Qiu Chen, Diao Zhen-Hua, Qi Hui, Li Na, Ren Li-Li, Zhang Ting
Department of Respiratory Medicine, Second Clinical Medical College, Jinan University, Shenzhen 518020, China.
Zhonghua Jie He He Hu Xi Za Zhi. 2010 Nov;33(11):811-6.
to investigate the effect of gene silencing of integrin β1 on the proliferation and secretory function of airway smooth muscle cells (ASMC) in asthmatic mice.
integrin β1 gene was silenced by using RNAi technology in the fifth generation ASMC of normal and asthmatic mice. The integrin β1 mRNA expression and integrin β1 protein were analyzed by RT-PCR and Western blot respectively before and after the integrin β1 gene silencing in ASMC. The cell proliferation changes of ASMC were measured by MTT assay before and after gene silencing of integrin β1. The cell cycle distribution and apoptosis rate were analyzed by flow cytometry. The expression of interleukin (IL)-6 and RANTES were analyzed by ELISA assay.
(1) integrin β1 mRNA expression by RT-PCR (0.907 ± 0.041) was significantly higher in asthma control group compared with normal control group (0.527 ± 0.027) (t = 24.632, P < 0.05), and was significantly decreased in asthma siRNA intervention group (0.503 ± 0.034) compared with asthma control group (t = 24.079, P < 0.05). There was no significant difference in the normal group compared with the intervention group (t = 1.077, P > 0.05). (2) Integrin β1 protein expression (0.733 ± 0.067) was significantly higher (t = 13.622, P < 0.05) in the asthma control group compared with normal control group (0.386 ± 0.044), and was significantly reduced in the asthma siRNA intervention group (0.453 ± 0.074) compared with the control group (t = 8.880, P < 0.05). There was no significant change in the normal control group after the intervention (t = 1.908, P > 0.05). (3) The absorbance value was significantly enhanced in the asthma control group compared with the same period of normal control group (t = 9.528, 5.799, 3.372, all P < 0.05), and was significantly reduced in asthma siRNA intervention group compared with the same period of asthma control group (t = 2.684, 2.546, 2.897, all P < 0.05), and no significant changes in normal group after the intervention (t = 0.067, 1.198, 0.589, all P > 0.05). (4) The S + G(2)/M phase ratio of ASMC was significantly higher in asthma control group [(49 ± 4)%] compared with normal control group (34 ± 4)% (t = 8.035, P < 0.05), and was significantly lower in the asthma siRNA intervention group [(42 ± 7)%] compared with asthma control group (t = 2.212, P < 0.05), and no significant changes in normal control group after the intervention (t = 0.699, P > 0.05). (5) The apoptosis rate was significantly lower in asthma control group [(3.9 ± 1.4)%] compared with normal control group [(7.4 ± 0.5)%] (t = 7.465, P < 0.05), and was significantly higher in asthma siRNA intervention group [(12.6 ± 2.4)%] compared with asthma control group (t = 9.839, P < 0.05), but no significant changes in normal control group after the intervention (t = 2.094, P > 0.05). (6) The secreted IL-6 [(545 ± 28) ng/L] and RANTES [(345 ± 28) ng/L] were higher in asthma control group compared with the normal control group [(219 ± 26) ng/L, (138 ± 16) ng/L, respectively, t = 26.789, 20.451, all P < 0.05], and was significantly decreased in the asthma siRNA intervention group [(347 ± 26) ng/L, (250 ± 24) ng/L] compared with asthma control group (t = 6.192, 4.590, all P < 0.05), while there was no significant difference in the normal control group after the intervention.
gene silencing targeting integrin β1 inhibited proliferation and secretion, but promoted apoptosis of ASMC from asthmatic mice.
探讨整合素β1基因沉默对哮喘小鼠气道平滑肌细胞（ASMC）增殖及分泌功能的影响。
采用RNAi技术沉默正常及哮喘小鼠第5代ASMC中的整合素β1基因。分别于整合素β1基因沉默前后，采用RT-PCR和Western blot分析ASMC中整合素β1 mRNA表达及整合素β1蛋白。采用MTT法检测整合素β1基因沉默前后ASMC的细胞增殖变化。采用流式细胞术分析细胞周期分布及凋亡率。采用ELISA法分析白细胞介素（IL）-6和调节激活正常T细胞表达和分泌的趋化因子（RANTES）的表达。
（1）与正常对照组（0.527±0.027）相比，哮喘对照组RT-PCR检测的整合素β1 mRNA表达（0.907±0.041）显著升高（t=24.632，P<0.05）；与哮喘对照组相比，哮喘siRNA干预组（0.503±0.034）显著降低（t=24.079，P<0.05）。正常组与干预组相比无显著差异（t=1.077，P>0.05）。（2）与正常对照组（0.386±0.044）相比，哮喘对照组整合素β1蛋白表达（0.733±0.067）显著升高（t=13.622，P<0.05）；与对照组相比，哮喘siRNA干预组（0.453±0.074）显著降低（t=8.880，P<0.05）。干预后正常对照组无显著变化（t=1.908，P>0.05）。（3）与同期正常对照组相比，哮喘对照组吸光度值显著升高（t=9.528、5.799、3.372，均P<0.05）；与同期哮喘对照组相比，哮喘siRNA干预组显著降低（t=2.684、2.546、2.897，均P<0.05），干预后正常组无显著变化（t=0.067、1.198、0.589，均P>0.05）。（4）与正常对照组（34±4）%相比，哮喘对照组ASMC的S+G(2)/M期比例[(49±4)%]显著升高（t=8.035，P<0.05）；与哮喘对照组相比，哮喘siRNA干预组[(42±7)%]显著降低（t=2.212，P<0.05），干预后正常对照组无显著变化（t=0.699，P>0.05）。（5）与正常对照组[(7.4±0.5)%]相比，哮喘对照组凋亡率[(3.9±1.4)%]显著降低（t=7.465，P<0.05）；与哮喘对照组相比，哮喘siRNA干预组[(12.6±2.4)%]显著升高（t=9.839，P<0.05），但干预后正常对照组无显著变化（t=2.094，P>0.05）。（6）与正常对照组[(219±26) ng/L、(138±16) ng/L]相比，哮喘对照组分泌的IL-6[(545±28) ng/L]和RANTES[(345±28) ng/L]更高（t=26.789、20.451，均P<0.05）；与哮喘对照组相比，哮喘siRNA干预组[(347±26) ng/L、(250±24) ng/L]显著降低（t=6.192、4.590，均P<0.05），而干预后正常对照组无显著差异。
靶向整合素β1的基因沉默抑制了哮喘小鼠ASMC的增殖和分泌，但促进了其凋亡。
