High affinity neurotrophin receptors in the human pre-term newborn, infant, and adult cerebellum

The immunohistochemical occurrence of the high affinity neurotrophin (NT) receptors trkA, trkB, and trkC is shown in the pre-term newborn, infant, and adult human post-mortem cerebellum. Immunoreactive neuronal perikarya and processes were observed in all specimens examined, where they appeared unevenly distributed in the cerebellar cortical layers and deep nuclei, and showed regional differences among cerebellar lobules and folia. The trk receptor-antibodies, tested by Western blot on human cerebellum homogenates, revealed multiple immunoreactive bands for trkA and single bands for trkB and trkC. The results obtained show the tissue localization of the trk receptor-like immunoreactivity in the human cerebellum from prenatal to adult age. The analysis for codistribution of the receptors with the relevant ligand and among the receptors in discrete cortical and deep nuclei tissue fields shows a wide variety of conditions, from a good similarity in terms of type and density of labeled structures, to a lack of correspondence, and suggests the possibility of colocalization of trk receptors with the relevant neurotrophin and among them in the cerebellar cortex. These results sustain the concept that the neurotrophin trophic system participates in the development, differentiation, and maintenance of the human cerebellar connectivity and support the possibility of a multifactorial trophic support for the neurotrophins through target-derived and local mechanisms.     

胰腺癌组织中BRAF表达的初步研究

目的:了解BRAF基因在正常胰腺和胰腺癌组织中的表达水平。方法:6例胰腺癌标本取自胰腺癌手术患者,3例正常胰腺组织取自外伤性胰腺损伤需切除部分胰腺者。用Trizol法抽提每份组织的总RNA,分别进行紫外分光光度计测定、琼脂糖凝胶电泳及Lab on ch ip检测,对总RNA进行质控;应用实时定量PCR技术检测正常胰腺和胰腺癌组织BRAF的mRNA水平;应用W estern b lot检测BRAF的蛋白表达水平。结果:BRAF mRNA水平在正常胰腺组织中为0.001 016 7±0.000 25,胰腺癌组织中为0.002 606 2±0.001 09,上调2.56倍;W estern b lot显示,正常胰腺组织BRAF蛋白表达为6.70±2.50,胰腺癌组织为21.23±5.60,上调3.17倍。结论:胰腺癌组织中BRAF mRNA和蛋白表达水平均明显上调,其可能的调控机制及生物学效应还有待进一步研究。     

光生物素DNA探针的制备及其检测恶性疟原虫DNA的研究

本研究使用光生物素(PHOTOPROBE~(TM) Biotin)标记恶性疟原虫(P.f.)全基因组DNA,含P.f.cDNA片段的重组质粒DNA(C7)及克隆的P.f.cDNA片段(P9-25)等作为探针,并以斑点杂交试验检测靶DNA。实验结果显示,光生物素标记法简单、快速和安全;标记的探针活性较高,于-20℃贮存10个月无明显降低。用生物素化的P.f.全基因组DNA及C7探针分别可检出低至125pg和250pg的纯化的P.f.DNA。     

Nucleic acid spot hybridization with nonradioactive labeled probes in screening for human papillomavirus DNA sequences

We examined 161 human tissue samples using the spot hybridization technique with nonradioactive labeled DNA probes of human papillomavirus (HPV). Whole cells were spotted on nitrocellulose filters; DNA of the cells was denatured and fixed to the filter. Then the DNA spots were hybridized to nonradioactive labeled DNA and monitored by a sandwich immunoenzymatic reaction. This technique is simple, sensitive, specific, requires no special equipment, and can be used in clinical settings. HPV DNA was found in 92% of samples in which, on the basis of histologic and colposcopic criteria, its presence was suspected, as well as in 31 samples where it was not suspected.     

人乳头瘤病毒与呼吸道肿瘤关系的研究：Ⅰ.HPV与喉癌的关系

应用α－３２Ｐ－ｄＣＴＰ标记ＨＰＶ－１１及ＨＰＶ－１６ＤＮＡ为探针，以Ｓｌｏｔｂｌｏｔ及Ｓｏｕｔｈｅｒｎｂｌｏｔ两种核酸杂交技术，检测了取自青岛及北京两个地区的３７例喉鳞癌组织ＤＮＡ中ＨＰＶ－１１及ＨＰＶ－１６ＤＮＡ相关序列。结果表明，Ｓｌｏｔｂｌｏｔ杂交中，与ＨＰＶ－１１探针杂交阳性者占８６％（３２／３７），与ＨＰＶ－１６探针杂交阳性者占８１％（３０／３７）。在同一癌组织中，与两型探针同时呈现阳性者占８１％（３０／３７）。Ｓｏｕｔｈｅｒｎｂｌｏｔ杂交中，与所用探针ＤＮＡ的内切酶ＰｓｔⅠ酶切图谱比较，有４６％（１３／２８）与ＨＰＶ－１６ＤＮＡ的酶切图谱相同，可定为ＨＰＶ－１６型。当用ＨＰＶ－１１探针时无一例出现与ＨＰＶ－１１相同的图谱。提示喉癌的发生与ＨＰＶ－１６有关。     

Correlation between the induction of an immediate early gene, zif/268, and long-term potentiation in the dentate gyrus.

Expression of the immediate early gene zif/268 (also termed NGFI-A, Krox 24, TIS8 and Egr-1) was investigated in awake rats following various long-term potentiation (LTP) induction protocols.zif/268 mRNA (Northern blots) and protein (immunohistochemistry) levels sharply increased following LTP, and followed a time course characteristic of other immediate early genes. When measured across 3 tetanization protocols known to produce differing degrees of LTP persistence,zif/268 induction was found to be more highly correlated with LTP duration than with the magnitude of initial LTP. These data support the hypothesis that the immediate early gene zif/268 plays a role as a third messenger in the cascade of cellular and nuclear events that govern the persistence of LTP.     

链置换式扩增检测羊水中巨细胞病毒DNA

目的：介绍一种简便快速准确检测羊水中ＣＭＶ－ＤＮＡ的改良ＰＣＲ－链置换式扩增用于诊断胎儿先天感染ＣＭＶ。方法，将组成套式ＰＣＲ的外内两对引物按照一定比例（外：内＝１：５０－１００）加在同一试管中一次扩增羊水和胎儿组织中ＣＭＶ－ＤＮＡ。结果：９０例异常孕产史的孕妇羊水检测ＣＭＶ－ＤＮＡ，阳性率为３８．９％（３５／９０），其中合并染色数目异常２例（４７，ＸＹＹ和４７，ＸＸ，＋２１）（已引产）核型及染色     

与Graves眼病有关的眼肌自身抗原抗体的分子免疫学研究

为探讨Ｇｒａｖｅｓ眼病（ＧＯ）的发病机理，我们对新近发现的眼肌自身抗原及存在于ＧＯ患者血清的眼肌抗体进行了研究。血清取自１８例正常人、１８例活动性ＧＯ、１０例无限征的Ｇｒａｖｅｓ甲亢（ＧＨ）和３例桥本甲状腺炎（ＨＴ）患者。人眼肌膜蛋白经ＧＯ患者混合ＩｇＧ亲和层析后进行ＳＤＳ－聚丙烯酰胺凝胶电泳，显示分子量为４５、２８、５５和６４ｋＤ等蛋白条带。经正常人混合ＩｇＧ亲和层析未能显示上述结果。Ｗｅｓｔｅｒｎ印迹杂交虽然未能证实仅与患者血清作用的独特的眼肌抗原的存在，但６４ｋＤ印迹存在于６１％的ＧＯ、３０％的ＧＨ、０％的ＨＴ患者，正常人仅２２％阳性。抗６４ｋＤ阳性率ＧＯ组明显高于对照组（Ｐ＜０．０５）。眼肌抗体（ＥＭＡｂ）特别是抗６４ｋＤ抗体对于ＧＯ发病机理的作用值得进一步研究。     

In vivo and in vitro models demonstrate a role for caveolin-1 in the pathogenesis of ischaemic acute renal failure

Caveolae and their proteins, the caveolins, transport macromolecules; compartmentalize signalling molecules; and are involved in various repair processes. There is little information regarding their role in the pathogenesis of significant renal syndromes such as acute renal failure (ARF). In this study, an in vivo rat model of 30 min bilateral renal ischaemia followed by reperfusion times from 4 h to 1 week was used to map the temporal and spatial association between caveolin-1 and tubular epithelial damage (desquamation, apoptosis, necrosis). An in vitro model of ischaemic ARF was also studied, where cultured renal tubular epithelial cells or arterial endothelial cells were subjected to injury initiators modelled on ischaemia-reperfusion (hypoxia, serum deprivation, free radical damage or hypoxia-hyperoxia). Expression of caveolin proteins was investigated using immunohistochemistry, immunoelectron microscopy, and immunoblots of whole cell, membrane or cytosol protein extracts. In vivo, healthy kidney had abundant caveolin-1 in vascular endothelial cells and also some expression in membrane surfaces of distal tubular epithelium. In the kidneys of ARF animals, punctate cytoplasmic localization of caveolin-1 was identified, with high intensity expression in injured proximal tubules that were losing basement membrane adhesion or were apoptotic, 24 h to 4 days after ischaemia-reperfusion. Western immunoblots indicated a marked increase in caveolin-1 expression in the cortex where some proximal tubular injury was located. In vitro, the main treatment-induced change in both cell types was translocation of caveolin-1 from the original plasma membrane site into membrane-associated sites in the cytoplasm. Overall, expression levels did not alter for whole cell extracts and the protein remained membrane-bound, as indicated by cell fractionation analyses. Caveolin-1 was also found to localize intensely within apoptotic cells. The results are indicative of a role for caveolin-1 in ARF-induced renal injury. Whether it functions for cell repair or death remains to be elucidated.     

抗早孕因子单克隆抗体的制备与鉴定

目的建立分泌抗EPF(Early pregnancy factor,早孕因子)单克隆抗体的杂交瘤细胞株,纯化单抗并鉴定.方法用本实验室已纯化的早孕和肿瘤源性EPF作为抗原刺激Balb/c小鼠,用免疫后的小鼠脾细胞与同系小鼠骨髓瘤细胞(NS-1)融合,经4次克隆化,获得可稳定分泌抗EPF单克隆抗体的细胞株,注入Balb/c小鼠腹腔制备腹水型单抗,Protein-A亲和层析纯化,SDS电泳和Western-blot等方法分析纯化结果.结果融合后获得一株稳定分泌抗EPF抗体的细胞株(C3D11),克隆化后,获得稳定分泌抗EPF单克隆抗体的细胞株,将增殖后的细胞注射Balb/c小鼠腹腔获得腹水型单抗,以亲和层析法纯化,SDS-PAGE分析显示纯化后去掉了大部分杂蛋白,免疫印迹分析抗体纯度较高,与抗原匹配性良好.结论本研究制备的EPF单克隆抗体为特异性抗EPF抗体.