目的 观察辛伐他汀对脂多糖(LPS)诱导的急性肺损伤小鼠保护作用及对高迁移率族蛋白B1(HMGB1)/Toll样受体4(TLR4)/核因子κB(NF-κB)信号通路的影响.方法 将48只12～15周龄的SPF级Wistar雄性大鼠随机分为4组,模型组尾部静脉注射0.5 ml的LPS(5 mg/kg)构建急性肺损伤(ALI)模型,对照组给予等体积的生理盐水,地塞米松(DXM)干预组和辛伐他汀干预组则在LPS注射前24 h和10 min时分别给予腹腔注射1ml的DXM(2 mg/kg)和1ml辛伐他汀溶液(5 mg/kg)进行预处理,对照组和模型组则给予等体积的生理盐水代替.4h后苏木精-伊红染色法(HE)染色观察大鼠肺部组织病变,并进行评分,ELISA法检测支气管灌洗液(BALF)中肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)和白细胞介素-1β(IL-1β)因子水平,同时显微镜下观察并统计总细胞和嗜中性粒细胞的个数,计算肺组织湿干重比(W/D),ELISA检测肺组织中的髓过氧化物酶(MPO),蛋白免疫印迹检测HMGB 1/TLR4/NF-κB信号通路中的HMGB1、TLR4及NF-κB蛋白表达.结果 和对照组相比,模型组大鼠在受到LPS刺激后,W/D比值、BALF中的嗜中性粒细胞、总细胞数量及MPO活力出现明显升高(P＜0.05),与模型组相比,DXM组和辛伐他汀组上述指标均出现明显下降(P＜0.05).和对照组相比,模型组的肺泡腔明显变小,间隔出现增厚,肺泡壁出现充血水肿,且发现了大量的炎性细胞浸润,而DXM组、辛伐他汀组和模型组相比,肺组织结构接近于正常,炎性细胞浸润得到缓解,对照组、模型组、DXM组以及辛伐他汀组的HE评分分别为0.79±0.13、2.58±0.21、1.15±0.19和1.21±0.21,模型组HE评分明显高于对照组(t=25.106,P＜0.05),DXM组和辛伐他汀组HE评分明显低于模型组,差异有统计学意义(t=17.492、15.980,均P＜0.05).和对照组相比,模型组中IL-1β、IL-6和TNF-α水平出现明显的升高(P＜0.05);与模型组相比,DXM组和辛伐他汀组大鼠BALF中IL-1β、IL-6和TNF-α水平出现明显的下降,差异有统计学意义(P＜0.05).和对照组相比,模型组中HMGB1、TLR4及NF-κB蛋白表达明显升高(P＜0.05);和模型组相比,DXM组和辛伐他汀组大鼠肺组织中HMGB1、TLR4及NF-κB蛋白表达明显下降,差异有统计学意义(P＜0.05).结论 辛伐他汀可以抑制LPS所致肺损伤,其作用机制可能与HMGB1/TLR4/NF-κB信号通路有关.

目的 探讨滑石粉胸膜固定术治疗慢性阻塞性肺疾病(COPD)并发难治性气胸的作用机制.方法 回顾性分析25例COPD合并难治性气胸患者,经胸腔镜胸腔内喷洒滑石粉混悬液后,观察患者术前及术后静脉血白细胞、中性粒细胞、血沉、C-反应蛋白(CRP)、动脉血气分析、肝功能、肾功能及心肌酶谱的变化.结果 胸腔内喷洒滑石粉混悬液治疗后,患者血WBC、中性粒细胞、血沉、CRP均显著升高(P＜0.05),但动脉血气分析、肝、肾功能及心肌酶谱无显著变化(P＞0.05).结论 滑石粉胸膜固定术治疗COPD并发难治性气胸,对患者动脉血气分析、肝肾功能及心肌酶谱等无明显影响,安全性好.滑石粉胸膜固定术后患者静脉血中炎性指标明显升高,提示炎症反应可能是其治疗和发生并发症的机制之一.

Fibromodulin (FMOD), an ECM small leucine-rich proteoglycan (SLRP), was reported to promote angiogenesis not only during wound healing, but also in optical and cutaneous angiogenesis-dependent diseases. However, whether it plays important roles in tumor angiogenesis remains unclear. To explore the role of FMOD in tumor angiogenesis of human small cell lung cancer (SCLC), initially the study analyzed the relationship of FMOD expression in cancer tissues of SCLC with clinical characteristics. The analysis revealed that the positive FMOD expression was significantly associated with extensive stage of SCLC and higher vascular density. In mouse models, xenograft tumors developed with FMOD-silenced H446 cells (H446-shFMOD) exhibited slowed growth rate, decreased microvessel density, and reduced blood perfusion related to that of controls (H446-shCON). Additionally, compared with that of controls, the decreased secretion of FMOD in conditioned medium (CM) from H446-shFMOD inhibited proliferation, migration, and invasion of human umbilical vessel endothelial cells (HUVECs). Moreover, the decreased secretion of FMOD downregulated the expression of VEGF, TGF-β1, FGF-2, and PDGF-B in HUVECs. The findings strongly suggested that the autocrine FMOD of cancer cells may promote tumor angiogenesis of SCLC by upregulating the expression of angiogenic factors that act in concert to facilitate the angiogenic phenotype of endothelial cells as a proangiogenic factor. Therefore, silencing FMOD may be a potentially clinical therapy for repressing tumor angiogenesis.

It has been well established that besides environmental factors, genetic factors are also associated with lung cancer risk. However, to date, the prior identified genetic variants and loci only explain a small fraction of the familial risk of lung cancer. Hence it is vital to investigate the remaining missing heritability to understand the development and process of lung cancer. In the study, to test our hypothesis that the previously identified breast cancer risk-associated genetic polymorphisms at the TOX3/LOC643714 locus might contribute to lung cancer risk, 16 SNPs at the TOX3/LOC643714 locus were evaluated in a Han Chinese population based on a case-control study. Pearson's chi-square test or Fisher's exact test revealed that rs9933638, rs12443621, and rs3104746 were significantly associated with lung cancer risk (P < 0.001, P < 0.001, and P = 0.005, respectively). Logistic regression analyses displayed that lung cancer risk of individuals with rs9933638(GG+GA) were 1.89 times higher than that of rs9933638AA carriers (OR = 1.893, 95% CI = 1.308-2.741, P = 0.001). Similar findings were manifested for rs12443621 (OR = 1.824, 95% CI = 1.272-2.616, P = 0.001, rs12443621(GG+GA) carriers vs. rs12443621AA carriers) and rs3104746 (OR = 1.665, 95% CI = 1.243-2.230, P = 0.001, rs3104746TT carriers vs. rs3104746(TA+AA) carriers). The study discovered for the first time that three SNPs (rs9933638, rs12443621, and rs3104746) at the TOX3/LOC643714 locus contributed to lung cancer risk, providing new evidences that lung cancer and breast cancer are linked at the molecular and genetic level to a certain extent.

难治性气胸是指自发性气胸后患者经胸腔闭式引流或持续负压吸引超过1周仍存在漏气，或复发性气胸（2次以上）［1］。其多发生于老年气胸、肺大泡患者，尤其是合并有慢性阻塞性肺疾病（COPD）的患者［2-3］。当COPD患者合并难治性气胸，常因其心肺功能差而无手术条件，因此胸膜固定术是其常见的治疗方法［4］。作者近年来采用经纤维支气管镜胸腔内肺表面直接喷洒滑石粉混悬液治疗COPD并发难治性气胸患者21例，取得了较好的疗效。

Although aberrant DNA methylation has been implicated in the pathophysiology of lung cancer, the role of methylation in multidrug resistance (MDR) of lung cancer has remained unclear. To investigate whether certain distinct DNA methylation pattern is associated with acquired MDR of lung adenocarcinoma, methylated-DNA immunoprecipitation-chromatin immunoprecipitation (MeDIP-ChIP) was utilised to compare the genome-wide promoter methylation of the human lung adenocarcinoma MDR A549/cisplatin (A549/DDP) cells with its progenitor A549 cells. The comparison identified 3617 genes with differentially methylated promoter, of which 1581 were hypermethylated and 2036 were hypomethylated. Then, bisulphite sequencing polymerase chain reaction (PCR) (BSP) and quantitative reverse transcription (RT)-PCR (Q-PCR) were used to validate the promoter methylation of five candidate genes and to determine whether the expression of genes was associated with the promoter methylation. BSP confirmed that the promoter methylation incidence of the hypermethylated genes, G protein-coupled receptor 56 isoform 3 (GPR56), metallothionein 1G (MT1G), and RAS association domain family gene 1 (RASSF1), was significantly higher in A549/DDP cells compared with A549 cells (p<0.001, p=0.0099, and p=0.0165), whereas no significant difference was found in that of the other two genes, CCNL2 and BAD (p=0.0594 and p=0.5546). Additionally, Q-PCR showed that the mRNA expression of the three hypermethylated genes was significantly lower in A549/DDP cells compared with A549 cells (all p<0.001). In conclusion, this study reported for the first time that a distinct promoter methylation pattern is associated with MDR of lung adenocarcinoma A549/DDP cells and suggested that GPR56, MT1G, and RASSF1 might be the potential methylation markers associated with acquired MDR of lung adenocarcinoma.

The interplay of a complex genetic basis with the environmental factors of chronic obstructive pulmonary disease (COPD) may account for the differences in individual susceptibility to COPD. Mitochondrial DNA (mtDNA) contributes to an individual's ability to resist oxidation, an important determinant that affects COPD susceptibility. To investigate whether mtDNA haplogroups play important roles in COPD susceptibility, the frequencies of mtDNA haplogroups and an 822-bp mtDNA deletion in 671 COPD patients and 724 control individuals from southwestern China were compared. Multivariate logistic regression analysis revealed that, whereas mtDNA haplogroups A and M7 might be associated with an increased risk for COPD (OR=1.996, 95% CI=1.149-2.831, p=0.006, and OR=1.754, 95% CI=1.931-2.552, p=0.021, respectively), haplogroups F, D, and M9 might be associated with a decreased risk for COPD in this population (OR=0.554, 95% CI=0.390-0.787, p=0.001; OR=0.758, 95% CI=0.407-0.965, p=0.002; and OR=0.186, 95% CI=0.039-0.881, p=0.034, respectively). Additionally, the increased frequency of the 822-bp mtDNA deletion in male cigarette-smoking subjects among COPD patients and controls of haplogroup D indicated that haplogroup D might increase an individual's susceptibility to DNA damage from external reactive oxygen species derived from heavy cigarette smoking. We conclude that haplogroups A and M7 might be risk factors for COPD, whereas haplogroups D, F, and M9 might decrease the COPD risk in this Han Chinese population.

目的:探讨支气管肺泡灌洗术治疗社区获得性肺炎的疗效。方法:将60例社区获得性肺炎随机分为治疗组和对照组。每组各30例,对照组给予常规抗感染、祛痰、体位引流、支持治疗,治疗组在对照组治疗的基础上行经纤支镜肺泡治疗术。观察两组患者的临床症状、体征改善情况以及胸部X线下病变吸收情况。结果:治疗组临床症状缓解快、住院时间缩短、无严重并发症发生。结论:支气管肺泡治疗术治疗社区获得性肺炎疗效较好。

Objective To investigate the influence of hypoxia on the methylation level of genomic DNA in human small cell lung cancer H446 cells.Methods H446 cells were cultured under the hypoxic condition(3%O2) for different time periods(0,12,24,48 and 72 h respectively).Those cultured under normal condition served as control.The genomic DNA methylation level was detected by methylation sensitivity random primers PCR(MA-AS-PCR) after DNA extraction.Results Compared with the normoxic control group,the quantity and fluorescence intensity of amplified products of hypoxic experimental group were much higher,especially after 24 h.This indicated that the genomic DNA methylation level in the H446 cells was significantly changed when were treated by hypoxia,and represented mainly as multifocal abnormal methylation.Conclusion Hypoxia might be one of the key causer inducing unbalanced methylation in cancer cells.

<正>临床资料患者男性,64岁。因"反复咳嗽、咳痰16年,活动后气促9年,双下肢水肿2年,再发3 d",于2010年6月10日入院。既往有明确的"慢性阻塞性肺疾病(COPD)、慢性肺源性心脏病、双肺肺大疱、右侧自发性气胸"病史。入院查体:体温36.4℃,脉搏130次/min,呼吸

Objective As a reversible regulation progress,demethylation therapy has become the most promising strategy of tumor prevention and treatment.5-Aza-CdR has showed certainly antitumor activity in several tumors including non-small cell lung cancer,but its effect in small cell lung cancer needs further research.Methods After H446 cells treated with 5-Aza-CdR in vitro,the cell morphological changes was observed with light microscope.the proliferation ability and drug sensitivity were measured with MTT assay.The activity of caspase-3/7,efficacy of drug exclusion and cell cycle was assessed by luminometer and FACs.The genome DNA methylation status was examined by methylation-sensitive arbitrarily primed PCR(MS-AP-PCR).Results H446 cells genome DNA methylation level apparently decreased after 5-Aza-CdR treatment.5-Aza-CdR could inhibit growth of H446 cells in the dose-dependent manner,induce the arrest of H446 cells in G1 phase(from 56.3±2.5 to 73.75±2.47,P0.05),increase the caspase-3/7 activity(from 257510.5±12861.5 to 538585.5±10897.2) and efficacy of drug exclusion(from 11.5±2.3 to 23.8±3.0,P0.05).Conclusion 5-Aza-CdR has a strong antitumor activity to H446 cells in vitro.The low methylation cytotoxic effect including cell growth-inhibit,G1 phase arrest and apoptosis and expression restoration of tumor suppressor gene might play an important role.

Objective To study the correlation between genomic DNA methylation and acquired multidrug-resistance of human small cell lung cancer (SCLC) cell line H446/CDDP. Methods Genomic DNA was extracted from H446/CDDP cells and their parental cells H446. Genomic DNA methylation was detected by methylation-sensitive arbitrarily primed PCR (MS-AP-PCR). Results The amplified bands of H446/CDDP cells were significantly more than those of H446 cells and their fluorescence intensity was higher than that of H446 cells after enzymolysis, indicating that the genomic DNA methylation level is higher in H446/CDDP cells than in H446 cells. Conclusion The acquired multidrug-resistance of H446/CDDP cells is correlated with the high genomic DNA methylation level.

Objective To investigate the apoptosis-inducing effect and its potential mechanisms of docetaxel on human lung adenocarcinoma cells A549 and multidrug resistant cell subline A549/CDDP.Methods Light microscope and TEM were used to observe cell morphological changes.FACs was used to detect the cell cycle arrest effect of docetaxel.TEM and Annexin V/PI staining were employed to measure apoptosis induced by docetaxel.Immuocytochemistry was carried out to confirm the expression of Bcl-2,Bax,Fas and other apoptosis related protein before and after treatment of A549 and A549/CDDP cells with docetaxel.Results After the treatment of docetaxel,vacuoles increased,microvilli decreased,and mitochondria swelled.A few apoptosis bodies and cytoplasm without organelles were observed both in A549/CDDP and A549 cells.Multimicronucleation appeared in A549/CDDP cells.FACs showed that docetaxel induced the arrest of A549 and A549/CDDP cells in G2 phase.Docetaxel could induce apoptosis,necrosis and autoschizis of A549 and A549/CDDP cells,with the latter two dominated.After treatment of docetaxel,Bax upregulated in A549/CDDP cells and Fas noticeably upregulated in both cells.The expression of other apoptosis related proteins had no visible changes.Conclusion Docetaxel could induce the arrest of A549 and A549/CDDP cells in G2 phase and mainly results in cell necrosis and autuschizis;meanwhile,apoptosis also plays a role in it.The upregulation of Fas might be one of the mechanisms of docetaxel.

Objective To predict the B-cell epitope for lipopolysaccharide-binding protein (LBP) antigen Methods Using biotic software and network servers,the secondary structure of LBP was predicted,and the surface properties of LBP,such as hydrophilicity,flexibility,accessibility and antigenicity,were analyzed Then the average antigen index (AI) was calculated by using Wu’s method Results Many distinct antigenic epitopes on LBP were identified: AI of sequence 247-260 (FKGEIFHRNHRSPV) on LBP was 0040 7,significantly higher than that of other sequences AI of sequence 370-379 (LPSSSKEPVF) and sequence 303-325 (ITDDMIPPDSNIRLTTKSFRPFV) was 0033 6 and 0029 5,respectively Sequence 247-260 was the prevalent antigen epitope of B-cell on LBP Conclusion The B-cell epitopes of LBP is predicted successfully

Background and purpose:Regulation of MMR activity under hypoxia may play an important role in genetic instability of cancer,but the mechanism is still unclear.We investigated the expression of DNA mismatch repair genes MLH1 and MSH2 in human SCLC cell line H446 under hypoxic condition and explore the role of promoter methylation of genes in hypoxia.Methods:RT-PCR and Western blot were applied to detect MLH1 and MSH2 expression in human SCLC cell line H446 at the mRNA and the protein level,respectively,under either hypoxic condition or after 5-Aza-CdR treatment.Meanwhile,methylation-specific PCR(MSP)was used to determine promoter methylation of MLH1 and MSH2.Results:The expression of MLH1 and MSH2 in H446 cells significantly decreased both at the mRNA and the protein level under hypoxic condition.5-Aza-CdR treatment led to the restoration of MLH1 and MSH2 expression,while,both MLH1 and MSH2 were down-regulated again after removing 5-Aza-CdR.Conclusions:The promoter methylation of MLH1 and MSH2 may play an important role in its defective expression in H446 cells under hypoxic condition.And 5-Aza-CdR could restore MLH1 and MSH2 expression.

Objective To investigate the effects of perfluorocarbon(PFC) on lipopolysaccharide(LPS)-induced intercellular adhesion molecule-1(ICAM-1) expression in cultured pulmonary microvascular endothelial cells(PMVECs).Methods Rat PMVECs cultured by peripheral lung tissue-sticking method were seeded in Transwell chambers and divided randomly into 4 groups: group C,remaining untreated as blank control;group F,incubated with PFC;group L,stimulated with 100 ng/ml LPS;group LF,treated with 100 ng/ml LPS and PFC.After treatment for 3,8 and 24 h,the cells in each group were harvested respectively.The cell viability of normal rat PMVECs treated by PFC was determined by MTT assay.Then the mRNA expression of ICAM-1 was semi-quantified by reverse transcription-polymerase chain reaction and the level of ICAM-1 protein expression was detected by flow cytometry.Results The cell viability of normal rat PMVECs was not impaired significantly by treatment of PFC.No signifant difference in mRNA and protein expressions of ICAM-1 was found between group F and group C(P0.05).Treatment of cells in group L with 100 ng/ml LPS significantly induced the mRNA and protein expressions of ICAM-1 in a time-dependent manner(P0.05,P0.01).In comparison with those in group L correspondingly,the mRNA and protein expressions of ICAM-1 in group LF were not inhibited significantly at 3 h after treatment(P0.05),but were inhibited at 8 and 24 h after treatment(P0.01,P0.05).Conclusion Perfluorocarbon suppresses the mRNA and protein expressions of ICAM-1 induced by LPS in cultured PMVECs,suggesting a potential mechanism that PFC protects PMVECs from LPS-induced injury via its anti-inflammatory effect.

Objective To investigate the role of methylated DNA mismatch repair gene MLH1 and MSH2 in the acquired multidrug-resistance of human small cell lung cancer cells H446.Methods The reverse transcription polymerase chain reaction(RT-PCR)and Western blot were applied to measure MLH1 and MSH2 mRNA and protein expressions of the multidrug-resistant cells H446/DDP and its parental cells H446.The promoter methylation status of the genes was assessed by methylation-specific PCR(MSP).Results The expressions of MLH1 and MSH2 significantly decreased both in mRNA level and protein level.Promoter methylation of MLH1 was observed in H446/DDP cells but not in H446 cells.Promoter semi-methylation of MSH2 in H446 cells was transformed to methylation in H446/DDP cells.Conclusion The downregulation of DNA mismatch repair gene MLH1 and MSH2 induced by its promoter methylation may play an important role in the acquired multidrug resistance of human small cell lung cancer.

Background and purpose:Recent studies show that apoptosis induced by chemotherapeutic drugs may be involved in the different steps of nuclear metabolism.Whether small cell lung cancer(SCLC) cells apoptosis induced by cisplatin(DDP) associated with the same molecular mechanism is still under investigation.The purpose of our study was to discover the mechanism of human SCLC cell line H446 cell apoptosis induced by DDP.Methods:Apoptosis induced by DDP in human SCLC cell line H446 was detected by cell morphology,flow cytometry analysis and DNA end labeling,respectively.Results:H446 cells were treated with the concentrations of 0.1, 0.5,1.0,2.0 and 5.0 μg/ml DDP for 48 hr,respectively.Apoptotic index were 1.47±0.03,8.28±0.54,11.02±1.05,12.74±1.08,27.16±1.15,respectively and apoptosis was induced in both a concentration dependent manner and gradually in a time dependent pattern from 12 to 72 hr with a final concentration of 2.0 μg/ml;H446 cells were blocked either at G2 by the final concentration of 0.1,0.5,1.0 and 2.0 μg/ml DDP or at S phase with the final concentration of 5.0 μg/ml DDP.Conclusions:Cell apoptosis induced by DDP may play an important role in the mechanism of DDP antitumor effect.

Autoschizis(胞质自切)是近年来文献提到的一种与典型凋亡截然不同的新的细胞死亡方式,主要表现为严重的膜损伤和通过自行切除造成不含细胞器的胞浆进行性丢失。本文拟对其形态学特征、生化改变、信号调控的初步研究进展及其与凋亡的区别进行综述,以利于进一步加深对细胞死亡方式的认识。

目的探讨多西紫杉醇对肺腺癌多药耐药细胞A549/CDDP及其亲代细胞的生长抑制作用及其抗肿瘤作用机制.方法应用MTT比色法和透射电镜等方法,观察A549/CDDP及其亲代细胞在多西紫杉醇作用后,细胞生长曲线、生长抑制率及形态学等方面的变化.结果MTT比色结果表明,多西紫杉醇对A549/CDDP细胞及其亲代细胞具有明显的生长抑制作用,并且在0.1～2 μg/ml剂量范围内存在剂量及时间依赖关系.A549/CDDP细胞在多西紫杉醇作用12～24h较亲代细胞敏感(P＜0.01),在48 h开始表现出其药物耐受性(P＜0.01),在72 h则与亲代细胞间无显著性差异(P＞0.05);透射电镜观察结果发现,A549/CDDP细胞及其亲代细胞的胞质内空泡均明显增多,微绒毛减少甚至消失,线粒体出现肿胀甚至空泡化,同时,还存在少数凋亡小体和脱落的不含细胞器成分的细胞质.结论多西紫杉醇对肺腺癌多药耐药细胞及其亲代细胞均具有明显的生长抑制作用,并可能通过诱导肿瘤细胞坏死、凋亡和胞质自切,从而发挥其细胞毒作用.