Pathogenesis of guinea pig adenovirus infection

BACKGROUND AND PURPOSE: The existence of guinea pig adenovirus (GPAdV) has been suspected on the basis of histopathologic findings, but the virus has not yet been isolated. In susceptible animals, it may cause severe bronchopneumonia and death. Adenovirus-like inclusion bodies have been observed in the lungs of animals with clinical disease. Prevalence of the infection is unknown. Recently, a polymerase chain reaction (PCR) assay was described that was able to selectively detect GPAdV. METHODS: To investigate the pathogenesis of GPAdV, we inoculated eight guinea pigs with GPAdV; eight control animals were sham inoculated. The PCR assay was used to trace the infection. In a second experiment, transmission of GPAdV from an experimentally infected animal to five immune-naive cohorts was examined. RESULTS: None of the infected animals developed clinical disease. The GPAdV could be detected by PCR analysis of nasal-swab specimens on days 6 through 15 after infection. Infective virus could be recovered from the nasal mucosa during this period (as determined by inoculation of immune-naive animals). The virus was transmitted from an experimentally infected animal to two of five immune-naive cage mates. CONCLUSION: The GPAdV may cause transient subclinical upper respiratory tract infection that may descend to the lungs.     

Cardiac and skeletal muscle mitochondria have a monocarboxylate transporter MCT1.

To evaluate the potential role of monocarboxylate transporter-1 (MCT1) in tissue lactate oxidation, isolated rat subsarcolemmal and interfibrillar cardiac and skeletal muscle mitochondria were probed with an antibody to MCT1. Western blots indicated presence of MCT1 in sarcolemmal membranes and in subsarcolemmal and interfibrillar mitochondria. Minimal cross-contamination of mitochondria by cell membrane fragments was verified by probing for the sarcolemmal protein GLUT-1. In agreement, immunolabeling and electron microscopy showed mitochondrial MCT1 in situ. Along with lactic dehydrogenase, the presence of MCT1 in striated muscle mitochondria permits mitochondrial lactate oxidation and facilitates function of the "intracellular lactate shuttle."     

Optical and TDPAC spectroscopy of Hg(II)-rubredoxin: model for a mononuclear tetrahedral [Hg(CysS)4]2− center

Rubredoxins possess a well-defined mononuclear tetrahedral tetrathiolate metal binding site, a feature exploited by several investigations to study the spectroscopic characteristics and the coordination chemistry of different metal ions at this binding site. In the present work, Hg(II)-substituted rubredoxin (Rd) from Desulfovibrio gigas has been studied by electronic absorption, circular dichroism (CD), magnetic circular dichroism (MCD), and time differential perturbed angular correlation of γ-rays (TDPAC) spectroscopies. The TDPAC spectrum of ^199mHg-Rd at pH 8 exhibits a prevailing nuclear quadrupole interaction (NQI) with a precession frequency of ω₁=0.09 Grad/s and an asymmetry parameter η=0, features characteristic of a slightly distorted tetrahedral tetrathiolate metal coordination, i.e, a HgCysS₄ center. In addition, three minor populated NQIs have also been detected. They may represent a trigonal HgS₃ (ω₁=1.13 Grad/s, η=0.21), a digonal HgS₂ (ω₁=1.34 Grad/s, η=0.20), and a digonal Hg(II) coordination (ω₁=1.58 Grad/s, η= 0.18) with unidentified ligands. Since similar studies at pH 2.5 revealed a time-dependent increase of the HgCysS₄ population, the low populated sites may represent intermediate Hg(II) complexes formed prior to the generation of the thermodynamically stable structure. The metal-induced absorption envelope of Hg-Rd reveals three distinct transitions with Gaussian-resolved maxima located at 230, 257, and 284 nm, which are paralleled by dichroic features in the corresponding difference CD spectrum of Hg(II)-Rd versus apo-Rd. Based on the optical electronegativity theory of Jørgensen, the lowest energy transition has been attributed to a CysS-Hg(II) charge-transfer excitation. The T _d type of metal coordination in Hg-Rd is supported by the presence of an unresolved A-term with a negative lobe at 295 nm in the difference MCD spectrum. These results point to the usefulness of optical and TDPAC spectroscopies for studying Hg(II) sites in other proteins.     

Oscillatory coupling in writing and writer's cramp.

Writing is a highly skilled and overlearned movement. In patients suffering from writer's cramp, a focal task-induced dystonia, writing is impaired or even impossible due to involuntary muscle contractions and abnormal posture, which occur as soon as the person picks up a pen or within writing a few words. The underlying pathophysiological mechanisms of this movement disorder are not fully understood up to now. The aim of the present study was to unravel the oscillatory network underlying physiological writing in healthy subjects and dystonic writing in writer's cramp patients. Using whole-head magnetoencephalography (MEG) and the analysis tool dynamic imaging of coherent sources (DICS) we studied oscillatory neural coupling during writing in eleven healthy subjects and eight patients suffering from writer's cramp. Simultaneous recording of brain activity with MEG and activity of forearm and hand muscles with surface electromyography (EMG) was performed while subjects were writing for five minutes with their dominant right hand. Applying DICS sources of strongest cerebro-muscular coherence and cerebro-cerebral coherence during writing were identified, which consistently included six brain areas in both, the control subjects and the patients: contralateral and ipsilateral sensorimotor cortex, ipsilateral cerebellum, contralateral thalamus, contralateral premotor and posterior parietal cortex. Coherence between cortical sources and muscles appeared primarily in the frequency of writing movements (3-7 Hz) while coherence between cerebral sources occurred primarily around 10 Hz (8-13 Hz). Interestingly, consistent coupling between both sensorimotor cortices was observed in patients only, whereas coupling between ipsilateral cerebellum and the contralateral posterior parietal cortex was found in control subjects only. These results are consistent with the often described bilateral pathophysiology and impaired sensorimotor integration in writer's cramp patients.     

Cell analysis with the new Leipzig high-energy ion nanoprobe

The high-energy ion nanoprobe LIPSION at the University of Leipzig has been in operation since 1998. The ultrastable, 3.5 MV SINLETRON accelerator supplies the H+ or He+ ion beam. A magnetic scanning system moves the focused beam across the sample. At present, a resolution of 41 +/- 4 nm in the low current mode and 300 nm at 5 pA can be achieved. The experimental chamber is equipped with electron-, energy dispersive X-ray-, and particle detectors. They can be used simultaneously to analyse the sample by means of PIXE (particle induced X-ray emission), RBS (Rutherford backscattering), and in the case of thin sections or monolayer samples STIM (scanning transmission ion microscopy). A goniometer allows the application of channeling measurements in single crystals in combination with these methods. In contrast to previous publication describing microbeam facility at LIPSION, the current biomedical research has concentrated on microscopy and tomography on chondrocytes in pig cartilages and fixed single endothelial cells (HUVEC). For the irradiation of single living cells, an external beam facility with irradiation platform, fast beamgate and mini-Petri dishes is under construction.     

The human ubiquitin-conjugating enzyme Cdc34 controls cellular proliferation through regulation of p27Kip1 protein levels

Ubiquitin-mediated degradation of the cyclin-dependent kinase inhibitor p27Kip1 was shown to be required for the activation of key cyclin-dependent kinases, thereby triggering the onset of DNA replication and cell cycle progression. Although the SCFSkp2 ubiquitin ligase has been reported to mediate p27Kip1 degradation, the nature of the human ubiquitin-conjugating enzyme involved in this process has not yet been determined at the cellular level. Here, we show that antisense oligonucleotides targeting the human ubiquitin-conjugating enzyme Cdc34 downregulate its expression, inhibit the degradation of p27Kip1, and prevent cellular proliferation. Elevation of p27Kip1 protein level is found to be the sole requirement for the inhibition of cellular proliferation induced upon downregulation of Cdc34. Indeed, reducing the expression of p27Kip1 with a specific antisense oligonucleotide is sufficient to reverse the anti-proliferative phenotype elicited by the Cdc34 antisense. Furthermore, downregulation of Cdc34 is found to specifically increase the abundance of the SCFSkp2) ubiquitin ligase substrate p27Kip1, but has no concomitant effect on the level of IkBalpha and beta-catenin, which are known substrates of a closely related SCF ligase.