核转录因子EBF3 mRNA在肝癌和食管癌组织中表达水平的研究

目的 研究核转录因子EBF3 mRNA在肝癌和食管癌组织中表达水平及其在肿瘤发生中的意义.方法 肝及食管组织分别以β2微球蛋白(β2M)和18S rRNA为内参,实时荧光定量RT-PCR方法 检测肝癌及食管癌及其远端组织中EBF3 mRNA的含量.结果 18例肝癌和配对远端肝组织中EBF3 mRNA与岛β2 mRNA的对数比值分别为0.52士0.17和0.28士0.23,差异有统计学意义(t=3.56,P=0.001 1).12例食管癌和配对远端食管组织中EBF3 mRNA与18S rRNA的对数比值分别为0.58士0.054和0.22士0.24,差异有统计学意义(t=5.07,P=0.000 0).结论 EBF3mRNA在肝癌和食管癌组织中表达显著增高.核转录因子EBF3可能与肝和食管肿瘤的发生有关.     

半定量RT-PCR检测原发性肝癌cyclin B1的表达和意义

目的探讨cyclin B1在肝癌组织的表达和意义.方法用逆转录聚合酶链反应(RT-PCR)检测35例肝癌组织、配对癌旁组织及5例正常肝组织cyclin B1 mRNA,以GAPDH为内参照.结果 31例(88.6%)肝癌组织及相应癌旁组织、3例(60%)正常肝组织cyclin B1 mRNA呈阳性表达,三者比较无统计学意义(P＞0.05).癌组织cyclin B1 mRNA表达水平-x±s(0.517±0.015)显著高于癌旁组织(0.282±0.023)和正常肝组织(0.173±0.011).但癌旁组织与正常肝cyclin B1mRNA表达水平无显著性差异(P＞0.05).结论相对于癌旁组织和正常肝组织,肝癌组织cyclin B1 mRNA表达水平明显增强,cyclinB1可能在肝细胞癌变过程中发挥作用.     

实时荧光定量PCR检测肝癌组织中早期B细胞因子3 mRNA表达水平

目的 建立SYBR Green Ⅰ实时荧光定量RT-PCR(FQ-RT-PCR)检测肝组织中早期B细胞因子3(early B-cellfactor 3,EBF3)tuRNA含量的方法,探讨EBF3 mRNA在肝癌中的检测意义.方法 在EBF3基因第85,第9外显子区,在内参β2M第1和第2外显子区设计特异性引物,实时检测PCR产物的荧光强度,以各自标准品建立标准曲线,由软件自动计算出待测样本中EBF3及β2M mRNA含量,以EBF3 mRNA和β2M mRNA含量的比值进行EBF3 mRNA表达水平评价.结果 FQ-RT-PCR检测EBF3 mRNA含量线性范围为3.08×107～3.08×1012copies/L,批内和批间变异系数分别为8.6%和13.7%.18例肝癌和配对远端肝组织中EBF3 mRNA与β2M mRNA的对数比值分别为0.52±0.17和0.28±0.23,差异有统计学意义(t=3.56,P=0.0011).结论 FQ-RT-PCR检测EBF3 mRNA含量的方法具有较好的检测灵敏度和重复性,适用于临床研究.     

survivin mRNA和NF-κB mRNA在原发性肝癌中的表达及其意义——附30例检测结果

目的:探讨存活素信使核糖核酸(survivin mRNA)和核因子-κB信使核糖核酸(nuclear factor-κB mRNA,NF-κB mRNA)在原发性肝细胞癌(肝癌)癌组织和相应癌旁组织中的表达及其意义.方法:收集肝癌病人的肝癌组织标本及癌旁组织标本各30份,肝血管瘤病人的瘤旁正常肝组织标本(对照组)10份,采用实时荧光定量逆转录PCR技术检测标本中的survivin mRNA和NF-κB mRNA的表达水平,以及研究两者在肝癌组织中的相关性.结果:肝癌癌组织中survivin mRNA的表达水平均高于癌旁组织和正常组织,差异均有统计学意义(P＜0.05～P＜0.01),而在癌旁组织的表达水平与正常组织的表达水平比较,差异无统计学意义(P＞0.05).癌组织中NF-κB mRNA的表达均高于癌旁组织和正常组织,差异均有统计学意义(均为P＜0.05),在正常肝组织、癌旁组织及癌组织中,该因子的表达呈逐渐升高的趋势.survivin mRNA和NF-κB mRNA在肝癌癌组织中的表达呈正相关(r=0.36,P＜0.05).结论:survivin mRNA和NF-κB mRNA在肝癌癌组织中呈高表达,且两者在癌组织中的表达呈正相关,提示两者可能与肝癌的发生、发展有关.     

原发性肝癌组织中LASP-1 mRNA和HPA-1 mRNA表达水平及其临床意义

目的定量检测LIM和SH3蛋白1(LASP-1)mRNA和乙酰肝素酶-1(HPA-1)mRNA在原发性肝癌组织中的表达水平,探讨二者与肝癌发生、侵袭和转移的关系。方法利用实时荧光定量PCR检测法分别检测62例原发性肝癌癌组织、配对癌旁组织(距癌2 cm)和正常肝脏组织(距癌大于5 cm)中LASP-1mRNA和HPA-1 mRNA的表达水平,并分析二者与肝癌临床病理特征的关系。结果肝癌组织、癌旁组织、正常肝组织中HPA-1 mRNA表达水平分别为47.25±16.65、23.15±14.89、7.56±5.15,LASP-1 mRNA的表达水平分别为51.66±20.12、35.73±15.07、10.17±8.29。HPA-1 mRNA及LASP-1 mRNA在癌组织中的表达高于癌旁组织和正常组织(P均<0.05)。HPA-1及LASP-1的表达与肿瘤分化程度、浸润深度、淋巴结转移及TNM分期密切相关(P<0.05,P<0.01),与患者年龄、性别及肿瘤大小无关(P均>0.05)。Spearman等级相关分析显示LASP-1 mRNA和HPA-1 mRNA的表达呈正相关(OR=0.421,P<0.05)。结论肝癌组织中LASP-1 mRNA与HPA-1 mRNA的表达与肿瘤浸润深度、分化程度、淋巴结转移及TNM分期等密切相关。二者联合表达水平的增加可能促进肝癌的侵袭转移。     

gp96、Mcl-1在肝硬化和肝癌组织中表达及意义

目的通过研究gp96、髓样细胞白血病-1(Mcl-1)蛋白在肝硬化和肝癌组织中的表达及临床病理学意义,初步探讨其与肝硬化和肝癌发生、发展的关系。方法采用免疫组织化学ENVISION法分别检测19例肝硬化组织(癌旁肝硬化组织)、32例肝癌组织和21例对照肝组织(癌旁非硬化肝组织)中gp96、Mcl-1的表达,并分析其各自表达与肝癌临床病理学特征的关系。取来源于同一患者的肝癌组织、癌旁肝硬化组织或癌旁非肝硬化组织,分别配对检测gp96、Mcl-1,比较两者的阳性表达率。结果对照组、肝硬化组和肝癌组gp96阳性表达率逐渐递增,差异有统计学意义(P0.05);gp96的阳性表达与肿瘤有无包膜、TNM分期有关(P0.05),与患者的性别、年龄、肿瘤大小、血清甲胎蛋白(AFP)值、组织学分级及临床分期无关(P0.05)。肝癌组和肝硬化组Mcl-1阳性表达率明显高于对照组(P0.05),但肝癌组和肝硬化组之间差异无统计学意义(P0.05);Mcl-1阳性表达与肿瘤有无坏死和TNM分期有关(P0.05)。肝癌组gp96阳性表达率明显高于配对癌旁肝硬化组及配对癌旁非肝硬化组(P均0.05);而肝癌组Mcl-1阳性表达率明显高于配对癌旁非肝硬化组,与配对癌旁肝硬化组比较差异无统计学意义(P0.05)。结论 gp96、Mcl-1过表达可能与肝癌的发生、发展有关。gp96可能参与了肝硬化的发生、发展及向肝癌的恶性转化,有助于判断肝癌患者的预后。     

肝细胞肝癌中PED/PEA-15 mRNA及蛋白表达的临床意义

目的:探讨PED/PEA-15 mRNA及蛋白在肝癌组织、癌旁组织及正常肝组织中的表达情况及其临床意义.方法:应用半定量逆转录.聚合酶链反应检测40例肝细胞肝癌、相应癌旁组织和12例正常肝组织中PED/PEA.15基因的相对表达量.并通过免疫组化检测PED/PEA-15蛋白的表达.结果:半定量RT-PCR显示,PED/PEA-15基因在肝癌组织中均呈高表达,癌旁组织及正常肝组织呈低表达或不表达,相对表达量分别为1.201±0.301、0.64±0.101和0.565±0.077,PED/PEA-15 mRNA在肝癌组织中表达较癌旁组织、正常肝组织显著升高(P<0.01).免疫组织化学显示,肝癌组织、癌旁组织及正常肝组织中PED/PEA-15蛋白的表达率分别为72.5%、32.5%和16.7%.肝癌组织中PED/PEA-15蛋白表达较癌旁组织及正常肝组织显著升高(P<0.01).且PED/PEA-15 mRNA及蛋白的表达与肝癌病理分级、临床分期有关(P<0.05),而与发病年龄、性别、肿瘤大小、肿瘤数目差异无显著性(P>0.05).结论:PED/PEA-15 mRNA及蛋白在肝癌组织中表达水平明显升高,可能在肝癌的发生及发展过程中发挥重要作用.     

TLR4和NF-κB p65表达与结肠癌的关系

目的 评价结肠癌和癌旁正常组织中TLR4 mRNA和NF-κB p65蛋白表达水平与结肠癌发生发展的相关性.方法 取手术切除的新鲜结肠癌组织和距癌组织2 cm以上的癌旁组织各63份,其中:结肠癌高分化型组23份,中分化型组17份,低分化型组20份,其他分化类型3份;伴淋巴结转移组27份,不伴淋巴结转移组36份;Dukes A期组18份,Dukes B期组14份,DukesC期组22份,Dukes D期组9份.采用实时荧光定量RT-PCR检测结肠癌组织和癌旁组织中TLR4 mRNA的表达水平;用WB法检测NF-κB p65蛋白的相对表达水平.结果 结肠癌组织中TLR4 mRNA表达量为86.42±15.16,明显高于癌旁组织的32.74±9.44,差异有统计学意义(t=22.354,P＜0.01).高、中、低分化型结肠癌TLR4 mRNA表达量分别为69.58±11.27、64.57±13.91、97.12±15.44,低分化型结肠癌表达量高于高、中分化型结肠癌(t值分别为11.304、12.223,P均＜0.01).伴淋巴结转移组TLR4 mRNA表达量(89.91±13.33)与不伴淋巴结转移组(81.16±13.59)比较,差异无统计学意义(t=0.959,P＞0.05).Dukes A期组TLR4 mRNA表达量(59.05±11.66)低于Dukes B～D期组的表达量(分别为92.32±17.51、91.41±15.21、101.46±17.43),差异有统计学意义(t值分别为8.708、9.664、9.525,P均＜0.05);结肠癌组织和癌旁组织NF-κB p65蛋白表达量分别为0.63±0.11、0.34±0.08,结肠癌组织NF-κB p65表达量高于癌旁组织(t=18.266,P＜0.01),高、中、低分化型结肠癌组织中NF-κB p65蛋白表达量分别为0.46±0.09、0.72±0.11、0.77±0.14,中低分化型组表达高于高分化型(t值分别为11.223、10.875,P均＜0.01),伴淋巴结转移组与不伴淋巴结转移组NF-κB p65表达量分别为0.82±0.17、0.57±0.12,且差异有统计学意义(t=18.269,P＜0.05).Dukes A期的NF-κB p65表达量(0.39±0.06)低于Dukes B～D期(分别为0.72±0.12、0.69±0.14、0.76±0.13),且差异有统计学意义(t值分别为10.442、9.889、9.721,P均＜0.01).结论 TLR4/NF-κB p65信号通路的表达水平升高与结肠癌的临床进展和病理分级密切相关;NF-κB p65可能是结肠癌转移的分子指标之一,TLR4/NF-κB p65升高可促进结肠癌的发生发展.     

Ⅰ型钙黏蛋白基因异常甲基化及其功能蛋白表达与肺癌的相关性

目的 探讨Ⅰ型钙黏蛋白(cadherin 1,CDH1)基因甲基化及其功能蛋白表达与肺癌的相关性.方法 分别提取基因组DNA、总RNA和总蛋白,用半定量荧光甲基化特异PCR(MSP)、巢式逆转录PCR(nRT-PCR)和免疫印迹法(WB)检测30例肺癌患者的癌、癌旁和远癌肺组织及5例肺良性疾病对照肺组织中CDH1甲基化、mRNA相对表达和上皮钙黏蛋白(E-cadherin,E-cad)表达情况,并对部分表达阴性标本用免疫组化法进行验证.结果 半定量荧光MSP检测CDH1启动子甲基化相对比值,癌组织为0.13%～450.67%、中位数33.61%,癌旁组织为0.00%～177.02%、中位数18.04%,远癌组织为0.00%～51.68%、中位数13.69%.经配对秩和检验分析癌与癌旁组织和远癌组织中CDH1甲基化程度差异有统计学意义(Z分别为-2.355和-3.527,P分别为＜0.05和＜0.01).nRT-PCR检测CDH1 mRNA表达的相对比值,癌组织为1.33±0.48,癌旁组织为1.61±0.55,远癌组织为1.93±0.45、肺良性疾病对照肺组织为2.09±0.09,经方差分析,癌与癌旁和远癌组织的差异有统计学意义(F=9.081,P＜0.01).WB结果显示,癌组织E-cad阳性率为36.7%(11/30)、癌旁组织为70.0%(21/30)、远癌组织为96.7%(29/30),经确切概率法分析癌与癌旁和远癌组织的差异有统计学意义(X2分别为6.70、24.30和7.68,P均＜0.01).而肺良性疾病对照肺组织阳性率为100%(5/5).结论 肺癌患者癌组织CDH1启动子发生甲基化后,可抑制该基因的转录,进而影响E-cad蛋白的表达,CDH1异常甲基化与肺癌的发生有相关性.     

肝细胞核因子3β在人类肺癌组织中的表达及意义

目的 探讨肝细胞核因子3β蛋白(HNF-3β)在非小细胞肺癌(NSCLC)组织和癌旁正常肺组 织中的表达情况,探讨其临床意义.方法 分别采用免疫组化法和RT-PCR 方法检测50 例NSCLC 组织及癌 旁正常肺组织中HNF-3β蛋白和mRNA 的表达.结果 HNF-3β蛋白在NSCLC 和正常癌旁组织中的半定量 评分分别为0.42 ±0.13,2.28 ±0.28,差异有统计学意义(P <0.05).HNF-3βmRNA 在NSCLC 和正常癌旁 组织中的半定量评分分别为1.32 ±0.23,3.68 ±0.32,差异有统计学意义(P <0.05).结论 初步认定HNF- 3β对人类非小细胞肺癌的形成起抑制作用.     

微管相关蛋白1轻链3在新生期大鼠惊厥后海马的表达

Objective To explore the repetitive expressions of autophagy marker protein-rnicrotubule-associ-ated protein 1 light chain 3 (LC3) in hippocampus in newborn rats with recurrent seizure and the influence of 3-methyladeine (3-MA) on LC2 expressions. Method Seventy-two 6-day-old SD rats were randomly (random nam-ber) divided into the recurrent neonatal seizure group (RS group, n = 24), the 3-MA-treated seizure group (3-MA group, n = 24) and control group (n = 24). Rats in RS group were subjected to 55 attacks of seizure induced by flurothyl in 9 successive days from the 6th postnatal day (P6). In 3-MA group, 2 μL of 3-MA was injected every day till seizure induced. Western blot analysis was used to determine LC3 protein level in hippocampus at different intervals of 1.5 h,3 h,6 h and 24 h after the last convulsion. The LC3 protein level was analyzed with Dunnett test after ANOVA. Results LC3 protein levels in RS group at the different intervals were significantly higher than those in the control group and in 3-MA group (F =4.70,5.28,8.51 and 5.89, respectively, P <0.05), and there were no significant differences in LC3 protein level between 3-MA group and control group at those intervals (P > 0.05). Conclusions The autophagy/lysosomal pathway is immediately activated after recurrent seizure evidenced by the elevated expressions of LC3 in hippocampus. The 3-MA is involved in the regulation of autophagy/ lysosomal pathway by down-regulating the expressions of LC3.     

微管相关蛋白1轻链3在新生期大鼠惊厥后海马的表达

Objective To explore the repetitive expressions of autophagy marker protein-rnicrotubule-associ-ated protein 1 light chain 3 (LC3) in hippocampus in newborn rats with recurrent seizure and the influence of 3-methyladeine (3-MA) on LC2 expressions. Method Seventy-two 6-day-old SD rats were randomly (random nam-ber) divided into the recurrent neonatal seizure group (RS group, n = 24), the 3-MA-treated seizure group (3-MA group, n = 24) and control group (n = 24). Rats in RS group were subjected to 55 attacks of seizure induced by flurothyl in 9 successive days from the 6th postnatal day (P6). In 3-MA group, 2 μL of 3-MA was injected every day till seizure induced. Western blot analysis was used to determine LC3 protein level in hippocampus at different intervals of 1.5 h,3 h,6 h and 24 h after the last convulsion. The LC3 protein level was analyzed with Dunnett test after ANOVA. Results LC3 protein levels in RS group at the different intervals were significantly higher than those in the control group and in 3-MA group (F =4.70,5.28,8.51 and 5.89, respectively, P <0.05), and there were no significant differences in LC3 protein level between 3-MA group and control group at those intervals (P > 0.05). Conclusions The autophagy/lysosomal pathway is immediately activated after recurrent seizure evidenced by the elevated expressions of LC3 in hippocampus. The 3-MA is involved in the regulation of autophagy/ lysosomal pathway by down-regulating the expressions of LC3.     

容积重建成像3D-CTA与3D-DSA在诊断急性破裂性颅内微小动脉瘤的研究

目的 对比研究容积重建成像三维CT血管造影与三维DSA(3D-DSA)在颅内微小动脉瘤诊疗中的临床应用价值.方法 对广东省人民医院2007年5月至2008年11月收治的174例蛛网膜下腔出血患者首先采用采用GE公司的Light Speed Plus 64排容积螺旋CT机获得原始图像,采用容积重建成像技术(VR)进行三维重建.并辅助运用多轴面重建(MPR),然后再行全脑血管造影术,并行3D-DSA成像.结果 本组174例蛛网膜下腔出血患者诊断为颅内微小动脉瘤11例,均经开颅手术证实;其中CTA诊断11例,3D-DSA诊断10例.容积重建成像CTA清晰显示颅内微小动脉瘤、载瘤动脉、动脉瘤的形状和大小及其与邻近结构的解剖关系,与3D-DSA差异无统计学意义.结论 容积重建成像CTA是一种可靠、无创的快速诊断颅内微小动脉瘤的方法,为急症手术提供了详实的影像学资料,可帮助制定治疗方案.     

日本血吸虫信号蛋白14-3-3在虫卵的定位及其免疫诊断价值初探

目的探讨日本血吸虫信号蛋白14—3—3（sj14—3—3）在虫卵内的定位和重组Sj14—3-3（rsj14—3—3）的免疫诊断价值。方法从日本血吸虫尾蚴感染42d的兔肝脏中分离虫卵,用逆转录聚合酶链反应（1it—PCR）检测sj14．3．3基因在虫卵期的转录水平;兔肝组织石蜡切片免疫组化染色,观察内源性Sj14—3—3在虫卵内的分布和表达丰度。利用纯化的rSj14-3-3和可溶性虫卵抗原（SEA）,采用间接酶联免疫吸附试验（ELISA）检测急、慢性血吸虫病患者和正常人血清。结果RT-PCR从日本血吸虫成熟虫卵中检出760bp左右的基因片断;免疫组化显示Sj14-3-3主要分布在虫卵内毛蚴的体壁和两边的侧腺。检测急性、慢性患者和正常人血清抗rSj14-3—3抗原的抗体阳性率分别为91．0％、78．9％、0．0％;上述标本中抗SEA抗体的阳性率分别为97．4％、88．9％和2．5％。结论用ELISA检测抗SEA抗体和rSj14-3-3抗体诊断血吸虫病具有高度的特异性和敏感性,而Sj14—3-3在成熟虫卵阶段高表达,可能是SEA的主要组分,具有实用价值。     

N-甲基天冬氨酸受体拮抗剂AP-5在海马对兔P3波电位的作用

目的推测Ｎ甲基天冬氨酸受体（ＮｍｅｔｈｙｌＤａｓｐａｒａｔｅｒｅｃｅｐｔｏｒ,ＮＭＤＡＲ）的兴奋性突触后电位（ｅｘｃｉｔａｔｏｒｙｐｏｓｔｓｙｎａｐｔｉｃｐｏｔｅｎｔｉａｌｓ,ＥＰＳＰ）活动对兔Ｐ３波的作用。方法采用ＮＭＤＡＲ竞争性拮抗剂ＡＰ５（３．１２５、６．２５、１２．５ｍｍｏｌ／Ｌ）在海马ＣＡ１、ＣＡ３微量注入,观察Ｐ３波电位变化。结果ＡＰ５在ＣＡ１区呈一定量效依赖延长Ｐ３ａ波潜伏期,在ＣＡ１、ＣＡ３为明显量效依赖延长Ｐ３ｂ波潜伏期,ＡＰ５对Ｐ３ａ、Ｐ３ｂ波电压振幅无明显影响。结论ＣＡ１区ＮＭＤＡＲＥＰＳＰ活动是Ｐ３ａ波发生相关神经元兴奋的易化因素,可能是海马实施对Ｐ３ａ波潜伏期调节的重要机制。ＣＡ１、ＣＡ３区ＮＭＤＡＲＥＰＳＰ可能共同对Ｐ３ｂ波发生相关神经元兴奋起易化作用,故可影响其潜伏期。     

Optimal growth of human blood hematopoietic progenitor cells collected by apheresis for autografts

For safe autografts with peripheral blood hematopoietic cells (PBSCT), better methods for determining the kinetics of stem cell populations and predicting engraftment speed after PBSCT need to be established. Current methods include culture in semi-solid medium and measurement of CD34 cell surface antigen. In this study with only partially purified blood cells obtained from children with cancer in remission, we compared the effects of phytohemagglutinin-stimulated lymphocyte-conditioned medium (PHA-LCM) and recombinant human cytokines on the growth of progenitor cells in a methylcellulose culture system. Interleukin-3 (IL-3) alone supported more progenitor growth than standard PHA-LCM by a factor of 1.54 for colony-forming unit granulocyte/macrophages (CFU-GM) and by a factor of 1.84 for burst-forming unit/erythroids (BFU-E). No significant change, in terms of the number of growing colonies, was observed by adding granulocyte/macrophage colony-stimulating factor (GM-CSF), granulocyte-CSF (G-CSF), or IL-1 to IL-3. However, the addition of G-CSF resulted in increased colony size. A further increase in CFU-GM growth was observed by the addition of IFN-γ to the combination of cytokines. No significant effect was observed when stem cell factor (SCF) was added to the combination of cytokines containing IL-3, G-CSF, and IFN-γ. This analysis suggests that the combination of IL-3, G-CSF, and IFN-γ may provide sufficient stimulation for the growth of human blood cells. The effects of different oxygen tensions on progenitor growth in the presence of IL-3, G-CSF, and IFN-γ were also evaluated. Both CFU-GM and BFU-E formation were increased when the culture was grown at 5% O₂, as compared with an ambient 20% O₂ tension. A small number of infused cells were grown in culture incorporating either IL-3, G-CSF, and IFN-γ at 5% O₂ or PHA-LCM at 20% O₂, and the number of infused cells was correlated to the speed of hematopoietic recovery after PBSCT. Although a significant negative correlation was observed between the number of infused CFU-GM per kilogram of the patient's body weight and the recovery of hematopoiesis under both culture conditions, a better correlation was found when the former method was applied (P lt; .001 vs. P lt; .05). These findings suggest that a culture containing IL-3. G-CSF, and IFN-γ at low O₂ tension provides satisfactory conditions for the proliferation of blood progenitors, and that this mixture of recombinant cytokines may enable a standardized hematopoietic progenitor assay for PBSCT.     

FLT3靶向短发夹状干扰RNA体外转录合成及作用

FMS样酪氨酸激酶3（FLT3）是一种酪氨酸激酶受体,在大多数急性髓系白血病（AML）患者中持续激活,与患者预后差密切相关。为探讨3种FLT3靶向短发夹状干扰RNA（shRNA）对急性髓系白血病细胞株THP-1的沉默效应,设计和体外转录合成3个FLT3靶向shRNA（shRNA1、shRNA2、shRNA3）,体外转染THP-1细胞;以RT-PCR法检测FLT3mRNA水平表达,用流式细胞术、免疫荧光测定法检测FLT3蛋白的表达。结果显示：shRNA1、shRNA3可显著下调FLT3mRNA的表达,其中shRNA1的抑制作用较强。25nmol/LshRNA1转染48小时对FLT3mRNA的抑制率是（72.95±2.07）%,作用可达72小时。5nmol/L及以上浓度的shRNA1对FLT3mRNA表达有下调作用,作用存在量-效关系。15nmol/LshRNA1的抑制率是（67.53±0.66）%。FLT3蛋白位于细胞膜上,shRNA1对其有较强的抑制作用,转染72小时蛋白抑制率达（79.67±0.66）%。结论：FLT3-shRNA1具有较好的FLT3基因靶向抑制作用,可作为进一步研究该基因作用机制及靶向治疗可能性的工具。     

Role of antioxidant enzymes in brain tumours

Previous reports of patients with 3-hydroxy-3-methylglutaric aciduria have described the occurrence of di-trimethylsilyl (TMS) and tri-TMS derivatives of 3-hydroxy-3-methylglutaric acid on analysis using gas chromatography and mass spectrometry, leading to difficulty in quantification and ambiguity in diagnosis. We have extracted organic acids from the urine of patients with 3-hydroxy-3-methylglutaric aciduria using a variety of procedures. Solvent extraction combined with hydrochloric acid/sodium chloride resulted in production of both di-TMS and tri-TMS derivatives of 3-hydroxy-3-methylglutaric acid and also mono-TMS and di-TMS derivatives of 3-hydroxyisovaleric acid. The effects were not abolished by heating. Use of sulphate-based reagents minimised artefact formation and use of DEAE–Sephadex anion exchange extraction resulted in single fully trimethylsilylated derivatives. Artefact formation during use of chloride-based reagents was abolished by pyridine added prior to trimethylsilylation. Chloride ions form adducts with hydroxyl groups in these 3-hydroxy-3-methyl carboxylic acids that prevent complete trimethylsilylation. Chloride-based reagents should be avoided in the solvent extraction of organic acids from physiological fluids or, if used, pre-treatment of the dried extract with pyridine is essential to avoid partial trimethylsilylation of 3-hydroxy-3-methyl carboxylic acids.     

三维超声软件系统Ultra 3DTM的设计及临床应用

三维超声成像技术作为科研课题已经存在多年,随着计算机和图像处理技术的进步,开发具有临床应用价值的三维超声影像软件系统吸引了越来越多的注意力.本文基于开发Ultra 3DTM三维超声软件的经验,对三维超声成像系统的特点,设计和实现进行了分析和介绍.临床应用的实例验证了Ultra 3DTM系统的稳定性和有效性.     

Evaluation of the role of platelet integrins in fibronectin-dependent spreading and adhesion.

BACKGROUND: Recent studies have shown that platelet adhesion and subsequent aggregation can occur in vivo in the absence of the two principal platelets adhesive ligands, von Willebrand factor and fibrinogen. These results highlight a possible role for fibronectin in supporting thrombus formation. OBJECTIVE AND METHODS: To evaluate the platelet integrins and subsequent activation pathways associated with fibronectin-dependent platelet adhesion utilizing both human and murine platelets. RESULTS: Platelets can adhere to fibronectin via the integrin alpha(IIb)beta(3), leading to formation of lamellipodia. This is mediated through an interaction with the tenth type III domain in fibronectin. Spreading on fibronectin promotes alpha(IIb)beta(3)-mediated Ca(2+) mobilization and tyrosine phosphorylation of focal adhesion kinase and phospholipase C gamma2. In contrast, studies with blocking antibodies and mice demonstrate that alpha(5)beta(1) and alpha(v)beta(3) support adhesion and promote formation of filopodia but not lamellipodia or tyrosine phosphorylation of these proteins. Further, neither alpha(5)beta(1) nor alpha(v)beta(3) is able to induce formation of lamellipodia in the presence of platelets agonists, such as collagen-related-peptide (CRP). CONCLUSIONS: These observations demonstrate that integrins alpha(5)beta(1) and alpha(v)beta(3) support platelet adhesion and the generation of filopodia but that, in contrast to the integrin alpha(IIb)beta(3), are unable to promote formation of lamellipodia.