Effect of HIF-1α expression inhibition by RNA interference on radiosensitivity and autophagy of hypoxic human lung adenocarcinoma cell line A549

邹燕梅; 熊华; 肖志平; 于世英; 袁响林

doi:10.3969/j.issn.1007-3969.2013.06.003

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Effect of HIF-1α expression inhibition by RNA interference on radiosensitivity and autophagy of hypoxic human lung adenocarcinoma cell line A549

更新时间：2025-12-31

- Effect of HIF-1α expression inhibition by RNA interference on radiosensitivity and autophagy of hypoxic human lung adenocarcinoma cell line A549
- China Oncology Vol. 23, Issue 6, Pages: 413-419(2013)
- 作者机构：
  
  华中科技大学同济医学院附属同济医院肿瘤中心,湖北,武汉,430030
- 作者简介：
- 基金信息：
- DOI：10.3969/j.issn.1007-3969.2013.06.003
  CLC：
- Published Online：13 November 2014，
  
  Published：2013
- 稿件说明：
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邹燕梅,熊华,肖志平,于世英,袁响林. RNAi沉默HIF-1α基因调控乏氧肺腺癌A549细胞放射敏感性和自噬能力[J]. 中国癌症杂志, 2013, 23(6): 413-419.

邹燕梅, 熊华, 肖志平, et al. Effect of HIF-1α expression inhibition by RNA interference on radiosensitivity and autophagy of hypoxic human lung adenocarcinoma cell line A549[J]. China Oncology, 2013, 23(6): 413-419.
邹燕梅,熊华,肖志平,于世英,袁响林. RNAi沉默HIF-1α基因调控乏氧肺腺癌A549细胞放射敏感性和自噬能力[J]. 中国癌症杂志, 2013, 23(6): 413-419. DOI： 10.3969/j.issn.1007-3969.2013.06.003.

邹燕梅, 熊华, 肖志平, et al. Effect of HIF-1α expression inhibition by RNA interference on radiosensitivity and autophagy of hypoxic human lung adenocarcinoma cell line A549[J]. China Oncology, 2013, 23(6): 413-419. DOI： 10.3969/j.issn.1007-3969.2013.06.003.

摘要

背景与目的：乏氧导致肿瘤细胞放射敏感性下降是引起肿瘤放疗抵抗、复发转移的根源。HIF-1α基因在乏氧调控中起关键作用，但HIF-1α基因在乏氧肺腺癌放射敏感性中的作用及其与自噬的关系尚未阐明。本研究建立利用RNAi沉默HIF-1α基因的乏氧肺腺癌A549细胞模型，以此探讨HIF-1α对乏氧细胞放射敏感性和自噬的影响。方法：构建靶向抑制HIF-1α的shRNA表达质粒，转染乏氧A549细胞，筛选稳定表达的克隆细胞，将其命名为A549/HIF-1α-shRNA，同时设阴性对照A549/Neg-shRNA。克隆形成实验检测细胞D0、SF2、SER等值。Western-blot检测细胞照射前后HIF-1α、LC3、c-parp蛋白的表达。结果：乏氧A549细胞的SF2为0.62，高于常氧A549细胞，SER为1.45；乏氧A549细胞照射后LC3Ⅱ增加，c-parp下调。乏氧A549细胞HIF-1α表达增加；A549/HIF-1α-shRNA细胞HIF-1α表达降低；A549/HIF-1α-shRNA细胞的SF2为0.45，SER为0.72，低于A549/Neg-shRNA细胞；A549/HIF-1α-shRNA 细胞照射后LC3Ⅱ降低，c-parp上调。结论：稳定转染及RNAi技术建立的HIF-1α表达抑制克隆可以成为简单实用的细胞模型；shRNA抑制HIF-1α的表达可提高乏氧A549细胞的放射敏感性，降低自噬活性。

Abstract

Background and purpose: Hypoxia induced the decreased radiosensitivity of tumor cells

which was the cause of tumor radioresistance and relapse and metastasis. During the course

HIF-1a played the most important role in the regulation of hypoxia. However

it’s still unknown about the effect of HIF-1a on the radiosensitivity of hypoxia tumor cells and the relationship with autophagy. This study was to inhibit HIF-1α expression in hypoxic lung adenocarcinoma cell line A549 with RNA interference (RNAi)

and explore its effect on hypoxic cell radiosensitivity and autophagy. Methods: Plasmids pHIF-1α-shRNA and Neg-shRNA (negative control) were constructed and transfected into hypoxic A549 cells

this positive clone was named A549/HIF-1α-shRNA. Clone formation array was applied to calculate the value of D0

SF2

SER. The expression of HIF-1α

LC3

c-parp was detected by Western blot. Results: The SF2 of hypoxic A549 cell was 0.62

which was higher than that of normoxic A549 cell

SER was 1.45. The level of LC3Ⅱ increased significantly and the level of c-parp decreased after the radiation of hypoxic A549 cell. The level of HIF-1a increased in hypoxic A549 cells. The expression of HIF-1α in hypoxic A549 cells was suppressed markedly after transfection of HIF-1α-shRNA; this clone was named A549/HIF-1α-shRNA. The SF2 and SER were significantly lower in A549/HIF-1α-shRNA cells

0.45 and 0.72 respectively. Under the hypoxic condition and after the inhibition of HIF-1α

the expression of LC3Ⅱ decreased significantly and the expression of c-parp increased. Conclusion: We successfully established a cell model that HIF-1α expression was suppressed almost completely by RNAi. The inhibition of HIF-1α by shRNA may raise the radiosensitivity and decrease the autophagy of hypoxic A549 cells in vitro.

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