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Introduction We previously showed that Nestorone(R) (NES), a synthetic progestin structurally related to progesterone, stimulated remyelination of the corpus callosum in a Cuprizone (CUP) mouse model of demyelination in intact females by promoting replenishment with mature oligodendrocytes (OL) (Glia. 2015;63:104-117). Here, we further investigated the underlying mechanisms of this promyelinating effect. Methods We explored whether NES, applied subcutaneously through Alzet mini-osmotic pumps, regulates specific transcription factors involved in oligodendrocyte progenitor cell (OPC) proliferation and their differentiation into mature OL, using RT-qPCR and Western Blot analysis. Results Our present data show that in comparison to controls, a one-week treatment with NES, through Alzet mini-osmotic pumps, enhanced the production of three relevant transcription factor mRNAs encoding Olig2, Myt1, and Sox17. After 3 weeks, NES treatment reversed the effect of CUP on the levels of corresponding Olig2, Myt1, and Sox17 proteins. Moreover, in mice receiving NES + Estradiol (E2) co-treatment, levels of Olig2, Myt1, and Sox17 proteins did not change as compared to NES alone. Conclusion NES alone or with E2 increased the levels of transcription factors, essential for myelin synthesis.

Yellow fever virus (YFV) live attenuated vaccine can, in rare cases, cause life-threatening disease, typically in patients with no previous history of severe viral illness. Autosomal recessive (AR) complete IFNAR1 deficiency was reported in one 12-yr-old patient. Here, we studied seven other previously healthy patients aged 13 to 80 yr with unexplained life-threatening YFV vaccine-associated disease. One 13-yr-old patient had AR complete IFNAR2 deficiency. Three other patients vaccinated at the ages of 47, 57, and 64 yr had high titers of circulating auto-Abs against at least 14 of the 17 individual type I IFNs. These antibodies were recently shown to underlie at least 10% of cases of life-threatening COVID-19 pneumonia. The auto-Abs were neutralizing in vitro, blocking the protective effect of IFN-?2 against YFV vaccine strains. AR IFNAR1 or IFNAR2 deficiency and neutralizing auto-Abs against type I IFNs thus accounted for more than half the cases of life-threatening YFV vaccine-associated disease studied here. Previously healthy subjects could be tested for both predispositions before anti-YFV vaccination.

During affinity maturation, germinal center (GC) B cells alternate between proliferation and somatic hypermutation in the dark zone (DZ) and affinity-dependent selection in the light zone (LZ). This anatomical segregation imposes that the vigorous proliferation that allows clonal expansion of positively selected GC B cells takes place ostensibly in the absence of the signals that triggered selection in the LZ, as if by "inertia." We find that such inertial cycles specifically require the cell cycle regulator cyclin D3. Cyclin D3 dose-dependently controls the extent to which B cells proliferate in the DZ and is essential for effective clonal expansion of GC B cells in response to strong T follicular helper (Tfh) cell help. Introduction into the Ccnd3 gene of a Burkitt lymphoma?associated gain-of-function mutation (T283A) leads to larger GCs with increased DZ proliferation and, in older mice, clonal B cell lymphoproliferation, suggesting that the DZ inertial cell cycle program can be coopted by B cells undergoing malignant transformation.

Purpose: To develop and characterize a porcine model of liver cancer that could be used to test new locoregional therapies. Materials and Methods: Liver tumors were induced in 18 Oncopigs (transgenic pigs with Cre-inducible TP53R167H and KRASG12D mutations) by using an adenoviral vector encoding the Cre-recombinase gene. The resulting 60 tumors were characterized on multiphase contrast-enhanced CT, angiography, perfusion, micro-CT, and necropsy. Transarterial embolization was performed using 40?120 ?m (4 pigs) or 100?300 ?m (4 pigs) Embosphere microspheres. Response to embolization was evaluated on imaging. Complications were determined based on daily clinical evaluation, laboratory results, imaging, and necropsy. Results: Liver tumors developed at 60/70 (86%) inoculated sites. Mean tumor size was 2.1 cm (range, 0.3?4 cm) at 1 week. Microscopically, all animals developed poorly differentiated to undifferentiated carcinomas accompanied by a major inflammatory component, which resembled undifferentiated carcinomas of the human pancreatobiliary tract. Cytokeratin and vimentin expression confirmed epithelioid and mesenchymal differentiation, respectively. Lymph node, lung, and peritoneal metastases were seen in some cases. On multiphase CT, all tumors had a hypovascular center, and 17/60 (28%) had a hypervascular rim. After transarterial embolization, noncontrast CT showed retained contrast medium in the tumors. Follow-up contrast-enhanced scan showed reduced size of tumors after embolization using either 40?120 ?m or 100?300 ?m Embosphere microspheres, while untreated tumors showed continued growth. Conclusions: Liver tumors can be induced in a transgenic pig and can be successfully treated using bland embolization.

Purpose To report four cases of life-threatening COVID-19 pneumonia in patients with high blood concentrations of neutralizing autoantibodies against type I interferons (IFNs), who were treated with plasma exchange (PE) as a rescue therapy. Methods Prospective case series, which included patients, diagnosed with RT-PCR-confirmed SARS-CoV-2 infection and positive autoantibodies against type I IFNs in two French intensive care units (ICUs) between October 8 and November 14, 2020. Six critically ill COVID-19 patients with no anti-IFN antibodies were used as controls. Anti-IFN autoantibodies and IFN concentrations, together with the levels of anti-SARS-CoV-2 antibodies, were measured sequentially in serum. Viral load was determined in the upper and lower respiratory tract. Patients were followed during hospital stay. Results Three men and one woman were included. Three of the patients had four PE sessions each, while another had three PE sessions. PE decreased the concentrations of autoantibodies against type I IFN in all four patients, whereas anti-SARS-CoV-2 antibody levels remained stable. Autoantibodies against type I IFN levels were high in tracheal aspirates of one patient and decreased after three PE sessions. By contrast, anti-IFN autoantibodies were not detected in tracheal aspirates from five control patients without detectable anti-IFN autoantibodies in serum. During PE, serum IFN-alpha levels slightly increased in three out of four patients, and upper respiratory tract viral load decreased in all patients. All patients were alive at day 28 of ICU admission. Two patients eventually died in the ICU, while the two survivors were discharged from the ICU at days 50 and 66. Conclusions PE efficiently removes autoantibodies against type I IFNs, including those detected in tracheal aspirates, without affecting anti-SARS-CoV-2 antibody levels, in patients with life-threatening COVID-19 pneumonia. The clinical benefit of PE in patients with autoantibodies against type I IFNs should be tested in a larger study.

Aeromonas hydrophila is an important pathogenic bacterium that causes foodborne illness worldwide. In this study, virulent phages from the sediment of a fish farm were propagated and isolated on a multidrug-resistant strain of A. hydrophila, ZYAH75. One phage, designated as ZPAH7, featured a unique turbid halo around a clear plaque on the bacterial lawn (indicative of potential depolymerase activity), and was selected for further analysis. ZPAH7 was classified as podophage by morphological and genomic methods. Further comparisons of genome nucleotide similarity, ratios of homologous proteins and phylogenetic relatedness among the terminase large subunit and major capsid proteins of similar phage deposited in GENBANK, led us to propose a new genus, ZPAH7virus, in the Autographivirinae subfamily of Podoviridae. ZPAH7 had an adsorption rate of 79% in 5 min, an eclipse period of 15 min, a latent period of 25 min, and a burst size of 148 +/- 9 PFU/cell. Antimicrobial application experiments showed that ZPAH7 lead to significantly reduction on A. hydrophila on lettuce. Additionally, ZPAH7 was able to inhibit biofilm formation, as well as degrade and kill bacteria in established biofilms. Furthermore, lytic activity of ZPAH7 remained stable across a wide range of temperatures and pH measurements. These results suggest ZPAH7 could be used as a potential biological control agent against A. hydrophila on food and/or biofilms on food contact surfaces.

Pumilio paralogs, PUM1 and PUM2, are sequence-specific RNA-binding proteins that are essential for vertebrate development and neurological functions. PUM1&2 negatively regulate gene expression by accelerating degradation of specific mRNAs. Here, we determined the repression mechanism and impact of human PUM1&2 on the transcriptome. We identified subunits of the CCR4-NOT (CNOT) deadenylase complex required for stable interaction with PUM1&2 and to elicit CNOT-dependent repression. Isoform-level RNA sequencing revealed broad coregulation of target mRNAs through the PUM-CNOT repression mechanism. Functional dissection of the domains of PUM1&2 identified a conserved amino-terminal region that confers the predominant repressive activity via direct interaction with CNOT. In addition, we show that the mRNA decapping enzyme, DCP2, has an important role in repression by PUM1&2 amino-terminal regions. Our results support a molecular model of repression by human PUM1&2 via direct recruitment of CNOT deadenylation machinery in a decapping-dependent mRNA decay pathway.

Horizontal gene transfer and mutation are the two major drivers of microbial evolution that enable bacteria to adapt to fluctuating environmental stressors(1). Clustered, regularly interspaced, short palindromic repeats (CRISPR) systems use RNA-guided nucleases to direct sequence-specific destruction of the genomes of mobile genetic elements that mediate horizontal gene transfer, such as conjugative plasmids(2) and bacteriophages(3), thus limiting the extent to which bacteria can evolve by this mechanism. A subset of CRISPR systems also exhibit non-specific degradation of DNA(4,5); however, whether and how this feature affects the host has not yet been examined. Here we show that the non-specific DNase activity of the staphylococcal type III-A CRISPR-Cas system increases mutations in the host and accelerates the generation of antibiotic resistance in Staphylococcus aureus and Staphylococcus epidermidis. These mutations require the induction of the SOS response to DNA damage and display a distinct pattern. Our results demonstrate that by differentially affecting both mechanisms that generate genetic diversity, type III-A CRISPR systems can modulate the evolution of the bacterial host.

SARS-CoV-2 is responsible for an ongoing pandemic that has affected millions of individuals around the globe. To gain further understanding of the immune response in recovered individuals, we measured T cell responses in paired samples obtained an average of 1.3 and 6.1 mo after infection from 41 individuals. The data indicate that recovered individuals show persistent polyfunctional SARS-CoV-2 antigen-specific memory that could contribute to rapid recall responses. Recovered individuals also show enduring alterations in relative overall numbers of CD4(+) and CD8(+) memory T cells, including expression of activation/exhaustion markers, and cell division.

Several studies have analyzed antiviral immune pathways in late-stage severe COVID-19. However, the initial steps of SARS-CoV-2 antiviral immunity are poorly understood. Here we have isolated primary SARS-CoV-2 viral strains and studied their interaction with human plasmacytoid predendritic cells (pDCs), a key player in antiviral immunity. We show that pDCs are not productively infected by SARS-CoV-2. However, they efficiently diversified into activated P1-, P2-, and P3-pDC effector subsets in response to viral stimulation. They expressed CD80, CD86, CCR7, and OX40 ligand at levels similar to influenza virus-induced activation. They rapidly produced high levels of interferon-alpha, interferon-lambda 1, IL-6, IP-10, and IL-8. All major aspects of SARS-CoV-2-induced pDC activation were inhibited by hydroxychloroquine. Mechanistically, SARS-CoV-2-induced pDC activation critically depended on IRAK4 and UNC93B1, as established using pDC from genetically deficient patients. Overall, our data indicate that human pDC are efficiently activated by SARS-CoV-2 particles and may thus contribute to type I IFN-dependent immunity against SARS-CoV-2 infection.