Die Zellkernveränderungen unter der Einwirkung von Ribonuklease und β-Merkaptoäthanol in der Zellkultur

Zusammenfassung Nach der Einwirkung von RNase und -Merkaptoäthanol auf Zellkulturen treten außer unspezifischen Veränderungen des Cytoplasmas charakteristische Kernumwandlungen auf. Diese ähneln bei der RNase den Kernveränderungen bei mit Herpes-Virus inokulierten Kulturen und den bei Herpes-Infektionen auftretenden Kerneinschlußkörpern. — Das -Merkaptoäthanol führt zu Chromatinverklumpung, die aus dem Kerninnenraum ausgestoßen werden. Es wird auf den verschiedenen Angriffsort dieser beiden Substanzen im Zellstoffwechsel hingewiesen.     

Significance of the hydrophobic residues 225–230 of apoA-I for the biogenesis of HDL

We studied the significance of four hydrophobic residues within the 225–230 region of apoA-I on its structure and functions and their contribution to the biogenesis of HDL. Adenovirus-mediated gene transfer of an apoA-I[F225A/V227A/F229A/L230A] mutant in apoA-I^−/− mice decreased plasma cholesterol, HDL cholesterol, and apoA-I levels. When expressed in apoA-I^−/− × apoE^−/− mice, approximately 40% of the mutant apoA-I as well as mouse apoA-IV and apoB-48 appeared in the VLDL/IDL/LDL. In both mouse models, the apoA-I mutant generated small spherical particles of pre-β- and α4-HDL mobility. Coexpression of the apoA-I mutant and LCAT increased and shifted the-HDL cholesterol peak toward lower densities, created normal αHDL subpopulations, and generated spherical-HDL particles. Biophysical analyses suggested that the apoA-I[225–230] mutations led to a more compact folding that may limit the conformational flexibility of the protein. The mutations also reduced the ability of apoA-I to promote ABCA1-mediated cholesterol efflux and to activate LCAT to 31% and 66%, respectively, of the WT control. Overall, the apoA-I[225–230] mutations inhibited the biogenesis of-HDL and led to the accumulation of immature pre-β- and α4-HDL particles, a phenotype that could be corrected by administration of LCAT.     

Circadian changes in Drosophila motor terminals

In Drosophila melanogaster, as in most other higher organisms, a circadian clock controls the rhythmic distribution of rest/sleep and locomotor activity. Here we report that the morphology of Drosophila flight neuromuscular terminals changes between day and night, with a rhythm in synaptic bouton size that continues in constant darkness, but is abolished during aging. Furthermore, arrhythmic mutations in the clock genes timeless and period also disrupt this circadian rhythm. Finally, these clock mutants also have an opposing effect on the nonrhythmic phenotype of neuronal branching, with tim mutants showing a dramatic hyperbranching morphology and per mutants having fewer branches than wild-type flies. These unexpected results reveal further circadian as well as nonclock related pleiotropic effects for these classic behavioral mutants.     

Mosquito population structure,pathogen surveillance and insecticide resistance monitoring in urban regions of Crete,Greece

BackgroundIn Greece vector borne diseases (VBD) and foremost West Nile virus (WNV) pose an important threat to public health and the tourist industry, the primary sector of contribution to the national economy. The island of Crete, is one of Greece’s major tourist destinations receiving annually over 5 million tourists making regional VBD control both a public health and economic priority.MethodologyUnder the auspices of the Region of Crete, a systematic integrative surveillance network targeting mosquitoes and associated pathogens was established in Crete for the years 2018–2020. Using conventional and molecular diagnostic tools we investigated the mosquito species composition and population dynamics, pathogen infection occurrences in vector populations and in sentinel chickens, and the insecticide resistance status of the major vector species.Principal findingsImportant disease vectors were recorded across the island including Culex pipiens, Aedes albopictus, and Anopheles superpictus. Over 75% of the sampled specimens were collected in the western prefectures potentially attributed to the local precipitation patterns, with Cx. pipiens being the most dominant species. Although no pathogens (flaviviruses) were detected in the analysed mosquito specimens, chicken blood serum analyses recorded a 1.7% WNV antibody detection rate in the 2018 samples. Notably detection of the first WNV positive chicken preceded human WNV occurrence in the same region by approximately two weeks. The chitin synthase mutation I1043F (associated with high diflubenzuron resistance) was recorded at an 8% allelic frequency in Lasithi prefecture Cx. pipiens mosquitoes (sampled in 2020) for the first time in Greece. Markedly, Cx. pipiens populations in all four prefectures were found harboring the kdr mutations L1014F/C/S (associated with pyrethroid resistance) at a close to fixation rate, with mutation L1014C being the most commonly found allele (≥74% representation). Voltage gated sodium channel analyses in Ae. albopictus revealed the presence of the kdr mutations F1534C and I1532T (associated with putative mild pyrethroid resistance phenotypes) yet absence of V1016G. Allele F1534C was recorded in all prefectures (at an allelic frequency range of 25–46.6%) while I1532T was detected in populations from Chania, Rethymnon and Heraklion (at frequencies below 7.1%). Finally, no kdr mutations were detected in the Anopheles specimens included in the analyses.Conclusions/SignificanceThe findings of our study are of major concern for VBD control in Crete, highlighting (i) the necessity for establishing seasonal integrated entomological/pathogen surveillance programs, supporting the design of targeted vector control responses and; ii) the need for establishing appropriate insecticide resistance management programs ensuring the efficacy and sustainable use of DFB and pyrethroid based products in vector control.     

Type III protein translocase: HrcN is a peripheral ATPase that is activated by oligomerization

Type III protein secretion (TTS) is catalyzed by translocases that span both membranes of Gram-negative bacteria. A hydrophilic TTS component homologous to F1/V1-ATPases is ubiquitous and essential for secretion. We show that hrcN encodes the putative TTS ATPase of Pseudomonas syringae pathovar phaseolicola and that HrcN is a peripheral protein that assembles in clusters at the membrane. A decahistidinyl HrcN derivative was overexpressed in Escherichia coli and purified to homogeneity in a folded state. Hydrodynamic analysis, cross-linking, and electron microscopy revealed four distinct HrcN forms: I, 48 kDa (monomer); II, approximately 300 kDa (putative hexamer); III, 575 kDa (dodecamer); and IV, approximately 3.5 MDa. Form III is the predominant form of HrcN at the membrane, and its ATPase activity is dramatically stimulated (>700-fold) over the basal activity of Form I. We propose that TTS ATPases catalyze protein translocation as activated homo-oligomers at the plasma membrane.     

Immunotargeting of catalase to the pulmonary endothelium alleviates oxidative stress and reduces acute lung transplantation injury

Vascular immunotargeting may facilitate the rapid and specific delivery of therapeutic agents to endothelial cells. We investigated whether targeting of an antioxidant enzyme, catalase, to the pulmonary endothelium alleviates oxidative stress in an in vivo model of lung transplantation. Intravenously injected enzymes, conjugated with an antibody to platelet-endothelial cell adhesion molecule-1, accumulate in the pulmonary vasculature and retain their activity during prolonged cold storage and transplantation. Immunotargeting of catalase to donor rats augments the antioxidant capacity of the pulmonary endothelium, reduces oxidative stress, ameliorates ischemia-reperfusion injury, prolongs the acceptable cold ischemia period of lung grafts, and improves the function of transplanted lung grafts. These findings validate the therapeutic potential of vascular immunotargeting as a drug delivery strategy to reduce endothelial injury. Potential applications of this strategy include improving the outcome of clinical lung transplantation and treating a wide variety of endothelial disorders.     

Primary Malignant Melanoma of the Lung: A Case Report

BACKGROUND: Primary melanoma of the lung is an extremely rare pathological entity and sparsely reported in the literature. CASE PRESENTATION: A case of primary malignant melanoma of the lung in a 41-year-old female is reported. The clinical, radiological and histopathological features are discussed. The initial symptom was cough, whereas the chest radiography showed a round opacity of the right lung. The computed tomography of the chest revealed a well-demarcated mass lesion in the right upper lobe. Endobronchial mass causing obstruction of the upper lobar bronchus was the bronchoscopic finding. Patient underwent pneumonectomy. A diagnosis of melanoma was confirmed postoperatively after the immunohistochemistry. Primary nature of the tumour in the lung results from the demonstration of characteristic junctional pattern of melanoma cells beneath the bronchial epithelium on histopathology, and from exclusion of other potential primary sites in the clinical, paraclinical and laboratory examination. CONCLUSIONS: Primary melanoma of the lung represents a rare pathological entity. Careful interpretation of histopathological information in correlation with all other findings from clinical and paraclinical studies can establish a diagnosis. Follow-up is necessary in order to diagnose potential dissemination or secondary sites of the disease. Due to the small number of cases reported in the literature, there is no experience on the management and the prognosis of the disease, but surgical resection remains the cornerstone of the treatment.     

Investigating centre of mass stabilisation as the goal of posture and movement coordination during human whole body reaching

In the light of experimental results showing significant forward centre of mass (CoM) displacements within the base of support, this study investigated if whole body reaching movements can be executed whilst keeping the CoM fixed in the horizontal axis. Using kinematic simulation techniques, angular configurations were recreated from experimental data imposing two constraints: a constant horizontal position of the CoM and an identical trajectory of the hand to grasp an object. The comparison between recorded and simulated trials showed that stabilisation of the CoM was associated with greater backward hip displacements, which became more marked with increasing object distance. This was in contrast to recorded trials showing reductions in backward hip displacements with increasing distance. Results also showed that modifications to angular displacements were necessary only at the shoulder and hip joints, but that these modifications were within the limits of joint mobility. The analysis of individual joint torques revealed that the pattern and timing of simulated trials were similar to those recorded experimentally. Peak joint torque values showed particularly that keeping the CoM at a constant horizontal position resulted in significantly smaller ankle peak flexor and extensor torques. It may be concluded from this study that `stabilising' the CoM during human whole body reaching represents a feasible strategy, but not the one chosen by subjects under experimental conditions. Our results also do not support the idea of the CoM as the stabilised reference value for the coordination between posture and goal-directed movements. Received: 22 September 1998 / Accepted in revised form: 2 June 1999     

Understanding Market Size and Reporting Gaps for Paediatric TB in Indonesia,Nigeria and Pakistan: Supporting Improved Treatment of Childhood TB in the Advent of New Medicines

MethodologyExploratory assessments were carried out in Indonesia, Nigeria and Pakistan, reaching a range of facility types in two selected areas of each country. Record reviews and interviews of healthcare providers were carried out to assess numbers of unreported paediatric TB cases, diagnostic pathways followed and treatment regimens prescribed.

Main Findings

A total of 985 unreported diagnosed paediatric TB cases were identified over a three month period in 2013 in Indonesia from 64 facilities, 463 in Pakistan from 35 facilities and 24 in Nigeria from 20 facilities. These represent an absolute additional annualised yield to 2013 notifications reported to WHO of 15% for Indonesia, 2% for Nigeria and 7% for Pakistan. Only 12% of all facilities provided age and sex-disaggregated data. Findings highlight the challenges of confirming childhood TB. Diagnosis patterns in Nigeria highlight a very low suspicion for childhood TB. Providers note the need for paediatric medicines aligned to WHO recommendations.

Conclusion: How Market Data Can Support Better Public Health Interventions

This study emphasises the impact of incomplete reporting on the estimation of disease burden and potential market size of paediatric TB medicines. Further studies on “hubs” (facilities treating large numbers of childhood TB cases) will improve our understanding of the epidemic, support introduction efforts for new treatments and better measure markets for new paediatric medicines.     

Circadian rhythm gene regulation in the housefly Musca domestica

The circadian mechanism appears remarkably conserved between Drosophila and mammals, with basic underlying negative and positive feedback loops, cycling gene products, and temporally regulated nuclear transport involving a few key proteins. One of these negative regulators is PERIOD, which in Drosophila shows very similar temporal and spatial regulation to TIMELESS. Surprisingly, we observe that in the housefly, Musca domestica, PER does not cycle in Western blots of head extracts, in contrast to the TIM protein. Furthermore, immunocytochemical (ICC) localization using enzymatic staining procedures reveals that PER is not localized to the nucleus of any neurons within the brain at any circadian time, as recently observed for several nondipteran insects. However, with confocal analysis, immunofluorescence reveals a very different picture and provides an initial comparison of PER/TIM-containing cells in Musca and Drosophila, which shows some significant differences, but many similarities. Thus, even in closely related Diptera, there is considerable evolutionary flexibility in the number and spatial organization of clock cells and, indeed, in the expression patterns of clock products in these cells, although the underlying framework is similar.