慢病毒介导siRNA干扰尿路上皮癌相关基因1表达对人肺腺癌细胞系A549恶性行为的影响

目的探究干扰尿路上皮癌相关基因(UCA)1的表达对人肺腺癌细胞系A549恶性行为的影响。方法实时荧光定量PCR(RT-PCR)检测人肺腺癌细胞系A549、H358、H1299、H460及人支气管上皮细胞系HBE中UCA1的表达情况;慢病毒转染表达针对UCA1的小干扰RNA(siRNA)的载体,同时转染空白载体于A549细胞中,分别作为干扰组及对照组细胞,RT-PCR检测两组细胞UCA1的表达情况,MTS实验检测两组细胞0、12、24、48及72 h的增殖能力,Transwell侵袭及迁移实验检测两组细胞转移能力,免疫印迹实验检测上皮间充质转化(EMT)相关分子E-钙黏蛋白(cadherin)及波形蛋白(Vimentin)的表达。结果 A549、H358、H1299、H460细胞系中UCA1的表达均显著高于HBE细胞系,差异有统计学意义(P0.05),且A549细胞系表达高于其他肺腺癌细胞系。干扰组细胞UCA1的表达显著低于对照组(P0.05),干扰效率73.2%。干扰组细胞48及72 h增殖能力显著低于对照组细胞(P0.05)。干扰组侵袭及迁移细胞数均显著低于对照组,差异有统计学意义(P0.05)。干扰组细胞E-cadherin表达相比对照组显著上调,而Vimentin表达相比对照组显著下调,差异有统计学意义(P0.05)。结论体外干扰UCA1的表达可通过下调A549细胞EMT进程抑制增殖及转移能力,UCA1可能是潜在的肺腺癌靶向治疗位点。     

HPS-1对人肺腺癌A549细胞凋亡诱导作用的机制研究

目的 评价HPS-1对人肺腺癌A549细胞凋亡蛋白Caspase 3、8、9以及Bax、Bcl-2mRNA表达的影响,探究HPS-1抗肺肿瘤可能机制.方法 人肺腺癌A549细胞经低、中、高剂量HPS-1处理不同时间后,噻唑蓝(MTT)法检测各组细胞的增殖情况,流式细胞术(FCM)法检测各组细胞凋亡率及细胞周期比例,比色法检测各组细胞凋亡蛋白Caspase-3、8、9的表达,RT-PCR法检测各组细胞Bax、Bcl-2 mRNA的表达.结果 MTT结果显示,低、中、高剂量HPS-1对人肺腺癌A549细胞的增殖具有明显抑制作用,且具有时间、剂量依赖性;FCM法检测发现,低、中、高剂量HPS-1对人肺腺癌A549细胞的凋亡具有促进作用,且与剂量呈正相关;比色法检测各组细胞发现,HPS-1促进凋亡蛋白Caspase-3、8、9的上调,且HPS-1高剂量组凋亡蛋白上调更明显.结论 HPS-1具有抑制人肺腺癌A549细胞增殖、诱导其凋亡作用,且该作用可能与HPS-1阻滞细胞周期G1期,上调凋亡蛋白Caspase-3、8、9及Bax mRNA表达,下调Bcl-2 mRNA表达有关.     

启动子区域甲基化对肺腺癌GPC5表达的调控作用及意义

目的 研究启动子区域CpG岛的甲基化对肺腺癌细胞GPC5表达的调节作用.方法 在线预测GPC5启动子区域CpG岛的分布,合成甲基化特异性聚合酶链反应(Methylation specific polymerase chain reaction,MSP)引物,甲基化酶特异性抑制剂5-氮杂-2'-脱氧胞嘧啶核苷(5-AZA)处理肺腺癌细胞系A549、SPC-A1及永生化支气管上皮细胞HBE,检测处理前后GPC5表达的变化.同时用MSP方法检测肺腺癌组织标本中GPC5基因启动子区域CpG岛甲基化情况,以探讨其可能的临床意义.结果 GPC5在正常的支气管上皮中不表达,而在肺腺癌细胞系中均有明显表达.5-AZA处理后,GPC5在A549及SPC-A1中的表达均上调,提示启动子区域CpG岛的甲基化参与了GPC5基因表达的调节.但肺腺癌组织标本中GPC5基因启动子区域CpG岛呈现低甲基化状态,提示这种低甲基化状态参与了肺腺癌细胞中GPC5表达的上调.结论 GPC5基因启动子区域的甲基化参与了其表达的调控,可能是导致肺腺癌中GPC5上调的一个重要因素.     

iTRAQ联合LC-MS/MS技术在肺腺癌血浆生物标志物筛选中的应用

目的应用同位素标记相对和绝对定量(iTRAQ)联合液相色谱串联质谱(LC-MS/MS)技术检测正常者及肺腺癌患者血浆中的差异蛋白,从而筛选出潜在的肺腺癌血浆生物标志物,并探讨其临床意义。方法收集10例健康体检者血浆(正常组),10例肺腺癌患者血浆(腺癌组),应用iTRAQ标记联合LC-MS/MS技术对两组血浆进行差异蛋白组学分析。结果质谱共鉴定到蛋白369种,其中差异表达的蛋白有35种;腺癌组较正常组上调的蛋白有21种、下调的蛋白有14种,其中SCGB3A2、SFTPB蛋白表达显著上调。结论筛选出多种与肺腺癌相关的差异蛋白,其中SCGB3A2、SFTPB有望成为潜在的肺腺癌血浆生物标志物,提示iTRAQ联合LC-MS/MS技术可用于肺腺癌血浆肿瘤标志物的筛选。     

三氧化二砷对肺腺癌化学治疗敏感性的影响及其机制

目的　研究三氧化二砷 (As2 O3)对肺腺癌细胞生长过程及化学治疗 (简称化疗 )敏感性的影响。方法　应用四甲基偶氮唑蓝 (MTT)比色、免疫组织化学、流式细胞仪、逆转录 聚合酶链反应(RT PCR)等方法 ,观察不同浓度As2 O3 对人肺腺癌A549细胞株生长、化疗敏感性、细胞周期 ,及凋亡基因Fas/FasL、抗凋亡基因B淋巴细胞白血病 2 (bcl 2 )和多药耐药相关蛋白 (MRP)、肺耐药蛋白 (LRP)表达水平的影响。结果　 1、2 μmol/L的As2 O3 对人肺腺癌A549细胞株的生长抑制作用较弱 ,但能显著提高A549细胞对顺铂的敏感性 (P <0 .0 5)、提高A549在G1期细胞的比率 ,上调Fas、下调bcl 2及MRP、LRP的表达水平。 5μmol/L的As2 O3 对人肺腺癌A549细胞有一定的抑制作用 ,并可上调bcl 2、MRP、LRP的表达水平 ,但不能提高A549细胞对顺铂的敏感性。结论　低浓度的As2 O3 可能通过提高A549细胞在G1期的比率 ,上调Fas基因和下调bcl 2、MRP、LRP基因的表达水平 ,提高肺腺瘤细胞对化疗的敏感性     

糖尿病大鼠海马超微结构及蛋白质组学研究

目的 研究早期糖尿病大鼠海马超微结构及蛋白质表达谱变化,探寻糖尿病对中枢神经系统早期损害可能的机制.方法 将15只6～8周龄、体重200～250g雄性sD大鼠按随机数字表法分为对照组及糖尿病组.腹腔注射链脲佐菌素(STZ)55mg/kg制备糖尿病模型,5周后用透射电镜观察海马超微结构,采用双向凝胶电泳技术分离海马总蛋白,基质辅助激光解析电离飞行时间质谱(MALDI-TOF-MS)鉴定差异蛋白质.结果 超微病理学观察结果:成模5周时糖尿病组海马神经元固缩,毛细血管管周足突水肿明显;对照组大鼠神经元、毛细血管未见明显异常改变.蛋白质组学研究结果:建立早期糖尿病大鼠海马蛋白质的双向凝胶电泳图谱;质谱鉴定8个差异表达大于2倍的蛋白质,其中线粒体细胞色素b-cl复合物亚型、14-3-3蛋白β/α、胶质纤维酸性蛋白(GFAP)、鸟苷酸结合蛋白g(0)α亚基表达上调,线粒体ATP合成酶d亚基、线粒体硫氧还蛋白依赖性过氧化物还原酶、内质网蛋白(ERP29)表达下调,内吞蛋白只在对照组表达.结论 早期糖尿病大鼠海马已出现病理改变,这种改变可能与氧化应激损伤导致能量代谢异常有关,与老化及神经变性疾病相关的蛋白质可能参与了糖尿病中枢神经系统损害的发病.     

诺帝对肺腺癌细胞系A549细胞凋亡的诱导作用

目的探讨化学合成物诺帝对人肺腺癌细胞系A549细胞凋亡和凋亡调控基因bcl-2蛋白表达的影响。方法采用自行合成的化合物诺帝（终浓度为100umol／L）处理A549细胞,于处理后12、24、48和72h用细胞培养、流式细胞仪、免疫组化及图像分析等方法检测诺帝对A549细胞凋亡和bcl-2蛋白的表达。结果一定浓度的诺帝处理A549细胞12～48h,均可诱导A549细胞发生凋亡,且随着作用时间的延长,凋亡细胞数增加越明显;诺帝处理A549细胞后,出现细胞bcl-2蛋白表达水平显著降低。结论诺帝诱导了A549细胞的凋亡,其作用可能与其下调了bcl-2蛋白的表达有关。     

羟基喜树碱诱导肝癌细胞凋亡前后线粒体蛋白质组表达差异的定量分析

目的 对羟基喜树碱诱导人肝癌细胞SMMC-7721凋亡前后的线粒体进行定量蛋白质组学研究.方法 用羟基喜树碱诱导肝癌细胞SMMC-7721凋亡,提取线粒体并鉴定其纯度;用可裂解的同位素标记的亲和标签技术标记线粒体蛋白质,利用二维液相色谱串联质谱分析鉴定蛋白质.结果 获得了羟基喜树碱诱导细胞凋亡前后的高纯度线粒体;分析鉴定了细胞凋亡前后,相对表达量差异有统计学意义的蛋白质74种,其中42种线粒体蛋白质在细胞凋亡后的表达量下调,32种蛋白质的表达量在凋亡细胞中上调.这些差异蛋白的分子功能主要表现为能量代谢和核酸的翻译、转录、复制以及细胞骨架等.结论 获得了羟基喜树碱诱导肝癌细胞凋亡前后的线粒体差异蛋白信息,将促进对癌细胞凋亡机制及羟基喜树碱药理作用的深入理解,同时,实验所采用的整合技术平台将有助于亚细胞定量蛋白质组学研究.     

人参皂甙Rg3对肺腺癌A549细胞株信号传导相关基因表达的影响

目的探讨人参皂甙Rg3[20(R)-Rg3]对人肺腺癌A549细胞株的信号传导相关基因差异表达的影响。方法A549细胞株分2组进行培养，实验组培养液中含有10^-6mol／L的20(R)-Rg3，对照组不加药物，干预培养72h后用BiostarH-40s型基因芯片测定其基因差异表达。结果共检测到有效基因点数量3490个，20(R)-Rg3干预后有24个基因发生差异表达，其中2个细胞信号和传递蛋白基因以及2个细胞骨架和运动基因上调，原癌基因和抑癌基因下调的有3个，蛋白翻译合成的基因2个下调、1个上调，DNA合成修复重组和代谢相关基因1个、免疫相关基因1个下调，代谢基因1个上调，其他功能基因也受到一定影响。结论20(R)-Rg3可显著影响人肺腺癌A549细胞株信号传导等相关基因的差异表达。     

亚硒酸钠、砒霜对肺腺癌细胞周期、凋亡、耐药的影响

目的通过比较低浓度亚硒酸钠、砒霜对肺腺癌细胞A549的周期、凋亡、耐药及线粒体的影响,探讨肺腺癌药物治疗的新方法。方法体外培养肺癌A549细胞,分为对照组及5.0μmol/L亚硒酸钠（A组）、5.0μmol/L砒霜（B组）处理组,药物处理24 h后以流式细胞仪检测细胞周期;Western blot检测三组促凋亡因子Caspase-3、抗耐药基因RARα的表达,RT-PCR检测三组凋亡抑制基因Bcl-2、细胞周期素CyclinD3表达;电子显微镜观察细胞线粒体结构与凋亡小体。结果与对照组相比,A、B组G0/G1期增加、S和G2/M期减少,出现细胞周期阻滞,凋亡细胞比例增加。Caspase-3、RARα表达上调,Bcl-2、cyclinD3表达下调。线粒体肿胀、线粒体嵴模糊不清,出现凋亡小体。结论亚硒酸钠、砒霜治疗肺腺癌均有良好的效果,因为氧化还原能力不同,作用各有侧重。     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Changes in connexin43, the gap junction protein of astrocytes, during development of the rat pineal gland

Abstract: The abundance of gap junctions between rat pineal astrocytes formed by connexin43 (Cx43) was studied during development. Levels and distribution of Cx43 were measured by immunoblotting and indirect immunofluorescence, respectively. The amount of Cx43 in cells located within the gland was low until about the 7th postnatal day and increased to adult values between the 14th and 21st days postpartum. Although astrocytes, recognized by their vimentin immunoreactivity, were scarce before birth, they were abundant by the 7th postnatal day suggesting that the low levels of Cx43 found at this age corresponded to a low expression of this protein. Localization of the immunoreactivity to Cx43 and vimentin showed a close correlation, indicating that mature or immature pineal astrocytes form gap junctions made of Cx43. Since Cx43 levels attained their adult values at about the time the innervation and the functional state of the gland reached maturity (2–3 weeks after birth), it is proposed that astrocyte gap junctions are involved in the function of the adult rat pineal gland.     

Conservative management of perforated duodenal diverticulum： A case report and review of the literature

Duodenal diverticula are a relatively common condition. They are asymptomatic, unless they become complicated, with perforation being the rarest but most severe complication. Surgical treatment is the most frequently performed approach. We report the case of a patient with a perforated duodenal diverticulum, which was diagnosed early and treated conservatively with antibiotics and percutaneous drainage of secondary retroperitoneal abscesses. We suggest this method could be an acceptable option for the management of similar cases, provided that the patient is in good general condition and without septic signs.     

Presence of immunoreactive growth hormone and prolactin in the ovine pineal gland

Abstract: The use of antisera raised against bovine growth hormone (GH) and ovine prolactin (PRL) enabled the detection of related immunoreactive (ir) sequences of proteins in ovine pineal tissue. The isolation of PRL-like ir-material was accomplished using a 0.25 M ammonium sulphate (pH 5.5) extraction followed by ethanol precipitation, whereas the resulting 2.0 M ammonium sulphate (pH 7.0) precipitate contained a GH-like immunoreactivity. Gel chromatography of the GH-like immunoreactivity (Sephadex G-100) indicated the presence of several GH-like fragments ranging in the M_r range of 7,000 to 55,000. Analyses of the PRL-like ir-material found in pineal tissue on HPLC using a TSK 545-DEAE column led to the resolution into a single peak of immunoreactivity. A single peak of activity was also observed following chromatofocusing and hydrophobic interaction chromatography of the ir-peak from the TSK 545-DEAE column. The PRL-like ir-material inhibited the binding of [¹²⁵I]ovine PRL-S14 to anti-ovine PRL antibodies without showing an affinity for binding to anti-rat PRL or anti-bovine GH antibodies. Scatchard analysis of the binding of pineal PRL-like ir-material and pituitary ovine PRL-S14 to liver membranes from day-20 pregnant rats revealed similar affinity constants (K_a of 4.7 ± 0.2 × 10⁹ M^-1). In addition, the replication of Nb 2 Node rat lymphoma cells was stimulated by pineal PRL-like ir-material, an effect known to be specific for lactogenic hormones. The pineal PRL-like immunoreactivity appeared on sodium dodecyl sulfate polyacrylamide gels as a single major band of M_r 24,000. The functional status of PRL-and GH-like ir-material in the ovine pineal remains to be determined, but evidence is presented that the overall protein synthesis rate of the rat pineal responded to circulating concentrations of PRL.     

N-acetyl-L-cysteine combined with mesalamine in the treatment of ulcerative colitis： Randomized,placebo-controlled pilot study

AIM： To evaluate the effectiveness and safety of oral N-acetyl-L-cysteine （NAC） co-administration with mesalamine in ulcerative colitis （UC） patients.
METHODS： Thirty seven patients with mild to moderate UC were randomized to receive a four-wk course of oral mesalamine （2.4 g/d） plus N-acetyl-L-cysteine （0.8 g/d） （group A） or mesalamine plus placebo （group B）. Patients were monitored using the Modified Truelove-Witts Severity Index （MTWSI）. The primary endpoint was clinical remission （MTWSI ≤ 2） at 4 wk. Secondary endpoints were clinical response （defined as a reduction from baseline in the MTWSI of ≥ 2 points） and drug safety. The serum TNF-α, interleukin-6, interleukin-8 and MCP-1 were evaluated at baseline and at 4 wk of treatment. RESULTS： Analysis per-protocol criteria showed clinical remission rates of 63% and 50% after 4 wk treatment with mesalamine plus N-acetyl-L-cysteine （group A） and mesalamine plus placebo （group B） respectively （OR = 1.71; 95% CI： 0.46 to 6.36; P = 0.19; NNT = 7.7）. Analysis of variance （ANOVA） of data indicated a significant reduction of MTWSI in group A （P = 0.046） with respect to basal condition without significant changes in the group B （P = 0.735） during treatment. Clinical responses were 66% （group A） vs 44% （group B） after 4 wk of treatment （OR = 2.5; 95% CI： 0.64 to 9.65; P = 0.11; NNT = 4.5）. Clinical improvement in group A correlated with a decrease of IL-8 and MCP-1. Rates of adverse events did not differ significantly between both groups.
CONCLUSION： In group A （oral NAC combined with mesalamine） contrarily to group B （mesalamine alone）, the clinical improvement correlates with a decrease of chemokines such as MCP-1 and IL-8. NAC addition not produced any side effects.     

Delorme's transrectal excision for internal rectal prolapse

Surgical therapy of functional outlet obstruction in patients with internal rectal intussusception may include abdominal, perineal, or transrectal procedures. Because abdominal procedures often result in significant physiologic impact but unrelieved constipation, the authors have elected Delorme's transrectal excision for management of these patients. Since a short-term placebo effect attends many therapies, this report describes results of transrectal excision only after a threeyear postoperative period. Delorme's transrectal excision of internal intussusception accomplished sustained symptomatic relief in over 70 percent of otherwise refractory constipated patients. The association of internal intussusception with other abnormalities underscores the importance of defining both anatomic and functional components when selecting patients whose constipation may require surgical therapy. Critical technical elements, surgical pitfalls, and potential complications of the procedure are discussed.Poster presentation at the meeting of The American Society of Colon and Rectal Surgeons, Toronto, Canada, June 11 to 16, 1989.     

The oral glucose tolerance test: A comparison of the time points on the basis of limit values,normal dispersion,and reproducibility

Summary Time points in the glucose tolerance test (GTT) are compared on the basis of limit values, dispersion within a reference population, and reproducibility. We suggest using the distance between a limit value and the median reference value as a measure of the magnitude of abnormality. The distance between 140 mg/100 ml and the median fasting plasma glucose value is chosen as a standard distance and limits for other points in the GTT are calculated to equal this standard distance of abnormality. We suggest that the probability of correctly interpreting an inividual result is directly related to the reproducibility of the test and inversely related to the percentage of the total range of values which is dispersed among the normal population. The ratio of reproducibility to percentage normal dispersion is proposed as an index of the probability of correctly interpreting an individual result. According to this index, the probability of correct interpretation varies in order: fasting plasma glucose concentration>3-h>2-h>0.5-h>1-h plasma glucose concentration.