Recruitment kinetics of DNA repair proteins Mdc1 and Rad52 but not 53BP1 depend on damage complexity.

The recruitment kinetics of double-strand break (DSB) signaling and repair proteins Mdc1, 53BP1 and Rad52 into radiation-induced foci was studied by live-cell fluorescence microscopy after ion microirradiation. To investigate the influence of damage density and complexity on recruitment kinetics, which cannot be done by UV laser irradiation used in former studies, we utilized 43 MeV carbon ions with high linear energy transfer per ion (LET = 370 keV/µm) to create a large fraction of clustered DSBs, thus forming complex DNA damage, and 20 MeV protons with low LET (LET = 2.6 keV/µm) to create mainly isolated DSBs. Kinetics for all three proteins was characterized by a time lag period T(0) after irradiation, during which no foci are formed. Subsequently, the proteins accumulate into foci with characteristic mean recruitment times τ(1). Mdc1 accumulates faster (T(0) = 17 ± 2 s, τ(1) = 98 ± 11 s) than 53BP1 (T(0) = 77 ± 7 s, τ(1) = 310 ± 60 s) after high LET irradiation. However, recruitment of Mdc1 slows down (T(0) = 73 ± 16 s, τ(1) = 1050 ± 270 s) after low LET irradiation. The recruitment kinetics of Rad52 is slower than that of Mdc1, but exhibits the same dependence on LET. In contrast, the mean recruitment time τ(1) of 53BP1 remains almost constant when varying LET. Comparison to literature data on Mdc1 recruitment after UV laser irradiation shows that this rather resembles recruitment after high than low LET ionizing radiation. So this work shows that damage quality has a large influence on repair processes and has to be considered when comparing different studies

Anopheles stephensi p38 MAPK signaling regulates innate immunity and bioenergetics during Plasmodium falciparum infection.

BackgroundFruit flies and mammals protect themselves against infection by mounting immune and metabolic responses that must be balanced against the metabolic needs of the pathogens. In this context, p38 mitogen-activated protein kinase (MAPK)-dependent signaling is critical to regulating both innate immunity and metabolism during infection. Accordingly, we asked to what extent the Asian malaria mosquito Anopheles stephensi utilizes p38 MAPK signaling during infection with the human malaria parasite Plasmodium falciparum.MethodsA. stephensi p38 MAPK (AsP38 MAPK) was identified and patterns of signaling in vitro and in vivo (midgut) were analyzed using phospho-specific antibodies and small molecule inhibitors. Functional effects of AsP38 MAPK inhibition were assessed using P. falciparum infection, quantitative real-time PCR, assays for reactive oxygen species and survivorship under oxidative stress, proteomics, and biochemical analyses.ResultsThe genome of A. stephensi encodes a single p38 MAPK that is activated in the midgut in response to parasite infection. Inhibition of AsP38 MAPK signaling significantly reduced P. falciparum sporogonic development. This phenotype was associated with AsP38 MAPK regulation of mitochondrial physiology and stress responses in the midgut epithelium, a tissue critical for parasite development. Specifically, inhibition of AsP38 MAPK resulted in reduction in mosquito protein synthesis machinery, a shift in glucose metabolism, reduced mitochondrial metabolism, enhanced production of mitochondrial reactive oxygen species, induction of an array of anti-parasite effector genes, and decreased resistance to oxidative stress-mediated damage. Hence, P. falciparum-induced activation of AsP38 MAPK in the midgut facilitates parasite infection through a combination of reduced anti-parasite immune defenses and enhanced host protein synthesis and bioenergetics to minimize the impact of infection on the host and to maximize parasite survival, and ultimately, transmission.ConclusionsThese observations suggest that, as in mammals, innate immunity and mitochondrial responses are integrated in mosquitoes and that AsP38 MAPK-dependent signaling facilitates mosquito survival during parasite infection, a fact that may attest to the relatively longer evolutionary relationship of these parasites with their invertebrate compared to their vertebrate hosts. On a practical level, improved understanding of the balances and trade-offs between resistance and metabolism could be leveraged to generate fit, resistant mosquitoes for malaria control

Comparison of Michaelis-Menten kinetics modeling alternatives in cancer chemotherapy modeling

Model-based optimization and personalization of tumor therapies require tumor growth models that reliably describe the effect of the drug used during the therapy. A key phenomenon in the drug effect mechanism is the pharmacodynamics of the drugs which limits the maximal effect of the drug. The pharmacodynamics can be modeled with Michaelis-Menten kinetics, that can be realized in the differential equations of the model as a Hill function or bilinear functions with one extra state variable if we consider the quasi steady-state approximation or use triplet motifs, respectively. We use experimental data for a chemotherapeutic drug and carry out parametric identification of our tumor model with both Michealis-Menten kinetics models. The results show that the quasi steady-state approximation has better modeling power and less complexity

Survival of tumor cells after proton irradiation with ultra-high dose rates

<p>Abstract</p> <p>Background</p> <p>Laser acceleration of protons and heavy ions may in the future be used in radiation therapy. Laser-driven particle beams are pulsed and ultra high dose rates of >10⁹Gy s^-1may be achieved. Here we compare the radiobiological effects of pulsed and continuous proton beams.</p> <p>Methods</p> <p>The ion microbeam SNAKE at the Munich tandem accelerator was used to directly compare a pulsed and a continuous 20 MeV proton beam, which delivered a dose of 3 Gy to a HeLa cell monolayer within < 1 ns or 100 ms, respectively. Investigated endpoints were G2 phase cell cycle arrest, apoptosis, and colony formation.</p> <p>Results</p> <p>At 10 h after pulsed irradiation, the fraction of G2 cells was significantly lower than after irradiation with the continuous beam, while all other endpoints including colony formation were not significantly different. We determined the relative biological effectiveness (RBE) for pulsed and continuous proton beams relative to x-irradiation as 0.91 ± 0.26 and 0.86 ± 0.33 (mean and SD), respectively.</p> <p>Conclusions</p> <p>At the dose rates investigated here, which are expected to correspond to those in radiation therapy using laser-driven particles, the RBE of the pulsed and the (conventional) continuous irradiation mode do not differ significantly.</p

Poor Clinical Sensitivity of Rapid Antigen Test for Influenza A Pandemic (H1N1) 2009 Virus

Influenza A pandemic (H1N1) 2009 virus RNA was detected by reverse transcription–PCR in 144 clinical samples from Bonn, Germany. A common rapid antigen–based test detected the virus in only 11.1% of these samples. The paramount feature of rapid test–positive samples was high virus concentration. Antigen-based rapid tests appear unsuitable for virologic diagnostics in the current pandemic

Genomic determinants of Furin cleavage in diverse European SARS-related bat coronaviruses

The furin cleavage site (FCS) in SARS-CoV-2 is unique within the Severe acute respiratory syndrome-related coronavirus (SrC) species. We re-assessed diverse SrC from European horseshoe bats and analyzed the spike-encoding genomic region harboring the FCS in SARS-CoV-2. We reveal molecular features in SrC such as purine richness and RNA secondary structures that resemble those required for FCS acquisition in avian influenza viruses. We discuss the potential acquisition of FCS through molecular mechanisms such as nucleotide substitution, insertion, or recombination, and show that a single nucleotide exchange in two European bat-associated SrC may suffice to enable furin cleavage. Furthermore, we show that FCS occurrence is variable in bat- and rodent-borne counterparts of human coronaviruses. Our results suggest that furin cleavage sites can be acquired in SrC via conserved molecular mechanisms known in other reservoir-bound RNA viruses and thus support a natural origin of SARS-CoV-2

Az ökológiai és a konvencionális joghurtfogyasztás lehetséges szerepe a nem alkoholos zsírmájbetegség kezelésében

Napjainkban az egészséges életmódra törekvés fontos ösztönző tényező mind a bio-, mind a funkcionális élelmiszerek fogyasztására. A bioélelmiszerek kevesebb növényvédőszer-maradékot és szignifikánsan több, egészségre kedvező vegyületeket tartalmaznak, mint például a növényi termékek polifenoljait, valamint a tejben és a húskészít- ményekben található, többszörösen telítetlen zsírsavakat. A legújabb tanulmányok szerint a bél–máj tengely fontos szerepet játszik a nem alkoholos zsírmájbetegség (NAFLD) patogenezisében, így a pre- és probiotikumok terápiás eszközök lehetnek. Az ökológiai gazdálkodásból származó joghurtok egészségre gyakorolt hatásait az úgynevezett hagyományos joghurtokkal azért nehéz összehasonlítani, mert nincs kellően érzékeny biomarker. Célkitűzés: Munkánk során azt vizsgáltuk, hogy különböző biomarkerek mennyire használhatók annak kimutatására, hogy hogyan hat a hagyományos és az ökológiai joghurt a NAFLD-ben szenvedő betegek állapotára. Betegek és módszer: Prospektív, kohorszos vizsgálatot végeztünk 37 (életkor = 51,73 ± 11,82, férfi = 21, nő = 16), nem alkoholos zsírmájbetegségben szenvedő beteg bevonásával. A NAFLD diagnosztizálása ultrahangvizsgálaton és egyéb etiológia kizárásán alapult. A betegeket shear wave elasztográfiával is megvizsgáltuk a májfibrosis stádiumának meghatározására. A betegeket véletlenszerűen két csoportra osztottuk. A betegek egyenként 8 héten keresztül napi 300 gramm ökológiai (n = 21) vagy hagyományos (n = 16) gazdálkodásból származó joghurtot fogyasztottak. 37 ru- tin laboratóriumi adatot vizsgáltunk, mértünk 4 citokinkoncentrációt, 3 redoxhomeosztázis-markert és 14 testössze- tétel-értéket a joghurtok fogyasztása előtt, befejezéskor, valamint 12 héttel a befejezés után. Eredmények: Mindkét joghurt fogyasztását követően enyhén növekedett a D-vitamin-szint, és minimálisan csökkent a LDL-szint, a többi 35 rutin laboratóriumi adatban statisztikailag nem volt különbség. Az adiponektin (8793,64 ± 5365,98 pg/ml – 13 161,33 ± 8346,24 pg/ml) és a leptin (161,70 ± 158,41 – 277,94 ± 188,15 pg/ml) szintje megemelkedett a 8 hetes joghurtfogyasztás után a „hagyományos csoportban”. Ezzel szemben a kezelés után az adiponektinszintek szignifikáns csökkenését tapasztaltuk az „ökológiai csoportban” (12 017,57143 ± 7122,12 – 8833,5 ± 5216,17). A két csoportban csak az adiponektin tendenciája különbözött. Az indukált szabad gyökök mennyisége statisztikailag szintén alacsonyabb (10,05 ± 16,67 – 5,58 ± 4,41) volt közvetlenül a joghurtok fogyasz- tása után. A testösszetétel-mérés eredményei (testsúly, teljes testvíz, fehérje, ásványi anyagok, testzsírtömeg, vázizom- tömeg, testtömegindex, testzsírszázalék, visceralis zsír területe, alapanyagcsere, derék-csípő arány, a csontok ásványi- anyag-tartalma, testsejttömeg) között nem volt szignifikáns különbség. Következtetés: Ezek az adatok arra utalnak, hogy az adiponektin lehetséges biomarkerként szerepelhet a nem alkoho- los zsírmájbetegségben alkalmazott probiotikus kezelés hatékonyságának értékeléséhez. Munkánk az alapját képez- heti a jövőbeli tanulmányoknak, amelyek a (bio)joghurt fogyasztása és az egészségre gyakorolt hatások közötti összefüggéseket vizsgálják

Three-dimensional calibration targets for optical coherence tomography

The recent expansion of clinical applications for optical coherence tomography (OCT) is driving the development of approaches for consistent image acquisition. There is a simultaneous need for time-stable, easy-to-use imaging targets for calibration and standardization of OCT devices. We present calibration targets consisting of three-dimensional structures etched into nanoparticle-embedded resin. Spherical iron oxide nanoparticles with a predominant particle diameter of 400 nm were homogeneously dispersed in a two part polyurethane resin and allowed to harden overnight. These samples were then etched using a precision micromachining femtosecond laser with a center wavelength of 1026 nm, 100kHz repetition rate and 450 fs pulse duration. A series of lines in depth were etched, varying the percentage of inscription energy and speed of the translation stage moving the target with respect to the laser. Samples were imaged with a dual wavelength spectral-domain OCT system and point-spread function of nanoparticles within the target was measured

Sloths host Anhanga virus‐related phleboviruses across large distances in time and space

Sloths are genetically and physiologically divergent mammals. Phleboviruses are major arthropod-borne viruses (arboviruses) causing disease in humans and other animals globally. Sloths host arboviruses, but virus detections are scarce. A phlebovirus termed Anhanga virus (ANHV) was isolated from a Brazilian Linnaeus's two-toed sloth (Choloepus didactylus) in 1962. Here, we investigated the presence of phleboviruses in sera sampled in 2014 from 74 Hoffmann's two-toed (Choloepus hoffmanni, n = 65) and three-toed (Bradypus variegatus, n = 9) sloths in Costa Rica by broadly reactive RT-PCR. A clinically healthy adult Hoffmann's two-toed sloth was infected with a phlebovirus. Viral load in this animal was high at 8.5 × 107 RNA copies/ml. The full coding sequence of the virus was determined by deep sequencing. Phylogenetic analyses and sequence distance comparisons revealed that the new sloth virus, likely representing a new phlebovirus species, provisionally named Penshurt virus (PEHV), was most closely related to ANHV, with amino acid identities of 93.1%, 84.6%, 94.7% and 89.0% in the translated L, M, N and NSs genes, respectively. Significantly more non-synonymous mutations relative to ANHV occurred in the M gene encoding the viral glycoproteins and in the NSs gene encoding a putative interferon antagonist compared to L and N genes. This was compatible with viral adaptation to different sloth species and with micro-evolutionary processes associated with immune evasion during the genealogy of sloth-associated phleboviruses. However, gene-wide mean dN/dS ratios were low at 0.02–0.15 and no sites showed significant evidence for positive selection, pointing to comparable selection pressures within sloth-associated viruses and genetically related phleboviruses infecting hosts other than sloths. The detection of a new phlebovirus closely-related to ANHV, in sloths from Costa Rica fifty years after and more than 3,000 km away from the isolation of ANHV confirmed the host associations of ANHV-related phleboviruses with the two extant species of two-toed sloths