SARS-CoV-2 sensing by RIG-I and MDA5 links epithelial infection to macrophage inflammation

SARS-CoV-2 infection causes broad-spectrum immunopathological disease, exacerbated by inflammatory co-morbidities. A better understanding of mechanisms underpinning virus-associated inflammation is required to develop effective therapeutics. Here we discover that SARS-CoV-2 replicates rapidly in lung epithelial cells despite triggering a robust innate immune response through activation of cytoplasmic RNA-sensors RIG-I and MDA5. The inflammatory mediators produced during epithelial cell infection can stimulate primary human macrophages to enhance cytokine production and drive cellular activation. Critically, this can be limited by abrogating RNA sensing, or by inhibiting downstream signalling pathways. SARS-CoV-2 further exacerbates the local inflammatory environment when macrophages or epithelial cells are primed with exogenous inflammatory stimuli. We propose that RNA sensing of SARS-CoV-2 in lung epithelium is a key driver of inflammation, the extent of which is influenced by the inflammatory state of the local environment, and that specific inhibition of innate immune pathways may beneficially mitigate inflammation-associated COVID-19

Expansion of anti-AFP Th1 and Tc1 responses in hepatocellular carcinoma occur in different stages of disease

Copyright @ 2010 Cancer Research UK. This work is licensed under the Creative Commons Attribution-NonCommercial-Share Alike 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/3.0/.Background: α-Fetoprotein (AFP) is a tumour-associated antigen in hepatocellular carcinoma (HCC) and is a target for immunotherapy. However, there is little information on the pattern of CD4 (Th1) and CD8 (Tc1) T-cell response to AFP in patients with HCC and their association with the clinical characteristics of patients. Methods: We therefore analysed CD4 and CD8 T-cell responses to a panel of AFP-derived peptides in a total of 31 HCC patients and 14 controls, using an intracellular cytokine assay for IFN-γ. Results: Anti-AFP Tc1 responses were detected in 28.5% of controls, as well as in 25% of HCC patients with Okuda I (early tumour stage) and in 31.6% of HCC patients with stage II or III (late tumour stages). An anti-AFP Th1 response was detected only in HCC patients (58.3% with Okuda stage I tumours and 15.8% with Okuda stage II or III tumours). Anti-AFP Th1 response was mainly detected in HCC patients who had normal or mildly elevated serum AFP concentrations (P=0.00188), whereas there was no significant difference between serum AFP concentrations in these patients and the presence of an anti-AFP Tc1 response. A Th1 response was detected in 44% of HCC patients with a Child–Pugh A score (early stage of cirrhosis), whereas this was detected in only 15% with a B or C score (late-stage cirrhosis). In contrast, a Tc1 response was detected in 17% of HCC patients with a Child–Pugh A score and in 46% with a B or C score. Conclusion: These results suggest that anti-AFP Th1 responses are more likely to be present in patients who are in an early stage of disease (for both tumour stage and liver cirrhosis), whereas anti-AFP Tc1 responses are more likely to be present in patients with late-stage liver cirrhosis. Therefore, these data provide valuable information for the design of vaccination strategies against HCC.Association for International Cancer Research and Polkemmet Fund, London Clinic

The Duration of Antigen-Stimulation Significantly Alters the Diversity of Multifunctional CD4 T Cells Measured by Intracellular Cytokine Staining

The assessment of antigen-specific T cell responses by intracellular cytokine staining (ICS) has become a routine technique in studies of vaccination and immunity. Here, we highlight how the duration of in vitro antigen pre-stimulation, combined with the cytokine accumulation period, are critical parameters of these methods. The effect of varying these parameters upon the diversity and frequency of multifunctional CD4 T cell subsets has been investigated using a murine model of TB vaccination and in cattle naturally infected with Mycobacterium bovis. We demonstrate a substantial influence of the duration of the antigen pre-stimulation period on the repertoire of the antigen-specific CD4 T cell responses. Increasing pre-stimulation from 2 to 6 hours amplified the diversity of the seven potential multifunctional CD4 T cell subsets that secreted any combination of IFN-γ, IL-2 and TNF-α. However, increasing pre-stimulation from 6 to 16 hours markedly altered the multifunctional CD4 T cell repertoire to a dominant IFN-γ+ only response. This was observed in both murine and cattle models

The Effects of Wildfire on Mortality and Resources for an Arboreal Marsupial: Resilience to Fire Events but Susceptibility to Fire Regime Change

BACKGROUND: Big environmental disturbances have big ecological effects, yet these are not always what we might expect. Understanding the proximate effects of major disturbances, such as severe wildfires, on individuals, populations and habitats will be essential for understanding how predicted future increases in the frequency of such disturbances will affect ecosystems. However, researchers rarely have access to data from immediately before and after such events. Here we report on the effects of a severe and extensive forest wildfire on mortality, reproductive output and availability of key shelter resources for an arboreal marsupial. We also investigated the behavioural response of individuals to changed shelter resource availability in the post-fire environment. METHODOLOGY/PRINCIPAL FINDINGS: We fitted proximity-logging radiotransmitters to mountain brushtail possums (Trichosurus cunninghami) before, during and after the 2009 wildfires in Victoria, Australia. Surprisingly, we detected no mortality associated with the fire, and despite a significant post-fire decrease in the proportion of females carrying pouch young in the burnt area, there was no short-term post-fire population decline. The major consequence of this fire for mountain brushtail possums was the loss of over 80% of hollow-bearing trees. The types of trees preferred as shelter sites (highly decayed dead standing trees) were those most likely to collapse after fire. Individuals adapted to resource decline by being more flexible in resource selection after the fire, but not by increased resource sharing. CONCLUSIONS/SIGNIFICANCE: Despite short-term demographic resilience and behavioural adaptation following this fire, the major loss of decayed hollow trees suggests the increased frequency of stand-replacing wildfires predicted under climate change will pose major challenges for shelter resource availability for hollow-dependent fauna. Hollow-bearing trees are typically biological legacies of previous forest generations in post-fire regrowth forests but will cease to be recruited to future regrowth forests if the interval between severe fires becomes too rapid for hollow formation

Vesicular Stomatitis Virus Enters Cells through Vesicles Incompletely Coated with Clathrin That Depend upon Actin for Internalization

Many viruses that enter cells by clathrin-dependent endocytosis are significantly larger than the dimensions of a typical clathrin-coated vesicle. The mechanisms by which viruses co-opt the clathrin machinery for efficient internalization remain uncertain. Here we examined how clathrin-coated vesicles accommodate vesicular stomatitis virus (VSV) during its entry into cells. Using high-resolution imaging of the internalization of single viral particles into cells expressing fluorescent clathrin and adaptor molecules, we show that VSV enters cells through partially clathrin-coated vesicles. We found that on average, virus-containing vesicles contain more clathrin and clathrin adaptor molecules than conventional vesicles, but this increase is insufficient to permit full coating of the vesicle. We further show that virus-containing vesicles depend upon the actin machinery for their internalization. Specifically, we found that components of the actin machinery are recruited to virus-containing vesicles, and chemical inhibition of actin polymerization trapped viral particles in vesicles at the plasma membrane. By analysis of multiple independent virus internalization events, we show that VSV induces the nucleation of clathrin for its uptake, rather than depending upon random capture by formation of a clathrin-coated pit. This work provides new mechanistic insights into the process of virus internalization as well as uptake of unconventional cargo by the clathrin-dependent endocytic machinery

Search for Dark Matter Annihilation in the Galactic Center with IceCube-79

The Milky Way is expected to be embedded in a halo of dark matter particles, with the highest density in the central region, and decreasing density with the halo-centric radius. Dark matter might be indirectly detectable at Earth through a flux of stable particles generated in dark matter annihilations and peaked in the direction of the Galactic Center. We present a search for an excess flux of muon (anti-) neutrinos from dark matter annihilation in the Galactic Center using the cubic-kilometer-sized IceCube neutrino detector at the South Pole. There, the Galactic Center is always seen above the horizon. Thus, new and dedicated veto techniques against atmospheric muons are required to make the southern hemisphere accessible for IceCube. We used 319.7 live-days of data from IceCube operating in its 79-string configuration during 2010 and 2011. No neutrino excess was found and the final result is compatible with the background. We present upper limits on the self-annihilation cross-section, \left, for WIMP masses ranging from 30 GeV up to 10 TeV, assuming cuspy (NFW) and flat-cored (Burkert) dark matter halo profiles, reaching down to $\simeq 4 \cdot 10^{-24}$ cm$^3$ s$^{-1}$, and $\simeq 2.6 \cdot 10^{-23}$ cm$^3$ s$^{-1}$ for the $\nu\overline{\nu}$ channel, respectively.Comment: 14 pages, 9 figures, Submitted to EPJ-C, added references, extended limit overvie

Evolution of enhanced innate immune evasion by SARS-CoV-2

Emergence of SARS-CoV-2 variants of concern (VOCs) suggests viral adaptation to enhance human-to-human transmission1,2. Although much effort has focused on characterisation of spike changes in VOCs, mutations outside spike likely contribute to adaptation. Here we used unbiased abundance proteomics, phosphoproteomics, RNAseq and viral replication assays to show that isolates of the Alpha (B.1.1.7) variant3 more effectively suppress innate immune responses in airway epithelial cells, compared to first wave isolates. We found that Alpha has dramatically increased subgenomic RNA and protein levels of N, Orf9b and Orf6, all known innate immune antagonists. Expression of Orf9b alone suppressed the innate immune response through interaction with TOM70, a mitochondrial protein required for RNA sensing adaptor MAVS activation. Moreover, the activity of Orf9b and its association with TOM70 was regulated by phosphorylation. We propose that more effective innate immune suppression, through enhanced expression of specific viral antagonist proteins, increases the likelihood of successful Alpha transmission, and may increase in vivo replication and duration of infection4. The importance of mutations outside Spike in adaptation of SARS-CoV-2 to humans is underscored by the observation that similar mutations exist in the Delta and Omicron N/Orf9b regulatory regions

The Evolution of Compact Binary Star Systems

We review the formation and evolution of compact binary stars consisting of white dwarfs (WDs), neutron stars (NSs), and black holes (BHs). Binary NSs and BHs are thought to be the primary astrophysical sources of gravitational waves (GWs) within the frequency band of ground-based detectors, while compact binaries of WDs are important sources of GWs at lower frequencies to be covered by space interferometers (LISA). Major uncertainties in the current understanding of properties of NSs and BHs most relevant to the GW studies are discussed, including the treatment of the natal kicks which compact stellar remnants acquire during the core collapse of massive stars and the common envelope phase of binary evolution. We discuss the coalescence rates of binary NSs and BHs and prospects for their detections, the formation and evolution of binary WDs and their observational manifestations. Special attention is given to AM CVn-stars -- compact binaries in which the Roche lobe is filled by another WD or a low-mass partially degenerate helium-star, as these stars are thought to be the best LISA verification binary GW sources.Comment: 105 pages, 18 figure

Impact of digestive and oropharyngeal decontamination on the intestinal microbiota in ICU patients

Selective digestive microbial decontamination (SDD) is hypothesized to benefit patients in intensive care (ICU) by suppressing Gram-negative potential pathogens from the colon without affecting the anaerobic intestinal microbiota. The purpose of this study was to provide more insight to the effects of digestive tract and oropharyngeal decontamination on the intestinal microbiota by means of a prospective clinical trial in which faecal samples were collected from ICU patients for intestinal microbiota analysis. The faecal samples were collected from ICU patients enrolled in a multicentre trial to study the outcome of SDD and selective oral decontamination (SOD) in comparison with standard care (SC). Fluorescent in situ hybridization (FISH) was used to analyze the faecal microbiota. The numbers of bacteria from different bacterial groups were compared between the three regimens. The total counts of bacteria per gram faeces did not differ between regimens. The F. prausnitzii group of bacteria, representing an important group among intestinal microbiota, was significantly reduced in the SDD regimen compared to the SC and SOD. The Enterobacteriaceae were significantly suppressed during SDD compared to both SOD and SC; enterococci increased in SDD compared to both other regimens. The composition of the intestinal microbiota is importantly affected by SDD. The F. prausnitzii group was significantly suppressed during SDD. This group of microbiota is a predominant producer of butyrate, the main energy source for colonocytes. Reduction of this microbiota is an important trade-off while reducing gram-negative bacteria by SDD