木材烟雾凝聚物对人气道平滑肌细胞的弱增殖和强坏死效应  

Wood smoke condensate had weak proliferation and strong necrotic effects on human airway smooth muscle cells

作　　者：潘仪玲[1] 李冰[2] 冉丕鑫[1] 

机构地区：[1]呼吸疾病国家重点实验室(广州医学院第一附属医院),510120 [2]广州医学院实验医学研究中心

出　　处：《中华结核和呼吸杂志》2013年第8期581-586,共6页Chinese Journal of Tuberculosis and Respiratory Diseases

基　　金：国家自然基金（81170043）;国家“十二五”支撑课题（2012BA105801）;教育部博士点基金（20104423110001）;广东省自然科学基金（S2011020002789）

摘　　要：目的观察木材烟雾凝集物（WSC）对人气道平滑肌细胞（HASMC）增殖与坏死的影响。方法原代培养的HASMC取2至8代细胞，分为对照组、WSC组和烟草烟雾凝集物（CSC）组，每组4个复孔。用细胞毒性检测试剂盒检测细胞活性，用核酸类似物标记检测细胞增殖试剂盒和流式细胞术检测增殖期（s期）细胞比例并分析细胞周期，以荧光定量PCR和Westernblot法定量检测细胞周期蛋白D1（cyclinD1）的表达。用膜联蛋白-V／碘化丙啶双荧光染色法鉴别细胞坏死和凋亡。多组间比较采用单因素方差分析，两两比较采用LSD—t检验。结果细胞活性峰浓度时，WSC组和CSC组的细胞活性[（126±12）％和（142±11）％]均显著高于对照组[（100±0）％]，细胞活性抑制浓度时（WSC10mg／L、CSC60mg／L），WSC组和CSC组的细胞活性（86％和76％）均显著低于对照组[（100±0）％]，差异均有统计学意义（q值为3．63～9．33，均P〈0．05）。WSC1mg／L组细胞增殖比例和cyclinD1蛋白表达量[（124±20）％和1．31＋0．64]显著高于对照组[（100±0）％和1．0±0．0]．差异均有统计学意义（g值分别为5．85和5．91，均P〈0．05），S＋G：／M期细胞比例和cyclinD1的mRNA表达量[（103±4）％和1．18±0．21]与对照组[（100±0）％和1．0±0．0]比较，差异均无统计学意义（q值分别为1．16和2．05，均P〉0．05）；CSC10mg／L组细胞增殖比例、cyclinD1蛋白表达量、S＋G：／M期细胞比例和cyclinD1的mRNA表达量分别为（204±45）％、1．80±0．25、（140±6）％和1．48±0．2，均显著高于WSC1mg／L组和对照组[（100±0）％、1．0±0．0、（100±0）％和1．0±0．01。WSC10mg／L组坏死细胞比例[（13．39±0．15）％]均显著高于对照组[（1．57±0．41）％]和CSC60mg／L组[（6．61±1．91）％]，CSC40mg／L组凋亡细胞比例[（61．8±10．6）％]均显著高于对照组[（0．0±0．0）％]Objective To investigate the effect of wood smoke condensate （WSC） on proliferation and necrosis of human airway smooth muscle cells （ HASMCs ） . Methods Primary cultured HASMCs between passage 2 and 8 were divided into 3 groups： a control group, a WSC group and a cigarette smoke condensate （CSC） group. The viability of cells was examined by the CCK8 assays. The ratio of cellular proliferative stage （S phase） and cell cycle index were examined by fluorescent-labeled thymidine analogue uptake assays and flow cytometry. The expression of cyclin D1 was detected by quantitative reverse transcriptase polymerase chain reaction （Q-PCR） and Western blot. Cell apoptosis and necrosis were observed by the annexin-V and PI staining. Statistical analysis was performed by using the One-way ANOVA and LSD-t test. Results Cell viability reached peak at WSC 1 mg/L[ （126 ± 12）% ] and at CSC 10 mg/L exposure level [ （ 142 ± 11 ） % ] respectively. While at WSC 10 mg/L and CSC 60 mg,/L exposure levels, cell viability decreased significantly to 86% and 76% , respectively, as compared with that of the blank control group [ （ 100 ± 0） % ] （q = 3.63 - 9. 33,P 〈 0. 05 ）. In the WSC 1 mg/L group, the cell proliferation ratio and the expression of cyclin D1 protein were （ 124 ± 20）% and 1.31 ± 0. 64, respectively, the differences being significant as compared with the blank control group [ （ 100 ± 0 ） %, 1.0 ± 0.01 （ q = 5.85, 5.91, P〈0. 05）, while the expression of cyclin D1 mRNA and the percentage of S ± G2M phase were 1. 18 ± 0. 21 and （ 103 ± 4） % , respectively, not significantly different as compared to the control group [（100±0）%, 1.0 ±0.0], （q =1.16, 2. 05, P〉0.05）. In the CSC 10 mg/L group, the above-mentioned values were （204 ±45 ） % , 1.80 ± 0. 25, （ 140 ± 6 ） % , 1.48 ± 0. 2, respectively, which were significantly higher than those in the blank control group （ q = 5.38 - 16. 51, P 〈 0. 05 ） and in the WSC group （q =3.35 -15.35, P �

关 键 词：烟雾 肌细胞 平滑肌 细胞增殖 

分 类 号：R363[医药卫生—病理学]