解剖学报 ›› 2021, Vol. 52 ›› Issue (3): 410-417.doi: 10.16098/j.issn.0529-1356.2021.03.013

• 细胞和分子生物学 • 上一篇    下一篇

小泛素样修饰蛋白1假基因3通过蛋白激酶B信号通路影响非小细胞肺癌细胞系H1299增殖和凋亡

徐亮 杨剑烨 杨飞燕 杨国彪*   

  1. 绍兴市文理学院附属医院呼吸内科,浙江 绍兴 312000
  • 收稿日期:2020-03-11 修回日期:2020-07-01 出版日期:2021-06-06 发布日期:2021-06-06
  • 通讯作者: 杨国彪 E-mail:xbhaw49@163.com
  • 基金资助:
    浙江省医药卫生科技计划项目

Effect of small ubiquitin-like modifier 1 pseudogene 3 on the proliferation and apoptosis of non-small cell lung cancer cell line H1299 through protein kinase B signaling pathway

XU Liang  YANG Jian-ye  YANG Fei-yan  YANG Guo-biao*   

  1. Department of Respiratory Medicine, Affiliated Hospital of Shaoxing University of Arts and Sciences, Zhejiang Shaoxing 312000, China
  • Received:2020-03-11 Revised:2020-07-01 Online:2021-06-06 Published:2021-06-06
  • Contact: YANG Guo-biao E-mail:xbhaw49@163.com

摘要:

目的  探讨长链非编码RNA(lncRNA) 小泛素样修饰蛋白1假基因3(SUMO1P3)对非小细胞肺癌细胞增殖和凋亡的影响。   方法  用Real-time PCR方法测定非小细胞肺癌细胞系H1299中SUMO1P3表达变化。在H1299细胞中转染SUMO1P3小干扰RNA(siRNA),Real-time PCR方法测定下调效果, MTT法和5-乙炔基-2’-脱氧尿苷(EdU)法检测细胞增殖, PI单染法测定细胞周期,膜联蛋白 V-FITC(annexin V-FITC)/PI法检测细胞凋亡,TUNEL法检测细胞凋亡,Western blotting方法检测剪切的Caspase-3(c-Caspase-3)、细胞周期蛋白D1(cyclin D1)、P27、磷酸化磷脂酰肌醇-3激酶(p-PI3K)、磷酸化蛋白激酶B(p-Akt)蛋白表达。Akt信号激活剂处理转染SUMO1P3 siRNA后的H1299细胞,同样利用上述方法测定细胞增殖、凋亡和周期变化。样本数均为9。   结果  SUMO1P3在非小细胞肺癌细胞中表达上调。转染SUMO1P3 siRNA后的H1299细胞中SUMO1P3表达水平降低,细胞增殖能力下降,细胞G0/G1期比例升高,细胞凋亡率升高,细胞中c-Caspase-3和P27蛋白水平升高,cyclin D1、p-PI3K和p-Akt蛋白水平下降。Akt信号激活剂可以逆转SUMO1P3 siRNA对H1299细胞增殖抑制、周期阻滞和凋亡促进作用。   结论  下调SUMO1P3抑制非小细胞肺癌H1299细胞增殖并诱导细胞凋亡,作用机制与降低Akt信号通路激活水平有关。

关键词: 小泛素样修饰蛋白1假基因3, 蛋白激酶B, 免疫印迹法

Abstract:

Objective  To investigate the effect of long noncoding RNA (lncRNA) small ubiquitin-like modifier 1 pseudogene 3(SUMO1P3) on the proliferation and apoptosis of non-small cell lung cancer cell line 1299.   Methods  Determination of SUMO1P3 expression in non-small cell lung cancer cells by Real-time PCR. SUMO1P3 small interfering RNA(siRNA) was transfected into H1299 cells, the down regulation effect was determined by Real-time PCR.Cell proliferation was measured by MTT, 5-ethynyl-2’-deoxyuridine(EdU) method, the cell cycle was determined by PI single staining, apoptosis was detected by annexin Ⅴ-FITC/PI, detection of apoptosis by TUNEL,Western blotting was used to detect the expression of cleaved Caspase-3(c-Caspase-3), cyclin D1, P27, phosphorylated phospoinositide 3-kinase (p-PI3K)and phosphorylated protein kinase B(p-Akt). Akt signal activator treated H1299 cells transfected with SUMO1P3 siRNA, cell proliferation, apoptosis and cycle change were also measured by the above methods. The number of samples was 9.    Results  SUMO1P3 was up-regulated in non-small cell lung cancer cells. The expression of SUMO1P3 in H1299 cells decreased after transfection with SUMO1P3 siRNA, cell proliferation decreased, the ratio of G0/G1 phase increased, apoptosis rate increased, c-Caspase-3 and P27 protein in the cells increased, the protein levels of cyclin D1, p-PI3K and p-Akt decreased. Akt signal activator could reverse the inhibition of proliferation, cycle arrest and apoptosis of H1299 cells by SUMO1P3 siRNA.    Conclusion  Down-regulation of SUMO1P3 inhibits the proliferation of nonsmall cell lung cancer H1299 cells and induces apoptosis, the mechanism of action is related to the reduction of the activation level of the Akt signaling pathway.

Key words: Small ubiquitin-like modifier 1 pseudogene 3, Protein kinase B, Western blotting

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