Brain-derived neurotrophic factor in cerebrospinal fluid of patients with chronic daily headache: relationship with nerve growth factor and glutamate levels

Little has been done to investigate the biochemical basis of chronic daily headache (CDH). Our group has recently demonstrated an increase in the cerebrospinal fluid (CSF) levels of nerve growth factor (NGF) in CDH patients, supporting the involvement of this growth factor in the abnormal processing of head pain in this pathological condition. Other members of the neurotrophin family, especially brain-derived neurotrophic factor (BDNF), have been hypothesized as being involved in the development of chronic head pain in patients affected by CDH, but so far no data are available on this subject. BDNF, NGF and glutamate levels were determined in the CSF of 25 patients affected by CDH with a previous history of migraine. These levels were compared with those of a group of 20 control subjects, for whom the CSF examination and other instrumental investigations excluded diseases of the central and peripheral nervous systems. Significantly higher levels of BDNF, NGF and glutamate were found in CDH patients compared with control subjects (p<0.0001, p<0.0002 and p<0.001, respectively). A significant positive correlation emerged between CSF values of BDNF and those of NGF (r=0.61, p<0.001) and glutamate (r=0.44, p<0.025) in CDH patients. No significant differences were detected in BDNF, NGF and glutamate levels between CDH patients with analgesic overuse and those without. These results support the involvement of BDNF in CDH through the potentiation of glutamatergic transmission involved in the processing of head pain. The significant correlation between BDNF and NGF levels suggests that NGF-mediated up-regulation of BDNF in central sites involved in long-term sensitization plays a key role in persistent head pain in CDH patients. Correspondence to P. Sarchielli     

支气管扩张症肺组织内降钙素表达的临床意义

目的　了解支气管扩张症肺组织降钙素 (CT)蛋白表达水平与其病变特点的关系。方法　应用免疫组化方法定量分析 4 5例支气管扩张症患者手术切除的肺组织 ,对其气道上皮内神经内分泌细胞 (NEC)计数 ,并与 19例无肺部疾患的尸检肺组织进行比较。结果　 2组均可见被染成棕红色的CT阳性细胞 ,它们分布于肺内各级支气管上皮细胞间 ,以支气管分叉处较多见。支气管扩张症组肺组织内5 0 0 0个上皮细胞中CT阳性细胞数为 (6 7.4 8± 11.0 3)个 ,非支气管扩张症组肺组织内CT阳性细胞数仅为 (5 .0 2± 1.0 0 )个 ,2组比较有非常显著性差异 (P <0 .0 0 1)。结论　支气管扩张症肺组织高水平CT表达可能与其慢性炎症改变有关 ,其局部CT表达增高可能系机体对支气管扩张病变的一种局部性代偿或调节机制。     

Sperm ubiquitination positively correlates to normal morphology in human semen

BACKGROUND: Global ubiquitination in human semen has been found to negatively correlate with standard semen parameters, indicating that ubiquitination can be considered a marker of poor semen quality. However, the inclusion of all semen components in the analysis may be misleading on the biological significance of ubiquitination of sperm cells. We have recently demonstrated the variable presence of bodies of different size, with the highest concentration in oligoasthenoteratozoospermia. The purpose of the present study was to evaluate the relationship between ubiquitination and standard semen parameters, after distinguishing between ubiquitinated sperm and bodies in each sample. METHODS: Ubiquitination was evaluated by flow cytometric sperm ubiquitin tag immunoassay (SUTI) in sperm samples from 45 subjects. Semen analysis was performed according to WHO (1999) guidelines. RESULTS: When only ubiquitinated sperm were considered, a positive correlation with number, motility and normal morphology was found. When correlation was evaluated considering the percentage of ubiquitinated bodies, a negative correlation was found with good semen quality. CONCLUSIONS: Results indicate that the negative correlations previously found between global semen ubiquitination and parameters of semen analysis are mainly driven by components other than sperm. The positive correlation between sperm ubiquitination and good quality parameters suggests a previously unrecognized role for sperm ubiquitination.     

CYP17A1基因新突变致17α-羟化酶/17,20-裂解酶部分性缺陷症

目的研究1例17α-羟化酶/17,20-裂解酶部分性联合缺陷症患者CYP17A1基因突变特点,并结合患者的临床表现与基因突变类型初步探讨P450C17酶蛋白的结构与功能的关系。方法收集1例17α-羟化酶/17,20-裂解酶部分性联合缺陷症患者的临床资料及其亲属血标本,提取基因组DNA,设计7对引物扩增CYP17A1基因的8个外显子及外显子与内含子的连接区域,琼脂糖凝胶电泳鉴定PCR产物,产物胶回收后直接做为DNA双链模板测序。DNA双链模板不一致的PCR产物经克隆后测序。测序结果在核苷酸序列数据库进行比较分析。结果患者CYP17A1基因突变检测结果为5994-5995delAT/7541C>T复合杂合子。这两种突变均未见报道。推测5994-5995delAT导致I259H,274X,突变形成的截短蛋白质缺少血红素结合区域,因此是没有功能的;而通过人类P450C17酶计算机模型分析显示7541C>T导致的A398V远离酶的活性中心,推测突变可能使酶的活性减弱,而不是完全地丧失。患者临床表现为有自发不规则月经及轻度高血压、低血钾,结合激素测定结果提示肾上腺和性腺保留部分功能。因而患者的基因型与其临床表型是一致的。结论应进行突变P450C17酶的功能学研究来进一步明确结构改变对功能的影响。     

视黄酸依赖Fas/RARα融合基因转染HL-60细胞启动的凋亡效应研究

目的：检测视黄酸依赖Fas／RARα表达载体转染HL-60细胞启动的凋亡效应。方法：成功构建视黄酸依赖Fas／RARα表达载体，用脂质体转染HL-60白血病细胞，经四甲基偶氮唑盐[3-(4，5-dimethylthiaol-2-yl)-2，5-diphenyl tetrazolium bromide,MTT]、流式细胞、DNA梯形带和透射电镜等方法观测视黄酸处理后细胞凋亡效应的发生。结果：以一定剂量的视黄酸处理HL-60后，细胞增殖受抑，呈凋亡性变。结论：视黄酸及其依赖的融合基因表达可协同诱导HL-60细胞发生凋亡效应。     

Identification and disruption of the gene encoding the third member of the low-molecular-mass rhoptry complex in Plasmodium falciparum

The low-molecular-mass rhoptry complex of Plasmodium falciparum consists of three proteins, rhoptry-associated protein 1 (RAP1), RAP2, and RAP3. The genes encoding RAP1 and RAP2 are known; however, the RAP3 gene has not been identified. In this study we identify the RAP3 gene from the P. falciparum genome database and show that this protein is part of the low-molecular-mass rhoptry complex. Disruption of RAP3 demonstrated that it is not essential for merozoite invasion, probably because RAP2 can complement the loss of RAP3. RAP3 has homology with RAP2, and the genes are encoded on chromosome 5 in a head-to-tail fashion. Analysis of the genome databases has identified homologous genes in all Plasmodium spp., suggesting that this protein plays a role in merozoite invasion. The region surrounding the RAP3 homologue in the Plasmodium yoelii genome is syntenic with the same region in P. falciparum; however, there is a single gene. Phylogenetic comparison of the RAP2/3 protein family from Plasmodium spp. suggests that the RAP2/3 duplication occurred after divergence of these parasite species.     

体外分化胚胎干细胞为造血干细胞重建造血功能的研究

目的：探讨体外定向分化胚胎干细胞（ESCs）为造血干细胞（HSCs）对体内造血功能的重建作用。方法：将小鼠E14.1胚胎干细胞采用“三步诱导法”在体外分化发育为HSCs，造血克隆形成(CFU)实验观察其体外造血集落形成情况，免疫磁珠分选纯化HSCs移植给经亚致死剂量γ射线照射的雌性SCID小鼠，观察其植入及小鼠造血功能恢复情况。结果: 经过分阶段诱导，多种造血刺激因子联合应用能有效促进ESCs定向分化发育为HSCs,流式细胞仪检测HSCs特异性表面标志物CD34+/Sca-1+表达最高为（58.64±4.20）%，CFU培养能形成较多的红系、粒系/巨噬细胞系及混合细胞集落, Wright-Giemsa 染色显示为原始的造血细胞。此阶段的HSCs经分选纯化后移植给经γ射线照射后的小鼠，移植组小鼠+10 d造血功能开始恢复，观察40 d后除血小板恢复较慢外，白细胞、红细胞、血红蛋白等指标已接近正常，植入率为71.4%，存活率为43.0%，染色体检测证实已由受体鼠的XX转为供体鼠的XY。结论: 采用分阶段诱导的方法，可在体外定向诱导小鼠ESCs分化发育为HSCs，此来源的HSCs可以有效重建体内造血功能。