超声靶向破坏微泡技术介导shRNA抑制小鼠肝癌细胞株JNK1基因表达

目的 探讨应用超声靶向破坏微泡（UTMD）技术介导shRNA抑制小鼠肝癌细胞株JNK1基因表达的能力。方法 构建并筛选shRNA最佳表达载体。体外培养肝癌细胞Hca-F,共分为5组：A组为空白对照组;B组为shRNA质粒组;C组为脂质体组;D组为超声微泡+超声辐照组;E组为脂质体+超声微泡+超声辐照组。应用倒置荧光显微镜观察转染率;荧光定量PCR检测JNK1的mRNA水平;Western-Blot检测JNK1的蛋白质表达。结果 获得了shRNA干扰效果最好的表达载体。各组转染率比较：E组均大于B、C、D组（P均<0.05）;C、D组之间差异无统计学意义（P>0.05）。荧光定量PCR和Western-Blot检测各组JNK1mRNA和蛋白表达比较：E组的JNK1mRNA和蛋白表达水平均最低（P均<0.05）。结论 脂质体转染法与UTMD技术结合可以提高小鼠肝癌细胞株JNK1 shRNA的转染效率,并能够增强基因表达的抑制效果。     

超声微泡介导miR-34a联合DOX治疗小鼠U14皮下移植瘤的实验研究

目的研究超声介导载miR-34a、DOX微泡对小鼠U14宫颈癌皮下移植瘤联合治疗效果。方法制备载miR-34a、DOX高分子微泡,检测一般性质;建立小鼠皮下移植瘤模型,随机分为:模型组、空微泡组、DOX微泡组、miR-34a微泡组、miR-34a联合DOX微泡组;超声微泡治疗小鼠皮下移植瘤,绘制肿瘤生长曲线图,计算抑瘤率;RT-PCR检测各组小鼠肿瘤组织miR-34a基因表达情况;免疫组织化学染色法检测肿瘤微血管密度(MVD)和凋亡蛋白Bcl-2、Bax表达情况。结果自制高分子微泡呈球形,分布均一,平均粒径1.87μm,包封率为46.24%,载药量为10.58%;与模型组相比,各治疗组肿瘤生长减慢,肿瘤质量体积显著下降,联合治疗效果更佳并与单一作用比较具有显著差异(P0.05);聚合酶链式反应(RT-PCR)结果显示,miR-34a组和联合组肿瘤组织中miR-34a高表达;DOX组、miR-34a组和联合组免疫组织化学染色检测肿瘤微血管密度(MVD)降低,凋亡蛋白Bcl-2表达下调,Bax高表达,联合组更加明显(P0.05)。结论超声微泡介导的miR-34a和DOX的共同递送可以实现肿瘤抑制的协同效应。     

DWI评价正常核苷组合治疗小鼠结肠癌的疗效

目的 探讨DWI评价正常核苷组合治疗小鼠结肠癌的效果。方法 将24只结肠癌小鼠随机分为实验组和对照组,每组12只。对实验组每天腹腔注射正常核苷组合1000 mg/kg体质量,对照组每天以相同剂量注射生理盐水,持续用药14天。分别于用药前1天及用药第4、8、14天进行常规MR、DWI检查,后进行Western Blot检测,记录Bax、Bcl-2的表达水平,并进行统计学分析。结果 用药第4、8、14天,实验组肿瘤体积均较对照组减小(P均<0.05),实验组肿瘤ADC值均较对照组增大(P均<0.05)。与对照组比较,实验组肿瘤组织Bax蛋白表达明显增多(t=257.88,P<0.05),Bcl-2蛋白表达明显减少(t=-234.03,P<0.05)。ADC值与Bax蛋白表达呈正相关(r=0.738,P<0.05),与Bcl-2蛋白表达呈负相关(r=-0.736,P<0.05)。结论 DWI技术可实时、无创、准确地评价正常核苷组合对小鼠结肠癌的疗效。     

超声介导载阿霉素微泡对小鼠H22肝癌皮下移植瘤抑制作用的实验研究

目的探讨超声介导载阿霉素微泡对小鼠H22肝癌皮下移植瘤的抑制效果。方法 (1)制备载阿霉素微泡,并检测其一般性质。(2)建立小鼠H22肝癌皮下移植瘤模型,将荷瘤小鼠随机分为对照组、空微泡+超声组、阿霉素组及阿霉素微泡+超声组,治疗5次后,绘制各组肿瘤生长曲线,计算抑瘤率,冰冻切片观察阿霉素在肿瘤组织的聚集情况,RT-PCR及免疫组化检测肿瘤组织中Bax、Bcl-2基因和蛋白的表达情况,Tunel检测肿瘤组织的细胞凋亡情况。结果 (1)制备的载阿霉素微泡呈球形,平均粒径为(438.33±12.87)nm,包封率为43.65%±3.84%。(2)阿霉素微泡+超声组肿瘤组织内阿霉素的聚集量是阿霉素组的3.49倍。与对照组相比,空微泡+超声组、阿霉素组及阿霉素微泡+超声组肿瘤生长均受到抑制,肿瘤体积和质量抑瘤率均升高,Bax基因和蛋白表达上调,Bcl-2基因和蛋白表达下调,肿瘤组织出现不同程度的细胞凋亡,上述结果以阿霉素微泡+超声组表现更明显(P<0.01)。结论超声介导载阿霉素微泡可抑制小鼠H22肝癌皮下移植瘤的生长,促进肿瘤细胞的凋亡。     

超声空化联合恩度抑制大鼠Walker-256肿瘤血管新生

目的 观察微泡增强超声空化毁损肿瘤微血管联合恩度对肿瘤血管新生的抑制作用。方法 对健康雄性SD大鼠40只于单侧大腿内侧皮下种植Walker-256肿瘤，随机均分为超声空化联合恩度组（A组）、恩度组（B组）、超声空化组（C组）和超声假照组（D组），每组10只，每天1次、连续给予相应干预3天；干预前、干预后1天和4天分别行二维超声及超声造影（CEUS）检查。之后获取肿瘤组织，行血管内皮生长因子A（VEGFA）免疫组织化学染色，测定微血管密度（MVD）。比较各组造影峰值强度（PI）、曲线下面积（AUC）、肿瘤体积及MVD。结果 干预后1天，CEUS中A组PI和AUC值明显低于其余各组（P均<0.05），B组和C组则低于D组（P均<0.05）；干预后4天，各组中A组PI和AUC值仍最低（P均<0.05），B组仍显著低于D组（P<0.05）。干预后1天，A组肿瘤体积（1.94±0.64）cm³最低，B组、C组均低于D组（P均<0.05）。干预后4天，A组肿瘤体积（5.25±1.74）cm³仍为各组中最低（P均<0.05）。各组MVD存在差异（F=30.121，P<0.001）。A组（18.33±3.10）、B组（24.17±2.16）MVD明显低于C组（30.43±4.17）和D组（31.57±1.74）（P均<0.05）。结论 微泡超声空化联合恩度治疗可显著减少肿瘤血流灌注，从而抑制大鼠Walker-256肿瘤新生血管生成及肿瘤生长。     

过敏性哮喘患者高迁移率族蛋白1、Th1/Th2细胞亚群及相关细胞因子表达分析

目的：分析高迁移率族蛋白-1（high mobility group protein-1，HMGB-1）、辅助T淋巴细胞（Th）亚群以及相关细胞因子在不同严重程度的过敏性支气管哮喘患者中的表达意义。方法：选择98例轻中重度过敏性哮喘患者和30例同期体检的健康者（对照组），采用流式细胞术分析患者以及健康者外周血CD4⁺干扰素-γ⁺（IFN-γ⁺）T细胞、CD4⁺白细胞介素-4⁺（IL-4⁺）T细胞百分比。通过酶联免疫吸附测定法（ELISA）检测血清中HMGB-1、Th1细胞因子（IFN-γ）和Th2细胞因子（IL-4、IL-5）的表达水平。采用Spearman分析HMGB-1、Th1/Th2细胞亚群及其相关细胞因子与肺功能的相关性。结果：所有哮喘患者HMGB-1、CD4⁺IL-4⁺T细胞百分比、IL-4、IL-5均高于对照组；重度组患者HMGB-1、CD4⁺IL-4⁺T细胞百分比、IL-4、IL-5高于轻、中度患者。所有患者CD4⁺IFN-γ⁺T细胞百分比、IFN-γ表达均低于对照组；重度患者CD4⁺IFN-γ⁺T细胞百分比和IFN-γ表达低于轻、中度患者。HMGB-1、IL-4水平与肺功能负相关，IFN-γ水平与肺功能正相关。结论：HMGB-1和IFN-γ、IL-4可作为评价支气管哮喘严重程度的有效指标。     

微泡增强的超声空化阻断肿瘤微循环的病理观察

目的 探讨微泡增强的超声空化阻断肿瘤微循环的病理机制。方法 建立兔肝VX2转移瘤模型18只,随机分为超声微泡组(n=6)、单纯超声组(n=6)及假照组(n=6)。对超声微泡组经耳缘静脉注射微泡,并进行超声辐照;对单纯超声组以生理盐水代替微泡,并进行超声辐照;对假照组注射生理盐水,并进行超声假照。观察辐照后各组病理变化。结果 大体解剖观察,超声微泡组辐照面可见较多出血点;单纯超声组及假照组辐照后肿瘤组织均未见明显改变。光镜下观察,超声微泡组肿瘤组织可见大片出血,血管内皮细胞损伤、连续性中断,红细胞外溢;单纯超声组可见局灶性小片状出血,血管内皮尚完整;假照组肿瘤组织未见明显出血,血管内皮完整。电镜下观察,超声微泡组肿瘤血管内皮破损严重,可见大量红细胞渗出,线粒体肿胀及吞饮小泡;单纯超声组及假照组仅见内质网略水肿。结论 微泡增强的超声空化阻断兔VX2肝脏肿瘤微循环的机制可能为血管机械性损伤。     

体素内不相干运动成像在体检测人类结肠癌SW480裸鼠耐药性

目的 探讨体素内不相干运动（IVIM）成像活体评价人类结肠癌SW480裸鼠耐药性及其可行性。方法 对耐药组（n=5）和不耐药组（n=5）人类结肠癌SW480荷瘤裸鼠于肿瘤最长径大于1.50 cm后分别行IVIM DWI检查,测量肿瘤真扩散系数（D）、假扩散系数（D^*）及灌注分数（f）。处死荷瘤鼠,检测肿瘤坏死（HE染色）、凋亡（TUNEL法）及P-糖蛋白（P-gp）、多药耐药相关蛋白1（MRP1）、蛋白激酶C（PKC）蛋白表达（Western Blot）,对IVIM参数和肿瘤蛋白表达情况进行相关分析。结果 不耐药组D较耐药组增高（P<0.05）,2组D^*和f差异无统计学意义（P均>0.05）。2组肿瘤组织坏死区域范围相似;耐药组细胞核较不耐药组增大,且细胞间排列更紧密,细胞密度较大。2组肿瘤细胞凋亡指数差异无统计学意义（P>0.05）。耐药组PKC、P-gp、MRP1蛋白表达均较不耐药组增加（P均<0.05）。肿瘤D值与P-gp、MRP1、PKC表达均呈负相关（P均<0.05）。结论 IVIM成像参数D值可能成为评估小鼠人类结肠癌SW480耐药性的指标。     

超声评价微泡诱导超声空化联合血凝酶增强兔VX2肝癌微波热消融作用

目的 采用二维灰阶超声、CDFI和CEUS评价微泡诱导超声空化联合血凝酶对兔VX2肝癌微波热消融的增强作用。方法 将32只VX2肝癌荷瘤新西兰大白兔随机分为生理盐水组（空化假辐照+生理盐水）、血凝酶组（空化假辐照+生理盐水+血凝酶）、空化组（超声空化+微泡）和联合组（超声空化+微泡+血凝酶）4组,每组8只,分别给予相应空化治疗后行微波热消融治疗,并在治疗前后分别行二维灰阶超声、CDFI和CEUS检查,观察治疗前后超声表现,测量并比较肿瘤和消融区体积。结果 治疗前4组间肿瘤体积差异无统计学意义（P>0.05）,治疗后4组间消融区体积总体差异有统计学意义（P<0.001）;两两比较,联合组消融区体积大于其他3组（P均<0.05）,空化组消融区体积大于生理盐水组和血凝酶组（P均<0.05）,生理盐水组与血凝酶组体积差异无统计学意义（P>0.05）。治疗前肿瘤均可见丰富血流信号;消融后联合组肿瘤实质内CEUS均未显示增强,生理盐水组、血凝酶组和空化组部分肿瘤实质消融区边缘可见少量残余活性组织,呈典型"快进快出"表现,与CDFI显示的点状血流信号相对应。结论 微泡诱导超声空化联合血凝酶可增强兔VX2肝癌微波热消融效果。     

超声造影评价小鼠三阴性乳腺癌模型特征及EGFR靶向治疗效果

目的 以超声造影（CEUS）动态观察小鼠三阴性乳腺癌（TNBC）病灶及新生血管变化,评价表皮生长因子受体（EGFR）通路靶向治疗TNBC的价值。方法 建立30只TNBC小鼠模型,分为靶向组（靶向阻断EGFR通路）及对照组各15只。分别于建模后第5、10、15、20天行常规超声及CEUS检查,测量相关参数;每次检查结束后随机处死3~4只小鼠,以免疫组织化学检测肿瘤标本中EGFR及血管内皮生长因子（VEGF）表达。采用Spearman相关性分析评价CEUS参数与EGFR及VEGF的关系。结果 建模后第5天,靶向组与对照组肿瘤体积无明显差异（P>0.05）;建模后第10~20天,靶向组肿瘤体积均明显小于对照组（P均<0.05）。各时间点靶向组峰值强度（PI）均显著低于对照组（P均<0.05）;组间上升时间（RT）及峰值时间（TTP）差异均无统计学意义（P均>0.05）。建模后第10~20天靶向组EGFR均显著低于对照组（P均<0.05）;建模后第5~20天靶向组VEGF均显著低于对照组（P均<0.05）。EGFR与VEGF相关（r=0.629,P<0.001）;PI与EGFR及VEGF均相关（r=0.679、0.839,P均<0.001）。结论 CEUS可评估小鼠TNBC病灶及新生血管变化;靶向阻断EGFR通路可有效缩小小鼠TNBC模型肿瘤体积,降低其CEUS PI及EGFR、VEGF表达水平。     

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.     

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.     

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.     

手转胎头术失败的原因与分娩结局临床分析

目的 探讨手转胎头术失败的原因与分娩结局.方法 选择2008年1月至2010年12月于我院住院分娩的持续性枕横位、枕后位产妇198例,根据行手转胎头术后结果分为成功组126例、失败组72例.比较两组分娩结局,对比分析失败原因.结果 失败组胎儿体质量≥3500 g的发生率[76.4％(55/72)]明显高于成功组[31.7％(40/126)],差异有统计学意义(x2=30.177,P=0.001)、失败组宫缩乏力发生率[58.3％(42/72)]高于成功组[38.1％ (48/126)],差异有统计学意义(x2=7.569,P=0.006)、失败组骨盆临界或轻度狭窄发生率[38.9％ (28/72)]高于成功组[23.8％(30/126)],差异有统计学意义(x2 =5.030,P=0.002)、失败组手转胎头时机不当(宫口开大＜6 cm、胎头位于坐骨棘上及宫口开大8～10 cm、胎头位于坐骨棘下≥2 cm)发生率[61.1％(44/72)]高于成功组[38.9％(49/126)],差异有统计学意义(x2=9.084,P=0.003).失败组母儿并发症(产后出血、产褥病率、胎儿窘迫、新生儿窒息)发生率高于成功组(x2 =9.586,P=0.002、x2=9.334,P=0.002、x2=5.910,P=0.015、x2=5.240,P=0.022)、失败组剖宫产发生率[72.2％(52/72)]明显高于成功组[34.1 ％(43/126),x2=26.641,P=0.001)].结论 手转胎头术能使难产变顺产,降低剖宫产率,减少母儿并发症,但须积极预防、处理导致手转胎头术失败的原因,对矫正失败后继续矫正及试产应慎重.