神经再生素对背根神经节细胞GAP—43和NF—L基因表达的影响

目的：观察中药有效组分神经再生素作用于神经细胞生长过程中,相关基因的表达变化。探讨神经再生素促神经生长的分子生物学机制。方法：采用RT-PCR法,观察神经再生素在促大鼠背根神经节生长过程中（4h,12h,24h),生长相关蛋白43（GAP-43）和神经丝蛋白（NF-L）基因表达的变化。结果：神经再生素可使背根神经节细胞GAP-43和NF-LmRNA物表达量增加。结论：神经再生素可作用于体外培养的神经细胞,使神经生长相关基因表达上调。     

大鼠肾上腺切除后垂体前叶GAP-43免疫阳性神经纤维的发芽过程

目的　阐明大鼠双侧肾上腺切除后垂体前叶生长相关蛋白 43(GAP- 43)免疫阳性神经纤维表达的量与术后时间点的关系 ,即轴突发芽的时间过程 .方法　免疫组织化学ABC法 .结果　大鼠双侧肾上腺切除术后第 4日垂体前叶GAP- 43阳性表达显著增多 ,第 2周有所下降 ,至第 4周基本降至正常水平 .结论　大鼠在双侧肾上腺切除后 ,垂体前叶出现肽能神经纤维发芽 ,并围绕在腺细胞周围 ,该发芽过程于术后 4～ 7d达到高峰 ,第 2周开始下降 ,第 4周基本结束     

The Wolff-Parkinson-White syndrome: the cellular substrate for conduction in the accessory atrioventricular pathway

The longstanding quest for the anatomical basis of the Wolff-Parkinson-White syndrome has left many unanswered questions. The ultrastructuralmorphology of the myocytes comprising accessory atrioventricularpathways, which are capable of rapid and variable conduction,is central to understanding the development and behaviour ofthis congenital anomaly, but remains unknown. Examination ofthree surgically resected pathways was performed to determinetheir underlying cellular morphology and the pattern of intercellularcoupling, by correlative light microscopy, electron microscopyand confocal scanning laser microscopy combined with immunohistochemicallocalization of the cardiac gap-junctional protein, connexin43. Two left-sided pathways were composed of myocardium of ‘normalworking ventricular’ type. The right-sided pathway wascomposed almost entirely of highly abnormal myocytes characterizedby aberrant myofibril organisation, with a lack of A-band materialand abnormal mitochondria, but normal intact intercalated disksno different from those seen in left-sided pathways. The gapjunctions of all pathways were composed of connexin43 distributedas in ventricular myocardium, and not as found in atrial oratrioventricular nodal tissues. While myocytes of abnormal structure were present in one ofthe accessory atrioventricular pathways examined, all pathwayshad morphologically normal gap junctions, the structures responsiblefor efficient intercellular coupling, with a pattern of distributionsuggestive of working ventricular myocardium.     

Clusters of GABAA receptors on cultured hippocampal cells correlate only partially with functional synapses

We describe a method to label gamma-aminobutyric acid (GABA)A receptors on the surface of living hippocampal neurons in primary culture, and we compare the distribution of receptors with that of active synapses. To visualize GABAA receptors, the affinity-purified antibody beta3(1-13), recognizing the extracellular N-termini of the GABAA receptor beta2- and beta3-subunits, was used in combination with fluorescent secondary antibodies. The beta2- and beta3-subunits belong to the predominant GABAA receptor subunits in the hippocampus. As expected for aggregates of GABAA receptors in the somato-dendritic plasma membrane, a patchy staining pattern similar to that seen by labelling neurons after fixation was obtained. An antiserum recognizing an intracellular epitope of GABAA receptor beta3-subunits did not label the receptors in living neurons. Whole-cell recordings of GABA-evoked Cl - currents were not affected after decorating GABAA receptors with antibody beta3(1-13). Combining the staining of GABAA receptors with the labelling of active presynaptic terminals with the fluorescent dyes FM1-43 or FM4-64, consistently resulted in the detection of GABAA receptor clusters that were not located at active synapses. These amounted to approximately 50% of all labelled GABAA receptor clusters. GABAA receptor clusters that were not associated with active presynaptic terminals partially colocalized with the synaptic vesicle marker protein sv2, while another fraction had no presynaptic counterpart at all. These findings suggest the presence of presynaptically silent GABAergic synapses in cultured hippocampal neurons. They also indicate that for the maintenance of GABAA receptor aggregates, the release of GABA from an opposing active terminal is not essential.     

Propranolol and atenolol inhibit norepinephrine spillover rate into plasma in conscious spontaneously hypertensive rats

Summary Radiotracer techniques capable of measuring norepinephrine clearance and spillover rate into plasma were used to test the hypothesis that the antihypertensive effects of propranolol and atenolol in conscious spontaneously hypertensive rats are associated with an inhibition of norepinephrine release from postganglionic sympathetic neurons. The 10%–15% fall in mean arterial pressure produced over 4 h by propranolol (1, 3.3 and 10 mg/kg, s. c.) and atenolol (1, 3.3 and 10 mg/kg, s. c.) was not dose-related, and only the largest dose of propranolol caused a significant bradycardia. Each dose of atenolol significantly lowered heart rate. The decrease in blood pressure caused by propranolol and atenolol was not related to the decrease in heart rate. Both propranolol and atenolol inhibited norepinephrine clearance by 12% to 16%. The 1 mg/kg doses of propranolol and atenolol significantly suppressed norepinephrine spillover rate by 21 % and 32%, respectively, at 4 h postinjection. As the dose of propranolol was increased, the inhibition of norepinephrine spillover was reversed as plasma epinephrine concentration rose by 125%. The suppression of norepinephrine spillover rate caused by atenolol was more persistent but did diminish after the 10 mg/kg dose, when plasma epinephrine concentration was elevated by 55%. Both drugs suppressed plasma renin concentration, but the inhibition of norepinephrine spillover rate by propranolol and atenolol was not related to the fall in plasma renin concentration. By comparison, treatment with the adrenergic neuron blocking agent bretylium (5, 10, 20 and 40 mg/kg, s. c.) elicited a dose-related vasodepression with no change in heart rate or plasma renin concentration. The 10 mg/kg dose of bretylium inhibited norepinephrine spillover rate by 40%, but increasing the dose did not produce a further suppression of norepinephrine spillover rate. Bretylium caused a dose-related elevation of plasma epinephrine concentration (354% increase at 40 mg/kg). In a separate study, propranolol (10 mg/kg) and bretylium (40 mg/kg) significantly increased epinephrine spillover rate by 85% and 118%, respectively. Based on these data, we conclude that the -adrenoceptor antagonists lower blood pressure by inhibiting norepinephrine release from postganglionic sympathetic neurons. Send offprint requests to T. K. Keeton at the above address     

Accelerated Differentiation in Response to Retinoic Acid After Retrovirally Mediated Gene Transfer of GAP-43 into Mouse Neuroblastoma Cells

Although substantial evidence exists for the involvement of growth-associated protein-43 (GAP-43) in neuronal development and regeneration, the precise role of this protein in neurite outgrowth is currently debated. To investigate the role of GAP-43 in the initiation of neurite outgrowth, we transfected a full-length cDNA coding for GAP-43 into a mouse neuroblastoma cell line (Neuro-2a) which can be differentiated to a neuronal phenotype using retinoic acid (RA). We show that the consequent overexpression of GAP-43 results in a change in the basic morphology of these cells, but is not in itself sufficient to induce the extension of neurites. However, overexpression of GAP-43 results in a marked acceleration of neurite formation in response to RA. We propose that while GAP-43 does not trigger the initiation of neurite extension, its expression is rate-limiting for neurite outgrowth in response to differentiation agents such as RA.     

Large Scale Manufacturing of B43(Anti-CD19)-Genistein for Clinical Trials in Leukemia and Lymphoma

We have conjugated the murine monoclonal anti-CD 19 antibody B43 to the tyrosine kinase inhibitor genistein to construct an effective immunoconjugate against CD 19 antigen positive hematologic malignancies. The scaled-up production and purification of B43 antibody, genistein, and B43-Genistein immunoconjugate permitted the manufacturing of a highly purified clinical-grade B43-Genistein preparation. In clonogenic assays, B43-Genistein elicited selective and potent cytotoxicity against CD 19 antigen positive human leukemia cells. To our knowledge, this work represents the first effort of producing a clinical-grade genistein immunoconjugate for treatment of B-lineage leukemia and lymphoma.     

艾灸"肺俞""膏肓"影响BLMA5诱导肺纤维化大鼠TGF-β1表达实验研究

目的:探索针灸阻抑肺纤维化的效应机制,为针灸介入肺纤维化防治提供实验依据.方法:将SD大鼠140只随机分为4组:空白组、模型组、艾灸组、泼尼松组,每组35只.气管内注入博莱霉素制作大鼠肺纤维化模型,造模后7 d开始治疗,以5 mg艾绒灸其双侧"肺俞""膏肓",每穴3壮,每天1次,10 d为一疗程,共治疗3个疗程后处死动物,采用PCR检测其转化生长因子β1(TGF-β1)mRNA表达.结果:肺组织TGF-β1mRNA表达分别为:空白组1.0258±0.0057,模型组2.8104±0.2905,艾灸组1.6174±0.1136,泼尼松组1.7176±0.1079.艾灸组、泼尼松组与模型组比较,P<0.01,艾灸组与泼尼松组比较,P>0.05.结论:艾灸"肺俞""膏肓"与泼尼松治疗均能显著抑制肺纤维化大鼠肺组织TGF-β1mRNA表达.     

Targeted disruption of GAP-43 in P19 embryonal carcinoma cells inhibits neuronal differentiation. As well as acquisition of the morphological phenotype

GAP-43 is expressed in proliferating neuroblasts in vivo and in vitro, but its role during early neurogenesis has not been investigated. Here we show that neuroectodermal differentiation stimulated by retinoic acid (RA) in the embryonal carcinoma (EC) line P19 is accompanied by upregulation of GAP-43 expression in neuroepithelial precursor cells. In contrast, when upregulation of GAP-43 expression was prevented in 3 independent P19 lines because of a targeted insertion into the gene, generation of neuroepithelial precursors was inhibited. Consequently, neuronal number was significantly decreased, neuronal morphology was abnormal and fewer than 20% of all neurons were able to initiate neuritogenesis. Extracellular matrix (ECM) was unable to rescue initiation of neuritogenesis in the mutant cells, however those neurites that were extended responded normally to ECM-stimulated neurite outgrowth-promoting signals. These data suggest that GAP-43 function is required for commitment to a neuronal phenotype as well as initiation of neurite extension. However, stimulation of neurite outgrowth by ECM in P19s occurs independently of GAP-43.     

缝隙连接蛋白43和E-钙黏附素表达与喉癌生物学行为关系的研究

目的　研究缝隙连接蛋白基因 4 3(connexin4 3,Cx4 3)和E 钙黏附素 (E cadherin ,E cad)在喉癌组织中的表达与其生物学行为的关系 ,探讨喉癌发生、发展的机制。方法　应用免疫组织化学PicTureTM二步法 ,检测有 5年以上随访资料的 6 0例喉癌组织和 10例癌周正常喉组织中Cx4 3和E cad的表达 ,结果进行统计学处理。结果　Cx4 3和E cad蛋白主要定位于细胞膜上 ,10例癌周正常喉黏膜组织均呈阳性表达。 6 0例喉癌患者中 ,70 0 % (4 2 / 6 0例 )的喉癌Cx4 3呈阳性表达 ,按喉癌的临床分期、颈部淋巴结转移、喉癌复发情况分组差异有显著性意义 (χ2 =5 11、6 4 1、3 86 ,P值均 <0 0 5 ) ,与喉癌的组织学分级差异有极显著性 (χ2 =15 0 9,P <0 0 1) ,与喉癌患者的生存期无统计学意义 (χ2 =2 6 5 ,P >0 0 5 ) ;4 1 7% (2 5 / 6 0例 )的喉癌组织中E cad表达减低 ,按喉癌的临床分期、喉癌复发情况分组及生存期分组差异有显著性意义 (χ2 =6 0 9、5 35、5 14 ,P值均 <0 0 5 ) ,与喉癌的组织学分级、颈部淋巴结转移差异有极显著性意义 (χ2 =15 0 7、6 6 5 ,P均值 <0 0 1)。同一标本中Cx4 3和E cad基因表达有显著的相关性和一致性 (r=0 6 3,P <0 0 0 0 1)。结论　喉癌存在Cx4 3和E cad基因的失活及蛋