Summary Seizures are often the key manifestation of neurological diseases caused by pathogenic mutations in 169 of the genes that have so far been identified to affect mitochondrial function. ...Mitochondria are the main producers of ATP needed for normal electrical activities of neurons and synaptic transmission. Additionally, they have a central role in neurotransmitter synthesis, calcium homoeostasis, redox signalling, production and modulation of reactive oxygen species, and neuronal death. Hypotheses link mitochondrial failure to seizure generation through changes in calcium homoeostasis, oxidation of ion channels and neurotransmitter transporters by reactive oxygen species, a decrease in neuronal plasma membrane potential, and reduced network inhibition due to interneuronal dysfunction. Seizures, irrespective of their origin, represent an excessive acute energy demand in the brain. Accordingly, secondary mitochondrial dysfunction has been described in various epileptic disorders, including disorders that are mainly of non-mitochondrial origin. An understanding of the reciprocal relation between mitochondrial dysfunction and epilepsy is crucial to select appropriate anticonvulsant treatment and has the potential to open up new therapeutic approaches in the subset of epileptic disorders caused by mitochondrial dysfunction.
Emerging gene therapy approaches that aim to eliminate pathogenic mutations of mitochondrial DNA (mtDNA) rely on efficient degradation of linearized mtDNA, but the enzymatic machinery performing this ...task is presently unknown. Here, we show that, in cellular models of restriction endonuclease-induced mtDNA double-strand breaks, linear mtDNA is eliminated within hours by exonucleolytic activities. Inactivation of the mitochondrial 5'-3'exonuclease MGME1, elimination of the 3'-5'exonuclease activity of the mitochondrial DNA polymerase POLG by introducing the p.D274A mutation, or knockdown of the mitochondrial DNA helicase TWNK leads to severe impediment of mtDNA degradation. We do not observe similar effects when inactivating other known mitochondrial nucleases (EXOG, APEX2, ENDOG, FEN1, DNA2, MRE11, or RBBP8). Our data suggest that rapid degradation of linearized mtDNA is performed by the same machinery that is responsible for mtDNA replication, thus proposing novel roles for the participating enzymes POLG, TWNK, and MGME1.
Analysis of mitochondrial function is central to the study of intracellular energy metabolism, mechanisms of cell death and pathophysiology of a variety of human diseases, including myopathies, ...neurodegenerative diseases and cancer. However, important properties of mitochondria differ in vivo and in vitro. Here, we describe a protocol for the analysis of functional mitochondria in situ, without the isolation of organelles, in selectively permeabilized cells or muscle fibers using digitonin or saponin. A specially designed substrate/inhibitor titration approach allows the step-by-step analysis of several mitochondrial complexes. This protocol allows the detailed characterization of functional mitochondria in their normal intracellular position and assembly, preserving essential interactions with other organelles. As only a small amount of tissue is required for analysis, the protocol can be used in diagnostic settings in clinical studies. The permeabilization procedure and specific titration analysis can be completed in 2 h.
Olfactory testing is a useful tool in the differential diagnosis of Parkinson's Disease (PD). Although fast and easy to use, the high intercultural variability of odor detection limits the world-wide ...use of the most common test sets.
The aim of this study was to test one of the most commonly used olfactory tests (Sniffin' Sticks 12-identification test) in an adapted version for a Chinese population of healthy subjects and PD patients.
For this purpose, cohorts of 39 Chinese and 41 German PD patients as well as 70 Chinese and 100 German healthy subjects have been examined both with the original and the adapted version of the Sniffin' Sticks test, the latter being designed according to the regional culture.
The adapted Chinese version of the Sniffin' Sticks 12 identification test proved to discriminate Chinese PD patients from controls with a high specificity but relatively low sensitivity. Yet not all odor exchanges would have been necessary as the original odors including liquorice and coffee showed an equally high identification rate in the Chinese and German cohorts.
The results showed that the newly adapted test could be used as a screening test for PD related olfactory dysfunction in a Chinese population. However further investigation will be necessary to optimize the selection of odors for the Chinese version of the test.
Highlights • Challenging aspects of external postmortem examination are presented based on a case report of a shotgun suicide. • Possible reasons for unusual gunshot entrance and exit wounds are ...discussed. • A detailed forensic analysis is necessary in shotgun suicides, on the crime scene as well as during autopsy.
Sialic acid-binding Ig-like lectins (Siglecs) are members of the Ig superfamily that recognize sialic acid residues of glycoproteins. Siglec-E is a mouse CD33-related Siglec that preferentially binds ...to sialic acid residues of the cellular glycocalyx. Here, we demonstrate gene transcription and protein expression of Siglec-E by cultured mouse microglia. Siglec-E on microglia inhibited phagocytosis of neural debris and prevented the production of superoxide radicals induced by challenge with neural debris. Soluble extracellular Siglec-E receptor protein bound to the neural glycocalyx. Coculture of mouse microglia and neurons demonstrated a neuroprotective effect of microglial Siglec-E that was dependent on neuronal sialic acid residues. Increased neurotoxicity of microglia after knockdown of Siglece mRNA was neutralized by the reactive oxygen species scavenger Trolox. Data suggest that Siglec-E recognizes the intact neuronal glycocalyx and has neuroprotective function by preventing phagocytosis and the associated oxidative burst.
Mitochondrial respiratory chain complexes I and III have been shown to produce superoxide but the exact contribution and localization of individual sites have remained unclear. We approached this ...question investigating the effects of oxygen, substrates, inhibitors, and of the NAD+/NADH redox couple on H2O2 and superoxide production of isolated mitochondria from rat and human brain. Although rat brain mitochondria in the presence of glutamate+malate alone do generate only small amounts of H2O2 (0.04 ± 0.02 nmol H2O2/min/mg), a substantial production is observed after the addition of the complex I inhibitor rotenone (0.68 ± 0.25 nmol H2O2/min/mg) or in the presence of the respiratory substrate succinate alone (0.80 ± 0.27 nmol H2O2/min/mg). The maximal rate of H2O2 generation by respiratory chain complex III observed in the presence of antimycin A was considerably lower (0.14 ± 0.07 nmol H2O2/min/mg). Similar observations were made for mitochondria isolated from human parahippocampal gyrus. This is an indication that most of the superoxide radicals are produced at complex I and that high rates of production of reactive oxygen species are features of respiratory chain-inhibited mitochondria and of reversed electron flow, respectively. We determined the redox potential of the superoxide production site at complex I to be equal to –295 mV. This and the sensitivity to inhibitors suggest that the site of superoxide generation at complex I is most likely the flavine mononucleotide moiety. Because short-term incubation of rat brain mitochondria with H2O2 induced increased H2O2 production at this site we propose that reactive oxygen species can activate a self-accelerating vicious cycle causing mitochondrial damage and neuronal cell death.
Aims. As a sub-Uranus-mass low-density planet, GJ 3470b has been found to show a flat featureless transmission spectrum in the infrared and a tentative Rayleigh scattering slope in the optical. We ...conducted an optical transmission spectroscopy project to assess the impacts of stellar activity and to determine whether or not GJ 3470b hosts a hydrogen-rich gas envelop. Methods. We observed three transits with the low-resolution Optical System for Imaging and low-Intermediate-Resolution Integrated Spectroscopy (OSIRIS) at the 10.4 m Gran Telescopio Canarias, and one transit with the high-resolution Ultraviolet and Visual Echelle Spectrograph (UVES) at the 8.2 m Very Large Telescope. Results. From the high-resolution data, we find that the difference of the CaiiH+K lines in- and out-of-transit is only 0.67 + or - 0.22%, and determine a magnetic filling factor of about 10-15%. From the low-resolution data, we present the first optical transmission spectrum in the 435-755 nm band, which shows a slope consistent with Rayleigh scattering. Conclusions. After exploring the potential impacts of stellar activity in our observations, we confirm that Rayleigh scattering in an extended hydrogen-helium atmosphere is currently the best explanation. Further high-precision observations that simultaneously cover optical and infrared bands are required to answer whether or not clouds and hazes exist at high-altitude.
Representatives of
Bacillus amyloliquefaciens were shown to possess biocontrol activity against fire blight, a serious disease of orchard trees caused by
Erwinia amylovora. Genome analysis of
B. ...amyloliquefaciens FZB42 identified gene clusters responsible for synthesis of several polyketide compounds with antibacterial action. We show here that the antibacterial polyketides difficidin and to a minor extent bacillaene act efficiently against
E. amylovora. Surprisingly, a mutant strain blocked in the production of difficidin (CH8
Δdfn) inhibited growth of
E. amylovora and suppressed fire blight disease nearly in the same range as the wild type. In addition, a
sfp mutant (CH3
Δsfp) unable to synthesize non-ribosomally lipopeptides and polyketides did still suppress growth of
E. amylovora, suggesting that besides action of polyketides another antagonistic principle exist. A double mutant (RS06
Δsfp Δbac) devoid in polyketide and bacilysin synthesis was unable to suppress growth of
E. amylovora indicating that the additional inhibitory effect is due to production of bacilysin, a dipeptide whose synthesis does not depend on Sfp. We propose to use
B. amyloliquefaciens strains with enhanced synthesis of difficidin and/or bacilysin for development of biocontrol agents efficient against fire blight disease.
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