荧光定量逆转录-聚合酶链反应检测白血病bcr/abl mRNA

目的　建立荧光定量RT PCR法检测白血病融合基因bcr ablmRNA的方法 ,为白血病临床诊断及微量残留病监测提供有用工具。方法　RT PCR扩增培养的K5 6 2细胞bcr abl融合基因 ,A T载体克隆法构建定量标准模板 ,用PE770 0型检测仪 ,建立荧光定量RT PCR方法 ,对方法的灵敏性、稳定性、重复性进行测定 ;定量检测 14例慢性髓细胞白血病 (CML)患者和 4例急性淋巴细胞白血病(ALL)患者外周血样本。结果　建立的荧光定量RT PCR方法 ,可检测出约 10 - 5μgK5 6 2总RNA中的bcr abl融合基因和 10个bcr abl重组质粒拷贝 ,重复性的CT值 (Cyclethreshold)管间、批间变异系数 (CV)分别为 :2 0 % ,3.7%。 14例CML患者bcr abl融合基因表达量中位数为 5 .15× 10 4 拷贝 μgRNA ,产物经电泳分析 ,其中 11例为b2a2 ,3例为b3a2 ,1例 12× 10 4 拷贝 μgRNA的CML患者 ,骨髓移植后 1个月变为 0 .2 3× 10 4 拷贝 μgRNA。 4例ALL患者中 1例有bcr abl融合基因的表达 ,为b2a2 ,表达量为 8.2×10 5拷贝 μgRNA。 结论　建立的荧光定量RT PCR方法灵敏、特异、重复性好 ,结果用拷贝数表示 ,准确可靠 ,利于统一标准。该方法能检测出b2a2、b3a2两种类型的融合基因 ,可广泛用于CML的诊断和微量残留病的检测     

肝癌患者外周血甲胎蛋白mRNA表达水平与肝外转移及预后的关系

目的　探讨肝癌患者外周血甲胎蛋白mRNA表达水平与肝外转移及预后的关系。方法　采用realtime PCR(RT PCR)技术 ,定量检测 5 6例肝细胞癌患者外周血甲胎蛋白mRNA的表达水平。结果　5 6例肝癌患者外周血甲胎蛋白mRNA的平均表达水平为 (346 4 4 .6± 94 90 5 .4 )copies/ μgRNA ,伴有肝外转移的 13例 10 0 %为阳性 ,平均水平为 (1385 6 9.2± 15 94 99.3)copies/ μgRNA。 结论　伴有肝外转移的肝癌患者外周血甲胎蛋白mRNA呈高水平表达 ,甲胎蛋白mRNA可作为诊断肝癌远处转移的重要指标之一。     

TaqMan实时荧光定量逆转录聚合酶链反应检测新型趋化因子巨噬细胞炎症蛋白-2γmRNA表达水平

目的　建立TaqMan实时荧光定量逆转录聚合酶链反应 (RT PCR)法检测新型趋化因子巨噬细胞炎症蛋白 2γ(MIP 2γ)mRNA表达水平 ,并对方法进行评估。方法　TaqMan实时荧光定量RT PCR检测MIP 2γmRNA的表达。结果　线性检测范围达 6个数量级 ,最低检测下限为 6× 10 2 拷贝 ,最高检测上限为 6× 10 7拷贝 ,批间及批内变异 <12 2 %。正常Balb/c小鼠心脏、肝脏、肾脏组织均可检测到MIP 2γmRNA表达 ,以心脏表达水平最高。结论　采用TaqMan实时荧光定量RT PCR检测MIP 2γmRNA表达水平准确、特异、灵敏、快速、操作简便 ,具有临床推广价值。     

一氧化氮对急性出血坏死性胰腺炎肝脏中Toll样受体2/4 mRNA表达的影响

目的探讨一氧化氮(NO)对于急性出血坏死性胰腺炎(AHNP)时肝脏中T o ll样受体2/4(TLR 2/4)mRNA表达的影响。方法采用牛磺胆酸钠(TAC)逆行胰胆管注射制备AHNP肺损伤大鼠模型。动物分为假手术组、胰腺炎组和L精氨酸(L A rg)治疗组。假手术组于术后6 h,胰腺炎组和L A rg组分别于术后3、6、12 h取静脉血和肝组织,测定血清丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(A ST)、淀粉酶和肝组织NO,逆转录聚合酶链反应(RT PCR)方法检测不同时间点肝组织肿瘤坏死因子α(TNFα)、TLR 2/4mRNA表达变化。结果与假手术组比较,胰腺炎组大鼠3 h肝组织TLR 2/4 mRNA表达开始增高〔(1.970±0.362)×10-3,(175.000±0.111)×1-0 3比(1.150±0.725)×10-6,(11.450±1.724)×10-4,〕12 h表达达峰值〔(2.940±0.316)×1-0 3,(2 673.000±88.380)×1-0 3,P均<0.01〕;血清淀粉酶ALT、A ST 3 h后即升高,肝损伤加重,肝组织TNFαmRNA表达升高,NO浓度逐渐降低(P<0.05或P<0.01);给予L A rg治疗后,NO浓度升高(P<0.05),TLR 2/4 mRNA表达降低〔3 h:(0.351±0.153)×1-0 3,(135.000±22.310)×1-0 3;6 h:(2.100±0.535)×10-3,(187.000±26.850)×1-0 3;12 h:(2.620±0.208)×1-0 3,(1 959.000±270.000)×10-3;P<0.05或P<0.01〕,血清淀粉酶、ALT、A ST均降低,肝损伤减轻,肝组织TNFαmRNA降低(P<0.05或P<0.01)。结论AHNP时,肝组织内TLR 2/4 mRNA表达上调,肝组织损伤加重;NO可以明显抑制AHNP肝组织TLR 2/4 mRNA的表达。     

荧光定量RT-PCR检测胃癌患者外周血中CD44v6基因微小转移

目的探讨胃癌患者外周血中微转移的检测和意义。方法以CD44v6基因作为目的基因,以荧光定量RT-PCR方法检测40例胃癌患者手术前、后外周血及34例同期正常对照者的外周血标本。结果34例对照者外周血中未检出CD44v6mRNA表达,40例胃癌患者术前外周血中都检出有CD44v6mRNA基因的表达,强度在4.9×102~3.2×105拷贝/μg RNA,平均表达量3.9×104拷贝/μg RNA,术后CD44v6mRNA表达强度明显下降,强度在5.5×100拷贝/μg RNA~7.6×103拷贝/μgRNA,平均2.4×102拷贝/μg RNA,术后与术前比较有显著性差异(P<0.01)。结论检测胃癌患者外周血中CD44v6基因的定量表达,可作为胃癌负荷、疗效的参数指标,为临床分期、判断预后提供依据。     

脑缺血/再灌注后缺血核心区皮质内MCP-1mRNA和蛋白质表达

目的　观察大鼠脑缺血 /再灌注后缺血核心区皮质内单核细胞趋化蛋白 (MCP) - 1mRNA和蛋白质表达情况。方法　用半定量的逆转录PCR(RT -PCR)法测定缺血 2h再灌注不同时间缺血核心区脑皮质内MCP - 1mRNA的表达 ,ELISA法测定MCP - 1含量的变化。结果　缺血核心区脑皮质内MCP - 1的mRNA表达于缺血 2h明显升高 ,再灌注后 16h达高峰(均值是缺血 2h组的 2 2倍 ,与缺血 2h组相比差异显著 ,P <0 0 1) ,此后仍维持较高水平的表达直至再灌注后 4 8h(与假手术组相比 ,P <0 0 5 ) ;MCP - 1含量于再灌注后 6h开始升高 (均值 2 2 5ng/g组织 ,是假手术组的 17 0倍 ,P <0 0 5 ) ,此后逐渐升高 ,4 8h达到高峰 (均值 110 9ng/g组织 ,是假手术组的 83 7倍 ,P <0 0 1)。结论　脑缺血 /再灌注后缺血核心区皮质内MCP - 1的mRNA和蛋白质表达均明显增加 ,提示MCP - 1在脑缺血 /再灌注损伤中发挥重要作用     

C-反应蛋白测定对小儿急性上呼吸道感染的临床价值

目的 :探讨 C-反应蛋白 (CRP)在小儿急性上呼吸道感染诊断和治疗中的临床价值。方法 :选择 30 0例儿科的急性上呼吸道感染患儿 ,采集指端微量血用免疫比浊方法进行 C-反应蛋白检测 ,同时做血常规检查 ,将 C-反应蛋白结果与 WBC计数比较。结果 :C-反应蛋白 <8mg/ L 者 2 72例 (90 .7% ) ,C-反应蛋白 >8mg/ L 者 2 8例 (9.3% ) ,其中 C-反应蛋白为 8～ 16 mg/ L 者 5例 (1.7% ) ,C-反应蛋白 >16～ 10 0 mg/ L 者 16例 (5 .3% ) ,C-反应蛋白 >10 0 mg/ L 者 7例(2 .3% )。WBC计数 <10× 10 9/者 2 5 5例 (85 .0 % )。C-反应蛋白 <8mg/ L 2 72例中 ,WBC<10× 10 9/ L 2 4 6例 (90 .4 % ) ,WBC>10× 10 9/ L 2 6例 (9.6 % ) ;C-反应蛋白 >8mg/ L 2 8例中 ,WBC>10× 10 9/ L 19例 (6 7.9% ) ,WBC<10× 10 9/ L 9例 (32 .1% ) ,两者间差异有显著性意义 (P<0 .0 1)。 2 8例 C-反应蛋白增高经抗生素治疗后恢复正常。结论 :C-反应蛋白检测对急性上呼吸道细菌或病毒感染的判断及观察抗生素的疗效具有较高的临床价值     

脑脊液白细胞计数和蛋白浓度升高是儿童急性淋巴细胞白血病的独立危险因素

本研究的目的是了解测定儿童急性淋巴细胞白血病(ALL)的脑脊液(CSF)蛋白浓度和白细胞计数对患者中枢神经系统情况和预后是否能提供参考信息。作者研究随机儿童ALL患者160例,平均追踪72个月(25-143个月),CSF蛋白浓度为0.144.3g/L,平均0.26g/L,其中>0.45g/L者20例,5年存活率为18%,<0.45g/L者5年存活率为63%(P>0.05)。CSF白细胞计数为0-528×10~6/L,平均1×10~6/L,其中>10×10~6/L者10例,5年存活率为13%,其余患儿5年存活率为60%(P<0.001)。作者把患儿分为3组:Ⅰ组133例,CSF蛋白浓度≤0.45g/L,白细胞计数≤10×10~6/L,CSF中无原始淋     

定量RT-PCR测定冠心病患者外周血中Toll样受体4的基因表达水平

目的建立检测人Toll样受体4(Toll like receptor4,TLR4)mRNA含量的实时荧光定量RT-PCR的方法,并用来测定冠心病患者外周血单个核细胞(PBMCs)中TLR4的基因表达水平,探讨TLR4基因表达水平与冠状动脉粥样硬化发生发展的关系。方法基于TaqMan荧光探针技术,构建克隆载体pGEM-T-TLR4作为定量模板,建立实时荧光定量RT-PCR方法。在GeneAmp5700型检测仪上测定了50例冠状动脉粥样硬化性心脏病患者和40名健康对照者外周血中TLR4mRNA的含量。结果TLR4mRNA在50例冠心病患者的表达范围为7·7×105~2·3×107拷贝/μgRNA;40名健康对照者的表达范围为9·8×104~5·5×106拷贝/μgRNA,冠心病组TLR4mRNA的平均拷贝数显著高于健康对照组[(5·3±3·8)×106vs(1·8±1·4)×106;P<0·01]。而冠心病血脂正常组和血脂异常组的TLR4 mRNA平均拷贝数无明显差别[(5·1±4·7)×106vs(5·4±3·0)×106;P>0·05]。结论成功建立了人TLR4基因表达含量的荧光定量检测方法;冠心病患者TLR4 mRNA的含量显著高于健康对照组,且其含量高低与血脂无关,TLR4及其介导的天然免疫应答与动脉粥样硬化的发生发展存在一定的相关性。     

成人急性白血病生存10a以上15例分析

现将我院成人急性白血病生存 10 a以上的 15例分析如下。1　临床资料1.1　一般资料　本组男 10例 ,女 5例。年龄 16～ 5 8岁 ,其中16～ 4 0岁 12例 ,>4 0岁 3例。平均年龄 2 7.3岁。类型 :按 FAB分型 ,M1 1例 ,M2 4例 ,M33例 ,M53例 ,L1 1例 ,L2 1例 ,L32例。WBC(2 .2～ 92 )× 10 9/ L,平均 11.4 4× 10 9/ L。 Hb4 5～ 90 g/ L,平均 6 9.6 g/ L。PL T2 0～ 10 0× 10 9/ L,平均 4 2 .9× 10 9/ L。脑脊液检查 ,确诊后即做脑脊液检查 ,同时鞘内注射甲氨蝶呤 10mg,氟美松 5 mg。以后急性非淋巴细胞白血病 (AML) ,6～ 8周1次 ,做 …     

Guns and Knives in New Mexico: Patterns of Penetrating Trauma, 1978–1993

Objective: To identify patterns of nonfatal and fatal penetrating trauma among children and adults in New Mexico using ED and medical examiner data.
Methods: The authors retrospectively sampled in 5-year intervals all victims of penetrating trauma who presented to either the state Level-1 trauma center or the state medical examiner from a 16-year period (1978–1993). Rates of nonfatal and fatal firearm and stabbing injury were compared for children and adults.
Results: Rates of nonfatal injury were similar (firearm, 34.3 per 100,000 person-years; stabbing, 35.1). However, rates of fatal injury were significantly different (firearm, 21.9; stabbing, 2.7; relative risk: 8.2; 95% confidence interval: 5.4, 12.5). From 1978 to 1993, nonfatal injury rates increased for children (p = 0.0043) and adults (p < 0.0001), while fatal penetrating injury remained constant. The increase in nonfatal injury in children resulted from increased firearm injury rates. In adults, both stabbing and firearm nonfatal injury rates increased.
Conclusions: Nonfatal injury data suggest that nonfatal violence has increased; fatal injury data suggest that violent death rates have remained constant. Injury patterns vary by age, mechanism of trauma, and data source. These results suggest that ED and medical examiner data differ and that both are needed to guide injury prevention programs.     

Dissociable correlates of two classes of retrieval processing in prefrontal cortex

Although substantial evidence suggests that the prefrontal cortex (PFC) implements processes that are critical for accurate episodic memory judgments, the specific roles of different PFC subregions remain unclear. Here, we used event-related functional magnetic resonance imaging to distinguish between prefrontal activity related to operations that (1) influence processing of retrieval cues based on current task demands, or (2) are involved in monitoring the outputs of retrieval. Fourteen participants studied auditory words spoken by a male or female speaker and completed memory tests in which the stimuli were unstudied foil words and studied words spoken by either the same speaker at study, or the alternate speaker. On "general" test trials, participants were to determine whether each word was studied, regardless of the voice of the speaker, whereas on "specific" test trials, participants were to additionally distinguish between studied words that were spoken in the same voice or a different voice at study. Thus, on specific test trials, participants were explicitly required to attend to voice information in order to evaluate each test item. Anterior (right BA 10), dorsolateral prefrontal (right BA 46), and inferior frontal (bilateral BA 47/12) regions were more active during specific than during general trials. Activation in anterior and dorsolateral PFC was enhanced during specific test trials even in response to unstudied items, suggesting that activation in these regions was related to the differential processing of retrieval cues in the two tasks. In contrast, differences between specific and general test trials in inferior frontal regions (bilateral BA 47/12) were seen only for studied items, suggesting a role for these regions in post-retrieval monitoring processes. Results from this study are consistent with the idea that different PFC subregions implement distinct, but complementary processes that collectively support accurate episodic memory judgments.     

Interprofessional approaches to creating safe,high quality health care

Three supplementary perspectives are presented arguing that interprofessional collaboration is both necessary and desirable. Nonetheless, there are often too many serious intra-professional barriers and obstacles to interprofessional collaboration to make it successful. Some of these barriers, it is argued and illustrated, are found in the multiple ways in which professional identity is tacitly acquired and embodied in the practitioners' habitual, everyday practice. The paper then explores ways in which reflection, especially Second order reflection, can help to elucidate and overcome these obstacles, as well as increasing professional adaptability and competence.     

Systematic Reviews Published in the January 2011 Issue of The Cochrane Library

ABSTRACT

The Cochrane Library of Systematic Reviews is published quarterly as a DVD and monthly online. The January 2011 issue (first quarterly DVD for 2011) contains 4515 complete reviews, 1985 protocols for reviews in production, and 13,521 one-page summaries of systematic reviews published in the general medical literature. In addition, there are citations of 641,000 randomized controlled trials, and 14,018 cited papers in the Cochrane methodology register. The health technology assessment database contains over 9300 citations. One hundred and seven new reviews have been published in the last 3 months, of which five have potential relevance for practitioners in pain and palliative medicine.     

Modified micro-superior percutaneously-assisted total hip: early experiences & case reports

Because of the extensile nature and familiarity of the standard posterior-lateral approach to the hip, a family of "micro-posterior" approaches has been developed. This family includes the Percutaneously-Assisted Total Hip (PATH) approach, the Supercapsular (SuperCap) approach and a newer hybrid approach, the Supercapsular Percutaneously-Assisted Total Hip (SuperPATH) approach. Such approaches should ideally provide a continuum for the surgeon: from a "micro" (external rotator sparing) posterior approach, to a "mini" (external rotator sacrificing) posterior approach, to a standard posterior approach. This could keep a surgeon within his comfort zone during the learning curve of the procedure, while leaving options for complicated reconstructions for the more practiced micro-posterior surgeons. This paper details one author's experiences utilizing this combined approach, as well as permutations of this entire micro-posterior family of approaches as applied to more complex hip reconstructions.     

Determination of creatine kinase isoenzyme MB activity in serum using immunological inhibition of creatine kinase M subunit activity. Activity kinetics and diagnostic significance in myocardial infarction.

This is a new method for the determination of creatine kinase isoenzyme MB activity in serum. The method uses direct activity measurement of creatine kinase B subunit activity after blocking of CK-M subunit activity by inhibiting antibodies. The test takes no longer than 15 min. The method yields an intra-serial C.V. of 2.0-12.9%, and a C.V. from day to day of 5.5%. The detection limit is 3.4 U/l creatine kinase MB. In the 95 cases with proven myocardial infarction several types of creatine kinase MB activity kinetics could be determined. The percentage of creatine kinase MB of peak CK-total is 6-25%, with a mean of 11.1%. The amount of creatine kinase MB with respect to total CK activity after reinfarction is higher than the amount after initial infarction.     

Structure and function of "metalloantibiotics"

Although most antibiotics do not need metal ions for their biological activities, there are a number of antibiotics that require metal ions to function properly, such as bleomycin (BLM), streptonigrin (SN), and bacitracin. The coordinated metal ions in these antibiotics play an important role in maintaining proper structure and/or function of these antibiotics. Removal of the metal ions from these antibiotics can cause changes in structure and/or function of these antibiotics. Similar to the case of "metalloproteins," these antibiotics are dubbed "metalloantibiotics" which are the title subjects of this review. Metalloantibiotics can interact with several different kinds of biomolecules, including DNA, RNA, proteins, receptors, and lipids, rendering their unique and specific bioactivities. In addition to the microbial-originated metalloantibiotics, many metalloantibiotic derivatives and metal complexes of synthetic ligands also show antibacterial, antiviral, and anti-neoplastic activities which are also briefly discussed to provide a broad sense of the term "metalloantibiotics."