Data_Sheet_1_Altered metabolites in the periaqueductal gray of COVID-19 patients experiencing headaches: a longitudinal MRS study.pdf

BackgroundHeadache is one of the most common symptoms of acute COVID-19 infection. However, its mechanisms remain poorly understood, and there is a lack of studies investigating changes in the periaqueductal gray (PAG) in COVID-19 patients exhibiting headaches.PurposeThe study aimed to explore the alterations in metabolites of the PAG pre- and post-COVID-19 infection in individuals who suffered from headaches during the acute phase of the disease using proton magnetic resonance spectroscopy (1H-MRS).MethodsFifteen participants who experienced headaches during the acute phase of COVID-19 were recruited. All subjects underwent two proton magnetic resonance spectroscopy (1H-MRS) examinations focusing on the PAG before and after they were infected. Metabolite changes were assessed between the pre- and post-infection groups.ResultsThe combined glutamine and glutamate/total creatine ratio (Glx/tCr) was increased in the PAG following COVID-19 infection. The total choline/total creatine ratio (tCho/tCr) in the pre-infection group was negatively correlated with the duration of headache during the COVID-19 acute phase.ConclusionThe present study indicates that PAG plays a pivotal role in COVID-19 headaches, thereby supporting the involvement of trigeminovascular system activation in the pathophysiology of COVID-19 headaches.</p

Knockdown GNBP3 affected the locust phenotypic resistance to <i>M. anisopliae.</i>

<p>Adult male locusts of two phase were inoculated by <i>M. anisopliae</i> without septic injury under protonum after knockdown <i>gnbp1</i> and <i>gnbp3</i> expression for 48 hours by injection of dsRNA. (A) Log survival curves of gregarious locusts after GNBP1 and GNBP3 knockdown by dsRNAi injection (n_GNBP1 = 36, n_GNBP3 =44, n_GFP =32, n_control = 27; <i>P</i>_{GNBP1 vs. GFP} = 0.796, <i>P</i>_{GNBP3 vs GFP} = 0.006). (B) Log survival curves of solitary locusts after GNBP1 and GNBP3 knockdown (n_GNBP1 = 33, n_GNBP3 = 30, n_GFP =22, n_control = 27; <i>P</i>_{GNBP1 vs. GFP =}0.389, <i>P</i>_{GNBP3 vs. GFP =}0.941). Animals injected with dsGFP for infection were used as negative control; animals without RNAi knockdown and <i>M. anisopliae</i> infection were used as naïve group. Kaplan-Meier method in SPSS 13.0 was used to analysis locust survival data.</p

Altered Immunity in Crowded Locust Reduced Fungal (<em>Metarhizium anisopliae</em>) Pathogenesis

<div><p>The stress of living conditions, similar to infections, alters animal immunity. High population density is empirically considered to induce prophylactic immunity to reduce the infection risk, which was challenged by a model of low connectivity between infectious and susceptible individuals in crowded animals. The migratory locust, which exhibits polyphenism through gregarious and solitary phases in response to population density and displays different resistance to fungal biopesticide (<em>Metarhizium anisopliae</em>), was used to observe the prophylactic immunity of crowded animals. We applied an RNA-sequencing assay to investigate differential expression in fat body samples of gregarious and solitary locusts before and after infection. Solitary locusts devoted at least twice the number of genes for combating <em>M. anisopliae</em> infection than gregarious locusts. The transcription of immune molecules such as pattern recognition proteins, protease inhibitors, and anti-oxidation proteins, was increased in prophylactic immunity of gregarious locusts. The differentially expressed transcripts reducing gregarious locust susceptibility to <em>M. anisopliae</em> were confirmed at the transcriptional and translational level. Further investigation revealed that locust GNBP3 was susceptible to proteolysis while GNBP1, induced by <em>M. anisopliae</em> infection, resisted proteolysis. Silencing of <em>gnbp3</em> by RNAi significantly shortened the life span of gregarious locusts but not solitary locusts. By contrast, <em>gnbp1</em> silencing did not affect the life span of both gregarious and solitary locusts after <em>M. anisopliae</em> infection. Thus, the GNBP3-dependent immune responses were involved in the phenotypic resistance of gregarious locusts to fungal infection, but were redundant in solitary locusts. Our results indicated that gregarious locusts prophylactically activated upstream modulators of immune cascades rather than downstream effectors, preferring to quarantine rather than eliminate pathogens to conserve energy meanwhile increasing the “distance” of infectious and target individuals. Our study has obvious implications for bio-pesticides management of crowded pests, and for understanding disease epidemics and adaptiveness of pathogens.</p> </div

Locust phenotypic life span after lethal fungal infection (<i>M. anisopliae</i>).

<p>(A) Adult gregarious and solitary locusts were topically infected with fungi (<i>M. anisopliae</i>) under the protonum to prevent septic injury. The arrow indicates the inoculation site. Scale bar: 1 cm. (B) Gregarious (G) and solitary (S) locusts were randomly selected (female G:20, S:24 and male G:40, S:27) for fungal infection assay. The life span of the locusts was calculated by Kaplan-Meier methods, and Cox proportional hazards model analysis was used for assessing variables affecting locusts survival. The curves of control treatments (blue and black lines) largely overlapped and displayed as a single black line. (Three replicates for each locust phase, χ² = 25.959, <i>P<</i>0.001).</p

Transcriptome analysis of the fatbody of gregarious and solitary locusts before and after infection.

<p>The abundance and differential expression of transcripts were detected by software packages of Trinity and DEGseq respectively after assembling reference transcripts from raw reads from illumine Truseq experiments. (A) Hierarchical cluster analysis of fatbody transcripts that were significantly regulated (<i>P</i><0.001, q-value<0.05) in at least two samples of four experimental conditions, and two phase locusts displayed distinct response to fungal infection. Heatmap was calculated by implement package of Trinity software. (B) Differential expressed transcripts (<i>P</i><0.001, q-value<0.05) were classed by function (blast2go 1.0E-6). GC: fatbody sample of control gregarious locusts; SC: fatbody sample of control solitary locusts; GI: fatbody sample of gregarious locust infected by <i>M. anisopliae</i>; SI: fatbody sample of solitary locusts infected by <i>M. anisopliae</i>. IMM: immune defenses; MET: metabolism functions; OTHER: other biological process including unidentified. (C) Venn diagram representing unique and shared transcriptome regulation of prophylactic immunity and responsiveness of the two phases of locust to <i>M. anisopliae</i> (D) Prophylactically presented immune molecules of gregarious locust were showed as red in schematic immune pathways of resistance to fungal infection. PRPs: pattern recognition proteins; SP: serine protease; proPAP: pro-phenoloxidase activating proteinase; PPAE: proPO activating enzyme; PO: phenoloxidase; ROS: reactive oxygen species; Serpins: serine protease inhibitors; Grass: Gram-positive Specific Serine protease; SPE: Spätzle-processing enzyme; PSH: Persephone; PR1: fungal virulence protease PR1.</p

Locust GNBP3 affected <i>attacin</i> expression and was susceptible to humoral protease.

<p>(A) Immunofluorescence observed the locust GNBPs binding to fungi cell wall by confocal microscope. DIC: differential interference contrast; FITC: fluoresceinisothiocyanate staining; DAPI: 4′,6-diamidino-2-phenylindole staining; Ctr: <i>Drosophila</i> hemolymph injected with <i>M. anisopliae</i> was used as negative control for examining reactivity of anti-GNBP1 and anti-GNBP3 pAbs. White arrow indicated positive signals. (B) Locust <i>attacin</i> expression in fat body after RNAi knockdown of GNBP1 or GNBP3 was examined by quantitative real time PCR. Fold change was normalized to GFP knockdown locusts. (Mann-Whitney U test, n_GNBP1 = 9, P<0.001; n_GNBP3 = 9, P<0.001) (C) After 2-hours incubation of laminarin-stimulated hemolymph at room temperature, the proteolysis samples of GNBP1 and GNBP3 proteins in supernatant and clot were analyzed by immunoblot. Ctr: before incubation; In: 2-hours incubation; Pe: pellets after incubation. The black arrow indicates the degraded fragment of GNBP3 recognized by anti-N-terminal pAbs.</p

A model for prophylactic immunity in suppression of pathogens spread.

<p>A) Pathogens spread through entire population from the infective individuals to susceptible without any obstacles; B) The clustered individuals applied prophylactic immunity to increase the “distance” between the susceptible and infective individuals, as well as the probability of stochastic extinction of the pathogens.</p

p53 binding sites in normal and cancer cells are characterized by distinct chromatin context

<p>The tumor suppressor protein p53 interacts with DNA in a sequence-dependent manner. Thousands of p53 binding sites have been mapped genome-wide in normal and cancer cells. However, the way p53 selectively binds its cognate sites in different types of cells is not fully understood. Here, we performed a comprehensive analysis of 25 published p53 cistromes and identified 3,551 and 6,039 ‘high-confidence’ binding sites in normal and cancer cells, respectively. Our analysis revealed 2 distinct epigenetic features underlying p53-DNA interactions <i>in vivo</i>. First, p53 binding sites are associated with transcriptionally active histone marks (H3K4me3 and H3K36me3) in normal-cell chromatin, but with repressive histone marks (H3K27me3) in cancer-cell chromatin. Second, p53 binding sites in cancer cells are characterized by a lower level of DNA methylation than their counterparts in normal cells, probably related to global hypomethylation in cancers. Intriguingly, regardless of the cell type, p53 sites are highly enriched in the endogenous retroviral elements of the ERV1 family, highlighting the importance of this repeat family in shaping the transcriptional network of p53. Moreover, the p53 sites exhibit an unusual combination of chromatin patterns: high nucleosome occupancy and, at the same time, high sensitivity to DNase I. Our results suggest that p53 can access its target sites in a chromatin environment that is non-permissive to most DNA-binding transcription factors, which may allow p53 to act as a pioneer transcription factor in the context of chromatin.</p

Concentrations and BCFs of (-)-PCB149/(+)-PCB149 in embryo-larvae when exposed to atropisomers.

<p>Concentrations and BCFs of (-)-PCB149/(+)-PCB149 in embryo-larvae when exposed to atropisomers.</p

VIP-plot via PLS-DA on the expression level of indicated genes.

<p>A: differential genes through comparison of racemic and (+)- PCB149; B: differential genes through comparison of (-)- and (+)- PCB149.</p