Persephin基因转染对缺氧诱导的神经干细胞凋亡的影响

目的:探讨重组腺病毒介导Persephin基因治疗对缺氧状态下对神经干细胞凋亡的作用。方法:体外培养C17.2神经干细胞,建立神经干细胞缺氧模型;将携带人类Persephin基因的重组腺病毒感染C17.2神经干细胞;Western-blotting法分析Persephin蛋白的表达;TUNEL法检测凋亡指数,显微镜观察凋亡小体;流式细胞术测定细胞凋亡率的变化。结果:转染pAdPersephin的C17.2神经干细胞成功表达Persephin蛋白;缺氧后神经干细胞凋亡指数为(25.54±4.30)%,而转染pAdPersephin后的细胞凋亡指数显著减少至(10.04±1.32)%(P<0.01),而转染pAdCMVPersephin Persephin反义寡核脱氧核酸细胞凋亡指数为(24.05±3.05)%,与对照组无显著差异(P>0.05)。结论:腺病毒介导的Persephin基因能高效表达Persephin;外源性Persephin对C17.2神经干细胞具有抗凋亡作用,能够提高C17.2神经干细胞对缺氧的耐受性。     

缺氧对离体培养神经干细胞增殖、分化及凋亡的影响

为探讨缺氧对体外培养的神经干细胞生长、分化及凋亡的影响,本研究采用显微解剖、机械吹打、无血清悬浮培养方法分离培养神经干细胞,用巢蛋白(nestin)免疫荧光染色对其进行鉴定。三气培养箱予以缺氧干预,分为5%缺氧组、10%缺氧组和正常对照组,每组又依缺氧干预时间的不同,分为6、12、24、48、72、96和120h组。通地绘制细胞生长曲线(MTT法)和计数克降形成率检测缺氧对神经干细胞增殖的影响。缺氧培养后,再用含10%胎牛血清的培养基进行分化培养,用免疫荧光技术检测缺氧对神经干细胞分化、凋亡及形态变化的影响。结果显示:缺氧干预后神经干细胞的增殖率明显下降,细包凋亡数增加,分化的神经元和胶质细胞的突起变短变粗,数量减少,但神经元和胶质细胞的分化比例未见明显变化。结果提示:缺氧可严重影响神经干细胞的存活和正常分化、且影响程序与缺氧剂量有量效关系。     

胚鼠神经干细胞培养与鉴定

目的　体外获取高浓度神经干细胞 (NSC)。方法　加入bFGF培养E12天大鼠胚脑的全部细胞 ,用荧光免疫组化法显示。结果　加bFGF的神经干细胞 (Neuralstemcell,NSC)大量分裂 ,形成大量的细胞分裂球 ,传到 4代时 ,90 %以上的细胞表达nestin。对照组分裂的细胞少 ,维持到第二代细胞逐渐死亡。结论　bFGF对胚胎E12天大鼠脑细胞的NSC有明显的促分裂作用 ,应用此种方法可获得高浓度的NSC     

脊髓神经干细胞对小鼠视网膜移植的研究

目的　研究原代培养的脊髓神经干细胞在小鼠视网膜的整合和分化情况。　方法　利用细胞培养和体内移植技术 ,将原代脊髓神经干细胞 (NSC)移植到不同年龄小鼠的视网膜 ,并对移植后细胞的整合及分化情况进行了免疫组织化学分析。　结果　 1 移植的NSC对组织的整合能力随宿主年龄的增加而降低 ;2 移植的NSC在宿主视网膜内可以分化为星形胶质细胞、少突胶质细胞和神经元。　结论　脊髓原代NSC移植到小鼠视网膜后的整合和分化均受内外因素的调控 ,为NSC的体内分化研究提供了新的证据     

糖氧剥夺对神经干细胞增殖、分化及凋亡的影响

背景：缺氧作为神经系统疾病发展过程中诸多病理因素中最常见的因素之一,对于内源性神经干细胞以及移植后外源性神经干细胞的存活、迁移、分化、凋亡发挥着诸多调控作用。 目的：系统观察糖氧剥夺对神经干细胞增殖、分化、凋亡的影响。 方法：从新生鼠嗅球分离培养出神经干细胞,建立糖氧剥夺模型,免疫荧光技术检测糖氧剥夺后神经干细胞分化能力,MTT法检测糖氧剥夺24,48 h后恢复正常条件培养对神经干细胞增殖能力的影响,Hochest33258荧光染色检测糖氧剥夺24,48 h后神经干细胞凋亡状况。 结果与结论：第4代神经干细胞CD133免疫荧光鉴定结果呈阳性表达,MTT细胞增殖能力检测结果示：糖氧剥夺组增殖能力明显低于常氧组(P < 0.05),糖氧剥夺48 h组增殖能力低于糖氧剥夺24 h组(P < 0.05)。GFAP、β-Tubulin Ⅲ细胞免疫荧光染色结果示糖氧剥夺48 h后神经干细胞分化为星形胶质细胞、神经元总数明显低于常氧组(P < 0.05)。Hochest33258染色凋亡率检测结果示糖氧剥夺组细胞凋亡率明显高于常氧组(P < 0.01),糖氧剥夺48 h组神经干细胞凋亡率显著高于糖氧剥夺24 h组(P < 0.01)。上述结果表明糖氧剥夺对神经干细胞增殖、分化、凋亡造成不利影响,影响程度取决于缺氧浓度、缺氧时间及自身对于缺氧的耐受性。中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程全文链接：     

小鼠睾丸支持细胞对大脑皮质细胞的体外营养作用

目的通过观察睾丸支持细胞(SCs)对大脑皮质细胞体外的营养作用,探讨移植的SCs在体内对神经组织营养作用的方式。方法将原代分离的小鼠大脑皮质细胞单独培养(对照组)或与小鼠SCs共培养(实验组),比较两组中神经元、神经干细胞(NSC)及神经胶质细胞的生长情况。结果实验组的神经元数及其突起数、神经元特异性烯醇化酶光密度(P<0.01)和神经胶质细胞数均较对照组高,两组各时期的神经元胞体面积也不相同(P<0.05),实验组于第3天出现NSC的集落形成。结论SCs在体外对大脑皮质的神经元、NSC和神经胶质细胞均有较强的营养作用,在中枢神经系统疾病的细胞移植治疗中将具有很大的应用价值。     

2种诱导iPSC向神经干细胞分化方法的比较

目的:整体比较2种促进诱导性多能干细胞(induced pluripotent stem cells,i PSC)向神经干细胞(neural stem cells,NSC)分化的方法,确定一种稳定、高效的获得NSC的方法,并对NSC进行系统鉴定。方法:方法A:SB431542和drosomophorin的浓度均为5μmmol/L,诱导初始密度100%;方法 B:SB431542的浓度为5 mmol/L,drosomophorin的浓度为1 mmol/L,诱导初始密度为40%。比较及鉴定方法:镜下观察诱导获得NSC的状态;realtime PCR比较神经干细胞相关基因Pax6、nestin、Sox1、Sox2等表达量;流式细胞术分析诱导第16天Pax6阳性率;免疫荧光定性分析神经干细胞相关蛋白的表达及其自发分化的能力。结果:方法 A获得的NSC悬起后成球趋势明显,圆形,透明;方法 B诱导获得NSC形状不规则,色灰暗。Real-time PCR结果证明方法 A诱导获得的细胞神经干细胞相关基因的表达量高于方法 B。流式细胞术分析证明第16天,PAX6的阳性率,方法 A高于方法 B。经鉴定,方法 A获得的神经干细胞高表达Pax6、nestin、Sox2等基因自发分化30 d,形成明显的神经纤维束,表达TUJ-1、MAP2及GFAP等神经元和胶质细胞的特异性标志物。结论:方法 A整体优于方法 B,我们推荐方法 A作为诱导i PSC向神经干细胞分化的方法。     

大鼠胚胎脑组织神经干细胞的培养和鉴定

目的探讨从不同胎龄的大鼠脑组织中分离,培养神经干细胞(NSC)并对其鉴定,了解生物特性。方法通过采用机械分离和消化分离相结合的方法分离不同胎龄大鼠脑NSC。在无血清DMEM/F12(含20ng/m lbFGF,20ng/m lEGF及B27辅助培养液)中培养、传代和鉴定。诱导分化后采用SABC法对分化的细胞进行神经元特异烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)检测作细胞鉴定。结果从不同胎龄的胎鼠脑组织中成功培养出神经干细胞,胎龄为12.5天的胎鼠提取的神经干细胞集落最多,在上述条件下培养及传代的细胞不断分裂增殖,形成悬浮生长的呈巢素蛋白(nestin)阳性的神经球;用血清诱导分化为大量表达NSE阳性的神经元和GFAP阳性的星形胶质细胞。结论胎龄为12.5天胎鼠大脑皮质培养出的神经干细胞数量最多,可分化为神经元、神经胶质细胞及少突胶质细胞。     

人脐血间充质干细胞抗缺氧诱导心肌细胞凋亡的作用

背景:抗细胞凋亡成为心力衰竭生物治疗的一个新方向。近年研究发现成体间充质干细胞在一定条件下可转化成心肌样细胞,亦可通过分泌多种细胞因子,促进心脏血管形成和减少细胞凋亡。目的:观察人脐血间充质干细胞对缺氧诱人心肌细胞凋亡的保护作用。方法:将人心肌细胞细胞复苏后,接种在6孔细胞培养板(对照组)和Transwell3412培养板中;将人脐血间充质干细胞接种在Transwell插件的可渗透性滤膜上;将上述2组细胞置于体积分数95%N2+体积分数5%CO2缺氧环境下缺氧培2,4,12,24h;检测各组心肌细胞的凋亡率。ELLES法检测细胞条件培养基内胰岛素样生长因子1的浓度。结果与结论:第3代后人脐血间充质干细胞及原代心肌细胞均有胰岛素样生长因子分泌,但人脐血间充质干细胞条件培养基内胰岛素样生长因子1的浓度明显高于人心肌细胞。缺氧可以诱导心肌细胞凋亡,在短期和持续性缺氧的条件下,人脐血间充质干细胞对缺氧诱导的心肌细胞凋亡有保护作用,且人脐血间充质干细胞组心肌细胞凋亡率低于对照组(P0.05)。提示人脐血间充质干细胞对缺氧诱导的人心肌细胞凋亡有保护作用,这种保护作用可能与通过细胞间直接接触和旁分泌细胞因子有关。     

MPP+诱导的PC12细胞和星形胶质细胞共育对神经干细胞突触素和生长相关蛋白-43表达的影响

目的探讨星形胶质细胞和1-甲基-4苯基-吡啶离子(MPP+)诱导的PC12细胞共育对神经干细胞(NSC)突触素和生长相关蛋白-43表达的影响。方法 PC12细胞分别经MPP+诱导不同时间点(0h、24h、48h、72h、96h)后分为两部分,一部分应用流式细胞技术检测PC12细胞凋亡率;另一部分与星形胶质细胞共育2d,然后收集各组细胞条件培养液,对神经干细胞进行诱导分化,免疫荧光检测神经干细胞突触素(SYN)和生长相关蛋白-43(GAP-43)表达。结果在MPP+作用48h时间点,PC12细胞凋亡率达高峰(P0.05)。MPP+与PC12细胞和星形胶质细胞条件培养液共育48h可上调神经干细胞中突触素(A值=34.09±2.69)和GAP-43(A值=49.36±5.98)表达水平(P0.05)。结论星形胶质细胞与MPP+诱导凋亡的PC12细胞共育后,促进神经干细胞突触素和GAP-43表达。     

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.