侧脑室注射miR-7通过EGFR/STAT3途径抑制大鼠脑出血后脑损伤

目的观察微小RNA-7(miR-7)对大鼠脑出血后脑损伤的影响,并探讨其分子机制。方法选取75只SD大鼠,以Ⅶ型胶原酶注入苍白球构建脑出血模型,随机分为对照组、miR-7激动剂(miR-7 agomir)组、miR-7激动剂对照(agomir control)组、miR-7拮抗剂(miR-7 antagomir)组和miR-7拮抗剂对照(antagomir control)组,每组15只。造模后第2天通过侧脑室分别注射10μL的生理盐水、miR-7 agomir、agomir control、miR-7 antagomir、antagomir control,造模后第7天进行神经功能评分,然后处死动物,取出脑组织,HE染色观察脑组织病理改变,干湿重法测定脑组织含水量,荧光实时定量PCR检测血肿周围脑组织中miR-7、胶质纤维酸性蛋白(GFAP)和表皮生长因子受体(EGFR)表达,Western blot法分析血肿周围脑组织中GFAP、EGFR、信号转导子和转录激活子3(STAT3)、磷酸化STAT3(p-STAT3)水平。生物信息学预测miR-7与EGFR的靶向结合情况,构建野生型荧光素酶报告基因载体及相应的突变载体,与miR-7模拟物(mimic)共转染HEK293T细胞,检测荧光素酶活性。结果与对照组相比,miR-7 agomir明显增加血肿周围脑组织miR-7水平,减轻脑组织病理损害,减少神经功能评分和脑组织含水量,并降低血肿周围脑组织GFAP、EGFR表达水平及p-STAT3/STAT3比值;miR-7 antagomir的作用则相反。相应阴性对照物对上述指标无影响。生物信息学分析发现EGFR的3'-非翻译区存在miR-7结合位点,而且miR-7 mimic明显降低野生型EGFR荧光素酶活性,但对突变型无影响。结论 miR-7通过抑制EGFR/STAT3信号通路拮抗星形胶质细胞活化,从而减轻大鼠脑出血后脑损伤。     

亚低温对脑出血大鼠血肿周围神经营养因子和血管新生的影响

目的:研究亚低温治疗对脑出血大鼠血肿周围脑组织中胶质细胞源性神经营养因子(GDNF)、血管内皮生长因子(VEGF)的影响。方法:雄性SD大鼠随机分为3组:假手术组(Sham组)、脑出血组(ICH组)和亚低温组。采用立体定向注射自体血复制脑出血大鼠模型,造模后15 min给予持续4 h的亚低温治疗。应用修正神经功能缺损评分(mNSS)对各组大鼠进行神经行为学评分;干湿重法检测各组大鼠脑组织含水量;Real-time PCR检测各组大鼠血肿周围脑组织内G-DNF、VEGF的mRNA表达水平;免疫印迹检测各组大鼠血肿周围脑组织内GDNF、VEGF的蛋白表达含量。结果:与假手术组相应时问点相比,脑出血组大鼠神经功能评分明显升高,脑组织含水量显著增多,血肿周围脑组织GDNF、VEGF的mRNA和蛋白表达明显上调;与脑出血组对应时间点相比,亚低温组神经功能评分明显降低,脑组织含水量显著减少,GDNF、VEGF的mRNA和蛋白表达显著上调。结论:亚低温治疗可增加脑出血大鼠脑内神经营养因子和血管生长因子的表达,促进神经修复和新生血管的形成,改善神经功能缺损和脑水肿。     

亚低温治疗对大鼠脑出血后水通道蛋白4、磷酸化缝隙连接蛋白43表达的影响

目的:观察亚低温治疗对大鼠脑出血后血肿周围脑组织水通道蛋白4(AQP4)、磷酸化缝隙连接蛋白43(p-CX43)及脑水肿的影响。方法:成年雄性SD大鼠随机分为假手术组(Sham组)、脑出血模型组(ICH组)和亚低温组,采用注入自体血方法复制脑出血大鼠模型,成功后15 min给予亚低温治疗,使大鼠肛温在30 min内控制至30℃~31℃,维持4 h。采用干湿重法检测各组大鼠脑组织含水量变化;免疫组织化学检测AQP4和p-CX43的定位表达情况;免疫印迹检测AQP4和p-CX43的蛋白表达水平。结果:脑出血模型组大鼠与假手术组相比,脑组织含水量显著增多,血肿周围脑组织AQP4和p-CX43蛋白的表达明显上调。与脑出血模型组相比,亚低温组大鼠脑组织含水量明显减少,AQP4和p-CX43的蛋白表达量显著下调,但都高于假手术组。结论:亚低温治疗可以减轻大鼠脑出血后的脑水肿,这种保护性疗效可能与下调AQP4和p-CX43蛋白的表达相关。     

右美托咪定复合羟考酮对高血压脑出血大鼠模型血脑屏障的保护作用及机制研究

目的 探究右美托咪定复合羟考酮对高血压脑出血大鼠血脑屏障的保护作用及机制。方法 将50只SD大鼠随机分为假手术组（Sham）、高血压脑出血组（HICH）、右美托咪定组（Dex）、羟考酮组（Oxy）和右美托咪定复合羟考酮组（Dex+Oxy）,每组10只。参照Zea-Longa评分标准进行神经功能缺损评分,HE染色观察血肿周围脑组织病理形态变化;TUNEL方法检测血肿周围脑组织细胞凋亡水平;检测脑组织含水量、血肿直径并通过伊文思蓝渗透法检测BBB通透性;Western blot检测血肿周围脑组织紧密连接蛋白Occludin、Claudin-5和ZO-1及SIRT1和FoxO1蛋白表达。结果 右美托咪定复合羟考酮降低高血压脑出血大鼠神经功能评分;改善大鼠血肿周围脑组织病理损伤;降低脑组织细胞凋亡水平;降低脑含水量、血肿直径及BBB通透性;增加血肿周围脑组织Occludin、Claudin-5、ZO-1、SIRT1和FoxO1蛋白表达。结论 右美托咪定复合羟考酮保护高血压脑出血大鼠血脑屏障,改善神经功能,这可能与其激活SIRT1/FoxO1信号通路有关。     

AQP4表达对脑出血大鼠模型脑水肿与相关凋亡蛋白影响

目的:研究AQP4表达对脑出血大鼠模型脑水肿与相关凋亡蛋白影响。方法:选取40只雄性SD大鼠随机分为对照组、模型组、AQP4siRNA组、空载质粒组,每组10只。模型组、空载质粒组、AQP4siRNA组均采用脑室内注射凝血酶Ⅶ注射建立大鼠尾壳核出血模型。AQP4siRNA组造模后将载有siRNA的质粒(10μl)注射于大鼠脑室内;空载质粒组造模后将不含AQP4siRNA片段的空质粒(10μl)注射于大鼠脑室内,用于排除质粒与手术对结果的干扰。术后1d和3d分别采用Longa分级法进行神经功能评分,术后3d后处死大鼠,采用干湿重法测量脑组织含水量,RT-PCR法检测血肿周围脑组织AQP4 mRNA表达,Western blotting法检测脑组织Caspase-3和Bcl-2蛋白表达水平。结果:与对照组比较,模型组神经功能评分明显增加,脑水肿明显,AQP4 mRNA表达明显增加,脑组织Caspase-3蛋白表达增加,Bcl-2蛋白表达降低(P0.05);与模型组比较,AQP4siRNA干预组可明显降低神经功能评分,减轻脑水肿(P0.05);同时AQP4 mRNA表达明显降低,脑组织Caspase-3蛋白表达明显降低,Bcl-2蛋白表达增加(P0.05)。结论:抑制AQP4基因表达能抑制脑水肿、减轻脑出血大鼠神经功能损伤,可能与减少Caspase-3和增加Bcl-2表达有关。     

辛伐他汀对大鼠脑出血后热休克蛋白70及细胞凋亡的影响

目的:观察辛伐他汀对大鼠脑出血模型血肿周围热休克蛋白70(HSP70)及细胞凋亡的影响。方法:应用Ⅶ型胶原酶诱导法建立脑出血大鼠模型,72只SD大鼠按随机数字表法分为假手术组(Sham组)、脑出血组(ICH组)、辛伐他汀组(SIM组)。分别于6、12、24、48 h对各组大鼠进行神经功能评分后处死留取脑组织,采用干/湿重法检测脑组织含水量;免疫组织化学法检测HSP70、Bax和Bcl-2阳性产物的分布情况;Western Blot检测HSP70、Bax和Bcl-2蛋白表达量,并计算Bcl-2/Bax的比值。结果:与假手术组相比,脑出血组大鼠脑组织含水量及神经功能评分增加(P0.05),血肿周围HSP70、Bax和Bcl-2表达明显上调(P0.05),且HSP70、Bax和Bcl-2三种蛋白的表达高峰分别出现在48 h、24 h及12 h,Bcl-2/Bax比值降低。与脑出血组相比,辛伐他汀治疗组脑组织含水量与神经功能评分显著降低(P0.05),血肿周围HSP70和Bcl-2表达进一步上调(P0.05),而Bax表达下调(P0.05),但三种蛋白的表达趋势未发生改变,Bcl-2/Bax比值增加。结论:辛伐他汀可以促进脑水肿及神经功能的恢复,其可能通过增加血肿周围HSP70表达,抑制脑组织细胞凋亡而发挥神经保护作用。     

多巴胺对出血性脑卒中大鼠胶质纤维酸性蛋白和脑源性神经营养因子表达的影响及保护机制

目的：探讨多巴胺（DA）对出血性脑卒中大鼠的胶质纤维酸性蛋白（GFAP）和脑源性神经营养因子（BDNF） 表达的影响及保护机制。方法：建立出血性脑卒中大鼠模型,将大鼠随机分为假手术组、脑出血模型组（ICH 组）和DA 组（ 脑出血模型给予DA 组）。分别比较3 组大鼠的神经功能障碍评分、神经元形态、脑水肿、细胞凋 亡和脑组织形态变化;免疫组织化学染色法检测脑组织中GFAP 和BDNF 蛋白的表达。结果：与假手术组相比较, ICH 组神经功能障碍评分显著降低;与ICH 组相比,DA 组神经功能障碍评分显著升高。与假手术组比较,ICH 组尼氏小体数量显著减少;与ICH 组相比,DA 组尼氏小体数量明显增加;ICH 组脑组织含水量和细胞凋亡数量 显著高于假手术组;与ICH 组相比,DA 组脑组织含水量和细胞凋亡数量均显著降低。ICH 组脑组织出现了不同 程度的水肿,其中一部分神经细胞出现了明显肿胀,间质之间明显增宽,神经元结构受到了严重破坏,严重的 神经元出现坏死,且有大量的炎性细胞浸润;DA 组大鼠的脑组织形态得到明显的改善。与假手术组相比,ICH 组脑组织中GFAP 蛋白阳性表达显著增加,BDNF 蛋白阳性表达显著降低;与ICH 组大鼠相比,DA 组脑组织中 GFAP 阳性表达明显减少,BDNF 蛋白阳性表达明显增多。结论：DA 能抑制脑组织中GFAP 表达,促进BDNF 的 表达,改善脑出血的神经功能障碍,进而对出血性脑卒中受损脑组织起到保护作用。     

抑制小凹蛋白减轻大鼠脑缺血/再灌注诱导的神经功能损伤

目的 探讨小凹蛋白1(Cav1)的磷酸化在大鼠脑缺血/再灌注后神经功能损伤中的作用机制。方法 将大鼠随机分为假手术组(sham)、模型组(model)和抑制剂组(PP2)。用改良神经功能缺损评分(mNSS)评价大鼠神经功能,氯化三苯基四氮唑(TTC)染色检测脑梗死体积,Nissl染色检测脑缺血/再灌注后组织形态,Tunel染色检测梗死灶周围细胞凋亡数量,Western blot和免疫荧光检测脑梗死周围组织中p-Cav1、胶质纤维酸性蛋白(GFAP)和离子钙结合接头分子1(Iba1)的表达。结果 与假手术组相比,模型组大鼠mNSS评分增加(P<0.05);脑梗死体积明显增加(P<0.05),尼氏体数量减少(P<0.05);凋亡细胞数量增加(P<0.05);p-Cav1、GFAP和Iba1蛋白的表达量上调(P<0.05)。而抑制剂PP2能减轻上述指标的变化,保护脑缺血/再灌注损伤大鼠神经功能(P<0.05)。结论 抑制p-Cav1可以改善大鼠脑缺血/再灌注后神经功能,其机制可能与抑制胶质细胞激活及相互作用有关。     

神经调节素对大鼠脑缺血再灌注损伤细胞凋亡和STAT3及GFAP表达的影响

目的 探讨神经调节素-1β(NRG-1β)对大鼠脑缺血再灌注损伤细胞凋亡、信号转导和转录激活因子3(STAT3)及胶质纤维酸性蛋白(GFAP)表达的影响.方法 成年雄性Wistar大鼠,用线拴法经颈外-颈内动脉插线建立大脑中动脉闭塞再灌注(MCAO/R)动物模型,经颈内动脉单剂量注射1.5%NRG-1β5μl干预治疗.荧光DendEndTUNEL法检测神经细胞凋亡;免疫组织化学和免疫荧光染色枪测脑组织STAT3和GFAP表达.结果 脑缺血再灌注损伤可诱导脑组织细胞凋亡和STAT3及GFAP表达.对照组随缺血时间延长,皮质、纹状体和海马区细胞凋亡逐渐增多,STAT3和GFAP表达逐渐增强.治疗组在缺血不同时间点细胞凋亡较对照组相应脑氏显著减少(P<0.05),STAT3和GFAP表达水平较对照组相应脑区显著增强(P<0.05).结论 NRG-1β可能通过激活细胞JAK/STAT信号传导途径,促进星形细胞胶质化,启动神经细胞抗凋亡机制,对缺血性脑损伤起保护作用.     

辛伐他汀对实验性大鼠脑出血损伤中TNF-α表达及细胞凋亡的影响

目的:观察辛伐他汀对实验性大鼠脑出血(intracerebral hemorrhage,ICH)损伤中凋亡相关因子肿瘤坏死因子-α(TNF-α)表达的影响,探讨其通过抗凋亡机制发挥的脑保护作用。方法:采用注入新鲜自体不凝血复制脑出血模型,干湿重法检测脑组织含水量;免疫组化法检测TNF-α和caspase-3的表达,Western Blot检测脑组织TNF-α和caspase-3的蛋白表达水平。结果:大鼠脑出血后血肿周围基底节区脑组织TNF-α和caspase-3凋亡相关蛋白表达量、脑组织含水量及凋亡细胞数量明显上调。与模型组相比,辛伐他汀干预组的凋亡细胞数、TNF-α和caspase-3的表达与脑组织损伤周围含水量均显著降低(P0.05)。结论:辛伐他汀对大鼠脑出血损伤有一定的保护作用,可能通过抑制脑组织中TNF-α和caspase-3的表达,进而减少细胞凋亡。     

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.     

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.