Pyruvate kinase in human brain tumours. Its significance in the treatment of gliomas

Summary Pyruvate kinase isozyme distribution was studied in 101 intracranial tumours of various nature and origin, and in normal human brain (both foetal and adult). In foetal brain, five different forms could be detected by electrophoresis (K₄, K₃M, K₂M₂, KM₃, and M₄). In adult brain, the M₄ type, K₃M hybrid, and K₄ are present; the M type is largely predominant. Alanine inhibition of pyruvate kinase can be used to discriminate between M and K-type pyruvate kinase. The results obtained in an alanine inhibition test are in agreement with the electrophoretic pattern. Pyruvate kinase from foetal brain and brain of a newborn is more inhibited compared with pyruvate kinase from adult brain. In adult brain a high residual activity of pyruvate kinase is found in the presence of alanine. Well differentiated neuroepithelial tumours,i.e., astrocytomas, oligodendrogliomas, and ependymomas showed also relatively high residual activities, though less than in normal adult brain. On the contrary, in poorly differentiated gliomas low residual activity was found. Alanine inhibition of pyruvate kinase correlates well with degree of differentiation of these tumours. There is also a strong correlationship between alanine inhibition of pyruvate kinase and one year survival after total or subtotal resection of gliomas in adults.When in gliomas the residual activity is determined not in the centre of the tumour but more towards the periphery, much higher residual activity is found. It is suggested that brain biopsies in which a residual activity higher than 70% is found probably contain no tumour in the paraffin slides.Poorly differentiated gliomas were characterized by the presence of type K, and the hybrids K₃M. In well differentiated gliomas, besides K₄ and K₃M, M₄ was also present. Alanine inhibition was in agreement with the electrophoretic pattern in all tumours. In children (age 1–11 years) gliomas showed no correlation between the distribution of pyruvate kinase isozymes and the histological classification and grading. Of the non-neuro-epithelial tumours studied relatively high residual activities were found for pyruvate kinase in haemangioblastomas, chromophobe adenomas, and craniopharyngiomas. This was also found in an arteriovenous malformation. Other non-neuroepithelial tumours showed much less residual activity. These included benign tumours, meningiomas, neurilemmomas, malignant metastatic tumours, and fibrosarcomas. It was also found in cavernomas. The determination of pyruvate kinase activity in the presence of alanine may be useful for the diagnosis and treatment of intracerebral tumours, in particular gliomas of adults.The alanine inhibition test is a reliable quantitative procedure. It can be performed in 10 minutes, and may well fit in the scope of a surgical procedure.     

Different inhibitory effect of adrenaline on platelet adenylate cyclase in the presence of GTP plus cholera toxin and of stable GTP analogues

Summary GTP is generally required for hormonal stimulation of adenylate cyclase. On the other hand, the presence of GTP is essential for hormone-induced inhibition of adenylate cyclase in cell-free preparations of human platelets and other cells. In order to differentiate the dual roles of GTP in hormonal stimulation and inhibition of adenylate cyclase, we have studied the effect of adrenaline on the platelet enzyme under conditions where guanine mucleotides caused marked stimulation. In the presence of GTP (1 M), which by itself had no or only a small stimulatory effect on adenylate cyclase, adrenaline inhibited the basal and prostaglandin E₁-stimulated forms of the enzyme. In contrast, the stable GTP analogues, GMP-P(NH)P and GTP--S, which caused a time-dependent, persistent activation of the enzyme, reversed or prevented the inhibitory effect of adrenaline. Cholera toxin, which activates adenylate cyclase presumably by inhibition of a specific GTPase, increased cyclase activity in platelet membranes up to 4-fold, and GTP addition (0.1–30 M) augmented this activation about 2-fold. The -adrenergic component of adrenaline (0.1–100 M), in a concentration-dependent manner, prevented the GTP-induced increase in cholera toxin-stimulated activity. The inhibition was also observed in enzyme preparations that had been fully activated by pretreatment with cholera toxin. These data suggest that -adrenergic agonists may inhibit platelet adenylate cyclase through increased inactivation of the enzyme, possibly involving a stimulation of the GTPase connected to the adenylate cyclase system.Abbreviations GMP-P(NH)P guanylyl 5-imidodiphosphate - GTP--S guanosine 5-(-thio)triphosphate - GTPase guanosine 5-triphosphatase - P_i inorganic phosphate. Parts of the data were presented in preliminary form (Jakobs and Schultz, 1978)     

Effect of acute and chronic treatment of tandamine,a new heterocyclic antidepressant,on biogenic amine metabolism and related activities

Summary The effects of tandamine, a clinically effective heterocyclic antidepressant, administered either acutely (10 mg/kg i.p) or chronically (10 mg/kg i.p. daily for 21 days) on biogenic amine uptake and metabolism in the rat were determined and a comparison with desipramine was made. Tandamine, similarly to desipramine, blocked norepinephrine (NE) uptake in rat brain and heart following both acute and chronic administration. No effect of tandamine on dopamine (DA) or serotonin (5-HT) uptake was observed. Both drugs lowered endogenous brain NE when given chronically but not acutely. In contrast, no such effect on brain DA and 5-HT or heart NE was observed. Tandamine, like desipramine, administered chronically prior to an intraventricular injection of ³H-NE, produced increases in the decline of ³H-NE as indicated by decreased ³H-NE with increased levels of ³H-normetanephrine in brain stem of rats, suggesting an increased turnover of NE. No such effect was observed following acute treatment. Both drugs increased the behavioural effects of L-Dopa following an acute oral administration, with tandamine appearing superior to desipramine at the lower dose examined (10 mg/kg). Tandamine was 57–833 times less effective in binding to rat brain muscarinic receptors than desipramine, imipramine, butriptyline and amitriptyline, respectively. Thus, tandamine affects biogenic amine mechanism following either acute or chronic administration in a fashion similar to desipramine, but unlike desipramine, it exhibits relatively little anticholinergic properties, a further indication of the potential use of tandamine in the treatment of human depression, particularly where an increase in drive is desired.     

The extracellular matrix molecule tenascin-R and its HNK-1 carbohydrate modulate perisomatic inhibition and long-term potentiation in the CA1 region of the hippocampus

Perisomatic inhibition of pyramidal cells regulates efferent signalling from the hippocampus. The striking presence of HNK-1, a carbohydrate expressed by neural adhesion molecules, on perisomatic interneurons and around somata of CA1 pyramidal neurons led us to apply monoclonal HNK-1 antibodies to acute murine hippocampal slices. Injection of these antibodies decreased GABAA receptor-mediated perisomatic inhibitory postsynaptic currents (pIPSCs) but did not affect dendritic IPSCs or excitatory postsynaptic currents. The decrease in the mean amplitude of evoked pIPSCs by HNK-1 antibodies was accompanied by an increase in the coefficient of variation of pIPSC amplitude, number of failures and changes in frequency but not amplitude of miniature IPSCs, suggesting that HNK-1 antibodies reduced efficacy of evoked GABA release. HNK-1 antibodies did not affect pIPSCs in knock-out mice deficient in the extracellular matrix molecule tenascin-R which carries the HNK-1 carbohydrate as analysed by immunoblotting in synaptosomal fractions prepared from the CA1 region of the hippocampus. For control, HNK-1 antibody was applied to acute sections of mice deficient in the neural cell adhesion molecule NCAM, another potential carrier of HNK-1, and resulted in decrease of pIPSCs as observed in wild-type mice. Reduction in perisomatic inhibition is expected to promote induction of long-term potentiation (LTP) by increasing the level of depolarization during theta-burst stimulation. Indeed, LTP was increased by HNK-1 antibody applied before stimulation. Moreover, LTP was reduced by an HNK-1 peptide mimic, but not control peptide. These results provide first evidence that tenascin-R and its associated HNK-1 carbohydrate modulate perisomatic inhibition and synaptic plasticity in the hippocampus.     

注射用盐酸丁咯地尔细菌内毒素检查法的研究

研究注射用盐酸丁咯地尔对细菌内毒素检查试验的干扰情况 ,并建立其细菌内毒素检查的方法。按《中国药典》 2 0 0 0年版二部附录ⅪE细菌内毒素检查法和附录ⅪⅩF细菌内毒素检查法应用指导原则进行试验。本品在 0 33mg·ml-1稀释浓度下无干扰作用 ,L =1 5EU·mg-1。本品可用细菌内毒素检查法替代热原检查法     

丹参的体外抑菌作用研究

目的：探讨丹参的体外抑菌作用。方法：用K-B纸片扩散法。100％丹参浸出液滤纸片对大肠杆菌、金黄色葡萄球菌、白色葡萄球菌、变形杆菌、乙型链球菌抑菌作用进行了研究。结果：丹参对以上细菌均有抑菌作用。结论：丹参在体外有明显的抑菌作用。     

磷酯酶C对家兔血小板聚集和超微结构的影响

目的　应用电镜研究磷酯酶C(phospholipaseC ,PLC)对家兔血小板聚集和超微结构的影响 ,以探讨用药后PLC抗血小板聚集作用的机制。方法　家兔颈动脉插管取血 ,制备PRP ,将其分为 4组 :①空白对照组 ;②生理盐水组 ;③ 0 5UPLC组 ;④ 2 5UPLC组。 2～ 4组分别用ADP诱导聚集 ,测出其聚集抑制率 ,各组分别制成超薄切片样品进行电镜观察、摄片。结果　 0 5、2 5UPLC明显抑制血小板聚集反应 ,聚集抑制率分别为 86 0 3 %± 12 6%和 82 47%±5 49% ;0 5UPLC抑制血小板形成伪足 ,α颗粒和致密颗粒较多 ,OCS管腔较小 ,其超微结构较生理盐水组变化小 ;2 5UPLC处理血小板的形态结构和超微结构与空白对照组无差异 ,血小板呈圆形或椭圆形 ,边缘光滑 ,无伪足样突起 ,α颗粒、致密颗粒、OCS等正常分布于胞质中。结论　PLC明显抑制ADP诱导的血小板聚集反应及其超微结构的改变 ,这可能是PLC抗血小板聚集作用的重要原因之一     

Inhibition of β‐amyloid toxicity by short peptides containing N‐methyl amino acids

Abstract: Single N‐methyl amino acid‐containing peptides related to the central hydrophobic region β_16–20 (Lys‐Leu‐Val‐Phe‐Phe) of the β‐amyloid protein are able to reduce the cytotoxicity of natural β_1–42 in PC12 cell cultures. N‐methyl phenylalanine analogs yield statistically significant increments in cell viability (Student's t‐test < 0.01%) and are nontoxic in the same assay. These promising results indicate that these peptide molecules could be a starting point for the development of potential therapeutic compounds for the treatment of Alzheimer's disease.     

蛋白酶体抑制剂Z-LLL-CHO对人骨肉瘤细胞株MG-63作用的体外研究

目的　探讨蛋白酶体抑制剂Z LLL CHO对人骨肉瘤细胞株MG 63的作用 ,并探讨其作用机制。方法　将不同浓度的Z LLL CHO作用于骨肉瘤细胞株MG 63 ,采用荧光显微镜观察细胞形态改变、电镜观察细胞超微结构变化、MTT法检测细胞增殖活力、琼脂糖凝胶电泳检测细胞凋亡、流式细胞仪分析细胞凋亡率及细胞周期改变。结果　Z LLL CHO可有效抑制MG 63细胞株的生长 ,并诱导细胞发生凋亡。凋亡细胞表现为浓染致密的颗粒块状荧光 ,胞核固缩、染色质凝聚并边缘化 ,DNA呈“阶梯状”排列的条带。流式细胞仪分析显示 ,Z LLL CHO在 1.0 μmol/L作用 2 4h、3 6h、48h后 ,细胞凋亡率分别为 5 .4%、2 0 .5 %、5 2 .7%。随药物浓度增高及作用时间延长 ,细胞周期被阻滞于G2 /M期。结论　蛋白酶体抑制剂Z LLL CHO可有效抑制人骨肉瘤MG 63细胞株的生长增殖 ,阻滞细胞周期及诱导细胞凋亡可能起重要作用。     

菘蓝春化机理的初步研究

目的　探明菘蓝春化作用的机理 ,寻找预防菘蓝提早开花的有效方法。方法　将萌动的菘蓝种子和菘蓝幼苗分别低温处理 ,观察菘蓝生长后期的抽薹情况 ;用 ABA浸泡菘蓝种子后再进行低温春化 ,研究ABA浸种处理对菘蓝的萌发及后期抽薹开花的影响 ;用 ABA叶面喷施已通过春化作用的菘蓝幼苗 ,观测ABA对已春化菘蓝后期抽薹的抑制作用。结果　菘蓝可通过种子或幼苗进行春化 ,春化效果与春化时间在一定范围内呈正相关系 ;用低浓度的 ABA浸泡菘蓝种子对其发芽率无显著影响 ,且对菘蓝的后期春化无显著的抑制作用 ;用 ABA喷施已通过春化作用的菘蓝幼苗 ,可在一定程度上抑制菘蓝提前抽薹开花。结论　菘蓝属于种子和绿体春化类型 ;用 ABA喷施已春化的菘蓝幼苗 ,可以预防菘蓝提前抽薹开花。