Kupffer细胞CD14及Toll样受体4介导大鼠肝移植缺血再灌注损伤的机制

目的研究大鼠肝移植缺血再灌注后Kupffer细胞CD14和Toll样受体4(TLR4)的表达及其参与缺血再灌注损伤的机制。方法建立肝移植缺血再灌注模型,并分为正常对照组、缺血再灌注组、抗CD14抗体组,每组均为10只大鼠。分离培养大鼠肝移植缺血再灌注后的Kupffer细胞。检测Kupffer细胞CD14及TLR4的mRNA、蛋白表达、核转录因子κB(NFκB)活性以及培养上清TNFα的分泌量。结果再灌注后Kupffer细胞CD14及TLR4的mRNA和蛋白表达明显高于正常对照组(P<001),再灌注后核转录因子κB活性、培养上清TNFα表达量明显高于对照组(P<001)。用抗CD14抗体后NFκB活性,TNFα表达量明显下降(与再灌注组相比,P<005),但仍然高于对照组(P<001)。结论缺血再灌注后肠道内毒素(脂多糖)能够上调Kupffer细胞CD14及TLR4的表达,激活NFκB,启动细胞因子的转录和分泌,但除CD14和TLR4以外的其他信号途径参与了缺血再灌注损伤。     

还原型谷胱甘肽对大鼠肝脏缺血再灌注损伤后炎性细胞因子表达的影响

目的 探讨经门静脉注射还原型谷胱甘肽(GSH)对大鼠肝脏缺血再灌注损伤后TNF-α、IL-1β和巨噬细胞炎性蛋白-2(MIP-2)表达的影响及意义.方法 72只雄性SD大鼠平均分为假手术组(SO组)、生理盐水预处理组(IR组)和GSH预处理组(GPC组).建立肝脏缺血再灌注损伤模型,检测再灌注30、60和180 min血清TNF-α、IL-1β含量,以及肝组织中TNF-α mRNA、IL-1β mRNA和MIP-2mRNA表达水平.两独立样本采用t检验,多组比较采用方差分析.结果 GPC组血清TNF-α含量于缺血再灌注180 min后显著低于IR组(t=2.512,P<0.05).而肝组织TNF-αmRNA表达水平于缺血再灌注30 min后即显著低于IR组(t=2.427,P<0.05).GPC组血清中IL-1β含量和肝组织中IL-1βmRNA表达水平于缺血再灌注后各时相点均显著低于IR组(t=2.731,3.825,4.372,3.371,3.972,4.685,P<0.05).GPC组MIP-2 mRNA表达于缺血再灌注60 min和180 min显著低于IR组(t=2.593,5.429,P<0.05).结论 TNF-α、IL-1β和MIP-2等炎性因子在肝脏缺血再灌注损伤中发挥重要作用.GSH能够抑制炎性细胞因子如TNF-α、IL-1β和MIP-2的生成,并发挥抗肝脏缺血再灌注损伤的作用.     

RNA干扰Kupffer细胞肿瘤坏死因子-α减轻大鼠肝脏缺血再灌注损伤

目的 观察RNA干扰肝脏Kupffer细胞肿瘤坏死因子-α(TNF-α)对大鼠肝脏缺血再灌注损伤的保护作用.方法 构建针对大鼠TNF-α基因的短发夹状RNA(shRNA)真核表达载体.肝脏缺血再灌注损伤前48 h经门静脉注射磷酸盐缓冲液(PBS)、空载体或TNF-α shRNA.实验随机分为4组,假手术组、PBS组、空载体组和shRNA组.阻断大鼠70%入肝血流40 min,再灌注6 h检测血清谷丙转氨酶(ALT)、谷草转氨酶(AST)、肝脏Kupffer细胞TNF-α mRNA、血清TNF-α、肝组织中丙二醛(MDA)以及超氧化物岐化酶(SOD)含量.结果 与PBS组和空载体组比较,shRNA组再灌注6 h后血清ALT和AST水平显著降低(P＜0.05),Kupffer细胞TNF-α mRNA水平、血清TNF-α水平(56.6±6.7 pg/ml比87.8±8.7 pg/ml和96.5±7.3 pg/ml,P＜0.05)、肝组织中MDA含量(93.4±13.3 nmol/mg比133.5±12.4 nmo1/mg和136.7±13.6 nmol/mg,P＜0.05)显著降低,SOD活性显著升高(22.4±4.6 U/mg比12.2±3.1 U/mg和11.4±2.9 U/mg,P＜0.05).结论 RNA干扰Kupffer细胞TNF-α基因的表达可以减轻大鼠肝脏缺血再灌注损伤.     

利多卡因预先给药对肾脏缺血再灌注损伤大鼠肾组织CD44和TNF-α表达的影响

目的 评价利多卡因预先给药对肾脏缺血再灌注损伤大鼠肾组织CD44和TNF-α表达的影响.方法 健康雄性Wistar大鼠36只,体重300～350 g,随机分为3组(n=12):假手术组(S组)、肾脏缺血再灌注组(IR组)和利多卡因预先给药组(L组).采用无创动脉夹夹闭双侧肾动脉60min、恢复灌注4 h,建立大鼠肾脏缺血再灌注模型.L组于夹闭双侧肾动脉前5min时尾静脉注射利多卡因5 mg/kg;IR组于夹闭双侧肾动脉前5 min时尾静脉注射等容量生理盐水;S组不夹闭双侧肾动脉,于分离肾动脉后尾静脉注射等容量生理盐水.再灌注4 h时处死大鼠,取肾组织,光镜下观察病理学结果;采用免疫组化法测定肾组织CD44和TNF-α的表达水平.结果 与S组比较,IR组肾组织CD44和TNF-α的表达上调(P＜0.05),L组肾组织CD44和TNF-α的表达差异无统计学意义(P＞0.05);与IR组比较,L组肾组织CD44和TNF-α的表达下调(P＜0.05),肾组织损伤减轻.结论 利多卡因预先给药减轻大鼠肾脏缺血再灌注损伤与其抑制肾组织CD44和TNF-α的表达有关.     

核因子κB圈套寡核苷酸减轻大鼠移植肝缺血/再灌注损伤的作用和机制

目的探讨抑制枯否（Kupffer）细胞核因子κB（Nuclearfactor-kappaB，NF-κB）活性对减轻大鼠移植肝缺血／再灌注损伤（IRI）的作用和机制。方法建立大鼠肝移植缺血／再灌注损伤模型。实验分正常对照组、缺血／再灌注组和圈套寡核苷酸组，每组均为8只大鼠。圈套寡核苷酸组于移植术前2d经供者尾静脉注入120μg脂质体包裹的NF-κB圈套寡核苷酸。移植再灌注后2h，取各组受者移植肝分离枯否细胞。凝胶迁移变动分析法（EMSA）检测枯否细胞NF-κB蛋白结合活性，逆转录聚合酶链法（RT—PCR）观察枯否细胞肿瘤坏死因子α（TNF—α）和白细胞介素6（IL-6）mRNA的表达，同时观察肝组织病理及肝功能变化。结果缺血／再灌注组移植肝再灌注后2h，枯否细胞NF-κB活性及TNF-α、IL-6 mRNA表达量较对照组明显升高（P〈0．01）。光镜下肝细胞大量变性、坏死，伴有肝血窦明显淤血，血清丙氨酸转氨酶（ALT）和胆红素总量（TBIL）较对照组明显升高（P〈0．01）。相反，圈套寡核苷酸组枯否细胞NF-κB活性及细胞因子mRNA表达与缺血／再灌注组相比明显下降（P〈0．01），移植肝未见明显病理组织学改变，肝功能明显改善。结论NF-κB圈套寡核苷酸能高效抑制枯否细胞NF-κB活性，并抑制其下游有害细胞因子的产生，从而减轻缺血／再灌注损伤对移植肝的打击和损害。     

小鼠肝缺血/再灌注损伤时肝脏Kupffer细胞中TLR2的表达

目的　观察小鼠肝部分缺血 /再灌注损伤时肝脏Kupffer细胞表面TLR2mRNA的表达及蛋白质的合成情况。方法　制作小鼠肝部分缺血 /再灌注损伤模型。采用原位灌注消化法分离并纯化Kupffer细胞 ,用大鼠抗小鼠TLR 2IgG和异硫氰酸荧光素 (FITC )的二抗进行染色 ,流式细胞仪 (FCM )测定阳性细胞数 ;并测定Kupffer细胞的TLR2mRNA含量。结果　缺血 60min ,再灌注 4h时 ,实验组小鼠肝脏Kupffer细胞表面TLR2mRNA表达及蛋白的合成量均明显高于假手术组 ,且缺血叶高于非缺血叶。结论　小鼠肝缺血 /再灌注损伤时 ,肝脏Kupffer细胞表面的TLR2mRNA表达明显增高 ,其蛋白质的水平也明显升高     

活体/离体门静脉灌注转染IRAK-4-shRNA对大鼠移植肝脏再灌注损伤的影响

目的通过观察活体或冷保存期离体门静脉灌注供肝转染白细胞介素-1受体相关激酶-4（IRAK-4）特异短发夹RNA（shRNA）对再灌注后受体TNF-α产生的影响,判断以IRAK-4为肝移植缺血再灌注损害（I/RI）治疗靶点的可行性并探索可行的shRNA治疗途径.方法雄性SD大鼠,随机分为冷缺血转染组、活体转染组、对照组,以两袖套法建立同种异体肝移植模型.冷缺血转染组于冷缺血期经门静脉灌注转染携带染IRAK-4-shRNA的转染质粒pSIIRAK-4;活体转染组在门静脉袖套吻合完成后,经门静脉分支注入pSIIRAK-4;对照组不予任何处理.按门静脉血流恢复后第0 min、60 min及180 min分为三个亚组,逆转录-聚合酶链式反应及蛋白免疫印记法测定肝组织的染IRAK-4 mRNA和蛋白表达水平;酶连免疫吸附法检测肝组织NF-κB活性及血清TNF-α含量.结果再灌注后活体转染组、对照组的IRAK-4蛋白与mRNA表达水平、NF-κB活性以及TNF-α含量均高于冷缺血转染组（P＜0.01）;冷缺血转染组的IRAK-4表达明显抑制,再灌注后各时点差异无显著性（P＞0.05）.结论以IRAK-4为靶点的冷缺血shRNAs转染途径能有效减轻肝移植时的I/RI,但能否以IRAK-4作为预防其他器官的I/RI的治疗靶点尚需深入研究.     

七氟醚或缺血预处理对大鼠肺缺血再灌注时ERK和CaM表达的影响

目的 探讨七氟醚或缺血预处理对大鼠肺缺血再灌注时细胞外信号调节蛋白激酶(ERK)和钙调素(CaM)表达的影响.方法 健康雄性SD大鼠24只,体重270～320 g,随机分为4组(n=6):假手术组(S组)、肺缺血再灌注组(IR组)、缺血预处理组(IP组)和七氟醚预处理组(SP组).IR组采用夹闭左肺门45 min恢复灌注120 min的方法制备肺缺血再灌注损伤模型,IP组缺血前夹闭左肺门缺血5 min恢复灌注5 min,连续2次,SP组缺血前吸人2.1%七氟醚30 min.于再灌注120 min时取左肺组织,测定TNF-α和IL-6含量、ERK mRNA和CaM mRNA的表达水平.结果 与S组比较,IR组、IP组和SP组肺组织TNF-α和IL-6的含量、ERK mRNA和CaM mRNA的表达水平升高(P＜0.05);与IR组比较,IP组和SP组肺组织TNF-α和IL-6的含量和CaM mRNA的表达水平降低,ERK mRNA表达水平升高(P＜0.05);SP组和IP组各指标比较差异无统计学意义(P＞0.05).结论七氟醚预处理和缺血预处理均通过下调CaM表达和上调ERK表达减轻大鼠肺缺血再灌注损伤.     

银杏叶提取物预处理对大鼠移植肝的保护作用

目的: 探讨银杏叶提取物（EGb）预处理对大鼠移植肝的保护作用。方法:采用Kamada′s 袖套法建立大鼠原位肝移植模型。将大鼠随机分为银杏叶提取物预处理（EGb）组、生理盐水对照(NS)组和假手术组(SO)。分别于供肝再灌注后2，6，24h处死动物，检测血清ALT和AST；肝组织组织学检查，TUNEL法检测细胞凋亡；RT-PCR检测肝组织TNF-αmRNA及Bcl-2mRNA的表达。结果:供肝再灌注2，6，24h，EGb组血清ALT水平及细胞凋亡指数均明显低于生NS组（P<0.01）。血清AST水平在供肝再灌注2，6h时明显低于NS组（P<0.01）。供肝再灌注后2，6h时EGb组TNF-αmRNA的表达明显低NS组（P<0.05）。供肝再灌注后2，6，24hEGb组Bcl-2mRNA的表达明显高于NS组（P<0.01）。结论:经EGb预处理供可减轻大鼠肝移植供肝的缺血/再灌注损伤和细胞凋亡，影响TNF-αmRNA,Bcl-2mRNA的表达，对供肝有保护作用。     

大鼠肝缺血再灌注损伤NF-kB与ICAM-1在脑组织的表达及意义

目的：观察大鼠在肝脏缺血再灌注（IR）过程中肝脏炎性细胞因子TNF-α、IL-1βmRNA表达与脑组织NF-kB、ICAM-1表达之间的关系，探讨肝脏IR是否可导致脑组织损伤及其可能的机制。方法：选健康雄性Wister大鼠48只，随机分为对照组、缺血30min组（I组）、缺血30min再灌注组（IR组）、缺血30min再灌注后1h组（IR1h组）、缺血30min再灌注后2h组（IR2h组）、缺血30min再灌注后4h组（IR4h组），每组8只。应用免疫组化法分别检测各组大鼠肝脏TNF-α及脑皮质、海马和下丘脑区NF-kB、ICAM-1的表达情况，应用原位杂交的方法检测肝脏IL-1βmRNA表达情况。结果：肝脏IR导致肝脏明显的损伤，表现为血清GPT、AKP、γ-GT升高（P〈0．01），肝组织中TNF-α、IL-1βmRNA表达显著增N（P〈0．01），肝细胞水肿，炎性细胞浸润，甚至细胞坏死。随着再灌注时间的延长，脑组织HE染色可见脑细胞水肿，甚至个别脑细胞坏死。与对照组比较，I组和IR组大鼠脑皮质、海马和下丘脑中NF-kB、ICAM-1表达的差异无统计学意义（P〉0．05），IR1h组、IR2h组和IR4h组的差异有统计学意义（P〈0．05，P〈0．01）。IR1h、IR2h、IR4h组与对照组、I组、IR组比较，各部位脑组织NF-kB、ICAM-1的表达显著增N（P〈0．05或P〈0．01）。各组大鼠不同部位脑组织间NF-kB、ICAM-1表达无统计学意义（P〉0．05）。结论：肝脏IR对脑组织可产生损伤性影响，NF-kB、ICAM-1介导的炎性细胞反应，参与了脑组织损伤的发生机制。     

2015年乳腺癌辅助化疗进展

【摘要】〓乳腺癌是危害我国女性健康的头号杀手,尽管近年来辅助化疗的研究进展突飞猛进,但临床中仍有不少问题未能明确,如辅助化疗的合适人群、化疗的开始时间、蒽环及紫杉类的地位和用法、强化维持治疗的作用、疗效及预后的生物标志物等。本文结合乳腺癌辅助化疗在临床上的常见问题和2015年各大乳腺癌会议阐述乳腺癌辅助化疗的最新进展。     

Alterations in T-Cell Subset Frequency in Peripheral Blood in Obesity

Background: Obesity affects the regulation of immune and inflammatory responses. This study characterizes differences in peripheral blood lymphocyte phenotype in obese humans. Methods: Frequencies of lymphocyte subsets among peripheral blood mononuclear cells were compared between 10 obese (BMI ≥35) and 10 lean subjects, as determined by antibodies directed against cluster differentiation (CD) markers. Results: Obese patients demonstrated an increased frequency of CD3+CD4+ T-cells (mean difference 12%, P=0.004), a decreased frequency of CD3+CD8+ T-cells (mean difference 9.4%, P=0.016) and an increased frequency of CD3+CD8+CD95+ T-cells (mean difference 13.3%, P=0.032). No other differences among T-cell or monocyte subsets were noted. Conclusions: Obesity is associated with alterations in frequencies of peripheral CD4+ and CD8+ T-cells and aberrations in the expression of CD95 among CD8+ T-cells. These data suggest both CD4+ and CD8+ T-cell compartments, as well as the regulation of CD95 expression on CD8+ T-cells, as targets for further study into obesity's effects on the immune system.     

西宁地区烧伤病人动脉血气分析观察

对高海拔地区的27例烧伤病人动脉血气变化进行了分析和观察。结果证明:无论是存活病人还是死亡病人伤后均存在有低氧血症问题。并且在死亡病人和烧伤合并吸入性损伤病人其低氧血症的发生早于单纯烧伤病人。提示:吸入性损伤病人应立即行气管切开术以保障氧气供给,单纯烧伤病人可常规吸氧以维持正常血 PaO_2,ARDS 均发生在合并吸入性损伤的病人,高频喷射通气技术对纠正低氧血症有一定效果。     

Controversies in Fistula in Ano

Managing a complex fistula in ano can be a daunting task for most surgeons; largely due to the two major dreaded complications—recurrence & fecal incontinence. It is important to understand the anatomy of the anal sphincters & the aetiopathological process of the disease to provide better patient care. There are quite a few controversies associated with fistula in ano & its management, which compound the difficulty in treating fistula in ano. This article attempts to clear some of those major controversies.     

β-半乳糖苷酶在体内外成骨细胞中的表达

目的 研究β—半乳糖苷酶(β—gal)在成骨细胞中的表达状况，为阐明MorquioB综合征的发病机制提供依据。方法 裸鼠各器官和骨组织标本行X-gal染色检测。抽取羊和人骨髓行骨髓基质细胞(BMSCs)培养，分为4组：I：Adv-hBMP-2转染组；Ⅱ：Adv—β—gal转染组；Ⅲ：未转染组；Ⅳ：地塞米松诱导组。分别行X-gal染色和RT-PCR检测β—gal的表达。结果 裸鼠骺板两侧、骨膜内面及松质骨的成骨细胞和破骨细胞可见多量β—gal的表达。未转染BMSCs组有少量β—gal的表达，其他3组细胞的β—gal表达增高。结论成骨细胞和破骨细胞可表达多量β—gal，该两种细胞的β—gal缺乏可能是MorquioB综合征骨骼异常的直接原因。     

Fluid-phase transcytosis in the primate epididymis in vitro and in vivo

Ligated tubules from the corpus epididymidis of men and monkeys were incubated in medium containing horseradish peroxidase (HRP) as a marker for fluid-phase endocytosis. HRP was localized by light and electron microscopy after 0, 15, 30 and 60 min of incubation. Movement between the cells was prevented by tight junctions, but bypass of this barrier was apparently achieved by an intracellular vesicular mechanism leading to a time-dependent appearance of HRP in the lumen. Uptake of HRP into basal cells and capture by the lysosomal apparatus of principal cells were also observed. HRP-filled vesicles also appeared in the basal, mid and apical cytoplasm of epithelial cells in the caput 1 h after injection of the tracer into the epididymal circulation of the monkey, suggesting that this pathway also operates in vivo.     

Surgical management of ductal carcinoma in situ in Australia in 1995

Background: In the present paper we describe the presentation and management of ductal carcinoma in situ (DCIS) of the breast in women in Australia in 1995. This representative, national data set provides a historical comparator for studies examining DCIS management that follow. Methods: Surgeons identified by population‐based cancer registries as having treated a new diagnosis of DCIS between 1 April and 30 September 1995 completed a questionnaire on the presentation and management of each case. Results: Two hundred and five surgeons supplied treatment details on 418 DCIS tumours in 415 women . Half of all tumours were detected at BreastScreen clinics and a further 25% were detected at other mammography centres. Twenty‐six percent of tumours were palpable at presentation, 33% were multifocal and 55% were high grade (including comedocarcinoma). Breast conserving therapy (BCT) rather than mastectomy was utilized in 260 (62%) of cases. Tumours that were of low grade, small in size and not multifocal were more likely to be treated by BCT. Surgeons seeing six or more DCIS cases in the 6‐month period were more likely to utilize BCT. Of the conservatively treated cases, 22% were referred for a radiation oncology consultation. The most common reasons for treating DCIS with mastectomy were that the tumour was too extensive or multifocal (63%), it extended to margins of the specimen (42%), or patient concerns about recurrence (34%). Conclusions: In 1995 the majority of DCIS was treated with breast conserving surgery alone. Surgeons treating more DCIS cases were more likely to perform conservative surgery than surgeons treating only one DCIS case in the study period.     

Smoking-Attributable mortality in Spain in 2016

IntroductionSmoking-attributable mortality (SAM) is a valuable indicator that can be used to characterize the course and health burden of the smoking epidemic. The aim of this paper was to estimate SAM in Spain in 2016 in the population aged 35 and over, using the best available evidence.MethodsA smoking prevalence-dependent analysis based on the estimation of population-attributable fractions was performed. Smoking prevalence (never, former, and current smokers) was calculated from a combination of the Spanish Health Survey (2016) and the European Health Survey (2014); the relative risk of death among current and former smokers was taken from the follow-up of various cohorts; and mortality rates were obtained from National Center for Statistics data. SAM estimates are presented globally, and by sex, age groups, and major disease categories: cancer, cardiometabolic diseases and respiratory diseases.ResultsIn 2016, 56,124 deaths were attributed to tobacco consumption, 84% in men (47,000), and 50% in the population aged over 74 (27,795). Overall, 50% of SAM was due to cancer (28,281), 65% of which was lung cancer. One in 4 attributable deaths (13,849) occurred before the age of 65.ConclusionsOne in 7 deaths in Spain in 2016 were attributable to smoking. This estimation of SAM clearly highlights the great impact of smoking on mortality in Spain, mainly due to lung cancer and chronic obstructive pulmonary disease.