Zhu Min, Sun Yuyu, Xue Huawei, Wu Gang, Wang Zhen, Shi Junfeng, Ma Jiye, Gu Baorong, Yan Xiaoling
Department of Spine Surgery, Nantong Third People's Hospital, Affiliated Nantong Hospital 3 of Nantong University, Nantong, People's Republic of China.
Department of Chemotherapy, Affiliated Hospital of Nantong University, Nantong, People's Republic of China.
Int J Gen Med. 2023 Jul 4;16:2831-2848. doi: 10.2147/IJGM.S413461. eCollection 2023.
Osteosarcoma is a malignant tumor originating from the skeletal system. There is no effective treatment other than surgery and chemotherapy, which seriously endangers the health of children and adolescents. NEK6 is a novel discovered Serine/Threonine protein kinase that can regulate cell cycle and activate several oncogenic pathways.
NEK6 expression in pan-cancer including sarcoma was evaluated using analysis tools of TIMER, UALCNA and GEPIA with TCGA database, and its association with overall survival in patients with sarcoma was also analyzed. TargetScan, tarbase, microT-CDS and Starbase online software were used to predict NEK6-targeted miRNAs, including miR-26a-5p. Tumor tissues from patients with osteosarcoma were collected for NEK6 and miRNA detection using RT-qPCR. NEK6 down-regulated by siRNAs or miR-26a-5p in osteosarcoma cells was detected by RT-qPCR, Western blot and Immunofluorescence staining assays. Effects of NEK6 knockdown on proliferation, migration, invasion and apoptosis of osteosarcoma cells were detected by CCK-8, wound healing, transwell and flow cytometry, respectively. The expressions of STAT3, metastasis and apoptosis-related genes were detected by Western blot.
High expression of NEK6 and low expression of miR-26a-5p were lowly expressed in osteosarcoma and they were negative correlation. NEK6 has been confirmed as a direct target for miR-26a-5p. In addition, NEK6 down-regulated by siRNAs or miR-26a-5p led to inhibition of cell proliferation, migration and invasion while promoting cell apoptosis. The levels of phosphorylated STAT3 and metastasis genes (MMP-2, MMP-9) were inhibited, while apoptotic gene Bax was promoted and Bcl2 was inhibited by miR-26a-5p upregulation.
NEK6 can promote osteosarcoma progression via activating STAT3 signaling pathway, which is inhibited by miR-26a-5p, suggesting that NEK6 is a potential oncogene and miR-26a-5p is a suppressor of osteosarcoma. The strategy of inhibiting of NEK6 by miR-26a-5p may be an effective approach for osteosarcoma therapy.
骨肉瘤是一种起源于骨骼系统的恶性肿瘤。除手术和化疗外，尚无有效的治疗方法，这严重危及儿童和青少年的健康。NEK6是一种新发现的丝氨酸/苏氨酸蛋白激酶，可调节细胞周期并激活多种致癌途径。
使用TIMER、UALCNA和GEPIA分析工具以及TCGA数据库评估NEK6在包括肉瘤在内的泛癌中的表达，并分析其与肉瘤患者总生存期的关系。使用TargetScan、tarbase、microT-CDS和Starbase在线软件预测NEK6靶向的miRNA，包括miR-26a-5p。收集骨肉瘤患者的肿瘤组织，采用RT-qPCR检测NEK6和miRNA。通过RT-qPCR、蛋白质免疫印迹法和免疫荧光染色法检测骨肉瘤细胞中siRNA或miR-26a-5p下调的NEK6。分别通过CCK-8、伤口愈合、transwell和流式细胞术检测NEK6敲低对骨肉瘤细胞增殖、迁移、侵袭和凋亡的影响。通过蛋白质免疫印迹法检测STAT3、转移和凋亡相关基因的表达。
NEK6高表达和miR-26a-5p低表达在骨肉瘤中表达较低，且呈负相关。NEK6已被确认为miR-26a-5p的直接靶点。此外，siRNA或miR-26a-5p下调NEK6可导致细胞增殖、迁移和侵袭受到抑制，同时促进细胞凋亡。miR-26a-5p上调可抑制磷酸化STAT3和转移基因（MMP-2、MMP-9）的水平，同时促进凋亡基因Bax的表达并抑制Bcl2的表达。
NEK6可通过激活STAT3信号通路促进骨肉瘤进展，而miR-26a-5p可抑制该通路，提示NEK6是一种潜在的癌基因，miR-26a-5p是骨肉瘤的抑制因子。miR-26a-5p抑制NEK6的策略可能是骨肉瘤治疗的有效方法。
