紫草多糖抑制LPS活化PBMC表达炎症相关因子的基因转录

目的:研究紫草多糖对LPS活化外周血单个核细胞(Peripheral blood mononuclear cells,PBMC)表达TNF-α、IL-10、IL-18相关因子的影响及其可能的信号通路。方法:LymphopreTM(1.077 g/ml)淋巴细胞分离液离心分离正常健康人PBMC,紫草多糖处理正常健康人的PBMC或LPS活化的PBMC,利用Real-time RT-PCR法检测TNF-α、IL-10、IL-18、IL-18BP、IL-18Rα、IL-18Rβ基因转录水平,通过Western blot检测PBMC中ERK磷酸化水平。结果:紫草多糖可增强PBMC TNF-α、IL-10、IL-18Rα、IL-18Rβ的转录,与LPS处理的PBMC相近。紫草多糖处理LPS活化的PBMC,在处理早期(4小时)可抑制LPS活化的PBMCTNF-α及IL-18的表达,抑制率达15%~50%(P<0.05);并可抑制LPS诱导的ERK磷酸化,抑制率达27%~61%(P<0.05)。结论:紫草多糖可能是通过抑制PBMC的MAPK信号通路进而抑制LPS诱导的炎症相关因子TNF-α等高表达。     

首发精神分裂症PBMCNF-κB活性及其mRNA表达与IL-1β、IL-6、TNF-αmRNA表达的关系

目的：探讨首发精神分裂症患者外周血单个核细胞（PBMC）NF-κB的活性及其mRNA表达与IL-β、IL-6、TNF-α mRNA表达的相互关系，为临床应用NF-κB调节剂提供理论依据。方法：应用NF-κB活性ELISA试剂盒，检测精神分裂症组和健康对照组PBMC中NF-κB活性，并采用RT-PCR方法，测定两组PBMC中NF-κB mRNA表达及IL-1β、IL-6、TNF-α mRNA表达。结果：①精神分裂症组PBMC中NF-κB的活性及NF-κB mRNA表达量与对照组相比均明显增高，差异有显著性（P〈0．05）；②精神分裂症组PBMC中IL-1β、IL-6、TNF-α mRNA表达量与对照组相比均明显增高，差异有显著性（均P〈0．05）；③在精神分裂症组及对照组中，PBMC中NF-κB的活性与其mRNA表达量无明显相关（均P〉0．05）；④在精神分裂症组和对照组中，PBMC中NF-κB的活性与IL-1β、TNF-α mRNA均呈正相关（均P〈0．05）；无论是在精神分裂症组或对照组中，PBMC中NF-κB的活性与IL-6 mRNA均无明显相关（均P〉0．05）。结论：精神分裂症患者PBMC中NF-κB mRNA表达及活性均增高，对NF-κB活性的检测更能反映NF-κB的转录调控功能情况；活化的NF-κB对IL-1β、TNF-α的基因转录起了重要的调控作用。     

下调miR-217 抑制类风湿性关节炎患者外周血单个核细胞炎症反应的实验研究

目的:探讨下调miR-217表达对类风湿性关节炎(RA)患者外周血单个核细胞(PBMC)炎症反应的影响及作用机制。方法:分离培养RA患者PBMC细胞,脂多糖(LPS)诱导PBMC细胞24 h,ELISA法检测炎症因子TNF-α、IL-1β、IL-6和IL-10的表达,qRT-PCR检测miR-217和Sirt1 mRNA表达,Western blot检测Sirt1蛋白表达。分别转染anti-miR-217和pcDNA3.1-Sirt1至PBMC细胞,构建miR-217低表达、Sirt1过表达的PBMC细胞,ELISA法检测PBMC细胞分泌TNF-α、IL-1β、IL-6和IL-10的变化。双荧光素酶报告基因验证miR-217与Sirt1的关系。结果:LPS诱导PBMC细胞24 h后,TNF-α、IL-1β和IL-6水平升高,IL-10水平降低。miR-217在LPS诱导的PBMC细胞中表达上调,Sirt1 mRNA和蛋白表达下调。下调miR-217表达或上调Sirt1表达均可抑制PBMC细胞TNF-α、IL-1β和IL-6表达,促进IL-10表达。miR-217在PBMC细胞中负向调控Sirt1表达,上调miR-217表达抑制了Sirt1过表达对LPS诱导的PBMC细胞中炎症因子表达的影响。结论:下调miR-217可能通过负向调控Sirt1表达抑制RA患者PBMC细胞炎症反应,可作为治疗RA的新靶点。     

过表达SIRT1抑制脂多糖诱导的胰岛β细胞炎症因子表达及NF-κB信号通路激活

目的研究沉默信息调节因子1(SIRT1)对脂多糖(LPS)诱导的胰岛β细胞炎症因子表达及核因子-κB(NF-κB)信号通路激活的影响。方法用LPS处理胰岛β细胞,qRT-PCR和Western blot测定细胞中SIRT1表达变化。用pcDNA3.1-SIRT1慢病毒感染胰岛β细胞,qRT-PCR检测细胞中白细胞介素-6(IL-6)mRNA和肿瘤坏死因子-α(TNF-α)mRNA表达水平,流式细胞术测定细胞凋亡变化,Western blot检测细胞中Bcl-2相关X蛋白(Bax)、活化型Caspase-3(cleaved Caspase-3)、核因子-κBp65亚型(NF-κBp65)、Toll样受体4(TLR4)蛋白表达水平。结果 LPS处理后的胰岛β细胞中SIRT1表达水平降低。pcDNA3.1-SIRT1慢病毒感染可以提高LPS条件下胰岛β细胞中SIRT1表达水平。LPS处理后的胰岛β细胞中IL-6 mRNA和TNF-αmRNA表达水平升高,凋亡率升高,细胞中Bax、Cleaved Caspase-3、NF-κBp65、TLR4蛋白表达升高。过表达SIRT1可以降低LPS条件下胰岛β细胞中IL-6 mRNA和TNF-αmRNA表达水平,抑制细胞凋亡,减少细胞中Bax、cleaved Caspase-3、NF-κBp65、TLR4蛋白表达。结论 SIRT1减少LPS诱导的胰岛β细胞炎症因子表达并下调细胞中NF-κB信号激活水平。     

内皮细胞在感染性休克炎症性细胞因子释放中的作用

目的 以感染性休克患者血清刺激人原代内皮细胞(HPAEC)和外周血单个核细胞(PBMC),观察内皮细胞在感染性休克细胞因子释放中的作用.方法 采用密度梯度离心法分离健康人PBMC;RT-PCR及免疫印迹检测HPAEC细胞表面标志CD144和血管假性血友病因子(von Will-ebrand factor,vWF)的表达;ELISA检测感染性休克患者和健康对照者血清中的IL-6、TNF-α、MCP-1的水平;两种血清及LPS刺激HPAEC和PBMC后,ELISA检测培养上清中IL-6、TNF-α、MCP-1的水平;流式细胞技术检测休克患者血清和LPS刺激后HPAEC膜表面ICAM-1的表达;S1P1受体激动剂CYM-5442预处理后,休克血清刺激HPAEC培养上清中IL-6、TNF-α、MCP-1的水平.结果 HPAEC表达内皮细胞标志性分子CD144和vWF;感染性休克患者血清中炎症因子IL-6、TNF-α、MCP-1水平明显高于健康对照(P＜0.01);休克血清和LPS处理PBMC和HPAEC后,相对于对照组,均有明显升高(P＜0.01);PBMC的休克组炎症因子与PBMC的LPS组均无明显差异(P＞0.05),而HPAEC的休克组炎症因子相对于HPAEC的LPS组均明显升高(P＜0.01);同样地,LPS刺激两种细胞后,HPAEC的IL-6、TNF-α明显低于PBMC(P＜0.01),MCP-1的水平无明显变化(P＞0.05),但是休克血清刺激后,HPAEC的IL-6、TNF-α与PBMC无明显变化(P＞0.05),MCP-1则明显升高(P＜0.01);休克患者血清刺激S1P1受体特异性激动剂CYM-5442预处理的HPAEC后,培养上清中炎症因子IL-6、TNF-α、MCP-1水平明显降低(P＜0.01).结论 内皮细胞在感染性休克细胞因子释放中起着关键作用.     

卡氏肺大鼠肺泡巨噬细胞TNF-α、IL-1β、IL-6基因表达的变化

目的探讨卡氏肺孢子虫肺炎(Pneumocysitis carinii pneumonia,PCP)大鼠肺泡巨噬细胞(Alveolar macrophage,AM)TNF-α、IL-1β、IL-6基因表达的变化。方法采用AM体外培养技术,应用RT-PCR法分别测定脂多糖(LPS)诱导的AM中TNF-α、IL-1β、IL-6基因表达的动态变化。结果肺泡巨噬细胞受LPS刺激后,PCP模型大鼠TNF-αmRNA在1、4 h表达高于正常组(P<0.05),IL-1βmRNA表达在4、8 h时表达高于正常组(P<0.01),IL-6mRNA表达在8 h时PCP高于正常(P<0.05),表达峰值都提前,并且在4 h达到峰值。结论LPS刺激后,PCP大鼠肺泡巨噬细胞在早期可能更易分泌TNF-α、IL-1β、IL-6作为免疫分子,起免疫防御和免疫损伤作用。     

下调TSPO表达不影响脂多糖诱导的小胶质细胞TNF-α和IL-1β以及IL-6的分泌

目的研究相对分子质量(Mr)18 000转运蛋白(TSPO)基因表达下调对脂多糖(LPS)诱导BV-2小胶质细胞分泌TNF-α,IL-1β和IL-6的影响。方法以RNA干扰技术建立TSPO基因表达下调的细胞模型,实时荧光定量PCR(qRT-PCR)和Western blot法检测转染TSPO siRNA BV-2细胞TSPO mRNA及蛋白水平表达的效果;用qRT-PCR法检测TSPO基因下调后小胶质细胞BV-2在LPS作用下分泌TNF-α、IL-1β和IL-6的mRNA水平表达的情况;ELISA检测TSPO基因下调小胶质细胞BV-2在LPS作用下分泌TNF-α、IL-1β和IL-6的蛋白水平表达的变化。结果成功建立了TSPO基因下调的细胞模型,稳定表达TSPO siRNA细胞的TSPO mRNA和蛋白水平表达均明显下降,TSPO基因下调后BV-2细胞在LPS作用下分泌TNF-α、IL-1β和IL-6的量无变化。结论下调TSPO的表达对LPS刺激引起的小胶质细胞TNF-α、IL-1β和IL-6的分泌无明显影响。     

诱骗受体DcR3对佐剂型关节炎大鼠模型的作用分析

目的 研究诱骗受体DcR3 对佐剂型关节炎(AA)大鼠模型的作用及其机理.方法 注射弗式完全佐剂建立大鼠佐剂型关节炎(AA)模型,尾静脉注射DcR3蛋白,观察大鼠关节肿胀度、间接 ELISA 检测血清和滑膜液中细胞因子IL-1 β、TNF-α、IFN-γ的变化.RT-PCR检测滑膜和淋巴细胞中DcR3、Fas、FasL mRNA的表达以及脾脏中TGF-β、IFN-γ、TNF-α、IL-4、IL-10 mRNA的表达.Western blot分析滑膜细胞中Caspase-8、Caspase-3、Caspase-9、Bcl-2蛋白的表达.结果 DcR3 治疗AA大鼠后,足肿胀度降低;血液和滑膜液的IL-1β、TNF-α、IFN-γ水平下降;脾脏中TGF-β、IFN-γ、TNF-α mRNA 表达下调,IL-4、IL-10 mRNA表达上调;滑膜细胞中Caspase-8、Caspase-3蛋白表达上调,Bcl-2蛋白表达下调. 结论 DcR3 可以用于实验性大鼠AA的治疗,其治疗机制与调节滑膜细胞FasL、Fas mRNA的表达和血液淋巴细胞中 Fas mRNA的表达,促进滑膜细胞和自身反应性淋巴细胞的凋亡;调节脾脏细胞Th1/Th2细胞因子平衡相关.本研究为进一步阐明RA的发病机理奠定了重要基础,为有效治疗RA提供了新思路.     

天抗(TK)对脂多糖诱导小鼠炎症模型的抗炎作用及其机制

目的研究天抗(TK)对脂多糖(LPS)诱导的小鼠炎症模型的抗炎作用及机制研究。方法将42只昆明小鼠随机分为正常对照(NC)组、模型对照(LPS)组、地塞米松(DXM)组、天抗低(TK-L)、中(TK-M)和高(TK-H)剂量组(0.2,0.8和3.2 g/kg)。各组分别灌胃给药7 d后,腹腔注射30 mg/kg的LPS诱导小鼠急性炎性模型,6 h后处死小鼠,检测小鼠脾脏指数,ELISA测定小鼠血清中IL-1β、IL-6和TNF-α的表达水平;生化法检测小鼠血清中SOD和MDA的表达;qRT-PCR检测小鼠脾脏TLR4、MyD88、TRAF6、p65、IL-1β、IL-6和TNF-αmRNA的表达水平;Western blot检测小鼠脾脏TLR4、MyD88、TRAF6、p-p65和p65蛋白表达水平。结果与LPS组相比,TK组小鼠的脾脏指数明显降低,血清和脾脏组织中IL-1β、IL-6、TNF-α和MDA水平显著下降,SOD水平明显升高,小鼠脾脏组织的TLR4、MyD88、TRAF6和p-p65等蛋白及mRNA表达水平均明显降低。结论天抗对LPS诱导的小鼠急性炎症模型具有抗炎作用,其作用机制可能是通过TLR4/MyD88/NF-κB(p-65)信号通路抑制炎症因子的释放。     

miRNA-130a-3p通过调控NF-κB信号通路影响损伤心肌细胞

目的 探讨miRNA-130a-3p在LPS诱导的心肌细胞损伤中的作用及可能机制。方法H9C2心肌细胞分为4组,即正常对照组、LPS模型组、miRNA阴性对照组、miRNA-130a-3p mimics组,利用RT-qPCR检测miRNA-130a-3p mRNA表达水平,CCK-8检测细胞活性,ELISA检测培养液中TNF-α、IL-6、IL-1β含量,Western blot检测NF-κBp65、IκBα、Bax、Bcl-2、Cleaved-Caspase-3蛋白表达水平。结果RT-qPCR结果显示LPS模型细胞中miRNA-130a-3p mRNA水平显著低于正常对照组;与正常对照组相比较,LPS组细胞活性显著降低,而与LPS组相比较,miRNA-130a-3p mimics组细胞活性显著升高;与正常对照组相比较,LPS组细胞培养液中TNF-α、IL-6、IL-1β含量显著升高,而与LPS组相比较,miRNA-130a-3p mimics组细胞培养液中TNF-α、IL-6、IL-1β含量显著降低;与正常对照组相比较,LPS组细胞中NF-κBp65、Bax、Cleaved-Casp...     

Renal dysplasia and asplenia in two sibs

A family is reported in which two sibs, one male and the other female, both died within 24 hours of birth with enlarged polycystic kidneys. Postmortem histology in the second child showed gross renal dysplasia. In both children the pancreas was enlarged, nodular and cystic but the liver appeared macroscopically normal. In the second child, histological examination confirmed pancreatic fibrosis with cystic dilation of ducts, but showed portal fibrosis with bile duct proliferation in the liver.
This combination of findings is very reminiscent of those in a girl and her brother reported by Ivemark et al. (1959). The children reported here also showed absence or hypoplasia of the spleen, cardiac anomalies and other features of the Ivemark syndrome (Ivemark 1955), a quite different, usually sporadic, congenital disorder. It is suggested that the children described here have a distinct lethal congenital disorder, probably inherited in an autosomal recessive manner.     

Schizophrenia in a North Swedish geographical isolate, 1900–1977. Epidemiology, genetics and biochemistry

Over 200 schizophrenic patients belonging to three major and interrelated pedigree complexes have been investigated over the past 30 years in a North Swedish geographically isolated population, presently numbering about 6,000. An intensive investigation of a number of biochemical correlates and genetic markers in a few selected families belonging to one of the major pedigrees has indicated new strategies for the current research program.
Schizophrenia, as defined operationally, is significantly associated with decreased activities of two enzymes (1) blood platelet monoamine oxidase, (2) plasma dopamine-β-hydroxylase, and (3) with the genetic marker Gc² (group specific antigen). Both enzymes are subject to genetic variation. A positive score for linkage between schizophrenia and low plasma DBH activity has been calculated, but, so far, available data are insufficient for discrimination between linkage and partial contribution of genetically controlled low plasma DBH to the pathogenesis of the disease. Alternatively, both mechanisms could be involved.
As a model for continued research, schizophrenia is explained as based on a double dominant-recessive genotype (Aabb), representing a vulnerability which in about 50 % of cases develops into clinical schizophrenia. It is suggested that the dominant mutation (A) operates on or affects MAO activity, and that the recessive genotype (bb) is instrumental in low variates of DBH activity and very likely such variates within the normal range of physiological variation. Moreover, it is suggested that the combined effects of MAO- and DBH-reduced efficiency on the metabolism of e.g. dopamine could be an essential pathogenic mechanism for the schizophrenic illness which is segregating in this population.     

The dominant diseases of modernized societies as omega-3 essential fatty acid deficiency syndrome: Substrate beriberi

About 1900, modern food selection and processing caused widespread $\text{epidemics}$ of the B vitamin deficiency diseases of beriberi and pellagra which, for genetic reasons, often expressed as different diseases ranging from bowel and heart disease to dermatoses and psychoses. But the B vitamins merely help convert essential fatty acids (EFA) into the prostaglandin (PG) tissue regulators and it now turns out that, through hydrogenation, milling and selection of w3-poor southern foods, we have also been systematically depleting, by as much as 90%, a newly discovered trace Nordic EFA (w3) of special importance to primates and sole precursor of the PG3(4) series, even as a concurrent fiber deficiency increases body demand for EFA. Since substrate EFA is processed by many B vitamin catalysts, an EFA deficiency will mimic a $\text{panh}$ypovitaminosis B, i.e., a mixture of $\text{substrate}$ beriberi and $\text{substrate}$ pellagra resembling vitamin beriberi and pellagra but exhibiting as even more diverse $\text{endemic}$ disease. This would consitute a second stage of the Modern Malnutrition and explain why some workers now hold the dominant diseases of modermized societies to be new, nutritionally based, pellagraform yet lipid-related and to range, once again, from heart disease to psychosis. It is an assumption that our dominant diseases are unrelated to each other or are merely revealed by our diagnostic acumen and therapeutic success; and that hydrogenating millions of tons of food oils annually, to destroy the rancidity producing w3-EFA, is safe for $\text{primates}$. Extensive beriberiform disease is reported here in 32 typical cases taken from medical practice which responds strikingly to linseed oil supplements (60% w3-EFA) in confirmation of identical results in Capuchins.     

What will studies of Fulani individuals naturally exposed to malaria teach us about protective immunity to malaria?

There are an estimated over 200 million yearly cases of malaria worldwide. Despite concerted international effort to combat the disease, it still causes approximately half a million deaths every year, the majority of which are young children with Plasmodium falciparum infection in sub-Saharan Africa. Successes are largely attributed to malaria prevention strategies, such as insecticide-treated mosquito nets and indoor spraying, as well as improved access to existing treatments. One important hurdle to new approaches for the treatment and prevention of malaria is our limited understanding of the biology of Plasmodium infection and its complex interaction with the immune system of its human host. Therefore, the elimination of malaria in Africa not only relies on existing tools to reduce malaria burden, but also requires fundamental research to develop innovative approaches. Here, we summarize our discoveries from investigations of ethnic groups of West Africa who have different susceptibility to malaria.     

'Very sore nights and days': the child's experience of illness in early modern England, c.1580-1720

Sick children were ubiquitous in early modern England, and yet they have received very little attention from historians. Taking the elusive perspective of the child, this article explores the physical, emotional, and spiritual experience of illness in England between approximately 1580 and 1720. What was it like being ill and suffering pain? How did the young respond emotionally to the anticipation of death? It is argued that children’s experiences were characterised by profound ambivalence: illness could be terrifying and distressing, but also a source of emotional and spiritual fulfilment and joy. This interpretation challenges the common assumption amongst medical historians that the experiences of early modern patients were utterly miserable. It also sheds light on children’s emotional feelings for their parents, a subject often overlooked in the historiography of childhood. The primary sources used in this article include diaries, autobiographies, letters, the biographies of pious children, printed possession cases, doctors’ casebooks, and theological treatises concerning the afterlife.     

Guidelines for the Validation and Use of Immunoassays for Determination of Introduced Proteins in Biotechnology Enhanced Crops and Derived Food Ingredients

Recent advancements in agricultural biotechnology have created a need for analytical techniques to determine introduced proteins in crops enhanced through modern biotechnology techniques. These proteins are expressed in plant tissues and may be present in food ingredients. Immunoassays are ideally suited for protein detection and may be used as both quantitative and threshold methods. Microplate ELISA and lateral flow devices are two of the most commonly used immunoassay formats for agricultural biotechnology applications. This paper provides general background information and a discussion of criteria for the validation and application of immunochemical methods to the analysis of proteins introduced into plants and food ingredients using biotechnology methods. It is the result of a collaborative effort of members of the Analytical Environmental Immunochemical Consortium. This collaborative effort represents the combined expertise of several organizations to reach consensus on establishing guidelines for the validation and use of immunoassays. Further, the paper offers developers and users a consistent approach to adopting the technology as well as aid in producing accurate and meaningful results.     

Ultracryotomy: development and application (with examples from the yeast cytology) (author's transl)

The preparation steps usually necessary for obtaining ultrathin frozen sections of biological material (chemical prefixation, enclosing, cryoprotective treatment, freezing, sectioning, and post-staining the sections for transmission electron microscopy) are submitted to a critical analysis. The application of cryo-ultramicrotomy, in particularly for cytochemical purposes, is reviewed. Fundamental considerations of chemical prefixation and poststaining are supported by examples from yeast cytology. Furthermore, the efficiency of the cryo-ultramicrotomy (electron optical resolution of ultrastructural details) is demonstrated on yeast cells and protoplasts.     

HLA in North Colombia Chimila Amerindians

HLA-A,-B,-C,-DRB1 and -DQB1 alleles have been studied in Chimila Amerindians from Sabana de San Angel (North Colombian Coast) by using high resolution molecular typing. A frequent extended haplotype was found:HLA-A*24:02-B*51:10-C*15:02-BRB1*04:07-DQB1*03:02 (28.7%) which has also been described in Amerinndian Mayos Mexican population (Mexico, California Gulf, Pacific Ocean). Other haplotypes had already been found in Amerindians from Mexico (Pacific and Atlantic Coast), Peru (highlands and Amazon Basin), Bolivia and North USA. A geographic pattern according to HLA allele or haplotype frequencies is lacking in Amerindians, as already known. Also, five new extended haplotypes were found in Chimila Amerindians. Their HLA-A*24:02 high frequencies characteristic is shared with aboriginal populations of Taiwan; also, HLA-C*01:02 high frequencies are found in New Zealand Maoris, New Caledonians and Kimberly Aborigines from Australia. Finally, this study may show a model of evolutionary factors acting and rising one HLA allele frequency (-A*24:02), but not in others that belong to the same or different HLA loci.     

The universal muscular chamber. A basic unit of the genitourinary, cardiovascular, gastro-intestinal, skeletal, cutaneous, respiratory and other systems, endocameral hypertension; and spasm, the key to their idiopathic diseases and arthropathies

Starting with the integument, we see many organs are contractile sacs or multiples thereof, which tubes or bags constitute the major part of the entire body. Recognition of this basic unit and its characteristics sheds new light, individually and collectively, on many disorders previously considered unrelated. Muscular tears and perforations develop in the walls of these chambers, being no way peculiar to those organs, wherein, hydrochloric acid occurs. So, it is not necessary to explain the absence of excessive acid from patients who exhibit holes in the gastric, uterine, aortic, duodenal, rectal, pulmonary, retina, and other walls. Muscle, not acid is the great common factor relating idiopathic disorders in the gastrointestinal tract to each other and to similar diseases in other systems. When the units are linked together, the lesions tend to appear as arthropathies, i.e. at the joints. Rephrasing common-place observations, frees us from conventional, conceptual cul-de-sacs. An observation is only as good as its interpretation, so all possibilities must be considered, otherwise, we will remain blinded by our misconceptions.