IL-12对BALB/c小鼠Th17细胞的分化的影响

目的: 观察细胞因子IL-12对Th17细胞分化的影响.方法: 小鼠脾淋巴细胞经抗CD3单克隆抗体(mAb)和不同浓度的重组小鼠IL-12刺激, 3 d后使用ELISA方法观察培养物上清液中IL-17的产生情况.并使用细胞内细胞因子染色的方法, 通过流式细胞术观察CD3 mAb和重组小鼠IL-12刺激对Th1和Th17细胞分化的影响.结果: Th17细胞不分泌IFN-γ、 IL-5、 IL-10等细胞因子, 不表达Foxp3, 是一个独立的细胞亚群.不同浓度的重组小鼠IL-12可以诱导抗CD3 mAb 的T细胞分泌IFN-γ, 并向Th1细胞方向分化.同时, IL-12可以抑制活化的T细胞分泌IL-17, 抑制T细胞向Th17细胞分化.结论: IL-12可以抑制Th17细胞的分化.     

慢性乙肝患者树突状细胞对Th1/Th2分化的影响

探讨慢性乙肝患者树突状细胞(dendritic cells,DC)对CD4+Th细胞亚群分化的影响。分离慢性乙肝患者外周血单个核细胞(PBMC),以rhIL-4(50 ng/ml)、rhGM-CSF(10 ng/ml)和rhTNF-α(100 u/ml)诱导培养DC。以流式细胞仪检测DC表面CD1a、CD83、CD80、CD86、HLA-DR分子表达情况。MTT法检测DC刺激同种异体淋巴细胞增殖能力。免疫磁珠分离外周血CD4+T细胞亚群,PMA+Ionomycin刺激后胞内荧光染色,流式细胞仪检测辅助性T细胞(helper T cell,Th)内特征性细胞因子IFN-γ/IL-4以判断Th1/Th2分化。ELISA法检测DC或Th细胞培养上清中IL-6、IL-12、IFN-γ和IL-4的含量。结果:慢性乙肝患者的DC表达CD1a、CD83、CD80、CD86、HLA-DR分子水平明显低于正常人(P<0.01);培养至第7天,慢性乙肝患者DC分泌的IL-12水平低于正常人(P<0.01),而分泌的IL-6水平增高(P<0.05)。与正常人相比,慢性乙肝患者外周血中Th1细胞占CD4+T细胞的百分比较低(P<0.01),其Th细胞培养上清中IFN-γ的量也较低(P<0.01)。患者DC与同种异体的健康人Th细胞共培养,刺激Th1型细胞因子IFN-γ产生的能力低于正常人(P<0.01)。慢性乙肝患者体内DC功能的异常可能导致了外周血Th1细胞分化不足。     

3-氧十二烷酰高丝氨酸内酯对淋巴细胞Toll样受体2和4的影响

目的:研究铜绿假单胞菌群体感应(QS)系统信号分子3-氧十二烷酰高丝氨酸内酯(3-O-C12-HSL)对人外周血淋巴细胞增殖、TLR2/4mRNA表达以及单个核细胞分泌TNF-α的影响。方法:自健康献血者血液中分离单个核细胞进行体外培养,并以0、1、10、50、100μmol/L3-O-C12-HSL作用12h,分离淋巴细胞,采用MTT法,观察不同浓度的3-O-C12-HSL分别作用对人淋巴细胞生长增殖的影响。用RT-PCR测定淋巴细胞TLR2/4mRNA表达。收集上清,以双抗体夹心酶联免疫吸附试验(ELISA)法测TNF-α。结果:3-O-C12-HSL刺激12h后淋巴细胞TLR2/4mRNA表达增高,且呈剂量依赖关系,TLR2/4mRNA在100μmol/L时最为显著(0.723±0.005,0.363±0.013),P0.05。TLR2变化较TLR4明显,P0.05。3-O-C12-HSL对淋巴细胞增殖有促进作用,呈剂量依赖关系,在100μmol/L时最为显著(0.463±0.052),P0.05。3-O-C12-HSL能抑制单个核细胞分泌TNF-α,且呈剂量依赖关系,在100μmol/L时最为显著(267.090±13.878),P0.05。结论:3-O-C12-HSL可以激发淋巴细胞细胞增殖,上调Toll样受体的表达水平,Toll样受体表达上调抑制TNF-α表达,获得性免疫对固有免疫有反馈作用。     

内吗啡肽1抑制人外周血来源树突状细胞成熟和功能

目的研究加入脂多糖(LPS)刺激后内吗啡肽1(EM-1)对外周血来源树突状细胞(PBDC)的表型、细胞因子分泌和刺激T细胞增殖等方面的影响,探讨EM-1调节DC功能的可能作用途径及机制。方法用重组人粒细胞巨噬细胞集落刺激因子(rh GM-CSF)和重组人白细胞介素4(rh IL-4)联合刺激经培养和离心处理后的PBDC,分组加入LPS或EM-1刺激,观察细胞生长形态变化;流式细胞术检测PBDC表面CD80、CD86、CD83、HLA-DR、CC趋化因子受体7(CCR7)的水平;ELISA检测PBDC分泌的IL-10、IL-12、γ干扰素(IFN-γ)的水平;羟基荧光素二乙酸盐琥珀酰亚胺脂(CFSE)染色检测EM-1诱导后PBDC对T细胞增殖的影响。结果 EM-1使LPS刺激诱导后PBDC表面CD80、CD86、CD83、HLA-DR、CCR7表达下调;EM-1抑制LPS诱导成熟的PBDC释放炎性因子IL-12和IL-10,抑制PBDC对T淋巴细胞的增殖反应能力,减少PBDC和T淋巴细胞共培养上清液中IL-12和IFN-γ的分泌。结论 EM-1可抑制PBDC的成熟和功能。     

树突状细胞上调MHC Ⅰ类相关抗原A表达对NK细胞活性的影响

目的:观察单核细胞、未成熟和成熟树突状细胞(DCs)表达MHC Ⅰ类相关抗原A(MICA)的情况,以及DCs表达MICA后对NK细胞活性的影响.方法:用流式细胞术分别检测单核细胞、未成熟DC(iDCs)以及LPS、TNF-α、CD40L、IL-15和IFN-α分别刺激的成熟DCs表面MICA的表达,并观察DCs表达MICA后对NK细胞表达CD69、细胞毒活性和分泌IFN-γ的影响.最后用流式细胞术检测重组NKG2D/Fc蛋白和抗IL-12单克隆抗体(mAb)对DCs激活NK细胞的影响.结果:单核细胞不表达MICA,iDCs低表达MICA.LPS、TNF-α和CD40L对DCs表达MICA无影响;但IFN-α和IL-15可促进DC上调MICA表达.DCs表达MICA后可促进NK细胞表达CD69、分泌IFN-γ、杀伤K562细胞.NKG2D/Fe蛋白可抑制NK细胞的杀伤活性和分泌IFN-γ;而抗IL-12mAb仅抑制IFN-γ分泌.结论:MICA在DCs表面的表达受局部微环境影响,且DCs表达MICA后可增强NK细胞的活性.     

IFN-γ促进人单核细胞向不典型成熟DC分化

目的: 研究IFN-γ对人单核细胞表型及分化的影响.方法: 采用免疫磁珠法从健康志愿者外周血单个核细胞(PBMCs)中特异性分选单核细胞, 以细胞因子IFN-γ、 TNF-α、 IFN-α刺激单核细胞后, 观察单核细胞生长特性, 并以流式细胞术检测单核细胞表面CD1a、 CD14、 CD80、 CD83、 CD86、 HLA-DR等分子表达.结果: 细胞因子IFN-γ、 TNF-α、 IFN-α对单核细胞形态、生长及表型转化有不同影响.IFN-γ促进单核细胞黏附聚集形成"细胞小岛结构", 刺激单核细胞形态向梭形及多角形转化; IFN-γ上调单核细胞表达CD80、 CD83、 CD86及HLA-DR分子, 同时下调CD14分子表达.结论: IFN-γ促进单核细胞向不典型的成熟DC方向分化.     

Bryostatin-1调节TNF-α激活的树突状细胞免疫功能

目的探讨Bryostatin-1对树突状细胞(DC)免疫功能的调节作用。方法联合应用GM-CSF和IL-4自人外周血单核细胞定向分化DC;以Bryostatin-1刺激TNF-α诱导成熟的DC,收集DC及其上清夜,以FACS和ELISA方法分析DC表面免疫分子(CD80、CD83、CD86、HLA-DR和B7-H1)和细胞因子(IFN-γ、IL-1β、IL-10和IL-12)表达水平;自DC提取RNA,以Northern blot分析DC的B7-h1 mRNA表达水平;以DC为诱导细胞,进一步检测DC的免疫诱导功能。结果Bryostatin-1通过降低DC表面B7-H1表达和增强B7.2表达,以及调节DC细胞因子(IL-1b、IL-12和IFN-γ)分泌,增强DC的免疫诱导功能,PKC通道特异性的抑制剂BI明显逆转Bryostatin-1的上述作用。结论Bryostatin-1增强DC免疫功能,其机制可能是激活PKC通道。     

灵芝多糖通过上调STAT4促进人外周血淋巴细胞中Th1细胞的分化

目的 探讨灵芝多糖(GLP)对外周血淋巴细胞免疫分群的影响及其作用机制.方法 取肿瘤患者和正常人的外周血,分离外周血单个核细胞(PBMC)后,用不同剂量的GLP(10 ng/ml、50ng/ml和100 ng/ml)刺激后,用流式细胞仪检测DC细胞表面分子(HLA-DR、CD83和CD11c)、Th1细胞、Th2细胞和NK(CD3-CD56+)细胞数;并进一步用免疫磁珠分选出正常人外周血CD4+ Th细胞后用不同浓度GLP刺激24h后,荧光实时定量Q-PCR检测Th1和Th2细胞因子的表达水平,Westernblot分析Th1分化相关的转录因子水平.结果 灵芝多糖可以在体外呈浓度依赖性增加外周血中Th1细胞亚群和DC共刺激分子的表达(P＜0.01),并且增加STAT4的表达和IL-12、IFN-γ和TNF-α的mRNA的表达水平(P＜0.01).结论 灵芝多糖可能通过增加Th细胞STAT4的表达水平,促进其向Th1细胞分化,并增加Th1的分泌细胞因子.     

脂多糖活化的单核细胞对人Th17细胞分化的影响

目的:拟建立体外人外周血CD4+T细胞与CD14+单核细胞共培养体系,观察脂多糖(LPS)活化的单核细胞,在联合或不联合抗CD3 mAb、不同浓度和时间条件下对诱导Th17细胞分化的影响。方法:采用免疫磁珠法分离纯化健康志愿者外周血CD4+T细胞与CD14+单核细胞,在两者共培养体系中分别单加LPS或抗CD3 mAb、抗CD3 mAb刺激下添加不同浓度LPS刺激培养3 d或同时加入LPS和抗CD3 mAb刺激培养3 10 d。以流式细胞术检测细胞表面分子CD4,胞内细胞因子IL-17及IFN-γ,确定各种刺激培养条件下,诱导生成的Th17及Th1等细胞亚群占CD4+T细胞的百分比。结果:单独采用LPS或抗CD3 mAb刺激,分别仅能诱导(1.30 0.19)%、(1.10 0.21)%CD4+T细胞分泌表达IL-17,而采用LPS联合抗CD3 mAb刺激显著增加Th17频率,可达(2.01 0.46)%(P<0.05)。在抗CD3 mAb刺激条件下,LPS浓度分别为0.1μg/mL,1μg/mL,10μg/mL时,诱导生成Th17细胞频率依次为(1.92 0.21)%、(1.300.37)%、(1.01 0.25)%,随LPS浓度增加,Th17细胞生成显著减少(P<0.05)。LPS体外刺激3 d可诱导(2.130.32)%Th17细胞生成,随时间推移,Th17细胞减少(P<0.05),Th1细胞在6 d达最高频率(17.45 3.04)%,10 d时下降到(11.34 1.29)%。结论:低剂量LPS联合抗CD3mAb活化的单核细胞可在短时间内诱导CD4+T细胞向Th17细胞分化,所建立的共培养体系更加贴近类风湿关节炎等炎症性疾病体内异常免疫应答的微环境。     

“核转染”技术对树突状细胞功能的影响

目的:研究"核转染"方法对DC活率、细胞表面分子及细胞因子分泌水平的影响。方法:体外诱导单核细胞分化为DC后,通过"核转染"方法将p/IRES-EGFP质粒转染至细胞核内,同时设立未转染DC为对照组。应用台盼蓝染色计算转染后细胞的活率;荧光显微镜下观察转染效率;流式细胞术检测细胞表面分子CD80、CD83、CD86、HLA-DR及CD11c等的表达;ELISA法检测DC培养上清IL-12及TNFα-的分泌水平。结果:DC转染后随时间推移,细胞活率下降;荧光蛋白在转染后4小时即有表达,表达量逐渐增加,24小时转染效率为56%±12%,48小时转染效率为45%±15%;转染后细胞表面分子CD80、CD83及CD86表达随时间推移逐渐降低,CD11c及HLA-DR表达水平稳定;转染可诱导DC分泌IL-12及TNFα-水平增加。结论:"核转染"是一种有效的转染DC的方法,并可诱导DC分泌多种细胞因子。     

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.     

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.     

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.     

'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.     

Der Einfluß von Nahrungsmitteln und Rauchen auf die klinisch-chemische Diagnostik von Phäochromozytom,Neuroblastom und Karzinoid-Syndrom

Zusammenfassung Der Einfluß von verschiedenen Nahrungsmitteln auf Methoden zur Bestimmung von Adrenalin (AD), Noradrenalin (NA), Vanillinmandelsäure (VMS), Metanephrinen (MN), Homovanillinsäure (HVS) und 5-Hydroxyindolessigsäure (5-HIE) im 24 h-Harn zur Diagnose des Phäochromozytoms bzw. Karzinoid-Syndroms wurde untersucht. Die in die Untersuchung einbezogenen Nahrungsmittel waren: Tee, Kaffee, Mandeln, Ananas, Käse, Walnüsse, Vanillepudding, Bananen, Tomaten und Milchschokolade. Außerdem wurde der Einfluß des Zigarettenrauchens auf die Bestimmung von AD, NA, VMS und MN untersucht.Walnüsse führten zu einer starken Erhöhung der 5-HIE-Ausscheidung. Bananen erhöhten die Ausscheidung von AD, NA, VMS, MN und 5-HIE. Kaffee und Ananas bewirkten eine geringe Zunahme der MN-Werte. Rauchen von 20–30 Zigaretten/Tag beeinflußte keine der vier Variablen.Wenn die beschriebenen Methoden benutzt werden, sollte lediglich auf den Verzehr von Bananen und Walnüssen vor und während der Harnsammelperioden verzichtet werden, da die oberen Normgrenzen im Harn überschritten werden könnten. Ein Verzicht auf Kaffee und Ananas in normalen Mengen ist nicht erforderlich. Es besteht kein Anlaß, weiterhin die bisherigen umfangreichen Restriktionen der übrigen Nahrungsmittel beizubehalten.