腺病毒介导的RNA干扰ERCC1基因表达对逆转卵巢癌顺铂耐药的效果

 目的 探讨腺病毒介导的RNA干扰ERCC1基因表达逆转卵巢癌顺铂耐药的效果。方法 采用直接感染法、台盼蓝活细胞计数、MTT、Hoechst染色法及流式细胞仪检测等方法,绘制顺铂耐药卵巢癌细胞株SKOV3/DDP细胞生长曲线及倍增时间,检测SKOV3/DDP细胞感染重组病毒前后其顺铂的IC₅₀值、细胞凋亡形态及细胞各周期的分布。结果 (1)顺铂浓度与SKOV3/DDP细胞抑制率呈线性正相关,相关系数r=0.9862(P<0.05),顺铂IC₅₀为(13.93±0.37)μg/ml,耐药指数为1.79。(2)重组腺病毒感染后干扰ERCC1基因表达,SKOV3/DDP细胞对顺铂的敏感性分别增加了11.1%、16.7%、26.4%、33.5%,呈剂量依懒性(r=0.9716,P<0.05),耐药指数降至1.19。(3)重组腺病毒联合顺铂作用后,SKOV3/DDP细胞株G₁期细胞比例增大,S期细胞比例增大,细胞凋亡率增高(P<0.05)。结论 干扰ERCC1基因表达可以通过诱导耐药肿瘤细胞SKOV3/DDP凋亡、调节细胞周期分布等途径逆转卵巢癌细胞的顺铂耐药。     

EZH2蛋白在非小细胞肺癌顺铂耐药中的作用及其机制研究

目的探讨Zeste基因增强子同源物2(enhancer of zeste homolog 2,EZH2)蛋白在非小细胞肺癌对顺铂耐药中的作用及其调控机制。方法采用实时荧光定量PCR（real-time quantitative PCR,qRT-PCR）检测人肺腺癌细胞株A549及其顺铂耐药株A549/DDP中EZH2、P21、Puma和Bad的mRNA表达差异。采用RNA干扰技术抑制细胞中的EZH2蛋白表达。通过细胞毒性实验检测细胞耐药性的改变。qRT-PCR检测抑制EZH2蛋白表达后A549/DDP细胞中P21、Puma和Bad的表达变化。流式细胞仪检测细胞周期的分布和变化。结果EZH2在A549/DDP中呈高表达。抑制EZH2蛋白表达后,顺铂对A549/DDP的IC50由（34.57±3.70）μmol/L降低至（18.91±2.07）μmol/L（P<0.05）;抑制EZH2蛋白表达后,A549/DDP中P21、Puma和Bad的mRNA表达均上调;抑制A549/DDP细胞EZH2蛋白表达后,G0/G1期细胞比例下降,G2/M期细胞增多(P<0.05),细胞发生G2/M期阻滞。结论EZH2蛋白参与介导了A549/DDP对顺铂的耐药;EZH2蛋白可能通过调控与细胞凋亡相关基因相关的凋亡通路诱导A549/DDP对顺铂的耐药;EZH2蛋白可能通过抑制P21表达、促进细胞周期进展及肿瘤增殖能力,诱导非小细胞肺癌对顺铂产生耐药。     

黄荆子乙酸乙酯提取物增强顺铂诱导卵巢癌细胞株COC1凋亡的作用

目的观察黄荆子乙酸乙酯提取物（the acetoacetate extract of Vitexnegundoseed,Evn-50）和顺铂（cisplatin,DDP）在体外对人卵巢癌细胞株COC1凋亡的影响,探讨Evn-50提高COC1细胞株对顺铂化疗敏感性的机制。方法Evn-50,DDP以及两药联合分别处理人卵巢癌COC1细胞株,台盼蓝染色计数法检测药物对细胞生长的抑制率,并用流式细胞术、AO/CB荧光双染色法测Evn-50,DDP以及两药联合对细胞凋亡的影响。结果细胞抑制率在联用组中随Evn-50剂量的增加而增加,且明显高于同剂量单用组（P〈0.01）;细胞凋亡率在联用组中随Evn-50剂量的增加而增加,且高于同剂量单用组（P〈0.01）,流式细胞仪分析结果显示Evn-50呈浓度依赖性将COC1细胞阻滞在G2/M期,而DDP将COC1细胞阻滞在G1/S期。结论Evn-50有增强顺铂诱导卵巢癌细胞株COC1凋亡的作用,其机制可能与二者对卵巢癌细胞株COC1周期阻滞点不同有关。     

顺铂诱导人小细胞肺癌多药耐药细胞系NCI-H446/CDDP的建立及其生物学特征分析

目的　建立人小细胞肺癌多药耐药细胞系 ,鉴定其细胞生物学特性。方法　以顺铂为诱导药物 ,人小细胞肺癌细胞系NCI H4 4 6为诱导对象 ,采用首次大剂量冲击和逐步增加剂量结合的方式 ,建立人小细胞肺癌多药耐药细胞系NCI H4 4 6 /CDDP。细胞计数法绘制生长曲线 ,计算其倍增时间 ;MTT法测定细胞IC50 ,计算其耐药指数 ;光学显微镜、扫描电镜和透射电镜观察其形态。结果　经过 8个月的诱导 ,成功建立了人小细胞肺癌多药耐药细胞系NCI H4 4 6 /CDDP ;生长曲线测定结果表明 ,亲代和耐药细胞增殖速度接近 ,其倍增时间分别为 4 0 72h和 4 1 80h;流式细胞仪检测细胞周期发现 ,耐药细胞系中S期细胞所占比例为 2 0 2 4 % ,亲代细胞系中S期细胞所占比例为18 4 2 % ,二者比较差异有统计学意义 (P <0 0 5 ) ,而G0 /G1和G2 /M期细胞无显著性变化 ;MTT结果表明 ,耐药细胞系对顺铂、羟基喜树碱、长春新碱、5 氟尿嘧啶和拓扑替康等 5种化疗药物有不同程度的耐受性 ,其耐药指数分别为 85 0 0、4 2 75、6 0 2 8、2 4 31、12 4 7;细胞超微结构显示 ,耐药细胞较其亲代体积增大 ,核浆比例降低 ,线粒体、高尔基体、粗面内质网和游离核糖体增多 ,细胞表面指状突起增多。结论　NCI H4 4 6 /CDDP细胞生物学性状稳定 ,是一     

人卵巢癌拓扑替康耐药细胞株SKOV3/TPT的建立及生物学特性的研究

目的建立人卵巢癌拓扑替康（TPT）耐药细胞株（SKOV3／TPT）,研究其生物学特性和耐药特点。方法模拟临床用药特点,采用大剂量TPT（2160ng／ml）冲击建成人卵巢癌SKOV3／TPT耐药细胞株。MTT法检测多药耐药性,流式细胞仪测定细胞周期、胞内药物的积聚及P糖蛋白（Pgp）表达,从生长曲线、光镜下形态、超微结构等方面比较耐药细胞和SKOV3细胞某些生物学特性的差异。结果成功建立了SKOV3/TPT耐药株,耐药指数为10．67,对喜树碱、米托蒽醌及长春新碱有明显的交叉耐药,对紫杉醇、顺铂、5氟尿嘧啶、足叶乙甙和甲氨蝶呤敏感。与SKOV3细胞相比,SKOV3/TPT耐药细胞倍增时间延长;G2＋M期的比例明显增加（P〈0．01）;细胞内TPT浓度明显减少（P〈0．05）;无Pgp过表达。结论SKOV3/TPT细胞株呈中等水平耐药,耐药性稳定,呈非典型的多药耐药特征;对TPT耐药的主要原因可能与细胞内药物浓度降低和细胞周期改变及凋亡有关,与Pgp的过表达无关。     

MRP在人涎腺腺样囊性癌细胞中的表达及意义

目的:通过检测MRP蛋白的表达,探讨人涎腺腺样囊性癌细胞产生多药耐药的机制。方法:用浓度为1μg/ml的顺铂作用于腺样囊性癌细胞,每次作用时间48 h,经6个月后形成在该药物浓度下生长良好的耐顺铂细胞。运用免疫组化、Western-blot法检测SACC和SACC/DDP细胞株中MRP蛋白的表达,RT-PCR检测SACC和SACC/DDP的MRP mRNA的表达。结果:诱导形成的顺铂作用环境下生长稳定的细胞株SACC/DDP,其MRP蛋白和MRP mRNA表达水平明显高于母本细胞SACC(P<0.01)。结论:SACC/DDP细胞的细胞浆中以及细胞膜上MRP蛋白的表达率很高,可能是涎腺腺样囊性癌细胞产生多药耐药的机制所在。     

人肝癌多药耐药细胞系 Bel-7402／DOX 两种模型比较

目的：比较体外浓度梯度递增诱导和裸鼠肝脏移植诱导两种方法建立人肝癌多药耐药细胞系Bel-7402/多柔比星（ Doxorubicin ,DOX）模型的生物学特性。方法分别采用体外浓度梯度递增诱导和裸鼠肝脏移植诱导建立人肝癌多柔比星多药耐药细胞亚系Bel-7402/DOXV 和Bel-7402/DOXL 后,利用相差显微镜观察细胞,MTT法检测两种细胞的耐药性,细胞计数法绘制生长曲线并用公式法计算倍增时间,流式细胞仪测定细胞DOX的摄入和外排以及P糖蛋白（ P-gp）、多药耐药相关蛋白（MRP）、谷胱甘肽硫转酶系统（GSH/GST）的表达。结果两组耐药细胞Bel-7402/DOXV 和Bel-7402/DOXL 对DOX、CD-DP均产生了交叉耐药性,较亲本Bel-7402 IC50值差异有统计学意义（P＜0．01）;较亲本细胞的倍增时间明显延长,分别为65 h和46 h;较亲本的DOX外排率明显升高,分别为81．06％、66．56％;两组耐药细胞P-gp、MRP表达较亲本细胞显著提高（ P＜0．01）,而GSH/GST的表达无明显变化。结论两种方法建立的耐药细胞系模型均有稳定的耐药性。肝脏移植法更能高度模拟人肝癌,它是具有近似人肝癌生物学和抗癌药物动力学特征的较理想模型。     

辐射诱导人鼻咽癌细胞CNE1、CNE2多药耐药的研究

目的研究辐射诱导及逆转药物环孢霉素A（CsA）、干扰素a（IFNa）对鼻咽癌细胞CNE1、CNE2多药耐药表达的影响。方法通过体外辐射处理诱导鼻咽癌细胞CNEl、CNE2，流式细胞术、RT-PCR分析辐射诱导前后P-gp、MRP、TOPH、GST-π在蛋白、mRNA水平表达的改变。同时，研究CsA、IFN-α逆转耐药处理对放疗诱导的多药耐药的影响。结果辐射诱导后CNE1、CNE2细胞P-gp、MRP、GST-π在蛋白、mRNA水平表达上调，辐射后给予CsA、IFN-处理可以逆转辐射诱导的P-gp、MRP蛋白表达上调。结论辐射可以诱导鼻咽癌细胞CNE1、CNE2出现多药耐药，而CsA、IFN-可以逆转辐射诱导的多药耐药。     

葡萄籽原花青素联合顺铂对喉癌细胞Hep-2生物学作用及机制研究

目的利用葡萄籽原花青素及其联合顺铂在体外作用于喉癌细胞Hep-2,观察其对喉癌细胞增殖、凋亡的影响,并探讨抗肿瘤作用的可能机制。方法 MTT法检测各组药物对喉癌细胞增殖的抑制作用;流式细胞仪检测细胞周期;TUNEL法检测各组药物对喉癌细胞凋亡率的影响;采用流式细胞仪分析凋亡细胞比例,Western blot法检测各组药物对喉癌细胞Bax/Bcl-2表达情况的影响。结果 MTT和TUNEL结果显示,GSPE+顺铂组的喉癌细胞抑制增殖作用明显高于单纯GSPE组和单纯顺铂组,差异均有统计学意义(P<0.05);细胞周期结果显示,GSPE+顺铂组处于S+G2M期的细胞数明显低于单纯GSPE和单纯顺铂组,差异均有统计学意义(P<0.05);蛋白表达结果显示,GSPE+顺铂组的Bax/Bcl-2的表达明显高于单纯GSPE组和单纯顺铂组,差异均有统计学意义(P<0.05)。结论葡萄籽原花青素可诱导喉癌Hep-2细胞凋亡;与单纯应用葡萄籽原花青素或顺铂比较,葡萄籽原花青素联合顺铂可以起到更好的增殖抑制并诱导喉癌细胞凋亡,两者具有协同作用。     

L1210／DDP细胞交叉耐药性及耐药机制的初步研究

目的观察小鼠淋巴细胞白血病顺铂耐药细胞亚系（L1210／DDP）与其它抗癌药物的交叉耐药性及对耐药机制进行初步研究。方法用拒染法测定细胞对不同抗癌药物的敏感性，用无火焰原子吸收光谱仪、谷胱甘肽还原酶循环法和微孔滤膜碱洗脱技术分别测定细胞内铂离子浓度、总谷胱甘肽含量、谷胱甘肽s-转移酶活性以及细胞DNA链间交联效应。结果L1210／DDP对卡铂有明显的交叉耐药性，对丝裂霉素、噻替派具有部分交叉耐药性，对阿霉素和5-氟尿嘧啶等无交叉耐药性。L1210／DDP细胞内的铂离子浓度和DNA链间交联指数明显低于L1210细胞，总谷胱甘肽含量、谷胱甘肽s-转移酶活性明显高于L1210细胞。结论L1210／DDP细胞亚系对DDP的结构类似物卡铂具有明显的交叉耐药性，其耐药机制与降低细胞内铂离子浓度、升高总谷胱甘肽含量和谷胱甘肽s-转移酶活性、降低DNA链间交联效应有关。     

Knee injury and Osteoarthritis Outcome Score (KOOS) - validation of a Swedish version

The Knee injury and Osteoarthritis Outcome Score (KOOS) is a self-administered instrument measuring outcome after knee injury at impairment, disability, and handicap level in five subscales. Reliability, validity, and responsiveness of a Swedish version was assessed in 142 patients who underwent arthroscopy because of injury to the menisci, anterior cruciate ligament, or cartilage of the knee. The clinimetric properties were found to be good and comparable to the American version of the KOOS. Comparison to the Short Form-36 and the Lysholm knee scoring scale revealed expected correlations and construct validity. Item by item, symptoms and functional limitations were compared between diagnostic groups. High responsiveness was found three months after arthroscopic partial meniscectomy for all subscales but Activities of Daily Living.     

Endovascular management of carotid-cavernous fistulas

Objective To investigate endovascular treatment of traumatic direct carotid-cavernous fistulas （CCF） and their complications such as pseudoaneurysms. Methods： Over a five-year period, 22 patients with traumatic direct CCFs were treated endovascularly in our institution. Thirteen patients were treated once with the result of CCF occluded, 8 twice and 1 three times. Treatment modalities included balloon occlusion of the CCF, sacrifice of the ipsilateral internal carotid artery with detachable balloon, coll embolization of the cavernous sinus and secondary pseudoaneurysms, and covered-stem management of the pseudoaneurysms. Results All the direct CCFs were successfully managed endovascularly. Four patients developed a pseudoaneurysm after the occlusion of the CCF with an incidence of pseudoaneurysm formation of 18.2% （4/22）. A total number of 8 patients experienced permanent occlusion of the ICA with a rate of ICA occlusion reaching 36.4% （8/22）. Followed up through telephone consultation from 6 months to 5 years, all did well with no recurrence of CCF symptoms and signs. Conclusion Traumatic direct CCFs can be successfully managed with endovascular means. The pseudoaneurysms secondary to the occlusion of the CCFs can be occluded with stent-assisted coiling and implantation of covered stents.     

The role of interventional radiology in trauma

Introduction Interventional Radiology has evolved into a specialty having enormous input into the care of the traumatized patient.In all hospitals,regardless of size,the Interventional Radiologist must consider their relationships with the trauma service in order to     

The acutely limping child

Acute limping may be the result of multiple pathologies in children. The differential diagnosis varies based on the age of the child. Irrespective of age, the initial imaging work-up includes AP and frog leg radiographs of the pelvis and ultrasound; MRI may sometimes be helpful. In children less than 3 years, infections and trauma are most frequent. MRI is the imaging modality of choice when osteomyelitis is clinically suspected. Between the ages of 3 and 10 years, transient synovitis of the hip and Legg-Calvé-Perthes disease are main considerations but infection, inflammation and focal bony lesions are also considered. In children over 10 years, slipped capital femoral epiphysis also is considered.     

Diagnóstico ecográfico del neumotórax

The ultrasonographic diagnosis of pneumothorax is based on the analysis of artifacts. It is possible to confirm or rule out pneumothorax by combining the following signs: lung sliding, the A and B lines, and the lung point. One fundamental advantage of lung ultrasonography is its easy access in any critical situation, especially in patients in the intensive care unit. For this reason, chest ultrasonography can be used as an alternative to plain-film X-rays and computed tomography in critical patients and in patients with normal plain films in whom pneumothorax is strongly suspected, as well as to evaluate the extent of the pneumothorax and monitor its evolution.     

Carbohydrate Supplementation during Exercise: Does It Help? How Much is Too Much?

KEY POINTS· Carbohydrate intake during exercise can delay the onset of fatigue and improve performance of prolonged exercise as well as exercise of shorter duration and greater intensity (e.g., continuous exercise lasting about 1h and intermittent high-intensity exercise), but the mechanisms by which performance is improved are different.