Comparison of Echinacea alkylamide pharmacokinetics between liquid and tablet preparations

The relative oral bioavailability of alkylamides from two different Echinacea dosage forms (liquid and tablet) were compared in a small two-way crossover study in humans (n=3). The liquid preparation investigated contained a mixture of Echinacea purpurea root (300 mg/ml) and Echinacea angustifolia root (200 mg/ml) extracted in 60% ethanol. The tablet preparation investigated was also a mixture of E. purpurea root (675 mg/tablet) and E. angustifolia root (600 mg/tablet), but was prepared from the dried 60% ethanolic extracts of these two Echinacea species. Alkylamides were found to be rapidly absorbed and measurable in plasma from both preparations. No significant differences in the tetraene alkylamide pharmacokinetic parameters for T(1/2), AUC(t-lin) and C(max) in the two different preparations were found. T(max) increased from 20 min for the liquid to 30 min for the tablet, which is not unexpected as the tablet required time for disintegration before absorption could occur. These results suggested that there was no significant difference in the bioavailability of alkylamides from the liquid and tablet Echinacea formulations. Furthermore, the results also indicated that the absorption site and any alkylamide loss due to digestive processes were similar in both preparations.     

Intranuclear Aggregation of Mutant FUS/TLS as a Molecular Pathomechanism of Amyotrophic Lateral Sclerosis

Dominant mutations in FUS/TLS cause a familial form of amyotrophic lateral sclerosis (fALS), where abnormal accumulation of mutant FUS proteins in cytoplasm has been observed as a major pathological change. Many of pathogenic mutations have been shown to deteriorate the nuclear localization signal in FUS and thereby facilitate cytoplasmic mislocalization of mutant proteins. Several other mutations, however, exhibit no effects on the nuclear localization of FUS in cultured cells, and their roles in the pathomechanism of fALS remain obscure. Here, we show that a pathogenic mutation, G156E, significantly increases the propensities for aggregation of FUS in vitro and in vivo. Spontaneous in vitro formation of amyloid-like fibrillar aggregates was observed in mutant but not wild-type FUS, and notably, those fibrils functioned as efficient seeds to trigger the aggregation of wild-type protein. In addition, the G156E mutation did not disturb the nuclear localization of FUS but facilitated the formation of intranuclear inclusions in rat hippocampal neurons with significant cytotoxicity. We thus propose that intranuclear aggregation of FUS triggered by a subset of pathogenic mutations is an alternative pathomechanism of FUS-related fALS diseases.     

UBE2E Ubiquitin-conjugating Enzymes and Ubiquitin Isopeptidase Y Regulate TDP-43 Protein Ubiquitination

Trans-activation element DNA-binding protein of 43 kDa (TDP-43) characterizes insoluble protein aggregates in distinct subtypes of frontotemporal lobar degeneration and amyotrophic lateral sclerosis. TDP-43 mediates many RNA processing steps within distinct protein complexes. Here we identify novel TDP-43 protein interactors found in a yeast two-hybrid screen using an adult human brain cDNA library. We confirmed the TDP-43 interaction of seven hits by co-immunoprecipitation and assessed their co-localization in HEK293E cells. As pathological TDP-43 is ubiquitinated, we focused on the ubiquitin-conjugating enzyme UBE2E3 and the ubiquitin isopeptidase Y (UBPY). When cells were treated with proteasome inhibitor, ubiquitinated and insoluble TDP-43 species accumulated. All three UBE2E family members could enhance the ubiquitination of TDP-43, whereas catalytically inactive UBE2E3^C145S was much less efficient. Conversely, silencing of UBE2E3 reduced TDP-43 ubiquitination. We examined 15 of the 48 known disease-associated TDP-43 mutants and found that one was excessively ubiquitinated. This strong TDP-43^K263E ubiquitination was further enhanced by proteasomal inhibition as well as UBE2E3 expression. Conversely, UBE2E3 silencing and expression of UBPY reduced TDP-43^K263E ubiquitination. Moreover, wild-type but not active site mutant UBPY reduced ubiquitination of TDP-43 C-terminal fragments and of a nuclear import-impaired mutant. In Drosophila melanogaster, UBPY silencing enhanced neurodegenerative TDP-43 phenotypes and the accumulation of insoluble high molecular weight TDP-43 and ubiquitin species. Thus, UBE2E3 and UBPY participate in the regulation of TDP-43 ubiquitination, solubility, and neurodegeneration.     

Mitochondrial Dysfunction and Decrease in Body Weight of a Transgenic Knock-in Mouse Model for TDP-43

The majority of amyotrophic lateral sclerosis (ALS) cases as well as many patients suffering from frontotemporal lobar dementia (FTLD) with ubiquitinated inclusion bodies show TDP-43 pathology, the protein encoded by the TAR DNA-binding protein (Tardbp) gene. We used recombinase-mediated cassette exchange to introduce an ALS patient cDNA into the mouse Tdp-43 locus. Expression levels of human A315T TDP-43 protein were 300% elevated in heterozygotes, whereas the endogenous mouse Tdp-43 was decreased to 20% of wild type levels as a result of disturbed feedback regulation. Heterozygous TDP-43^A315TKi mutants lost 10% of their body weight and developed insoluble TDP-43 protein starting as early as 3 months after birth, a pathology that was exacerbated with age. We analyzed the splicing patterns of known Tdp-43 target genes as well as genome-wide gene expression levels in different tissues that indicated mitochondrial dysfunction. In heterozygous mutant animals, we observed a relative decrease in expression of Parkin (Park2) and the fatty acid transporter CD36 along with an increase in fatty acids, HDL cholesterol, and glucose in the blood. As seen in transmission electron microscopy, neuronal cells in motor cortices of TDP-43^A315TKi animals had abnormal neuronal mitochondrial cristae formation. Motor neurons were reduced to 90%, but only slight motoric impairment was detected. The observed phenotype was interpreted as a predisease model, which might be valuable for the identification of further environmental or genetic triggers of neurodegeneration.     

丹蒌片治疗老年性主动脉瓣狭窄的临床疗效分析

目的:探讨丹蒌片对老年性主动脉瓣狭窄患者临床疗效及血清炎症因子水平的影响。方法:选取老年性主动脉瓣钙化患者共34例,将其随机分为丹蒌片治疗组(22例)及对照组(12例)。根据西雅图心绞痛问卷中文版、SF-36生活质量量表、中医临床证候评分比较两组的治疗效果,ELISA检测治疗前后患者血清肿瘤坏死因子α(TNF-α)、白介素6(IL-6)及同型半胱氨酸(HCY)的水平。结果:丹蒌片治疗组患者心绞痛程度及治疗满意度较常规治疗组均有明显统计学差异,丹蒌片可提高病人生活质量,改善中医部分证候,减轻血清炎症因子表达。结论:丹蒌片或可减缓主动脉瓣钙化进展,其原因可能和炎症反应减轻有关。     

正、反杂交二西蜂及其父、母本DNA基因组的多态性研究

使用RAPD-PCR(random amplified polymorphic DNA-polymerase chain reaction）技术,分析了正杂交松丹一号西蜂,反杂交松丹二号西蜂及其父,母本的DNA多态性,结果正杂交松丹一号西峰蜂,反杂交松丹二号西蜂及其父,母本的DNA多态性均有明显的差别,这说明通过正杂交松丹一号与反杂交松丹二号的DNA分子发生了变异。     

水稻花培品种单209抗病性的遗传学分析

单209是1977年由组合(IR26×农虎_6~2),经花药培养育成的粳稻品种。遗传学研究表明,该品种对稻瘟病和水稻白叶枯病的抗性受不同的单基因控制,而它们是由非轮回亲本IR26导入的。鉴于已发现的具白叶桔病抗性基因X(?)的材料均属籼稻。因此,单209可能是具该抗性基因的第1个粳稻品种。它已作为抗性亲本广泛应用于育种中。     

Usefulness of marine protected areas as tools for preserving the highly endangered limpet,Patella ferruginea,in the Mediterranean Sea

Patella ferruginea is the most endangered endemic marine invertebrate of western Mediterranean rocky shores. From 2012 to 2015, we performed a census and an ecological study for this gastropod mollusc in the Zembra Archipelago National Park in Tunisia, where a large population still remains. In 1986, a total of 20,000 individuals were estimated, with 0.7?ind/m², 4.4?cm adult mean size and low recruitment rate. In 2009, the estimated population reached 38,559 individuals with 2.65?ind/m², 5.42?cm adult mean size and high recruitment rate. During the present work, 43,790 individuals were estimated in 2012, 27,359 in 2013, 33,170 in 2014 and 40,404 in 2015, with an average density ranging from 0.1 to 8.5 individuals per square metre, but reaching as many as 25?ind/m² on some shores. This makes the archipelago one of the most important Mediterranean ‘hot spots’ for P. ferruginea, as it has a large and well-conserved population of this species. Variability of the population size is influenced by various biotic and abiotic factors, but is principally impacted by poaching of the species. The poachers target individuals larger than 6?cm in length, and the sex ratio study has shown that over 70% of individuals become females at that size. In addition, high recruitment rates were recorded in 2012 and 2013, but these decreased in 2014 and 2015, although population numbers show the opposite pattern. Moreover, growth rate is low and P. ferruginea may need many years to reach a large adult size. The loss of reproductive potential due to poaching inevitably causes a significant imbalance in the limpet population. Our results indicate the importance of the National Park for P. ferruginea and underline the need for this species to be protected and monitored over a long period to provide the most appropriate management measures.     

DJ-1 Is a Copper Chaperone Acting on SOD1 Activation

Lack of oxidative stress control is a common and often prime feature observed in many neurodegenerative diseases. Both DJ-1 and SOD1, proteins involved in familial Parkinson disease and amyotrophic lateral sclerosis, respectively, play a protective role against oxidative stress. Impaired activity and modified expression of both proteins have been observed in different neurodegenerative diseases. A potential cooperative action of DJ-1 and SOD1 in the same oxidative stress response pathway may be suggested based on a copper-mediated interaction between the two proteins reported here. To investigate the mechanisms underlying the antioxidative function of DJ-1 in relation to SOD1 activity, we investigated the ability of DJ-1 to bind copper ions. We structurally characterized a novel copper binding site involving Cys-106, and we investigated, using different techniques, the kinetics of DJ-1 binding to copper ions. The copper transfer between the two proteins was also examined using both fluorescence spectroscopy and specific biochemical assays for SOD1 activity. The structural and functional analysis of the novel DJ-1 copper binding site led us to identify a putative role for DJ-1 as a copper chaperone. Alteration of the coordination geometry of the copper ion in DJ-1 may be correlated to the physiological role of the protein, to a potential failure in metal transfer to SOD1, and to successive implications in neurodegenerative etiopathogenesis.