Higher RNA Levels Research Articles

881. Nasal and Plasma SARS-CoV-2 RNA Levels Predict Timing of Symptom Resolution in the ACTIV-2 Trial of Non-hospitalized Adults with COVID-19

Abstract Background Symptoms during acute COVID-19 can limit daily activities and delay return to work and school. Little is known about the association between SARS-CoV-2 burden in either the upper airway or plasma and the duration of COVID-19 symptoms. Methods ACTIV-2/A5401 is a platform trial for COVID-19 treatments in non-hospitalized symptomatic adults enrolled within 10 days of symptom onset. We included participants randomized to placebo from August 2020 to July 2021. Participants self-reported severity of 13 symptoms daily from day 0 (baseline) to 28 as Absent 0, Mild 1, Moderate 2, Severe 3; total symptom score was calculated as the sum of all scores. Anterior nasal (AN) and plasma SARS-CoV-2 RNA levels at day 0 were measured with a quantitative qPCR assay. The relationship between day 0 RNA and time to symptom improvement or resolution (first of 2 consecutive days of all symptoms improved or resolved from day 0, respectively) was evaluated using proportional hazards regression adjusted for time from symptom onset. Time to resolution of distinct symptoms was also assessed. Results Among 570 participants randomized to placebo, median age was 48 years, 51% were female, and median time since symptom onset at baseline was 6 days; 7% had prior COVID-19 vaccination. At day 0, AN RNA was detectable in 80% with a median of 4.1 log10 copies/ml (n=533, quartiles: 1.7, 6.0) and plasma RNA was detectable in 19% (91/476). Detectable plasma RNA at day 0, but not AN RNA, was associated with more severe symptoms at day 0 (2.4-point higher mean total symptom score, P=0.001). Both high AN (≥6 vs < 2 log10 copies/ml, adjusted hazard ratio [aHR] 0.63, P=0.001) and detectable plasma RNA (aHR 0.74, P=0.03) at day 0 predicted delayed symptom improvement. High AN RNA at day 0 also predicted a delay in symptom resolution (aHR 0.59, P=0.001). Both high AN RNA and detectable plasma RNA levels predicted delays in the resolution of cough and shortness of breath. Detectable plasma RNA also predicted delayed body pain resolution. Conclusion COVID-19 outpatients with high AN or detectable plasma SARS-CoV-2 RNA at day 0 are more likely to have prolonged symptoms, particularly respiratory symptoms. Additional studies are needed to determine whether the decline in viral load with early treatment impacts symptom duration. Disclosures Kara W. Chew, M.D., M.S., Merck Sharp & Dohme: Grant/Research Support|Pardes Bioscences: Advisor/Consultant Joseph J. Eron, MD, GSK: Advisor/Consultant|Merck: Advisor/Consultant Eric S. Daar, M.D., Gilead: Advisor/Consultant|Gilead: Grant/Research Support|Merck: Advisor/Consultant|ViiV: Advisor/Consultant|ViiV: Grant/Research Support David A. Wohl, M.D., Gilead: Advisor/Consultant|Gilead: Grant/Research Support|Lilly: Grant/Research Support|ViiV: Advisor/Consultant|ViiV: Grant/Research Support Judith S. Currier, M.D., MSc, Merck: Advisor/Consultant Davey M. Smith, M.D., M.A.S., Arena Pharmaceuticals: Advisor/Consultant|Bayer Pharmaceuticals: Advisor/Consultant|Brio Clinical.: Advisor/Consultant|Fluxergy: Advisor/Consultant|Kiadis: Advisor/Consultant|Linear Therapies: Advisor/Consultant|Matrix BioMed: Advisor/Consultant|Model Medicines: Advisor/Consultant|Signant Health: Advisor/Consultant|VxBiosciences: Advisor/Consultant Jonathan Z. Li, MD, MMSc, Abbvie: Advisor/Consultant|Merck: Grant/Research Support.

Detection of Prostate Cancer Biomarker PCA3 by Using Aptasensors.

Prostate cancer cells have very high PCA3 messenger RNA levels, which turns them into one of the new biomarkers for prostate cancer prognosis and diagnosis. Our goal here is to develop a new aptasensor to detect PCA3 release by the cancer cell. DNA hairpin containing PCA3 aptamer was thiolated, conjugated to methylene blue (MB) redox probe, and immobilized on gold electrode through self-assembly to detect label-free cancer cells. Our data have evidenced stable and sensitive sensors presenting a wide linear detection range (0-150ng/mL). In addition, monitoring PCA3 released by different types of prostate cells can provide in-depth knowledge about prostate cancer dynamics; therefore, it is a powerful platform for earlier clinical diagnostic. The released PCA3 can vary depending on the type of adopted prostate cells. PCA3 release was monitored in a group of cells for 2 h; it showed significantly higher expression in both LNCaP and PC-3 cells. This strategy provides a unique and simple methodology to achieve more sensitive and specific PCA3 detection; thus, it emerged as a promising tool for early cost-effective diagnosis.

Novel aldo-keto reductase AKR2E9 regulates aldehyde content in the midgut and antennae of the silkworm (Bombyx mori).

We studied the effects of green leaf volatiles (including reactive aldehydes) emitted by plants on insects that feed on these plants. The silkworm (Bombyx mori) is a model lepidopteran that eats mulberry leaves. Defense-related enzymes in silkworms can be targeted for developing new pest control methods. The aldo-keto reductase (AKR) superfamily catalyzes aldehyde reduction by converting a carbonyl group into an alcohol group. Here, we characterized a novel silkworm AKR, designated as AKR2E9. Recombinant AKR2E9 was overexpressed in Escherichia coli. The recombinant protein was used, along with nicotinamide adenine dinucleotide phosphate as a coenzyme, to reduce aldehydes present in mulberry (Morus alba) leaves. The catalytic efficiency of AKR2E9 toward various aldehyde substrates and its inhibitor sensitivity was lower than those of AKR2E8. High expression levels of akr2e9 messenger RNA (mRNA) were detected in the midgut and antennae of silkworms. In the antennae of adult silkworms, akr2e9 mRNA was more abundant than akr2e8 mRNA. The catalytic efficiency of AKR2E9 was low because of steric hindrance, due to which its active site is blocked. High expression levels of AKR2E9 in the midgut and antennae suggest that it may regulate the detoxification of toxic aldehydes in silkworms.

NSMCE2, a novel super-enhancer-regulated gene, is linked to poor prognosis and therapy resistance in breast cancer

BackgroundDespite today’s advances in the treatment of cancer, breast cancer-related mortality remains high, in part due to the lack of effective targeted therapies against breast tumor types that do not respond to standard treatments. Therefore, identifying additional breast cancer molecular targets is urgently needed. Super-enhancers are large regions of open chromatin involved in the overactivation of oncogenes. Thus, inhibition of super-enhancers has become a focus in clinical trials for its therapeutic potential. Here, we aimed to identify novel super-enhancer dysregulated genes highly associated with breast cancer patients’ poor prognosis and negative response to treatment.MethodsUsing existing datasets containing super-enhancer-associated genes identified in breast tumors and public databases comprising genomic and clinical information for breast cancer patients, we investigated whether highly expressed super-enhancer-associated genes correlate to breast cancer patients’ poor prognosis and to patients’ poor response to therapy. Our computational findings were experimentally confirmed in breast cancer cells by pharmacological SE disruption and gene silencing techniques.ResultsWe bioinformatically identified two novel super-enhancer-associated genes – NSMCE2 and MAL2 – highly upregulated in breast tumors, for which high RNA levels significantly and specifically correlate with breast cancer patients’ poor prognosis. Through in-vitro pharmacological super-enhancer disruption assays, we confirmed that super-enhancers upregulate NSMCE2 and MAL2 transcriptionally, and, through bioinformatics, we found that high levels of NSMCE2 strongly associate with patients’ poor response to chemotherapy, especially for patients diagnosed with aggressive triple negative and HER2 positive tumor types. Finally, we showed that decreasing NSMCE2 gene expression increases breast cancer cells’ sensitivity to chemotherapy treatment.ConclusionsOur results indicate that moderating the transcript levels of NSMCE2 could improve patients’ response to standard chemotherapy consequently improving disease outcome. Our approach offers a new avenue to identify a signature of tumor specific genes that are not frequently mutated but dysregulated by super-enhancers. As a result, this strategy can lead to the discovery of potential and novel pharmacological targets for improving targeted therapy and the treatment of breast cancer.

Nasal and Plasma Severe Acute Respiratory Syndrome Coronavirus 2 RNA Levels Are Associated With Timing of Symptom Resolution in the ACTIV-2 Trial of Nonhospitalized Adults With Coronavirus Disease 2019.

Acute Coronavirus Disease 2019 symptoms limit daily activities, but little is known about its association with severe acute respiratory syndrome coronavirus 2 viral burden. In this exploratory analysis of placebo recipients in the ACTIV-2/A5401 platform trial, we showed that high anterior nasal RNA levels and detectable plasma RNA were associated with delayed symptom improvement. Clinical Trials Registration. https://clinicaltrials.gov/ct2/show/NCT04518410.

S3066 Interferon Therapy for Chronic Hepatitis Delta Viral Infection

Introduction: Hepatitis Delta Virus (HDV) infects those that are already infected with the Hepatitis B Virus (HBV), resulting in a serious infection that may damage hepatocytes. We present a patient that undergoes interferon therapy for chronic hepatitis delta infection with an emphasis on efficacy and side effects of treatment. Case Description/Methods: A 20 year old previously healthy African male presents with progressive fatigue. Bloodwork demonstrates elevated liver enzymes, thrombocytopenia, positive Hepatitis B surface antigen and core antibody, and is negative for hepatitis B e-antigen. HBV DNA levels are 30 IU/mL (reference range < 10 IU/mL). Six months later, liver enzymes remain elevated (AST 100 U/L, ALT 186 U/L). Liver biopsy confirms chronic hepatitis and moderate fibrosis with Metavir grade A3, stage F2, and is consistent with viral hepatitis infection. The biopsy also reveals schistosomiasis of the liver, for which the patient is treated with two series of Praziquantel 1800 mg, 3 times per day. Schistosomiasis resolve following treatment, yet liver enzymes remain elevated. Subsequent bloodwork is positive for HDV, with very high levels of HDV RNA at 14,000,000 IU/mL. The patient is started on tenofovir 300mg/day and peginterferon 180 mcg/week. This therapy is maintained for over 4 years, with side effects of headache and fatigue. Liver biopsy near end of treatment reveals mild portal inflammatory infiltrate composed predominantly of lymphocytes with scattered foci of lobular inflammation; Metavir grade 1, stage 0. Bloodwork at this time shows improvement in liver enzymes with AST 57 U/L and ALT 39 U/L. Patient stopped interferon due to side effects after four years of suppressive treatment. The most recent bloodwork shows HBV and HDV viral suppression via PCR, and negative Hepatitis B surface antigen. Liver surveillance via ultrasound reveals no evidence hepatocellular carcinoma during the duration of therapy. Discussion: Hepatitis delta virus (HDV) is a blood-borne virus that infects human hepatocytes. The replication of HDV depends upon a simultaneous infection with the Hepatitis B virus (HBV). Chronic HDV infections manifest as a rapidly progressing form of viral hepatitis, and can lead to cirrhosis with an increased risk of hepatocellular carcinoma. Currently, HDV has no approved treatment options from the United States Food and Drug Administration, and the only option for treatment is pegylated interferon-alpha.

Immunogenicity and Protective Efficacy of a Recombinant Pichinde Viral-Vectored Vaccine Expressing Influenza Virus Hemagglutinin Antigen in Pigs.

Influenza A virus of swine (IAV-S) is an economically important swine pathogen. The IAV-S hemagglutinin (HA) surface protein is the main target for vaccine development. In this study, we evaluated the feasibility of using the recombinant tri-segmented Pichinde virus (rPICV) as a viral vector to deliver HA antigen to protect pigs against IAV-S challenge. Four groups of weaned pigs (T01–T04) were included in the study. T01 was injected with PBS to serve as a non-vaccinated control. T02 was inoculated with rPICV expressing green fluorescence protein (rPICV-GFP). T03 was vaccinated with rPICV expressing the HA antigen of the IAV-S H3N2 strain (rPICV-H3). T04 was vaccinated with the recombinant HA protein antigen of the same H3N2 strain. Pigs were vaccinated twice at day 0 and day 21 and challenged at day 43 by intra-tracheal inoculation with the homologous H3N2 IAV-S strain. After vaccination, all pigs in T03 and T04 groups were seroconverted and exhibited high titers of plasma neutralizing antibodies. After challenge, high levels of IAV-S RNA were detected in the nasal swabs and bronchioalveolar lavage fluid of pigs in T01 and T02 but not in the T03 and T04 groups. Similarly, lung lesions were observed in T01 and T02, but not in the T03 and T04 groups. No significant difference in terms of protection was observed between the T03 and T04 group. Collectively, our results demonstrate that the rPICV-H3 vectored vaccine elicited protective immunity against IAV-S challenge. This study shows that rPICV is a promising viral vector for the development of vaccines against IAV-S.

Modeling within-host and aerosol dynamics of SARS-CoV-2: The relationship with infectiousness.

The relationship between transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and the amount of virus present in the proximity of a susceptible host is not understood. Here, we developed a within-host and aerosol mathematical model and used it to determine the relationship between viral kinetics in the upper respiratory track, viral kinetics in the aerosols, and new transmissions in golden hamsters challenged with SARS-CoV-2. We determined that infectious virus shedding early in infection correlates with transmission events, shedding of infectious virus diminishes late in the infection, and high viral RNA levels late in the infection are a poor indicator of transmission. We further showed that viral infectiousness increases in a density dependent manner with viral RNA and that their relative ratio is time-dependent. Such information is useful for designing interventions.

Transcriptomic fingerprint of bacterial infection in lower extremity ulcers

Clinicians and researchers utilize subjective, clinical classification systems to stratify lower extremity ulcer infections for treatment and research. The purpose of this study was to examine whether these clinical classifications are reflected in the ulcer's transcriptome. RNA sequencing (RNA‐seq) was performed on biopsies from clinically infected lower extremity ulcers (n = 44). Resulting sequences were aligned to the host reference genome to create a transcriptome profile. Differential gene expression analysis and gene ontology (GO) enrichment analysis were performed between ulcer severities as well as between sample groups identified by k‐means clustering. Lastly, a support vector classifier was trained to estimate clinical infection score or k‐means cluster based on a subset of genes. Clinical infection severity did not explain the major sources of variability among the samples and samples with the same clinical classification demonstrated high inter‐sample variability. High proportions of bacterial RNA were identified in some samples, which resulted in a strong effect on transcription and increased expression of genes associated with immune response and inflammation. K‐means clustering identified two clusters of samples, one of which contained all of the samples with high levels of bacterial RNA. A support vector classifier identified a fingerprint of 20 genes, including immune‐associated genes such as CXCL8, GADD45B, and HILPDA, which accurately identified samples with signs of infection via cross‐validation. This study identified a unique, host‐transcriptome signature in the presence of infecting bacteria, often incongruent with clinical infection‐severity classifications. This suggests that stratification of infection status based on a transcriptomic fingerprint may be useful as an objective classification method to classify infection severity, as well as a tool for studying host–pathogen interactions.

Alzheimer's Biomarker Findings Cast Doubt on Dietary Supplement

Genetic Engineering & Biotechnology NewsVol. 42, No. 6 News & AnalysisFree AccessAlzheimer's Biomarker Findings Cast Doubt on Dietary SupplementPublished Online:8 Jun 2022https://doi.org/10.1089/gen.42.06.04AboutSectionsPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail KATERYNA KON/SCIENCE PHOTO LIBRARY/Getty ImagesScientists are still trying to understand how Alzheimer's disease (AD) is related to serine, which is known to modulate synaptic plasticity, and phosphoglycerate dehydrogenase (PHGDH), which is required for serine's synthesis. Although some researchers have suggested that a form of serine could serve as an AD therapeutic, other researchers have expressed doubts.The scientists with doubts include the researchers at the University of California, San Diego, led by neuroscientist Xu Chen and bioengineer Sheng Zhong. For these researchers, a key issue is whether PHGDH and serine expression levels rise or fall in AD. Two years ago, in Current Biology, the researchers reported that they had analyzed blood samples of older adults and found a steep increase in PHGDH gene expression in AD patients, and that the increase was evident in healthy individuals approximately two years before they were diagnosed with the disease.The results were promising, indicating that high blood levels of RNA produced by the PHGDH gene could serve as a blood biomarker for early detection of AD. To follow up on these results, the researchers investigated whether increases in PHGDH gene expression could be linked back to the brain. They measured PHGDH mRNA and protein levels in two mouse models of AD and in four human cohorts in AD brains compared to ageand sex-matched control brains.The researchers also compared the subjects' PHGDH expression levels with their scores on two different clinical assessments: the Dementia Rating Scale (which rates a person's memory and cognitive ability) and Braak Staging (which rates the severity of AD based on the brain's pathology).What did the scientists find? The answers appeared in Cell Metabolism, in an article titled, “PHGDH expression increases with progression of Alzheimer's disease pathology and symptoms.” The increases were found in experiments with mouse models of AD and/or tauopathy, and in analyses of genetic data collected from postmortem human brains.“PHGDH mRNA and protein levels,” the scientists reported, “are progressively increased in human brains with no, early, and late AD pathology, as well as in people with no, asymptomatic, and symptomatic AD.”“It's exciting that our previous discovery of a blood biomarker is now corroborated with brain data,” said Zhong. “Now we have strong evidence that the changes we see in human blood are directly correlated to changes in the brain in AD.”“The fact that this gene's expression level directly correlates with both a person's cognitive ability and disease pathology is remarkable,” Zhong added. “Being able to quantify both of these complex metrics with a single molecular measurement could potentially make diagnosing and monitoring the progression of AD much simpler.”The scientists also noted that in their work, they were careful to control for postmortem delay (PMD). “Considering that human PHGDH protein is sensitive to protease cleavage at room temperature,” the scientists wrote, “PMD-related protein degradation may explain the lower PHGDH levels” that were reported by another research team. In Cell Metabolism, the other team wrote, “Our findings … suggest oral L-serine as a ready-to-use therapy for AD.” In contrast, the team led by Chen and Zhong declared that oral L-serine “warrants precaution.”FiguresReferencesRelatedDetails Volume 42Issue 6Jun 2022 Information© 2022 by GEN PublishingTo cite this article:Alzheimer's Biomarker Findings Cast Doubt on Dietary Supplement.Genetic Engineering & Biotechnology News.Jun 2022.12-12.http://doi.org/10.1089/gen.42.06.04Published in Volume: 42 Issue 6: June 8, 2022PDF download

Allelic complexity of KMT2A partial tandem duplications in acute myeloid leukemia and myelodysplastic syndromes

AbstractKMT2A partial tandem duplication (KMT2A-PTD) is an adverse risk factor in acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), a potential therapeutic target, and an attractive marker of measurable residual disease. High initial KMT2A-PTD RNA levels have been linked to poor prognosis, but mechanisms regulating KMT2A-PTD expression are not well understood. Although KMT2A-PTD has been reported to affect only a single allele, it has been theorized but not proven that genomic gains of a monoallelic KMT2A-PTD may occur, thereby potentially driving high expression and disease progression. In this study, we identified 94 patients with KMT2A-PTDs using targeted DNA next-generation sequencing (NGS) and found that 16% (15/94) had complex secondary events, including copy-neutral loss of heterozygosity and selective gain involving the KMT2A-PTD allele. High copy numbers indicating complexity were significantly enriched in AML vs MDS and correlated with higher RNA expression. Moreover, in serial samples, complexity was associated with relapse and secondary transformation. Taken together, we provide approaches to integrate quantitative and allelic assessment of KMT2A-PTDs into targeted DNA NGS and demonstrate that secondary genetic events occur in KMT2A-PTD by multiple mechanisms that may be linked to myeloid disease progression by driving increased expression from the affected allele.

PCR-based analysis of PD-L1 RNA expression in lung cancer: comparison with commonly used immunohistochemical assays

BackgroundPD-L1 testing is currently performed by immunohistochemistry (IHC). We questioned whether the results of PCR-based measurement of PD-L1 RNA expression correlate with IHC scores obtained by different commercial assays. Materials and methods167 consecutive non-squamous non-small cell lung carcinomas (NSCLCs) were analyzed for PD-L1 RNA expression and 22C3, SP263, and SP142 IHC scoring using recommended cut-offs. RNA expression was divided into low, moderate, and high categories. ResultsRNA and protein expression demonstrated moderate correlation as continuous variables. Using prespecified RNA cut-offs, PCR testing showed a high negative predictive value towards the IHC analysis: the share of PD-L1 protein-negative tumors among cases classified as PD-L1-low by the PCR test reached 92–99% for all three antibodies. Meanwhile, about half of cases with moderate to high PD-L1 RNA expression had IHC staining in less than 1% tumor cells as determined by 22C3 or SP263 antibodies. Among the 51 discordant cases, which had <1% tumor staining by both 22C3 and SP263 clones but high RNA level, 29 (57%) showed ≥1% positive immune cells by SP263 and/or 22C3, 14 cases (27%) had detectable IHC expression in 0.1–0.9% tumor or immune cells by SP263 and/or 22C3, and 8 (16%) were entirely negative by IHC. ConclusionSome NSCLCs demonstrate readily detectable PD-L1 expression on the level of RNA, but fall below commonly accepted cut-offs by IHC. It remains to be studied whether these discrepancies are attributed to technical or biological reasons. Clinical sensitivity of these tumors to immune therapy deserves additional investigations.

Breakthrough Gastrointestinal COVID-19 and Intrahost Evolution Consequent to Combination Monoclonal Antibody Prophylaxis.

Breakthrough gastrointestinal COVID-19 was observed after experimental SARS-CoV-2 upper mucosal infection in a rhesus macaque undergoing low-dose monoclonal antibody prophylaxis. High levels of viral RNA were detected in intestinal sites contrasting with minimal viral replication in upper respiratory mucosa. Sequencing of virus recovered from tissue in 3 gastrointestinal sites and rectal swab revealed loss of furin cleavage site deletions present in the inoculating virus stock and 2 amino acid changes in spike that were detected in 2 colon sites but not elsewhere, suggesting compartmentalized replication and intestinal viral evolution. This suggests suboptimal antiviral therapies promote viral sequestration in these anatomies.

Inflammatory responses in the placenta upon SARS-CoV-2 infection late in pregnancy

SummaryThe effect of SARS-CoV-2 infection on placental function is not well understood. Analysis of placentas from women who tested positive at delivery showed SARS-CoV-2 genomic and subgenomic RNA in 22 out of 52 placentas. Placentas from two mothers with symptomatic COVID-19 whose pregnancies resulted in adverse outcomes for the fetuses contained high levels of viral Alpha variant RNA. The RNA was localized to the trophoblasts that cover the fetal chorionic villi in direct contact with maternal blood. The intervillous spaces and villi were infiltrated with maternal macrophages and T cells. Transcriptome analysis showed an increased expression of chemokines and pathways associated with viral infection and inflammation. Infection of placental cultures with live SARS-CoV-2 and spike protein-pseudotyped lentivirus showed infection of syncytiotrophoblast and, in rare cases, endothelial cells mediated by ACE2 and Neuropilin-1. Viruses with Alpha, Beta, and Delta variant spikes infected the placental cultures at significantly greater levels.

SARS-CoV-2 Delta Variant (AY.3) in the Feces of a Domestic Cat.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections have spilled over from humans to companion and wild animals since the inception of the global COVID-19 pandemic. However, whole genome sequencing data of the viral genomes that infect non-human animal species have been scant. Here, we detected and sequenced a SARS-CoV-2 delta variant (AY.3) in fecal samples from an 11-year-old domestic house cat previously exposed to an owner who tested positive for SARS-CoV-2. Molecular testing of two fecal samples collected 7 days apart yielded relatively high levels of viral RNA. Sequencing of the feline-derived viral genomes showed the two to be identical, and differing by between 4 and 14 single nucleotide polymorphisms in pairwise comparisons to human-derived lineage AY.3 sequences collected in the same geographic area and time period. However, several mutations unique to the feline samples reveal their divergence from this cohort on phylogenetic analysis. These results demonstrate continued spillover infections of emerging SARS-CoV-2 variants that threaten human and animal health, as well as highlight the importance of collecting fecal samples when testing for SARS-CoV-2 in animals. To the authors’ knowledge, this is the first published case of a SARS-CoV-2 delta variant in a domestic cat in the United States.

A double deletion prevents replication of the pestivirus bovine viral diarrhea virus in the placenta of pregnant heifers.

In contrast to wild type bovine viral diarhea virus (BVDV) specific double deletion mutants are not able to establish persistent infection upon infection of a pregnant heifer. Our data shows that this finding results from a defect in transfer of the virus from the mother animal to the fetus. Pregnant heifers were inoculated with such a double deletion mutant or the parental wild type virus and slaughtered pairwise on days 6, 9, 10 and 13 post infection. Viral RNA was detected via qRT-PCR and RNAscope analyses in maternal tissues for both viruses from day 6 p.i. on. However, the double deletion mutant was not detected in placenta and was only found in samples from animals infected with the wild type virus. Similarly, high levels of wild type viral RNA were present in fetal tissues whereas the genome of the double deletion mutant was not detected supporting the hypothesis of a specific inhibition of mutant virus replication in the placenta. We compared the induction of gene expression upon infection of placenta derived cell lines with wild type and mutant virus via gene array analysis. Genes important for the innate immune response were strongly upregulated by the mutant virus compared to the wild type in caruncle epithelial cells that establish the cell layer on the maternal side at the maternal–fetal interface in the placenta. Also, trophoblasts which can be found on the fetal side of the interface showed significant induction of gene expression upon infection with the mutant virus although with lower complexity. Growth curves recorded in both cell lines revealed a general reduction of virus replication in caruncular epithelial cells compared to the trophoblasts. Compared to the wild type virus this effect was dramtic for the mutant virus that reached only a TCID50 of 1.0 at 72 hours post infection.

Progression and Resolution of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection in Golden Syrian Hamsters

To catalyze severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) research, including development of novel interventive and preventive strategies, the progression of disease was characterized in a robust coronavirus disease 2019 (COVID-19) animal model. In this model, male and female golden Syrian hamsters were inoculated intranasally with SARS-CoV-2 USA-WA1/2020. Groups of inoculated and mock-inoculated uninfected control animals were euthanized at 2, 4, 7, 14, and 28 days after inoculation to track multiple clinical, pathology, virology, and immunology outcomes. SARS-CoV-2–inoculated animals consistently lost body weight during the first week of infection, had higher lung weights at terminal time points, and developed lung consolidation per histopathology and quantitative image analysis measurements. High levels of infectious virus and viral RNA were reliably present in the respiratory tract at days 2 and 4 after inoculation, corresponding with widespread necrosis and inflammation. At day 7, when the presence of infectious virus was rare, interstitial and alveolar macrophage infiltrates and marked reparative epithelial responses (type II hyperplasia) dominated in the lung. These lesions resolved over time, with only residual epithelial repair evident by day 28 after inoculation. The use of quantitative approaches to measure cellular and morphologic alterations in the lung provides valuable outcome measures for developing therapeutic and preventive interventions for COVID-19 using the hamster COVID-19 model.

Defining the transcriptional control of pediatric AML highlights RARA as a superenhancer-regulated druggable dependency

AbstractSomatic mutations are rare in pediatric acute myeloid leukemia (pAML), indicating that alternate strategies are needed to identify targetable dependencies. We performed the first enhancer mapping of pAML in 22 patient samples. Generally, pAML samples were distinct from adult AML samples, and MLL (KMT2A)–rearranged samples were also distinct from non–KMT2A-rearranged samples. Focusing specifically on superenhancers (SEs), we identified SEs associated with many known leukemia regulators. The retinoic acid receptor alpha (RARA) gene was differentially regulated in our cohort, and a RARA-associated SE was detected in 64% of the study cohort across all cytogenetic and molecular subtypes tested. RARA SE+ pAML cell lines and samples exhibited high RARA messenger RNA levels. These samples were specifically sensitive to the synthetic RARA agonist tamibarotene in vitro, with slowed proliferation, apoptosis induction, differentiation, and upregulated retinoid target gene expression, compared with RARA SE− samples. Tamibarotene prolonged survival and suppressed the leukemia burden of an RARA SE+ pAML patient-derived xenograft mouse model compared with a RARA SE− patient-derived xenograft. Our work shows that examining chromatin regulation can identify new, druggable dependencies in pAML and provides a rationale for a pediatric tamibarotene trial in children with RARA-high AML.

First detected case of rabbit Haemorrhagic disease virus 2 (RHDV2) in the Irish hare (Lepus timidus hibernicus)

BackgroundRabbit haemorrhagic disease virus (RHDV) is a Lagovirus, a subgroup of the family Caliciviridae. RHDV2 is a variant first described in France in 2010, and has since spread globally. It has been reported in several Lagomorph species (rabbits, hares, and their relatives) as well as other mammals including voles and shrews. The disease has raised international concerns for its potential impact on population abundance trajectories, particularly as 25% of Lagomorphs are currently Red-Listed by the International Union for the Conservation of Nature (IUCN). The Irish hare (Lepus timidus hibernicus) is a subspecies of the mountain hare, L. timidus, and is endemic to Ireland, making it an Evolutionarily Significant Unit of intrinsic value.Case presentationThe first case of RHDV2 was detected in a wild Irish hare in July 2019. The individual exhibited atypical neurological behaviour (running in circles) prior to death. On necropsy, pink tinged foam was seen in the trachea and congestion was noted in the lungs, but there was no evidence of haemorrhages in any other organ. Both the liver and spleen were tested by reverse transcription real time qPCR confirming high levels of RHDV2 RNA. Histopathology confirmed multifocal necrotising hepatitis.ConclusionThe Irish hare is susceptible to RHDV2 infection. Further investigation is warranted to explore the clinical, epidemiological, and population biology implications.

Exosomes Derived from ADSCs Attenuate Sepsis-Induced Lung Injury by Delivery of Circ-Fryl and Regulation of the miR-490-3p/SIRT3 Pathway.

Sepsis-induced lung injury is a clinical syndrome characterized by injury of alveolar epithelium cells (AECs). Previous investigations illustrate that exosomes secreted from adipose-derived stem cells (ADSCs) have therapeutic effects in a variety of disease treatments, but roles and mechanisms regarding ADSC-derived exosomes in sepsis-induced lung injury are unclear. In this study, high-throughput sequencing was used to explore the molecular delivery of ADSC exosomes. A sepsis-induced lung injury mouse model and a lipopolysaccharide-induced AEC damage model were used for mechanistic analysis. The results showed that ADSC exosomes have high levels of the circular RNA (circ)-Fryl. Downregulation of circ-Fryl suppressed ADSC protective effects exosomes against sepsis-induced lung injury by decreasing apoptosis and inflammatory factor expression. Bioinformatics and luciferase reporting experiments showed that miR-490-3p and SIRT3 are downstream targets of circ-Fryl. miR-490-3p overexpression or SIRT3 silencing reversed ADSC exosome protective effects. Studying the mechanism showed that overexpression of circ-Fryl promoted autophagy activation by inducing SIRT3/AMPK signaling. Autophagy activation can suppress sepsis-induced lung injury by decreasing apoptosis and inflammatory factor expression. Taken together, our results suggest that exosomes derived from ADSCs attenuate sepsis-induced lung injury by delivery of circ-Fryl and regulation of the miR-490-3p/SIRT3 pathway.