Smac调控Caspase-3对耐药肺腺癌细胞株生物活性及相关信号的研究

摘要 目的：探讨Smac基因调控Caspase-3表达对紫杉醇耐药肺腺癌细胞株生物活性及经典凋亡信号通路的作用机制。方法：取构建好的耐药A549细胞,将其分为A549细胞（LC）组、A549细胞+Smac-NC（SN）组、A549细胞+Smac抑制剂（SI）组、A549细胞+Smac激动剂（SM）组、A549细胞+Caspase-3-NC（CN）组、A549细胞+Caspase-3抑制剂（CI）组、A549细胞+Caspase-3激动剂（CM）组、A549细胞+Smac激动剂+Caspase-3激动剂（MM）组;Real-time PCR法检测正常肺上皮细胞及4种肺腺癌细胞系中Smac、Caspase-3表达水平,将阴性对照、Smac、Caspase-3类似物转染至紫杉醇耐药肺腺癌细胞株,MTT法检测细胞增殖,流式细胞仪检测细胞凋亡,免疫印迹法检测经典凋亡信号通路表达,并分析Smac与Caspase-3的相关性。结果：肺腺癌细胞系中的Smac、Caspase-3 mRNA表达量显著低于正常肺上皮细胞系BEAS-2B（P＜0.05）,其中A549的Smac、Caspase-3 mRNA值最小（P＜0.05）,因此选取其作为此次实验细胞;LC组与SN组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与SN组相比,SI组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显降低（P＜0.05）,增殖率、Bcl-2表达明显升高（P＜0.05）,与SI组相比,SM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;LC组与CN组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与CN组相比,CI组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显降低（P＜0.05）,增殖率、Bcl-2表达明显升高（P＜0.05）,与CI组相比,CM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;SM组与CM组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与CM组相比,MM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;Smac与Caspase-3呈现正相关（r=0.470,P=0.002）,组间具有显著差异。结论：Smac基因可显著改善紫杉醇耐药肺腺癌细胞株细胞生物活性,并激活经典凋亡信号通路,其作用机制可能与调控Caspase-3表达有关。     

低剂量螺旋CT联合肺癌自身抗体、肿瘤标志物CA12-5、CA19-9在肺癌诊断中的临床应用价值

摘要 目的：探讨低剂量螺旋CT联合肺癌自身抗体、肿瘤标志物糖类抗原（CA）12-5、CA19-9在肺癌诊断中的临床应用价值。方法：选择2019年1月至2021年12月在我院经低剂量螺旋CT检查发现肺部病灶的患者86例,以病理诊断结果分为肺癌组49例、肺部良性病变组37例,另选取同期健康体检志愿者30例作为对照组。观察肺癌的低剂量螺旋CT影像学特征,检测比较各组血清肺癌自身抗体[p53、SOX区域Y相关HMG蛋白家族成员2（SOX2）、G抗原7（GAGE7）、RNA解旋酶自身抗体4-5（GBU4-5）]及CA12-5、CA19-9水平,并对比阳性表达情况,分析低剂量螺旋CT联合肺癌自身抗体、CA12-5、CA19-9对肺癌的诊断价值。结果：49例肺癌患者经低剂量螺旋CT检查共发现51个病灶,其中实性结节或部分实性结节30个,非实性结节21个;病灶平均直径（10.92±1.17）mm,＜5 mm者9个,5～10 mm者17个,≥10 mm者25个;11个病灶伴有钙化,16个病灶有毛刺征,20个病灶有分叶征;有6例表现为空洞,空洞通常壁薄厚不均匀,呈中心性或偏心性。肺癌组血清p53、SOX2、GAGE7、GBU4-5及CA12-5、CA19-9水平均明显高于肺部良性病变组和对照组,肺部良性病变组上述指标水平均明显高于对照组（P<0.05）。肺癌组CA12-5、CA19-9、4项肺癌自身抗体单独及联合的阳性率均明显高于肺部良性病变组和对照组（P<0.05）,肺部良性病变组CA12-5、CA19-9及4项肺癌自身抗体联合的阳性率高于对照组（P<0.05）。低剂量螺旋CT联合4项肺癌自身抗体、CA12-5、CA19-9对肺癌诊断的灵敏度为91.84%,特异度为97.30%,准确度为94.19%。结论：低剂量螺旋CT联合肺癌自身抗体、肿瘤标志物CA12-5、CA19-9在肺癌诊断中有较好的临床应用价值。     

快速康复外科理念对单孔胸腔镜肺叶切患者术后应激因子、肺功能及疼痛评分的影响

目的:探讨快速康复外科理念(FTS)对单孔胸腔镜肺叶切除患者术后应激因子、肺功能及疼痛评分的影响。方法:选择2018年5月~2020年12月期间我院收治的单孔胸腔镜肺叶切除患者117例,根据随机数字表法将患者分为对照组(58例)和研究组(59例),对照组给予常规干预,研究组给予FTS干预。对比两组手术指标、应激因子、肺功能、疼痛评分及并发症。结果:研究组的胸管留置时间、术后排气时间、卧床时间、住院时间均短于对照组(P<0.05)。两组术后5 d C反应蛋白(CRP)、白细胞介素-6(IL-6)、皮质醇(Cor)升高,但研究组的升高幅度更低(P<0.05)。两组术后5 d第一秒最大呼气量(FEV1)、用力肺活量(FVC)、最大通气量(MVV)升高,且研究组的升高幅度更高(P<0.05)。研究组术后4 h、24 h、48 h的视觉疼痛模拟量表(VAS)评分低于对照组(P<0.05)。研究组的并发症总发生率低于对照组(P<0.05)。结论:FTS应用于单孔胸腔镜肺叶切除术可有效减轻术后应激,减轻患者疼痛,促进患者肺功能恢复,降低并发症发生风险。     

右美托咪定联合肺保护性通气策略对肺癌根治术患者氧化应激、炎症反应和免疫功能的影响

摘要 目的：探讨右美托咪定联合肺保护性通气策略对肺癌根治术患者氧化应激、炎症反应和免疫功能的影响。方法：选择我院行肺癌根治术患者110例,入选患者根据信封抽签法分为A组和B组,各为55例。A组接受右美托咪定联合传统通气策略,B组接受右美托咪定联合肺保护性通气策略,观察两组患者的氧合指数（OI）、氧化应激指标[超氧化物歧化酶（SOD）、丙二醛（MDA）]、炎症反应指标[白介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）]、免疫功能变化以及术后肺部并发症发生率。结果：B组单肺通气前即刻（T2）~恢复双肺通气15 min（T6）时间点OI高于A组（P<0.05）。B组术后72 h（T7）时间点CD3⁺、CD4⁺、CD4⁺/CD8⁺高于A组,CD8⁺低于A组（P<0.05）。B组T7时间点SOD水平高于A组,MDA水平低于A组（P<0.05）。B组T7时间点IL-6、TNF-α水平低于A组（P<0.05）。两组术后肺部并发症发生率组间对比无统计学差异（P>0.05）。结论：右美托咪定联合肺保护性通气策略可改善肺癌根治术患者氧合和炎症反应,有效减轻免疫抑制及氧化应激反应。     

肺癌相关基因甲基化谱研究进展

近年来,细胞基因组的表观遗传学(epigenetics)改变在肿瘤发生发展中的作用日益受到国内外学者的关注。基因启动子区域CpG岛的甲基化导致的基因表达的失活是表观遗传学改变的重要方式之一。大量的研究表明,肺癌中存在多种抑癌基因或促凋亡基因启动子甲基化,并证实这些基因的甲基化与肺癌的发生、发展相关。这些研究为原发性肺癌的早期诊断和疾病监测找到了新的分子标志物。现对与肺癌发生相关基因甲基化的研究现状作一综述。     

Alcohol consumption and lung cancer risk: A pooled analysis from the International Lung Cancer Consortium and the SYNERGY study

BackgroundThere is inadequate evidence to determine whether there is an effect of alcohol consumption on lung cancer risk. We conducted a pooled analysis of data from the International Lung Cancer Consortium and the SYNERGY study to investigate this possible association by type of beverage with adjustment for other potential confounders.MethodsTwenty one case-control studies and one cohort study with alcohol-intake data obtained from questionnaires were included in this pooled analysis (19,149 cases and 362,340 controls). Adjusted odds ratios (OR) or hazard ratios (HR) with corresponding 95% confidence intervals (CI) were estimated for each measure of alcohol consumption. Effect estimates were combined using random or fixed-effects models where appropriate. Associations were examined for overall lung cancer and by histological type.ResultsWe observed an inverse association between overall risk of lung cancer and consumption of alcoholic beverages compared to non-drinkers, but the association was not monotonic. The lowest risk was observed for persons who consumed 10–19.9 g/day ethanol (OR vs. non-drinkers = 0.78; 95% CI: 0.67, 0.91), where 1 drink is approximately 12–15 g. This J-shaped association was most prominent for squamous cell carcinoma (SCC). The association with all lung cancer varied little by type of alcoholic beverage, but there were notable differences for SCC. We observed an association with beer intake (OR for ≥20 g/day vs nondrinker = 1.42; 95% CI: 1.06, 1.90).ConclusionsWhether the non-monotonic associations we observed or the positive association between beer drinking and squamous cell carcinoma reflect real effects await future analyses and insights about possible biological mechanisms.     

急性呼吸窘迫综合征小鼠肺内源性干细胞表达水平的研究

目的探讨急性呼吸窘迫综合征（ARDS）小鼠肺组织中肺内源性干细胞的表达水平。 方法10只C57BL/6小鼠分成两组：实验组和对照组,实验组通过气管内注射脂多糖（LPS）构建小鼠ARDS模型,采用气管内注射PBS作为对照组;采用胶原酶、热消化法消化小鼠肺组织获取小鼠肺单细胞悬液;双重免疫荧光染色方法鉴定小鼠肺组织中sca-1⁺CD31^-CD45^-细胞;流式细胞术对肺sca-1⁺CD31^-CD45^-细胞进行分选。采用方差分析及独立t检验进行统计学分析。 结果通过气管内注入LPS成功制作小鼠急性ARDS模型;5只小鼠的全肺组织制备单细胞悬液总数目达5×10⁷个/ml,活细胞百分比为98﹪;肺内源性干细胞包括Ⅱ型肺泡上皮细胞、clara细胞以及支气管肺泡干细胞等,通过肺组织双重免疫荧光染色,验证小鼠肺组织Ⅱ型肺泡上皮细胞、clara细胞以及支气管肺泡干细胞;对照组及实验组各样本肺内源性干细胞数目占单细胞悬液细胞数比例呈正态分布,且实验组肺内源性干细胞数目水平（10.73±10.65）﹪较对照组水平（12.23±0.73）﹪降低（t = -3.405,P < 0.01）。 结论ARDS时,小鼠肺内源性干细胞（sca-1⁺CD31^-CD45^-）水平降低,减少的肺内源性干细胞具体去向尚不明确,其有可能参与机体急性炎症过程中气道上皮细胞的修复、再生过程。     

肺炎克雷伯菌致肺癌患者术后肺部感染的病因与耐药性分析

目的探讨肺癌术后并发肺炎克雷伯菌肺部感染的病因和耐药情况,为术后肺炎克雷伯菌感染的预防和治疗提供病原学依据。方法收集2011年1月1日至2014年6月30日本院肺癌术后合并肺部感染患者下呼吸道标本,常规分离培养肺炎克雷伯菌,K-B纸片法进行药敏试验,利用WHONET 5.6软件分析处理试验数据。结果从肺癌术后患者下呼吸道分离的肺炎克雷伯菌产ESBLs高,达68.1%;对头孢替坦、阿米卡星、亚胺培南和美罗培南都较敏感,耐药率分别为6.0%、21.4%、18.1%和20.9%,其余抗菌药物的耐药率均30.0%。结论肺癌术后并发肺炎克雷伯菌肺部感染耐药率高,对碳青霉烯类抗菌药物仍保持高度敏感性,临床应加强耐药性监测,并根据药敏试验结果合理选用抗菌药物。     

肠杆菌科细菌致恶性肿瘤患者肺部感染的临床特点与耐药性分析

目的探讨肠杆菌科细菌致恶性肿瘤患者肺部感染的临床特点、病原学构成和耐药特征,为今后的临床治疗提供参考。方法收集2009年7月至2014年6月本院住院恶性肿瘤患者下呼吸道标本,应用BD Phoenix100全自动微生物鉴定仪鉴定菌种,药敏试验采用纸片法(K-B)进行,并进行ESBLs检测,按CLSI标准判定药敏结果,用WHONET 5.6软件分析数据。结果从恶性肿瘤患者下呼吸道标本中共分离肠杆科细菌256株,主要为肺炎克雷伯菌和大肠埃希菌,分别占39.5%和33.2%;药敏试验表明常见肠杆菌科细菌除对碳青霉烯类、含酶抑制剂抗菌素耐药率较低外,对其余抗菌药物均为中高度耐药;肺炎克雷伯菌的ESBLs检出率最高为60.4%。结论从本院恶性肿瘤合并下呼吸道感染患者中分离出的肠杆菌科细菌多重耐药现象严重,产酶率高,治疗该类细菌感染,首选碳青霉烯类、含酶抑制剂抗菌素,临床应加强ESBLs检测和耐药性监测,防止多重耐药菌的产生。     

Fibroblast viability and phenotypic changes within glycated stiffened three-dimensional collagen matrices

BackgroundThere is growing interest in the development of cell culture assays that enable the rigidity of the extracellular matrix to be increased. A promising approach is based on three-dimensional collagen type I matrices that are stiffened by cross-linking through non-enzymatic glycation with reducing sugars.MethodsThe present study evaluated the biomechanical changes in the non-enzymatically glycated type I collagen matrices, including collagen organization, the advanced glycation end products formation and stiffness achievement. Gels were glycated with ribose at different concentrations (0, 5, 15, 30 and 240 mM). The viability and the phenotypic changes of primary human lung fibroblasts cultured within the non-enzymatically glycated gels were also evaluated along three consecutive weeks. Statistical tests used for data analyze were Mann–Whitney U, Kruskal Wallis, Student’s t-test, two-way ANOVA, multivariate ANOVA, linear regression test and mixed linear model.ResultsOur findings indicated that the process of collagen glycation increases the stiffness of the matrices and generates advanced glycation end products in a ribose concentration-dependent manner. Furthermore, we identified optimal ribose concentrations and media conditions for cell viability and growth within the glycated matrices. The microenvironment of this collagen based three-dimensional culture induces α-smooth muscle actin and tenascin-C fibroblast protein expression. Finally, a progressive contractile phenotype cell differentiation was associated with the contraction of these gels.ConclusionsThe use of non-enzymatic glycation with a low ribose concentration may provide a suitable model with a mechanic and oxidative modified environment with cells embedded in it, which allowed cell proliferation and induced fibroblast phenotypic changes. Such culture model could be appropriate for investigations of the behavior and phenotypic changes in cells that occur during lung fibrosis as well as for testing different antifibrotic therapies in vitro.

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