星形胶质细胞条件培养液对体外模拟脑缺血再灌注损伤神经元的作用

目的：探讨星形胶质细胞(Ast)条件培养液(ACM)对体外模拟脑缺血再灌注损伤神经元的影响。方法：分离纯化培养大鼠大脑皮质Ast和神经元,体外模拟脑缺血再灌注损伤模型,用ACM培养损伤后的神经元,测定其神经元的活性、存活率、死亡率、培养液乳酸脱氢酶(LDH)的漏出率和NOS强阳性细胞的表达量。结果：ACM能使脑缺血再灌注损伤神经元的活性和存活率明显提高,使死亡率、LDH的漏出率和NOS强阳性细胞的表达量显著降低。结论：(1)体外模拟脑缺血再灌注损伤神经元的变化具有一定的规律性;(2)ACM对受损的神经元具有重要的保护和修复作用;(3)Ast在脑缺血预适应中发挥重要的作用。     

激活的星形胶质细胞条件培养液对神经元内iNOS、CaMKⅡ及AC表达的影响

为探讨星形胶质细胞(Ast)在癫痫发病机制中的作用,本研究通过体外分离纯化培养分别获取大鼠大脑皮质Ast及神经元,用睫状神经营养因子(CNTF)激活Ast,用其培养液,即星形胶质细胞条件培养液(ACM)孵育神经元4、8、12h后,采用Westernblot法及图像分析技术观察ACM对神经元内表达钙调蛋白依赖性激酶II(CaMKⅡ)、诱导型一氧化氮合酶(iNOS)及腺苷酸环化酶(AC)的影响。结果表明,ACM作用神经元4h即可引起CaMKⅡ、iNOS及AC的表达增多并持续至12h(P<0.05),iNOS在8h最为明显,AC的表达在12h最为明显。上述结果提示ACM中含有致痫因子,NOS-NO-cGMP,Ca2+/CaM-CaMKⅡ及AC-CaMP-PKA三条信号通路可能参与其致痫机制中的信号转导。     

二苯乙烯苷对同型半胱氨酸诱导血管内皮细胞凋亡及bcl-2、bax、caspase-3表达的影响

 目的：研究何首乌二苯乙烯苷（TSG）对同型半胱氨酸（Hcy）诱导人脐静脉内皮细胞（HUVECs）凋亡及bcl-2、bax和caspase-3 mRNA表达的影响。方法：建立Hcy （3 mmol/L）所致培养HUVECs核损伤模型,TSG（1和10 μmol/L）提前2 h预孵育,然后再以Hcy处理,作为TSG保护组。用Hoechst 33342核染色法观察HUVECs细胞核损伤状态,以流式细胞术检测细胞凋亡情况,用实时荧光定量RT-PCR法检bcl-2、bax和caspase-3 mRNA的表达。结果：经3 mmol/L Hcy处理后,与正常培养的细胞相比,HUVECs核损伤加重,凋亡细胞比例升高,bcl-2表达降低（P＜0.01）,bax和caspase-3表达增加（P＜0.01）。TSG 1 μmol/L和10 μmol/L预孵育后再经3 mmol/L Hcy处理,与单经3 mmol/L Hcy损伤模型组相比,HUVECs细胞核损伤降低,凋亡率下降,bcl-2的表达增加（P＜0.05）,bax和caspase-3表达降低（P＜0.05）。结论：TSG具有降低Hcy所致HUVECs的细胞损伤和抑制凋亡的作用,其机制可能与影响bcl-2、bax和caspase-3的表达有关。     

TNFα刺激的星形胶质细胞条件培养液对海马神经元的兴奋作用

目的:探讨肿瘤坏死因子-α刺激的星形胶质细胞对神经元的作用。材料和方法:体外纯化培养大鼠海马星形胶质细胞,采用反相高效液相法测定TNFα刺激后细胞培养液内谷氨酸的含量;将TNFα刺激后的星形胶质细胞条件培养液（Astrocytic Conditioned Medium,ACM）作用于培养的海马神经元,运用免疫细胞化学方法和图像分析技术研究神经元NF-κBp65和谷氨酸的表达。结果:（1）TNFα可明显促进星形胶质细胞释放谷氨酸;（2）ACM作用15min即可诱导神经元NF-κBp65的核表达,30min达高峰,180min恢复至对照水平;（3）ACM作用60min可使谷氨酸免疫反应阳性神经元平均光密度明显升高,持续至240min。结论:TNFα刺激的星形胶质细胞可通过释放谷氨酸等可溶性物质使神经元快速激活、兴奋性升高。     

星形胶质细胞条件培养液对缺氧损伤神经元的保护作用

为了探讨星形胶质细胞条件培养液(ACM)对缺氧损伤神经元的保护作用,本实验将新生的KM小鼠的皮层星形胶质细胞分离、纯化并传代培养第三代第5d后,将星形胶质细胞(Ast)条件培养液,加入到缺氧损伤的神经元细胞培养液中,观测其对缺氧神经元的存活及其合成和释放一氧化氮(NO)、乳酸脱氢酶(LDH)和胞膜ATP酶(ATPase)的影响。结果显示:10%和20%ACM对神经元存活有明显促进作用;与对照组相比,20%ACM还有效地减少了缺氧神经元NO、LDH的生成和分泌,保护和提高了ATPase的活性。以上结果提示ACM促进缺氧神经元的存活,其机制可能与减少NO、LDH合成释放及保护细胞膜上ATPase的活性有关。     

星形胶质细胞条件培养液促进新生大鼠海马神经干细胞分化

目的探讨星形胶质细胞条件培养液(ACM)对神经干细胞(NSCs)体外分化的影响。方法行新生大鼠大脑皮质星形胶质细胞的纯化培养后收集ACM,将新生大鼠海马NSCs单克隆培养后行nestin免疫细胞荧光染色,诱导分化5d后行神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)、半乳糖脑苷脂(GALC)免疫细胞荧光染色;将单克隆培养的NSCs分为对照组和实验组,对照组为单纯NSCs培养液,实验组根据NSCs培养液与ACM比的不同分3组:A组(2:1),B组(1:1),C组(1:2)。各组培养1周,采用NSE免疫细胞荧光检测方法标记神经元并计数和统计学分析。结果纯化的星形胶质细胞胶质纤维酸性蛋白抗体标记阳性率为96.5%;单克隆培养的海马NSCs呈nestin阳性,诱导后呈NSE、GFAP和GALC阳性表达。ACM培养的海马NSCs各组分化为神经元的比例明显高于对照组(<0.01),其中A组的比例最高。结论ACM可以促进新生大鼠海马NSCs向神经元分化。     

硼替佐米上调fas、bcl2112、caspase-9和caspase-3基因表达

目的：探讨硼替佐米诱导慢性粒细胞白血病细胞株K562细胞凋亡及新基因bcl2H2在其中的作用。方法：MTT比色法观察硼替佐米对K562细胞的生长抑制作用;Annexin—V标记和线粒体跨膜电位（△ψm）分析细胞凋亡;RT—PCR方法检测0、6、12和24h fas、bcl2112、bcl-2、bim、bax、caspase-3和caspase-9基因表达变化。结果：硼替佐米抑制K562细胞生长呈时间和剂量依赖性,24h和48h半数抑制浓度分别为161．41nmol／L和96．33nmol／L;硼替佐米诱导K562细胞凋亡,12hAnnexin—V阳性细胞就开始增高,并呈时间依赖性,AtOm减低;RT—PCR显示fas、bcl2112、caspase-3和caspase-9表达增高,但bcl-2、bim和bax表达无明显改变。结论：硼替佐米可以抑制K562生长并诱导凋亡,上调如、bcl2112,使线粒体膜电位下降,激活caspase-9和caspase-3基因,促使DNA发生断裂可能是其诱导凋亡的机制之一。     

体外血小板活化因子对神经元和星形胶质细胞的作用

目的：研究血小板活化因子（PAF）对神经元活力及星形胶质细胞胶质纤维酸性蛋白（GFAP）表达的影响。 方法： 分别取BALB/c胎鼠和新生小鼠大脑皮层，纯化培养神经元和星形胶质细胞，设正常对照组和实验组，实验组中分别加入4、8和16 μmol/L的PAF并分别作用4 h、24 h和72 h,用MTT法和免疫组织化学方法测定神经元活力和星形胶质细胞表达GFAP的平均灰度值。 结果： PAF作用后，神经元活力降低；星形胶质细胞数量减少，但存活细胞GFAP表达增加。两者均呈浓度依赖关系。 结论： PAF不仅直接作用于神经元，且可通过作用于星形胶质细胞间接影响神经元的存活。     

星形胶质细胞条件培养液促进胚胎干细胞分化为神经元样细胞的实验研究

目的:探讨星形胶质细胞条件培养液在体外能否促进胚胎干细胞（ESCs）向神经元方向分化。 方法:在Bain等建立的4-/4+方案基础上，分成两组：单用ATRA组和ATRA联合胎脑星形胶质细胞条件培养液组，分3阶段诱导ESCs定向生成神经元样细胞。形态学观察及畸胎瘤形成试验对ESCs的全能性进行鉴定；免疫组化染色法检测nestin、GFAP、NSE和NF-200的表达对诱导过程进行动态监测。 结果:（1）体外培养的小鼠ESC-D3细胞在胚胎成纤维饲养层细胞上连续传代培养，仍保持向3胚层分化的能力。（2）在诱导分化阶段，联合诱导组分化细胞NF-200和NSE的表达阳性率高于单用ATRA组。（3） 联合诱导组最终诱导ESCs生成纯度高达73.5%神经元样细胞。 结论:采用胚胎脑星形胶质细胞条件培养液可较高产率及纯度地促进ESCs生成神经元样细胞，值得推广应用。     

TNFα激活的星形胶质细胞在慢性癫痫复发中的作用

目的：研究TNFα激活的星形胶质细胞在慢性癫痫复发中的作用。方法：分别用反相高效液相色谱法和免疫组织化学反应检测细胞释放谷氨酸（Glu）和NF－κBp65表达的变化。将TNFα激活的星形胶质细胞条件培养液（ACM）注射入慢性马桑内酯致痫发作间期大鼠之侧脑室，观察动物行为和脑电图的变化，并用免疫组织化学反应观察海马NF－kBp65和胶质原纤维酸性蛋白（GFAP）表达的变化。结果：1．TNFα可明显促进星形胶质细胞释放Glu，并快速诱导星形胶质细胞核内NF－kBp65的表达。2．侧脑室注射ACM可引起大鼠4级癫痫行为及典型的痫样脑电图表现；注射ACM后0．5h即可观察到海马回特别是CA1区细胞核内p65的表达，2－4h达高峰，8h恢复至对照水平；注射ACM后1h海马GFAP免疫反应阳性细胞数开始高于对照组，4h达高峰，8h仍明显高于对照组。结论：TNFα激活的星形胶质细胞可通过释放可溶性的神经活性物质引起慢性癫痫的复发。     

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.     

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.     

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.     

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.