Alterations of Red Cell Membrane Properties in Nneuroacanthocytosis

Neuroacanthocytosis (NA) refers to a group of heterogenous, rare genetic disorders, namely chorea acanthocytosis (ChAc), McLeod syndrome (MLS), Huntington’s disease-like 2 (HDL2) and pantothenate kinase associated neurodegeneration (PKAN), that mainly affect the basal ganglia and are associated with similar neurological symptoms. PKAN is also assigned to a group of rare neurodegenerative diseases, known as NBIA (neurodegeneration with brain iron accumulation), associated with iron accumulation in the basal ganglia and progressive movement disorder. Acanthocytosis, the occurrence of misshaped erythrocytes with thorny protrusions, is frequently observed in ChAc and MLS patients but less prevalent in PKAN (about 10%) and HDL2 patients. The pathological factors that lead to the formation of the acanthocytic red blood cell shape are currently unknown. The aim of this study was to determine whether NA/NBIA acanthocytes differ in their functionality from normal erythrocytes. Several flow-cytometry-based assays were applied to test the physiological responses of the plasma membrane, namely drug-induced endocytosis, phosphatidylserine exposure and calcium uptake upon treatment with lysophosphatidic acid. ChAc red cell samples clearly showed a reduced response in drug-induced endovesiculation, lysophosphatidic acid-induced phosphatidylserine exposure, and calcium uptake. Impaired responses were also observed in acanthocyte-positive NBIA (PKAN) red cells but not in patient cells without shape abnormalities. These data suggest an “acanthocytic state” of the red cell where alterations in functional and interdependent membrane properties arise together with an acanthocytic cell shape. Further elucidation of the aberrant molecular mechanisms that cause this acanthocytic state may possibly help to evaluate the pathological pathways leading to neurodegeneration.     

Diffusion- and Perfusion-Weighted Imaging in Acute Lacunar Infarction: Is There a Mismatch?

Purpose

Characterization of lacunar infarction (LI) by use of multimodal MRI including diffusion- and perfusion-weighted imaging (DWI, PWI) is difficult because of the small lesion size. Only a few studies evaluated PWI in LI and the results are inconsistent.

Methods

In 16 LI patients who underwent initial MRI within 6 hours after symptom onset and follow-up MRI within 1 week demographics, clinical presentation, and MRI findings were analyzed with special emphasis on DWI and PWI findings. Time to peak maps were classified as showing a normal perfusion pattern or areas of hypoperfusion which were further categorized in mismatch (PWI>DWI), inverse mismatch (PWI<DWI), and match (PWI=DWI). Quantitative perfusion maps were generated and analyzed by use of Signal Processing in NMR-Software (SPIN).

Results

Of the 16 patients (mean age 65.5±12.9 years), 14 (87.5%) were male. Clinical symptoms comprised dysarthria (50%), hemiparesis (81.3%), and hemihypaesthesia (18.8%). Intravenous thrombolysis was performed in 7 (43.8%) patients. Clinical improvement was observed in 12 patients (75 %), while 2 (12.5%) patients showed a deterioration and another 2 (12.5%) a stable course. Acute ischemic lesions (mean volume of 0.46±0.29 cm³) were located in the thalamus (n=8, 50%), internal capsule (n=4, 25%), corona Radiata (n=3, 18.8%) and the mesencephalon (n=1, 6.3%). Circumscribed hypoperfusion (mean volume 0.61±0.48 cm³) was evident in 10 (62.5%) patients. Of these, 3 patients demonstrated a match, 4 an inverse mismatch, and 3 a mismatch between DWI and PWI lesion. Mean CBF and CBV ratios were 0.65±0.28 and 0.84±0.41 respectively. Growth of DWI lesions was observed in 7 (43.8%) and reversal of DWI lesions in 3 (18.8%) patients.

Conclusions

MRI allows identification of different DWI and PWI patterns in LI, including growth and reversal of ischemic lesions. Consequently, it may serve for a better characterization of this stroke subtype and support treatment decisions in daily clinical practice.     

Cardiac Per2 Functions as Novel Link between Fatty Acid Metabolism and Myocardial Inflammation during Ischemia and Reperfusion Injury of the Heart

Disruption of peripheral circadian rhyme pathways dominantly leads to metabolic disorders. Studies on circadian rhythm proteins in the heart indicated a role for Clock or Per2 in cardiac metabolism. In contrast to Clock^−/−, Per2^−/− mice have larger infarct sizes with deficient lactate production during myocardial ischemia. To test the hypothesis that cardiac Per2 represents an important regulator of cardiac metabolism during myocardial ischemia, we measured lactate during reperfusion in Per1^−/−, Per2^−/− or wildtype mice. As lactate measurements in whole blood indicated an exclusive role of Per2 in controlling lactate production during myocardial ischemia, we next performed gene array studies using various ischemia-reperfusion protocols comparing wildtype and Per2^−/− mice. Surprisingly, high-throughput gene array analysis revealed dominantly lipid metabolism as the differentially regulated pathway in wildtype mice when compared to Per2^−/−. In all ischemia-reperfusion protocols used, the enzyme enoyl-CoA hydratase, which is essential in fatty acid beta-oxidation, was regulated in wildtype animals only. Studies using nuclear magnet resonance imaging (NMRI) confirmed altered fatty acid populations with higher mono-unsaturated fatty acid levels in hearts from Per2^−/− mice. Unexpectedly, studies on gene regulation during reperfusion revealed solely pro inflammatory genes as differentially regulated ‘Per2-genes’. Subsequent studies on inflammatory markers showed increasing IL-6 or TNFα levels during reperfusion in Per2^−/− mice. In summary, these studies reveal an important role of cardiac Per2 for fatty acid metabolism and inflammation during myocardial ischemia and reperfusion, respectively.     

Distribution and Ecophysiology of Calanoid Copepods in Relation to the Oxygen Minimum Zone in the Eastern Tropical Atlantic

Oxygen minimum zones (OMZs) affect distribution patterns, community structure and metabolic processes of marine organisms. Due to the prominent role of zooplankton, especially copepods, in the marine carbon cycle and the predicted intensification and expansion of OMZs, it is essential to understand the effects of hypoxia on zooplankton distribution and ecophysiology. For this study, calanoid copepods were sampled from different depths (0–1800 m) at eight stations in the eastern tropical Atlantic (3°47′N to 18°S) during three expeditions in 2010 and 2011. Their horizontal and vertical distribution was determined and related to the extent and intensity of the OMZ, which increased from north to south with minimum O₂ concentrations (12.7 µmol kg⁻¹) in the southern Angola Gyre. Calanoid copepod abundance was highest in the northeastern Angola Basin and decreased towards equatorial regions as well as with increasing depth. Maximum copepod biodiversity was observed in the deep waters of the central Angola Basin. Respiration rates and enzyme activities were measured to reveal species-specific physiological adaptations. Enzyme activities of the electron transport system (ETS) and lactate dehydrogenase (LDH) served as proxies for aerobic and anaerobic metabolic activity, respectively. Mass-specific respiration rates and ETS activities decreased with depth of occurrence, consistent with vertical changes in copepod body mass and ambient temperature. Copepods of the families Eucalanidae and Metridinidae dominated within the OMZ. Several of these species showed adaptive characteristics such as lower metabolic rates, additional anaerobic activity and diel vertical migration that enable them to successfully inhabit hypoxic zones.     

Equivalence of regression curves sharing common parameters

In clinical trials, the comparison of two different populations is a common problem. Nonlinear (parametric) regression models are commonly used to describe the relationship between covariates, such as concentration or dose, and a response variable in the two groups. In some situations, it is reasonable to assume some model parameters to be the same, for instance, the placebo effect or the maximum treatment effect. In this paper, we develop a (parametric) bootstrap test to establish the similarity of two regression curves sharing some common parameters. We show by theoretical arguments and by means of a simulation study that the new test controls its significance level and achieves a reasonable power. Moreover, it is demonstrated that under the assumption of common parameters, a considerably more powerful test can be constructed compared with the test that does not use this assumption. Finally, we illustrate the potential applications of the new methodology by a clinical trial example.     

Fungizide: Risiken der resistenzentwicklung und suche nach neuen targets

Trotz intensiver Bemühungen im Bereich des Pflanzenschutzes sind weltweit erhebliche Verluste im Bereich pflanzlicher Erntegüter zu verzeichnen. Rund ein Drittel der Verluste werden mikrobiellen Schaderregern, also Pilzen, Bakterien und Viren, zugeschrieben. Besonders schwierig ist der Tatsache zu begegnen, dass Pilze gegenüber spezifisch wirkenden Fungiziden Insensitivitäten bzw. Resistenzen entwickeln. Aus diesem Grunde muss über geeignete Strategien beim Fungizideinsatz, beim ‘Disease management’ und bei der Suche nach neuen, hochwirksamen Fungiziden nachgedacht werden. Der Erfolg bei der Suche nach neuen Fungiziden wird stark davon abhängen, ob neue Zielmoleküle für potentielle Fungizide entdeckt werden können. Dabei können moderne molekulargenetische Techniken von großem Nutzen sein.     

Robust RNAi enhancement via human Argonaute-2 overexpression from plasmids,viral vectors and cell lines

As the only mammalian Argonaute protein capable of directly cleaving mRNAs in a small RNA-guided manner, Argonaute-2 (Ago2) is a keyplayer in RNA interference (RNAi) silencing via small interfering (si) or short hairpin (sh) RNAs. It is also a rate-limiting factor whose saturation by si/shRNAs limits RNAi efficiency and causes numerous adverse side effects. Here, we report a set of versatile tools and widely applicable strategies for transient or stable Ago2 co-expression, which overcome these concerns. Specifically, we engineered plasmids and viral vectors to co-encode a codon-optimized human Ago2 cDNA along with custom shRNAs. Furthermore, we stably integrated this Ago2 cDNA into a panel of standard human cell lines via plasmid transfection or lentiviral transduction. Using various endo- or exogenous targets, we demonstrate the potential of all three strategies to boost mRNA silencing efficiencies in cell culture by up to 10-fold, and to facilitate combinatorial knockdowns. Importantly, these robust improvements were reflected by augmented RNAi phenotypes and accompanied by reduced off-targeting effects. We moreover show that Ago2/shRNA-co-encoding vectors can enhance and prolong transgene silencing in livers of adult mice, while concurrently alleviating hepatotoxicity. Our customizable reagents and avenues should broadly improve future in vitro and in vivo RNAi experiments in mammalian systems.     

Polymorphisms and genetic linkage of histamine receptors

Histamine is a biogenic amine that plays an essential role in controlling many physiological functions, both in the central nervous system (CNS) and the peripheral nervous system (PNS). Most of these physiological effects are mediated through interactions with four histamine receptor subtypes, all of which are members of the larger family of rhodopsin-like class A G-protein coupled receptors (GPCRs) (Leurs et al., 2011; Lim et al., 2009). Here, we focus on the genetic variations and polymorphisms localized on the genes encoding for human histamine receptors where it provides an up to date collection of all polymorphisms found on genes encoding the histamine receptor subtypes and their association to diseases.