钙蛋白酶抑制蛋白基因过表达Balb/C小鼠的繁育及鉴定

目的:建立钙蛋白酶抑制蛋白(calpastatin,CAST)基因过表达Balb/C小鼠的培育方法,并探讨碱性裂解液提取基因组DNA在子代鼠基因型鉴定中的价值。方法:将雄性C57BL/6-CAST~(+/-)转基因小鼠与雌性Balb/C野生型小鼠杂交,将雄性CAST~(+/-)杂合子代小鼠与雌性Balb/C野生型子代小鼠杂交,连续杂交至消除C57BL/6小鼠遗传背景。取子代小鼠鼠尾组织,采用碱性裂解液提取基因组DNA,PCR法扩增CAST基因片段,琼脂糖凝胶电泳鉴定结果。采用柯萨奇B3病毒(CVB3)感染Balb/C-CAST~(-/-)小鼠以建立急性病毒性心肌炎小鼠模型。结果:C57BL/6-CAST~(+/-)转基因小鼠与雌性Balb/C野生型小鼠连续杂交10代可消除C57BL/6小鼠遗传背景。采用碱性裂解液提取的基因组DNA数量、纯度能够满足后续实验需要,且产物大小与目的片段一致。成功建立Balb/C-CAST~(-/-)基因型小鼠急性病毒性心肌炎模型。结论:成功建立Balb/CCAST基因过表达小鼠,为后续研究奠定了基础;碱性裂解液提取基因组DNA简便、高效,值得推广。     

Shirley Clift modified-TG小鼠基因组DNA提取方法及其在Gene-Trap中的应用

目的：介绍一种简单、快速、高效同时从数十、甚至上百个小鼠尾巴中提取基因组DNA的方法。方法Shirley Clift modified-TG小鼠Gene-Trap DNA提取法。结果使用该法从小鼠尾巴中获取的基因组DNA的数量、纯度完全适用各种实验,如在C3H/HCJ-Mgrn1md转基因小鼠杂交C57BL/6小鼠Mahogunin基因变异传代培殖实验中,应用鼠尾抽提的DNA进行Gene-trap。类推该法到小鼠其它组织如肝、肾、肌肉和心脏,也同样得到了高质量的基因组DNA。结论该技术可以连续、高效、快速、批量从小鼠组织中提取基因组DNA,为国内利用小鼠进行基因遗传疾病的研究提供了基础方法。     

Shirley Clift modified-TG 小鼠基因组DNA提取方法及其在Gene -Trap中的应用

目的介绍一种简单、快速、高效同时从数十、甚至上百个小鼠尾巴中提取基因组DNA的方法.方法 Shirley Clift modified-TG 小鼠Gene-Trap DNA提取法.结果 使用该法从小鼠尾巴中获取的基因组DNA的数量、纯度完全适用各种实验,如在C3H/HCJ-Mgrn1md转基因小鼠杂交C57BL/6小鼠Mahogunin基因变异传代培殖实验中,应用鼠尾抽提的DNA 进行Gene-trap.类推该法到小鼠其它组织如肝、肾、肌肉和心脏,也同样得到了高质量的基因组DNA.结论 该技术可以连续、高效、快速、批量从小鼠组织中提取基因组DNA,为国内利用小鼠进行基因遗传疾病的研究提供了基础方法.     

利用Cre/loxp技术构建血管内敲除cdc42基因杂合子小鼠

目的:利用Cre/loxp条件性基因敲除技术构建血管内敲除ede42基因杂合子小鼠,并进行鉴定,为进一步在整体动物水平研究edc42在维持微血管屏障中的作用提供研究平台.方法:将引进的edc42fkxx/fkxx小鼠和血管内皮细胞特异性表达Cre重组酶的小鼠分别进行繁殖和鉴定,然后两种小鼠进行杂交,并对其子代小鼠的基因型进行鉴定,其子代基因型为Cdc42fkx/+Cre+/-的小鼠即为本实验所构建的血管内敲除edc42基因杂合子小鼠.结果:从引进这两种小鼠开始繁殖5个月,共繁殖edc42fkxx/fkx小鼠40只,取其21只进行鉴定,均为edc42flox纯合的子小鼠.血管内皮细胞特异性表达Ere重组酶的小鼠和野生型C57小鼠杂交后繁殖,共得到子代小鼠36只,其中表达Cre重组酶的小鼠17只,基因型为edc42fkxx/flox与表达Cre重组酶的杂合子小鼠杂交并繁殖,得到基因型为Cde42 fkxx/+Cre+/-的小鼠10只,基因型为Cde42fkox/Cre+/-13只.这两种基因型小鼠其身高、体重无明显差异.结论:本研究利用Cre/loxp技术成功构建了血管内敲除ede42基因杂合子小鼠.为进一步构建血管内敲除cdc42基因小鼠奠定基础,为在整体动物水平研究cdc42在维持微血管屏障中的作用提供研究平台.     

在小鼠骨髓移植中CCR5与急性移植物抗宿主病的相关性研究

本研究评价供者CCR5在经过强化预处理的骨髓移植动物模型受者体内的作用,为今后的异基因造血干细胞移植的临床应用提供科学依据.经过致死剂量照射的BALB/c小鼠接受异基因C57BL/6小鼠的骨髓移植.根据回输的细胞不同实验分为4组:B6 CCR5 KO组,受者接受C57BL/6 CCR5-/-小鼠骨髓和脾脏细胞;B6 WT组,受者接受野生型C57BL/6小鼠骨髓和脾脏细胞;B6 CCR5 KO BMC组,受者只接受C57BL/6 CCR5-/-小鼠骨髓细胞;B6 WT BMC组,受者只接受野生型C57BL/6小鼠骨髓细胞.结果表明:较之B6 WT组,B6 CCR5 KO组小鼠以更快的速度死于急性GVHD;其受者体内的CD8+T细胞更大量的增殖;其T细胞恢复后产生更多的INF-γ和TNF-α并且由于其T细胞有丝分裂原刀豆素水平处于较高水平,从而进一步促进T细胞的增殖,提示CCR5的作用之一是下调参与排异反应的供者CD8+T细胞的增殖.组织学评价提示,移植剔除CCR5基因受者细胞的小鼠肾脏出现了病理损伤并且肝脏存在有更为严重的病理变化.结论:剔除CCR5基因的异基因骨髓移植使GVHD发病率的增加,供者CD8+T细胞在受者体内增殖增加以及肝肾损害加重,这提示CCR5在异基因骨髓移植中起着重要作用.     

老年痴呆症模型小鼠蛋白磷酸化与短期丰富环境刺激的影响

背景:丰富环境刺激可提高对神经可塑性、学习记忆很重要的CAMKII、CREB蛋白的转录。目的:探讨短期丰富环境刺激对老年痴呆模型小鼠海马CAMKII和CREB蛋白磷酸化的影响。方法:实验分3组:长期环境组(以APP/PS1转基因C57/BL6小鼠作为老年痴呆症动物模型,从小鼠6月龄时开始进行长期的丰富环境刺激)、对照组(未进行长期环境刺激的APP/PS1转基因C57/BL6小鼠)与野生型组(非转基因的野生型C57/BL6小鼠)。在3组小鼠18月龄时,每组又随机分为基线组和刺激后两个亚组,对刺激后亚组施以为期1d的短期丰富环境刺激。结果与结论:野生型组的刺激后亚组海马CREB磷酸化水平显著高于基线亚组(P<0.05),CAMKII的磷酸化水平也有所升高;丰富环境组的刺激后亚组海马CAMKII磷酸化水平稍高于基线亚组,CREB磷酸化水平的区别不明显;对照组两个亚组海马的CAMKII和CREB磷酸化水平无明显区别。说明短期丰富环境刺激仅提高了丰富环境刺激后老年痴呆症小鼠的CAMKII磷酸化水平,但显著提高了野生型小鼠的CAMKII和CREB磷酸化水平。     

白细胞介素-6在ALD-DNA诱导的SLE模型小鼠SLE中的作用机制研究

目的探讨白细胞介素(IL)-6对ALD-DNA诱导的系统性红斑狼疮(SLE)模型小鼠SLE中的作用机制。方法在健康的雌性C57BL/6小鼠体内用活化淋巴细胞来源的DNA(ALD-DNA)免疫,从而诱导小鼠的SLE,未免疫的小鼠被用作对照。检测了疾病相关的一些发病指标,包括抗双链DNA抗体,尿蛋白和肾脏的病理学改变。结果 IL-6KO基因敲除的小鼠能抵抗ALD-DNA诱导的小鼠SLE模型。在IL-6KO免疫的小鼠中,CD4~+T细胞的活化状态低于野生型的免疫小鼠。胞内细胞因子染色结果表明,IL-6KO免疫的小鼠体内Foxp3的表达高于野生型免疫的小鼠。结论在ALD-DNA诱导的SLE模型中,IL-6可以抑制Treg细胞的分化,从而促进疾病的发展。     

雌激素诱导C57BL/6妊娠小鼠肝损伤的比较研究

目的比较雌激素对C57BL/6同种系和异种系妊娠小鼠肝脏的影响,探讨妊娠期肝损伤小鼠模型选择C57BL/6异种系妊娠小鼠的可能性。方法选择健康C57BL/6雄鼠、BALB/c雄鼠分别和C57BL/6雌鼠交配,得到C57BL/6同/异种系妊娠小鼠,并分为对照组、溶剂组和不同剂量给药组。给药组从怀孕10 d起连续4 d分别每天皮下注射不同剂量17-α-乙炔雌二醇。结果给药的异种系孕鼠血清中除碱性磷酸酶(ALP)指标外丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、总胆红素(TBIL)和甘胆酸(CG)升高水平与同种系孕鼠比较差异有统计学意义(P<0.05),其肝脏组织病理学结果也不同,且剂量为1.0μg/g体重的异种系孕鼠肝脏NK细胞出现集聚和活化现象。结论雌激素对C57BL/6同种系和异种系妊娠小鼠肝脏作用不同,选择雌激素诱导C57BL/6异种系妊娠小鼠建立妊娠期肝损伤模型能够反映该病的肝脏免疫学变化。     

腺苷A1受体敲除小鼠戊四氮点燃后脑组织病理形态学变化

目的:通过观察腺苷A1受体敲除小鼠戊四氮点燃后脑组织病理形态学变化,探讨腺苷A1受体的神经保护作用。方法:腺苷A1受体基因敲除纯合型(-/-)小鼠20只纳入敲除鼠组,腺苷A1受体基因敲除野生型小鼠20只纳入野生型组,C57BL/6小鼠10只纳入对照组。敲除鼠组和野生型组小鼠制作戊四氮点燃癫痫模型,于点燃成功后24 h、30 d采用尼氏染色观察各组小鼠海马及皮质神经元的形态结构变化。结果:野生型组小鼠点燃后皮质和海马神经元损伤较敲除鼠组出现晚、范围小,损伤程度轻。结论:腺苷A1受体对癫痫发作小鼠具有神经保护作用。     

在小鼠骨髓移植中CCRS与急性移植物抗宿主病的相关性研究

本研究评价供者CCR5在经过强化预处理的骨髓移植动物模型受者体内的作用，为今后的异基因造血干细胞移植的临床应用提供科学依据。经过致死剂量照射的BALB／c小鼠接受异基因C57BL／6小鼠的骨髓移植。根据回输的细胞不同实验分为4组：B6CCR5KO组，受者接受C57BL／6CCR5^-/-小鼠骨髓和脾脏细胞；B6WT组，受者接受野生型C57BL／6小鼠骨髓和脾脏细胞；B6CCR5KOBMC组，受者只接受C57BL／6CCR5^-/-小鼠骨髓细胞；B6、WT BMC组，受者只接受野生型C57BL／6小鼠骨髓细胞。结果表明：较之B6、WT组，B6CCR5KO组小鼠以更快的速度死于急性GVHD；其受者体内的CD8^＋T细胞更大量的增殖；其T细胞恢复后产生更多的INF-γ和TNF-α并且由于其T细胞有丝分裂原刀豆素水平处于较高水平，从而进一步促进T细胞的增殖，提示CCR5的作用之一是下调参与排异反应的供者CD8^＋T细胞的增殖。组织学评价提示，移植剔除CCR5基因受者细胞的小鼠肾脏出现了病理损伤并且肝脏存在有更为严重的病理变化。结论：剔除CCR5基因的异基因骨髓移植使GVHD发病率的增加，供者CD8^＋T细胞在受者体内增殖增加以及肝肾损害加重，这提示CCR5在异基因骨髓移植中起着重要作用。     

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.     

俯卧位通气对高海拔地区肺复张治疗无效急性呼吸窘迫综合征患者氧合的影响

目的 探讨俯卧位通气对高海拔地区肺复张术(RM)治疗无效急性呼吸窘迫综合征(ARDS)患者的治疗作用.方法 从海拔2260m的地区医院筛选RM治疗无效的41例ARDS患者[平均氧合指数( PaO2/FiO2)较RM前升高＜20％视为RM无效],依不同病因分为肺内源性ARDS组(ARDSp组)和肺外源性ARDS组(ARDSexp组),每组再按信封法随机分为俯卧位组和仰卧位组,即ARDSp俯卧位组(11例)、ARDSp仰卧位组(9例)、ARDSexp俯卧位组(10例)、ARDSexp仰卧位组(11例).在通气前及通气1、2、3、4h监测动脉血氧分压( PaO2)、PaO2/FiO2、静态顺应性(Cst)、气道阻力(Raw)的变化.结果 通气lh时,ARDSexp俯卧位组PaO2/FiO2( mm Hg,l mm Hg=0.133 kPa)即较通气前显著升高(157.4±40.6比129.3±48.7,P＜0.05),并随通气时间延长呈持续增高趋势,4h达峰值(219.1 ±41.1);且ARDSexp俯卧位组通气3h内PaO2/FiO2较其他3组显著增高,另3组间则差异无统计学意义.ARDSp俯卧位组、ARDSexp俯卧位组通气4h时PaO2/FiO2均较相应仰卧位组显著增高(208.8±39.7比127.4±47.1,219.1±41.1比124.9±50.8,均P＜0.05).4组通气前后Cst无显著改变,各组间差异也无统计学意义.ARDSp俯卧位组通气4h时Raw(cmH2O·L-1·s-1)较通气前显著降低(6.8±1.7比10.7±1.8,P＜0.05),且明显低于其他3组;其他3组各时间点Raw组内及组间比较差异均无统计学意义.结论 俯卧位通气作为ARDS机械通气重要策略之一,可以改善RM无效高原ARDS患者的氧合,为抢救患者赢得宝贵的时间.     

Digital imaging among medical facilities

The Department of Veterans Affairs (VA) in the USA operates a network of 172 medical centres which all utilize a hospital information system (HIS) which has been developed and is currently maintained by the VA. During the past several years, an image management and communication module has been developed, installed and clinically utilized at the Washington DC and Maryland VA Medical Centres. This image management and communication system, referred to as the decentralized hospital computer program (DHCP) imaging system, is fully integrated with a commercial picture archiving and communication system (PACS). The system is utilized to capture, archive, and display all images generated within the hospital including radiology, nuclear medicine, pathology, endoscopy, bronchoscopy, and dermatology, intraoperative photographs, ECG data, and a limited number of paper documents. The ultimate goal of the project is to have all patient text and image data available at any clinical workstation to any authorized user anywhere within the network of medical centres. Clinical requirements for an imaging workstation include ease of use, rapid and reliable access to the complete set of patient information, and images which are of acceptable quality to meet the requirements of the user and the subspecialty. Patient confidentiality and data security must be safeguarded at all times. Integration of the images with the remainder of the patient's database was found to be critical to the success of the project. The experience at the Washington and Maryland facilities suggests that an imaging system that is successfully integrated with a hospital information system can provide substantial clinical and economic benefits both within and among medical centres. Clinical acceptance and utilization of the system has been excellent, particularly in diagnostic radiology where DHCP Imaging has been interfaced to a commercial PAC system. Based upon this initial experience, the VA has begun to deploy the system throughout its large network of medical centres.     

Myocardial elastography at both high temporal and spatial resolution for the detection of infarcts

Myocardial elastography is a novel method for noninvasively assessing regional myocardial function, with the advantages of high spatial and temporal resolution and high signal-to-noise ratio (SNR). In this paper, in-vivo experiments were performed in anesthetized normal and infarcted mice (one day after left anterior descending coronary artery [LAD] ligation) using a high-resolution (30 MHz) ultrasound system (Vevo 770, VisualSonics Inc., Toronto, ON, Canada). Radiofrequency (RF) signals of the left ventricle (LV) in longitudinal (long-axis) view and the associated electrocardiogram (ECG) were simultaneously acquired. Using a retrospective ECG gating technique, 2-D full field-of-view RF frames were acquired at an extremely high frame rate (8 kHz) that resulted in high-quality incremental displacement and strain estimation of the myocardium. The incremental results were further accumulated to obtain the cumulative displacements and strains. Two-dimensional and M-mode displacement images and strain images (elastograms), as well as displacement and strain profiles as a function of time, were compared between normal and infarcted mice. Incremental results clearly depicted cardiac events including LV contraction, LV relaxation and isovolumetric phases in both normal and infarcted mice, and also evidently indicated reduced motion and deformation in the infarcted myocardium. The elastograms indicated that the infarcted regions underwent thinning during systole rather than thickening, as in the normal case. The cumulative elastograms were found to have higher elastographic SNR (SNR(e)) than the incremental elastograms (e.g., 10.6 vs. 4.7 in a normal myocardium, and 6.0 vs. 2.4 in an infarcted myocardium). Finally, preliminary statistical results from nine normal (m = 9) and seven infarcted (n = 7) mice indicated the capability of the cumulative strain in differentiating infracted from normal myocardia. In conclusion, myocardial elastography could provide regional strain information at simultaneously high temporal (>/=0.125 ms) and spatial ( approximately 55 microm) resolution as well as high precision ( approximately 0.05 microm displacement). This technique was thus capable of accurately characterizing normal myocardial function throughout an entire cardiac cycle, at the same high resolution, and detecting and localizing myocardial infarction in vivo.     

Clinical Pharmacokinetics of Morphine

Morphine, the most widely used mu-opioid analgesic for acute and chronic pain, is the standard against which new analgesics are measured. A thorough understanding of the pharmacokinetics of morphine is required in order to safely and effectively use this analgesic in a wide variety of patients with different levels of organ function. A MEDLINE search was conducted to identify literature published between 1966 and January 2002 relevant to the pharmacokinetics of morphine. These publications were reviewed and the literature summarized regarding unique and clinically important elements of morphine disposition relative to its parenteral administration (including intravenous, intramuscular, subcutaneous, epidural and intrathecal administration), absorption profile (immediate release, controlled release, and sublingual/buccal, and rectal administration), distribution, and its metabolism/ excretion. Special populations, including infants, elderly, and those with renal/liver failure, have a unique morphine pharmacokinetic profile that must be taken into account in order to maximize analgesic efficacy and reduce the risk of adverse events.     

Evidence-Based Pain Management and Palliative Care in Issue Four for 2009 of The Cochrane Library

ABSTRACT

The Cochrane Library of Systematic Reviews is published quarterly. Issue 4 for 2009 contains 4027 complete reviews, 1906 protocols for reviews in production, and 11447 one-page summaries of systematic reviews published in the general medical literature. In addition, there are citations of 600,000 randomized controlled trials, and 12,200 cited papers in the Cochrane methodology register. The health technology assessment database contains over 7500 citations. This edition of the Library contains 90 new reviews, of which 19 have potential relevance for practitioners in pain and palliative medicine.     

Molecular characterization of virulence and antimicrobial resistance profile of Shigella species isolated from children with moderate to severe diarrhea in northeastern Brazil

Molecular characterization of virulence and antimicrobial resistance profiles were determined for Shigella species isolated from children with diarrhea in Fortaleza, Brazil. Fecal specimens were collected along with socioeconomic and clinical data from children with moderate to severe diarrhea requiring emergency care. Shigella spp. were isolated by standard microbiological techniques, and we developed 4 multiplex polymerase chain reaction assays to detect 16 virulence-related genes (VRGs). Antimicrobial susceptibility tests were performed using disk diffusion assays. S. flexneri and S. sonnei were the predominant serogroups. S. flexneri was associated with low monthly incomes; more severe disease; higher number of VRGs; and presence of pic, set, and sepA genes. The SepA gene was associated with more intense abdominal pain. S. flexneri was correlated with resistance to ampicillin and chloramphenicol, whereas S. sonnei was associated with resistance to azithromycin. Strains harboring higher numbers of VRGs were associated with resistance to more antimicrobials. We highlight the correlation between presence of S. flexneri and sepA, and increased virulence and suggest a link to socioeconomic change in northeastern Brazil. Additionally, antimicrobial resistance was associated with serogroup specificity in Shigella spp. and increased bacterial VRGs.     

护理干预对面部中重度寻常型痤疮的临床疗效影响分析

目的研究护理干预对面部中重度寻常型痤疮的临床疗效影响。方法选取本院在2014年4月~2016年7月诊治的136例面部中重度寻常型痤疮患者,随机分为研究组与对照组,每组68例;所有患者均依据其情况给予对应的治疗,其中对照组在治疗期间给予常规护理,研究组在对照组的基础上再给予综合性护理干预,比较两组的治疗效果及护理满意度情况等。结果患者在接受治疗和护理后,研究组中度与重度患者的治疗效果较对照组均明显提高(P0.05),研究组护理满意度较对照组明显增高(P0.05)。结论对面部中重度寻常型痤疮患者在其治疗期间给予综合性护理干预,具有良好的效果。