MRP1及TopoⅡ的表达与肺癌耐药性的关系

目的探讨肺癌耐药机制及明确耐药基因与化疗药物耐药性的相关性研究。方法应用MTT法对顺铂（Cisplatin）,吉西他滨（Gemcitabine）,长春瑞滨（Vinorelbine）,紫杉醇（Paclitaxel）,畀环磷酰胺（Iphospamide）5种化疗药物针对肺癌细胞悬液进行药敏检测;应用免疫组化进行MRP,（多药耐药相关蛋白1）及TopoII（拓扑异构酶Ⅱ）表达的检测,并将其表达情况与对应的化疗耐药性进行相关分析。结果①MRP．、TopoⅡ在肺癌中总的阳性表达率分别为：72．5％、50．0％。鳞癌和腺癌MRP1、TopoⅡ相比的表达差异无统计学意义（P〉0．05）,但鳞癌或腺癌和小细胞肺癌在MRP1、TopoⅡ表达相比差异有统计学意义（P〈0．05）;②MRP,的表达与顺铂、吉西他滨、长春瑞滨的耐药性均呈显著的正相关（P〈0．05）,其表达与紫杉醇、异环磷酰胺的耐药性差异无统计学意义（P〉0．05）;TopoⅡ的表达与顺铂、吉西他滨、紫杉醇、异环磷酰胺的耐药性均呈显著的正相关（P〈0．05）,其表达与长春瑞滨的耐药性差异无统计学意义（P〉0．05）。结论①MRP1较高表达及TopoⅡ的较低表达共同介导参与了肺癌耐药的机制,且与检测的化疗药物有不同程度的相关性。②应用肿瘤细胞体外培养、进行体外药敏检测,并明确耐药基因的表达情况,对于识别可能的耐药个体,选择合理有效的化疗药物,可能将会最大限度地避免无效化疗和提高化疗疗效。     

积雪草酸通过抑制耐药相关蛋白表达增强U87MG胶质瘤细胞对紫杉醇 的敏感性

目的： 探索积雪草酸（asiatic acid,AA）对紫杉醇（paclitaxel, PTX）耐药性胶质瘤细胞的抑制作用及其可能的作用机 制。 方法： CCK-8实验、实时荧光定量PCR、Western blotting检测AA对成胶质细胞瘤U87MG细胞的增殖、凋亡的影响。浓度递 增法构建PTX耐药性细胞株PR-U87MG,以U87MG细胞为对照,CCK-8实验验证PR-U87MG细胞对PTX的耐药性,实时荧光定 量PCR、Western blotting检测PR-U87MG细胞中MDR1、LRP mRNA及蛋白的表达水平。AA和PTX单独或联合处理PR-U87MG 细胞,CCK-8实验、实时荧光定量PCR、Western blotting检测各组细胞增殖活力及凋亡的变化。 结果： 成功构建PTX耐药性细胞 株PR-U87MG。AA可以剂量依赖方式抑制U87MG细胞和PR-U87MG细胞的增殖活力（P<0.01）,并明显促进其凋亡（P<0.01）。 与AA或PTX单独处理组相比,联合处理组中PARP1的蛋白水平显著减少（P<0.01）,caspase 3的裂解量显著增加（P<0.01）,耐药 相关蛋白P-糖蛋白1 （P-dycoprotein 1, Pgp-1）和LRP表达水平显著减少（P<0.01）。 结论： AA可有效增强U87MG胶质瘤细胞株 对PTX的敏感性,其机制可能与AA抑制具有药物排出功能的耐药蛋白Pgp-1和LRP表达有关。     

Demethylzeylasteral (ZST93) inhibits cell growth and enhances cell chemosensitivity to gemcitabine in human pancreatic cancer cells via apoptotic and autophagic pathways

The overall 5‐year survival rate of patients with human pancreatic cancer remains less than 8% because of its aggressive growth, early metastasis and resistance to conventional chemoradiotherapy. It is essential to develop innovative and effective therapeutic agents to improve its prognosis. Demethylzeylasteral (ZST93) is a novel triterpenoid monomer extracted from the xylem of Tripterygium roots. Our study aimed to assess the effects of ZST93 on cell proliferation and its role in the chemosensitivity to gemcitabine in human pancreatic cancer cells. The effects of ZST93 on cancer cell proliferation, cell cycle distribution, apoptosis and autophagy were evaluated in various human pancreatic cancer cell lines, and the antitumor effects of ZST93 alone and in combination with gemcitabine were identified in a xenograft mouse model. The results showed that ZST93 could inhibit the proliferation of pancreatic cancer cells and arrest cell cycle at G0/G1 phase by regulating the expression of Cyclin D1 and Cyclin A2. Moreover, ZST93 killed pancreatic cancer cells through two different mechanisms: inducing autophagic cell death at low concentrations and apoptotic cell death at high concentrations. Furthermore, ZST93 could enhance the chemosensitivity of pancreatic cancer cells to gemcitabine both in vitro and in vivo through modulation of the cross talk between autophagy and apoptosis. ZST93 is a potential therapeutic agent for developing novel therapeutic strategies in human pancreatic cancer.     

微卫星不稳定的人类结直肠癌细胞系对5-氟尿嘧啶的敏感性

目的：探讨人类结直肠癌细胞系对化疗药物的敏感性。方法：用MTT方法测定LoVo,SW480和SW1116细胞对5－氟尿嘧啶（5－FU）的敏感性；用免疫细胞化学方法检测3细胞系hMSH2、hMLH1表达水平；用PCR－SSLP－银染法分别分析了3个细胞系基因组中10个微卫星位点状况。结果：比较3个细胞系MTT分析获得的IC50结果发现，对5－FU LoVo细胞系较SW480和SW1116更加敏感（分别为：0．8μmol/L，2.2μmol/L和1．9μmol/L，P＜0．01）。免疫细胞化学检测结果显示hMSH 2在SW480和SW1116阳性，而在LoVo阳性；hMLH1在3个细胞系均阳性。PCR－SSLP－银染法结果显示LoVo细胞系在8／10个位点和其余两个细胞系不同，电泳条带有不同程度的增加或位移，而另外的2／10个位点则具有相同的电泳带型。结论：该组微卫星位点和错配修复状况一起可以作为离体人类结直肠癌细胞系对5－FU敏感性的指标之一，为了深入阐明它们能否作为临床结直肠癌化疗病人药物筛选和预后评价的有效指标，进一步的在体研究和临床资料总结分析很有意义。     

Hypoxic ventilatory response is correlated with increased submaximal exercise ventilation after live high, train low

This study tested the hypothesis that live high, train low (LHTL) would increase submaximal exercise ventilation (_E) in normoxia, and the increase would be related to enhanced hypoxic ventilatory response (HVR). Thirty-three cyclists/triathletes were divided into three groups: 20 consecutive nights of hypoxia (LHTLc, n=12), 20 nights of intermittent hypoxia (4×5-night blocks of hypoxia interspersed by two nights of normoxia, LHTLi, n=10), or control (CON, n=11). LHTLc and LHTLi slept 8–10 h per night in normobaric hypoxia (2,650 m), and CON slept under ambient conditions (600 m). Resting, isocapnic HVR (_E/blood oxygen saturation) was measured in normoxia before (PRE) and after 15 nights (N15) hypoxia. Submaximal cycle ergometry was conducted PRE and after 4, 10, and 19 nights of hypoxia (N4, N10, and N19 respectively). Mean submaximal exercise _E was increased (P<0.05) from PRE to N4 in LHTLc [74.4 (5.1) vs 80.0 (8.4) l min^–1; mean (SD)] and in LHTLi [69.0 (7.5) vs 76.9 (7.3) l min^–1] and remained elevated in both groups thereafter, with no changes observed in CON at any time. Prior to LHTL, submaximal _E was not correlated with HVR, but this relationship was significant at N4 (r=0.49, P=0.03) and N19 (r=0.77, P<0.0001). Additionally, the increases in submaximal _E and HVR from PRE to N15–N19 were correlated (r=0.51, P=0.02) for the pooled data of LHTLc and LHTLi. These results suggest that enhanced hypoxic chemosensitivity contributes to increased exercise _E in normoxia following LHTL.     

耐碳青霉烯类弗劳地枸橼酸杆菌耐药机制及治疗策略研究

目的 探索耐碳青霉烯类弗劳地枸橼酸杆菌耐药机制及治疗对策.方法 收集来自该院的17株耐碳青霉烯类弗劳地枸橼酸杆菌临床资料,采用PCR方法扩增碳青霉烯类耐药基因;采用琼脂稀释法和肉汤稀释棋盘法测磷霉素单药及联合用药最低抑菌浓度(MIC)值,计算部分抑菌浓度指数(ΣFICI).结果 8株菌产blaNDM-1,9株菌产blaIMP,blaKPC、blaoxA-48、blasPM均未检出;磷霉素联合亚胺培南协同作用和相加作用占75.oo％,其中协同作用高达56.25％,磷霉素联合头孢哌酮/舒巴坦钠协同作用和相加作用占50.oo％.结论 磷霉素联合亚胺培南或头孢哌酮/舒巴坦钠体外有较好的抗菌活性,亚胺培南联合磷霉素效果可能更好.     

DcR3基因对胰腺癌裸鼠移植瘤化疗敏感性的影响

目的探讨RNA 干扰沉默诱骗受体-3（DcR3）对人胰腺癌细胞裸鼠移植瘤化疗敏感性的影响及其可能机制。方法取对数生长期的AsPC-1 细胞1×106个/ml,接种于5 周龄裸鼠右后肢腹股沟,当皮下肿瘤直径为8 mm 左右,随机分为4 组（ n=10）：对照组、化疗组、阴性质粒+ 化疗组、DcR3 siRNA+ 化疗组。观察DcR3 siRNA联合化疗药物对人胰腺癌AsPC-1细胞裸鼠移植瘤的治疗效果。应用ELISA和Western blot检测DcR3 蛋白的变化;TUNEL 分析肿瘤细胞凋亡;Western blot 和RT-PCR 检测FasL、Caspase-8 和Caspase-3 等凋亡因子的表达。结果化疗组、阴性质粒+ 化疗及DcR3 siRNA+ 化疗组均可抑制移植瘤的生长,与对照组比较,3 组抑瘤率平均值分别为（35.87±4.58）%、（40.68±4.16）%和（90.25±2.53）%,4 组间差异有统计学意义（F =47.736,P =0.000）,DcR3 siRNA+ 化疗组抑瘤率较化疗组增加;对照组、化疗组、阴性质粒+ 化疗以及DcR3 siRNA+ 化疗组移植瘤重量平均值分别为（0.95±0.03）、（0.63±0.04）、（0.67±0.02）和（0.17±0.06）g,4 组间差异有统计学意义（F =85.531,P =0.000）,DcR3 siRNA+ 化疗组瘤重较化疗组减轻;对照组、化疗组、阴性质粒+化疗组、DcR3 siRNA+化疗组凋亡率平均值分别为（6.3±2.21）%、（14.8±2.65）%、（14.5±3.06）%和（54.6±3.23）%,4 组间差异有统计学意义（F =104.225,P =0.000）,DcR3 siRNA+ 化疗组凋亡率较化疗组增加;DcR3siRNA+化疗组的FasL、Caspase-8 和Caspase-3 蛋白表达较其他各组上升（P =0.000）。结论沉默DcR3 可激活FasL/Caspase 凋亡途径,促进细胞凋亡,增加胰腺癌细胞移植瘤对化疗的敏感性。