可溶性支架转染c-myc基因反义寡核苷酸对静脉移植物c-myc及PCNA表达的影响

目的采用可溶性支架将原癌基因c—myc反义寡核苷酸转染静脉移植物，观察其对静脉移植物内c—myc蛋白和增殖细胞核抗原（PCNA）蛋白产物表达的影响。方法50只新西兰大耳白兔建立兔颈外静脉颈总动脉旁路移植模型后随机分成5组，每组10只。对照组：无支架；组1：植入单纯可溶性支架；组2：植入正义c—myc寡核苷酸可溶性支架；组3：植入反义c—myc寡核苷酸可溶性支架；组4：植入不匹配c—myc寡核苷酸可溶性支架。于静脉移植后7d、28d和90d取出静脉移植物，采用免疫组织化学方法检测其c—myc蛋白和PCNA蛋白的表达。结果对照组、组1、组2、组4静脉移植术后7d、28d、90d时静脉移植物内膜和中膜内可见大量c—myc蛋白和PCNA蛋白表达阳性的细胞，与同时间点组3比较差别均有统计学意义（P〈0．01）。28d、90d时5组静脉移植物内c—myc蛋白的表达与同组7d时比较明显增强（P〈0．01）。结论可溶性支架转染c—myc反义寡核苷酸可显著抑制静脉移植物内c—myc蛋白和PCNA蛋白的表达。     

反义寡核苷酸对血管平滑肌细胞c-myc、增殖细胞核抗原蛋白表达的影响

目的　研究反义c myc、增殖细胞核抗原 (PCNA)基因片断对血管平滑肌细胞 (VSMCs)增殖的作用。　方法　使正义、反义及错配反义c myc、PCNA ,分别作用于体外培养的VSMCs ,应用蛋白印迹和免疫组化法测定并经计算机图像分析 ,观察其对c myc、PCNA蛋白表达的影响。　结果　 10μmol/L浓度的反义c myc、PCNA作用于VSMCs后 1～ 5d ,相应c myc和PCNA蛋白表达减弱 ,计算机图像分析表明其表达产物与对照组相比 ,差异有显著意义 (P <0 0 5 ) ,而正义和错配反义寡核苷酸无此效应 (P >0 0 5 )。　结论　反义c myc、PCNA寡核苷酸可通过阻遏其相应基因表达进而抑制蛋白表达及VSMCs增殖。     

小干扰RNA抑制人端粒酶逆转录酶基因对胃癌细胞增殖的影响

目的 应用小干扰RNA(siRNA)技术特异性抑制人端粒酶逆转录酶(hTERT)基因在胃癌SGC7901细胞中的表达,观察其对细胞增殖的影响.方法 hTERT基因的RNA干扰表达重组体用脂质体介导方法转染SGC7901细胞.荧光定量聚合酶链反应(PCR)检测hTERT基因表达.TRAP-PCR法检测端粒酶活性、噻唑蓝(MTT)比色法检测细胞增殖.结果 SGC7901细胞转染前后hTERT基因的表达强度分别为:转染pU6组平均表达量为6.5×105拷贝/μg RNA;转染pU6-hTERT-siRNA Ⅰ组平均表达量为4.1 × 104拷贝/μg RNA;转染pU6-hTERT-siRNAⅡ组平均表达量为5.4×104拷贝/μg RNA.转染pU6组与其余两组P值<0.01;转染pU6-hTERT-siRNA Ⅰ组和转染pU6-hTERT-siRNAⅡ组P值>0.05.端粒酶活性下降,细胞生长速度明显减慢.结论 小干扰RNA能有效抑制SGC7901细胞中hTERT基因表达,hTERT基因表达下调影响SGC7901细胞增殖.     

P53蛋白对人瘢痕疙瘩成纤维细胞端粒酶活性的影响

目的:探讨P53蛋白(P53protein)对人瘢痕疙瘩成纤维细胞(Keloidfibroblast/KFB)端粒酶活性的影响,明确在人瘢痕疙瘩成纤维细胞中P53蛋白与端粒酶活性之间的相互关系。方法:采用腺病毒介导法将野生型P53基因(Ad-P53)转染至人瘢痕疙瘩成纤维细胞中;将来源于瘢痕疙瘩组织的成纤维细胞随机分成二组。实验组转染野生型P53基因至成纤维细胞中。非转染组的成纤维细胞未进行野生型P53基因转染。分别用间接免疫荧光法和Westernblot法检测成纤维细胞中P53蛋白的表达;TRAP-ELISA法检测成纤维细胞中端粒酶活性。结果:转染组细胞中P53蛋白表达明显高于非转染组细胞;而端粒酶活性明显低于非转染组细胞(P<0.01)。结论:P53蛋白能够抑制人瘢痕疙瘩成纤维细胞端粒酶活性。     

前列腺癌细胞凋亡与iNOS、Bcl-2的表达及意义

目的 :探讨细胞凋亡和诱生型一氧化氮合酶 (iNOS)、Bcl 2蛋白表达在前列腺癌中的意义以及两者之间的关系。　方法 :对 2 4例前列腺癌、15例前列腺增生及 5例正常前列腺组织行原位细胞凋亡检测及iNOS、Bcl 2蛋白免疫组化检测。　结果 :前列腺癌组的细胞凋亡率和iNOS的染色强度均显著高于前列腺增生组及正常前列腺组(P <0 .0 1) ,且Bcl 2蛋白表达阳性者的细胞凋亡率显著低于阴性者 (P <0 .0 1)。　结论 :前列腺癌的细胞凋亡率可反映其恶性程度 ;iNOS的表达与前列腺癌的分化程度无关 ,但与Bcl 2蛋白表达呈负相关 ;Bcl 2蛋白表达与前列腺癌细胞凋亡呈负相关。iNOS与Bcl 2均可通过影响前列腺癌细胞凋亡而在前列腺癌病理发生发展中起作用     

端粒酶反义RNA基因转染抑制裸鼠人膀胱癌移植瘤生长的研究

目的　观察端粒酶反义RNA基因转染对裸鼠膀胱癌移植瘤细胞生长的影响。方法将脂质体介导的端粒酶反义RNA真核表达载体 (pBBS hTR)、空载体 ( pBBS 2 12 )及生理盐水注入移植瘤体内 (每天 3 0 0 μl,共 7d) ,应用端粒酶活性聚合酶链反应 酶联免疫吸附试验 (PCR ELISA)、苏木素 伊红 (HE)染色、透射电镜等连续观察 6周移植瘤组织细胞端粒酶活性和生长变化。结果　注射转染pBBS hTR组和空载体组瘤体积抑制率分别为 48.7%和 2 .8% (P <0 .0 5 )。转染pBBS hTR组端粒酶活性降低 ,细胞生长受抑制。电镜观察见典型凋亡特征。 结论　端粒酶反义RNA基因瘤体内注射转染有效地抑制裸鼠人膀胱癌细胞生长 ,具有潜在临床应用价值。     

反义增殖细胞核抗原cDNA对人膀胱癌裸鼠异体移植瘤生长的抑制作用

目的　探讨增殖细胞核抗原 (PCNA)反义cDNA对人膀胱癌裸鼠移植瘤模型生长的影响。　方法　脂质体介导反义PCNA真核表达载体转染人膀胱癌EJ细胞后 ,接种到裸鼠皮下组织 ,观察体内致瘤及生长情况 ;免疫组化检测瘤体PCNA蛋白表达 ,RT PCR分析瘤体PCNA、wt p5 3、c myc基因mRNA水平 ;并进行瘤体病理学评估。 　结果　反义PCNAcDNA导入后 ,EJ细胞体内致瘤活性降低 ,同对照组比较瘤体体积缩小 5 4 .2 3% (P <0 .0 5 ) ,重量减轻 4 2 .5 4 % (P <0 .0 1) ,瘤组织PCNA蛋白及mRNA水平分别减少 6 1.6 2 % (P <0 .0 1)和 72 .13% (P <0 .0 1) ,c myc基因表达下调4 7.34% (P <0 .0 1) ,wt p5 3表达活性无显著性改变 (P >0 .0 5 ) ,瘤体病理学特征改善。 　结论　转导反义PCNAcDNA能有效逆转肿瘤的恶性表型 ,是膀胱癌基因治疗的合理策略之一     

异氟醚对转染APPsw基因SH-SY5Y细胞凋亡的影响和IP3受体在其中的作用

目的 评价异氟醚对转染APPsw基因SH-SY5Y细胞凋亡的影响和1,4,5-三磷酸肌醇(IP3)受体在其中的作用.方法 将转染APPsw基因的SH-SY5Y细胞以1.2×104个/cm2密度接种于培养瓶中,采用随机数字表法,将其分为4组(n=6):对照组(C组)、IP3受体拮抗剂组(Ⅹ组)、异氟醚组(Ⅰ组)和异氟醚+ IP3受体拮抗剂组(Ⅰ+Ⅹ组).继续培养24h细胞贴壁后,C组常规培养;Ⅹ组和Ⅰ+Ⅹ组加入IP3受体拮抗剂Xestospongin C 100 nmol/L; 30 min后Ⅰ组和Ⅰ+Ⅹ组给予1.2％异氟醚处理8h.收集细胞,电镜下观察超微结构,采用流式细胞术检测细胞凋亡率和胞浆内游离钙离子浓度([Ca2+]i),采用Western blot法测定IP3受体蛋白表达.结果 与C组比较,Ⅹ组细胞凋亡率、[Ca2+]i和IP3受体蛋白表达差异无统计学意义(P＞0.05),Ⅰ组和Ⅰ+Ⅹ组细胞凋亡率增加,[Ca2+]i升高,IP3受体蛋白表达上调(P ＜0.05或0.01);与Ⅰ组比较,Ⅰ+Ⅹ组细胞凋亡率和[Ca2+]i降低,IP3受体蛋白表达下调(P＜0.01).Ⅰ组和Ⅰ+Ⅹ组细胞出现病理学损伤,Ⅰ组损伤重于Ⅰ+Ⅹ组.结论 异氟醚可通过升高胞浆内游离Ca2+浓度、上调IP3受体蛋白表达,引起转染APPsw基因的SH-SY5Y细胞凋亡.     

雌激素对前列腺增生症移行区细胞Bcl-2、Bax和c-myc基因表达的影响

目的　探讨大剂量外源性雌激素对良性前列腺增生症 (BPH )移行区 (TZ)细胞Bcl 2、Bax和c myc基因表达的影响 ,籍此了解雌激素对BPH的治疗机制。 方法　对 3 0例经服用已烯雌酚的BPH患者 (处理组 )和 3 0例空白对照患者 (对照组 )的标本 ,采用免疫流式细胞光度术检测上述 3种基因在TZ的标记率和表达量。结果　雌激素处理后 ,TZ细胞Bcl 2的标记率和表达量分别由对照组的 2 5 .5 0 %和 73 .0 0降低为 13 .3 5 %和 44 .46(P <0 .0 1) ,Bax的表达量则由对照组的48.0 0升高为 5 6.97(P <0 .0 1)。结论　大剂量雌激素对BPH治疗作用可能与其抑制TZ细胞Bcl 2的表达和促进Bax的表达进而提高细胞凋亡率有关。     

凋亡抑制基因Survivin在非小细胞肺癌中的表达及其与Bcl-2、Bax蛋白表达的关系

目的　探讨Survivin基因的表达在肺癌发生、发展中的作用及其与Bax、Bcl 2蛋白表达的相互关系。方法　对 12例正常支气管粘膜上皮、9例不典型增生、3 4例肺癌及 12例淋巴结转移癌石蜡切片组织 ,应用原位分子杂交法检测SurvivinmRNA的表达 ;用免疫组织化学链霉菌抗生物素蛋白 过氧化酶法 (S P法 )检测Bcl 2、Bax蛋白的表达。结果　肺癌、淋巴结转移癌中SurvivinmRNA阳性率分别为 61.76%及 91.67% ,显著高于正常支气管粘膜上皮及不典型增生中的阳性率16.7%及 3 3 .3 % (P均 <0 .0 5 ) ;低分化、中分化肺癌中阳性率 88.89%、75 .0 0 %较高分化 11.11%明显升高 (P均 <0 .0 1)。TNM分期Ⅲ期病例中SurvivinmRNA表达阳性率 93 .3 %高于Ⅰ Ⅱ期病例的 3 6.8% (P <0 .0 1)。SurvivinmRNA表达与Bcl 2蛋白表达呈正相关 (P <0 .0 1)结论　Sur vivin基因在非小细胞肺癌中表达上调 ,提示其通过抑制细胞凋亡 ,在肺癌癌变发生、发展中起重要作用 ,可能成为肺癌基因治疗的新靶点 ,其阳性表达亦预示肿瘤有较高的侵袭性和不良预后 ,Sur vivin基因与凋亡抑制基因Bcl 2的共同表达可能在肺癌中起协同作用。     

Rivaroxaban is as efficient and safe as bemiparin as thromboprophylaxis in knee arthroscopy

Purpose

The aim of this study is to compare effectiveness and safety profile of rivaroxaban with bemiparin in 3-week extended prophylaxis after knee arthroscopy.

Methods

Four hundred and sixty-seven patients were included in this review divided in two groups. One followed prophylaxis with rivaroxaban and the other one with bemiparin. All patients were interviewed and explored at 1 and 3 months postoperatively, looking for symptomatic signs of deep-vein thrombosis (DVT). In case of suspicion, diagnostic tests were performed. Collected data were age, sex, gender, diagnosis, time with ischemia, body mass index, concomitant diseases, concomitant therapy, DVT signs, treatment satisfaction, minor and major complications, treatment adherence and tolerability.

Results

No thromboembolic events were observed in any of the groups. In one case treated with rivaroxaban, the drug had to be withdrawn due to epistaxis.

Conclusions

Our study showed that extended prophylaxis with 10 mg of rivaroxaban once daily for 3 weeks resulted as effective as bemiparin in knee arthroscopy thromboprophylaxis.

Level of evidence

IV.     

When to start: as soon as possible

Debate regarding “When to start” antiretroviral (ARV) therapy has raged since the introduction of zidovudine in 1987. Based on the entry criteria for the original Burroughs Wellcome (002) study, the field has been anchored to “CD4 counts” as the prime metric to indicate ARV treatment initiation for asymptomatic HIV‐positive individuals. The pendulum has swung back and forth, based mostly on the efficacy and toxicity of available regimens. In today's world, several factors have converged that compel us to initiate therapy as soon as possible: (i) The biology of viral replication (1 to 10 billion viruses/day) screams that we should be starting early. (ii) Resultant inflammation from unchecked replication is associated with earlier onset of multiple co‐morbid conditions. (iii) The medications available today are more efficacious and less toxic than in years past. (iv) Clinical trials have demonstrated benefit for all but the highest CD4 strata (>450 to 500 cells/µL). (v) Some cohort studies have demonstrated clear benefit of ARV therapy at any CD4 count, and almost all cohort studies have demonstrated no detrimental effects of early treatment. (vi) In addition to the demonstrated and inferred benefits to the individual patient, we now have a public health benefit of earlier intervention: treatment is prevention. Finally, from a practical/common sense perspective, we are talking about life‐long therapy. Whether we start at a CD4 count of 732 or 493/µL, the patient will be on therapy for over 40 to 50 years! There does not seem to be much benefit in waiting, and there is likely to be significant long‐term harm. Treat early!     

Percutaneous dilatational tracheostomy is as safe as open tracheostomy

Although percutaneous dilatational tracheostomy (PDT) has been advocated as an alternative to open tracheostomy (OT) its relative safety has been questioned repeatedly. This study prospectively compared the safety and complications of PDT and OT. Ninety-four patients underwent PDT and 252 patients underwent OT at this institution from December 1998 through April 2000 with the choice of procedure left to the operator. OT was performed in the operating room whereas PDT was performed in intensive care units (ICUs). PDT was performed by surgeons and medical intensivists under a strict institutional policy and procedure governing patient selection and conduct of the procedure. Complications were defined as bleeding, loss of airway, hypotension, hypoxia, tracheostomy tube malposition, subcutaneous emphysema, infection, and conversion of PDT to OT. All patients survived the operation. PDT and OT had similar complication rates: 2.1 per cent for PDT versus 2.8 per cent for OT (P = not significant). Postoperative bleeding, which was the most frequent complication, occurred in one PDT patient and four OT patients. One PDT patient required conversion to OT as a result of extensive tracheal fibrosis. Subcutaneous emphysema, soft-tissue infection, and a malpositioned tracheostomy tube were the remaining complications in the OT patients. We conclude that the complication rates of PDT and OT are comparable. The choice of PDT or OT should be dictated by the surgeon's training and experience, the patient's condition, neck anatomy, and stability for transfer to the operating room.