与BUB1、CCNA2、MELK等基因呈正相关。同时,KIF4A在卵巢癌中与有丝细胞核分裂等功能功能相关,并且可影响细胞周期等相关通路。结论:沉默KIF4A基因可抑制卵巢癌细胞的增殖,迁移能力并促进其细胞凋亡,影响其细胞周期,该基因有望成为卵巢癌治疗的新靶点。

Objective: To investigate the role of kinesin family member 4 A(KIF4 A) in prognosis of ovarian cancer patients, and its effect on biological function of ovarian cancer cells, and conduct its enrichment analysis. Methods: 372 ovarian cancer patients from TCGA database were selected for enrichment analysis of KIF4 A gene and related co-expressed genes. The prognosis of ovarian cancer patients was further investigated by using Kaplan-Meier method. Reverse transcription quantitative real-time PCR(RT-qPCR) and western blot were used to detect the expression of KIF4 A in different ovarian cancer cells. Lentiviral vectors containing KIF4 A-shRNA(sh-KIF4 A-1, sh-KIF4 A-2, sh-KIF4 A-3) and sh-NC(the negative control) were prepared, and KIF4 A-shRNA was transfected into SKOV3 cells with the highest relative expression of KIF4 A. RT-qPCR and western blot were used to detect the transfection effect, and the vector with the highest silencing efficiency was selected to establish stable silent cells;MTT assay and clonogenic assay were used to detect cell proliferation;scratch assay and transwell migration assay were used to detect cell migration ability;and flow cytometry was used to detect cell apoptosis and cycle distribution. Results: KIF4 A was significantly over-expressed in ovarian cancer clinical samples(P<0.05), and the overall survival of the KIF4 A high expression group was significantly lower than that of the KIF4 A low expression group(P<0.01). The expression level of KIF4 A in ovarian cancer cells was higher than that in human normal ovarian epithelial cell line IOSE80(P<0.05), among which the expression was the highest in SKOV3 cells. Silencing the expression of KIF4 A could effectively inhibit the proliferation and migration ability of ovarian cancer cells(P<0.05), promote the apoptosis of ovarian cancer cells(P<0.05), and mainly influence cell cycle in G0/G1 phase. Gene set enrichment analysis showed that KIF4 A was negatively correlated with MYH11, ACTG2, HSPB6, and positively correlated with BUB1, CCNA2, and MELK. KIF4 A was associated with mitotic nuclear division and other functions in ovarian cancer. And it could affect cell cycle and other related pathways. Conclusion: Silencing KIF4 A gene can inhibit the proliferation, migration ability and promote the apoptosis of ovarian cancer cells, which is expected to become a new target for ovarian cancer treatment.