TOP1 inhibition therapy protects against SARS-CoV-2-induced lethal inflammation.

The ongoing pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently affecting millions of lives worldwide. Large retrospective studies indicate that an elevated level of inflammatory cytokines and pro-inflammatory factors are associated with both increased disease severity and mortality. Here, using multidimensional epigenetic, transcriptional, in vitro, and in vivo analyses, we report that topoisomerase 1 (TOP1) inhibition suppresses lethal inflammation induced by SARS-CoV-2. Therapeutic treatment with two doses of topotecan (TPT), an FDA-approved TOP1 inhibitor, suppresses infection-induced inflammation in hamsters. TPT treatment as late as 4 days post-infection reduces morbidity and rescues mortality in a transgenic mouse model. These results support the potential of TOP1 inhibition as an effective host-directed therapy against severe SARS-CoV-2 infection. TPT and its derivatives are inexpensive clinical-grade inhibitors available in most countries. Clinical trials are needed to evaluate the efficacy of repurposing TOP1 inhibitors for severe coronavirus disease 2019 (COVID-19) in humans.

Hybrid Gene Origination Creates Human-Virus Chimeric Proteins during Infection.

RNA viruses are a major human health threat. The life cycles of many highly pathogenic RNA viruses like influenza A virus (IAV) and Lassa virus depends on host mRNA, because viral polymerases cleave 5'-m7G-capped host transcripts to prime viral mRNA synthesis ("cap-snatching"). We hypothesized that start codons within cap-snatched host transcripts could generate chimeric human-viral mRNAs with coding potential. We report the existence of this mechanism of gene origination, which we named "start-snatching." Depending on the reading frame, start-snatching allows the translation of host and viral "untranslated regions" (UTRs) to create N-terminally extended viral proteins or entirely novel polypeptides by genetic overprinting. We show that both types of chimeric proteins are made in IAV-infected cells, generate T cell responses, and contribute to virulence. Our results indicate that during infection with IAV, and likely a multitude of other human, animal and plant viruses, a host-dependent mechanism allows the genesis of hybrid genes.

Nuclear RNA catabolism controls endogenous retroviruses, gene expression asymmetry, and dedifferentiation.

Endogenous retroviruses (ERVs) are remnants of ancient parasitic infections and comprise sizable portions of most genomes. Although epigenetic mechanisms silence most ERVs by generating a repressive environment that prevents their expression (heterochromatin), little is known about mechanisms silencing ERVs residing in open regions of the genome (euchromatin). This is particularly important during embryonic development, where induction and repression of distinct classes of ERVs occur in short temporal windows. Here, we demonstrate that transcription-associated RNA degradation by the nuclear RNA exosome and Integrator is a regulatory mechanism that controls the productive transcription of most genes and many ERVs involved in preimplantation development. Disrupting nuclear RNA catabolism promotes dedifferentiation to a totipotent-like state characterized by defects in RNAPII elongation and decreased expression of long genes (gene-length asymmetry). Our results indicate that RNA catabolism is a core regulatory module of gene networks that safeguards RNAPII activity, ERV expression, cell identity, and developmental potency.

Non-coding RNA Generated following Lariat Debranching Mediates Targeting of AID to DNA.

Transcription through immunoglobulin switch (S) regions is essential for class switch recombination (CSR), but no molecular function of the transcripts has been described. Likewise, recruitment of activation-induced cytidine deaminase (AID) to S regions is critical for CSR; however, the underlying mechanism has not been fully elucidated. Here, we demonstrate that intronic switch RNA acts in trans to target AID to S region DNA. AID binds directly to switch RNA through G-quadruplexes formed by the RNA molecules. Disruption of this interaction by mutation of a key residue in the putative RNA-binding domain of AID impairs recruitment of AID to S region DNA, thereby abolishing CSR. Additionally, inhibition of RNA lariat processing leads to loss of AID localization to S regions and compromises CSR; both defects can be rescued by exogenous expression of switch transcripts in a sequence-specific manner. These studies uncover an RNA-mediated mechanism of targeting AID to DNA.

A Quantitative Genetic Interaction Map of HIV Infection.

We have developed a platform for quantitative genetic interaction mapping using viral infectivity as a functional readout and constructed a viral host-dependency epistasis map (vE-MAP) of 356 human genes linked to HIV function, comprising >63,000 pairwise genetic perturbations. The vE-MAP provides an expansive view of the genetic dependencies underlying HIV infection and can be used to identify drug targets and study viral mutations. We found that the RNA deadenylase complex, CNOT, is a central player in the vE-MAP and show that knockout of CNOT1, 10, and 11 suppressed HIV infection in primary T cells by upregulating innate immunity pathways. This phenotype was rescued by deletion of IRF7, a transcription factor regulating interferon-stimulated genes, revealing a previously unrecognized host signaling pathway involved in HIV infection. The vE-MAP represents a generic platform that can be used to study the global effects of how different pathogens hijack and rewire the host during infection.

NMDAR antagonists suppress tumor progression by regulating tumor-associated macrophages.

Neurotransmitter receptors are increasingly recognized to play important roles in anti-tumor immunity. The expression of the ion channel N-methyl-D-aspartate receptor (NMDAR) on macrophages was reported, but the role of NMDAR on macrophages in the tumor microenvironment (TME) remains unknown. Here, we show that the activation of NMDAR triggered calcium influx and reactive oxygen species production, which fueled immunosuppressive activities in tumor-associated macrophages (TAMs) in the hepatocellular sarcoma and fibrosarcoma tumor settings. NMDAR antagonists, MK-801, memantine, and magnesium, effectively suppressed these processes in TAMs. Single-cell RNA sequencing analysis revealed that blocking NMDAR functionally and metabolically altered TAM phenotypes, such that they could better promote T cell- and Natural killer (NK) cell-mediated anti-tumor immunity. Treatment with NMDAR antagonists in combination with anti-PD-1 antibody led to the elimination of the majority of established preclinical liver tumors. Thus, our study uncovered an unknown role for NMDAR in regulating macrophages in the TME of hepatocellular sarcoma and provided a rationale for targeting NMDAR for tumor immunotherapy.

The splicing regulator PTBP2 interacts with the cytidine deaminase AID and promotes binding of AID to switch-region DNA.

During immunoglobulin class-switch recombination (CSR), the cytidine deaminase AID induces double-strand breaks into transcribed, repetitive DNA elements called switch sequences. The mechanism that promotes the binding of AID specifically to switch regions remains to be elucidated. Here we used a proteomic screen with in vivo biotinylation of AID to identify the splicing regulator PTBP2 as a protein that interacts with AID. Knockdown of PTBP2 mediated by short hairpin RNA in B cells led to a decrease in binding of AID to transcribed switch regions, which resulted in considerable impairment of CSR. PTBP2 is thus an effector of CSR that promotes the binding of AID to switch-region DNA.

The uncharacterized SANT and BTB domain-containing protein SANBR inhibits class switch recombination.

Class switch recombination (CSR) is the process by which B cells switch production from IgM/IgD to other immunoglobulin isotypes, enabling them to mount an effective immune response against pathogens. Timely resolution of CSR prevents damage due to an uncontrolled and prolonged immune response. While many positive regulators of CSR have been described, negative regulators of CSR are relatively unknown. Using an shRNA library screen targeting more than 28,000 genes in a mouse B cell line, we have identified a novel, uncharacterized protein of 82kD (KIAA1841, NM_027860), which we have named SANBR (SANT and BTB domain regulator of CSR), as a negative regulator of CSR. The purified, recombinant BTB domain of SANBR exhibited characteristic properties such as homodimerization and interaction with corepressor proteins, including HDAC and SMRT. Overexpression of SANBR inhibited CSR in primary mouse splenic B cells, and inhibition of CSR is dependent on the BTB domain while the SANT domain is largely dispensable. Thus, we have identified a new member of the BTB family that serves as a negative regulator of CSR. Future investigations to identify transcriptional targets of SANBR in B cells will reveal further insights into the specific mechanisms by which SANBR regulates CSR as well as fundamental gene regulatory activities of this protein.

Role of Different Doses of Ketamine in Postoperative Neurocognitive Function in Aged Mice Undergoing Partial Hepatectomy by Regulating the Bmal1/NMDA/NF-Κb Axis.

BACKGROUND: The current study set out to probe the function of different doses of ketamine in postoperative neurocognitive disorder (PND) in aged mice undergoing partial hepatectomy (PH) with the involvement of the brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein 1 (Bmal1)/n-methyl-D-aspartate (NMDA)/nuclear factor-kappa B (NF-κB) axis. METHODS: First, aged mice were intraperitoneally injected with different doses of ketamine prior to surgery, followed by hepatic lobectomy. Afterward, mice cognitive function was assessed. In addition, Bmal1 mRNA expression patterns were quantified, while NMDA 2B receptor, NF-κB p65, synapsin 1, and postsynaptic density 95 (PSD95) levels were determined; the release of inflammatory factors was detected, and ionized calcium-binding adapter molecule-1 expression was measured to quantify microglia activation. In addition, Bmal1-knockout (Bmal1-KO) mice were intraperitoneally injected with a subanesthetic dose of ketamine to verify the mechanism of Bmal1 in regulating the NMDA 2B subunit (NR2B)/NF-κB axis to affect PH in aged patients. RESULTS: After PH, hippocampal-dependent memory was impaired, and synapsin 1 and PSD95 levels were downregulated. On the other hand, PH diminished Bmal1 expression but elevated NR2B and NF-κB p65 levels and anesthetic doses of ketamine further regulated the Bmal1/NMDA/NF-κB axis. In Bmal1-KO mice, the NMDA/NF-κB axis was activated, the release of inflammatory cytokines was promoted, and hippocampus-dependent memory was impaired, which were reversed by a subanesthetic dose of ketamine. CONCLUSION: Altogether, findings obtained in our study indicated that a subanesthetic dose of ketamine activated Bmal1, downregulated the NMDA/NF-κB axis, and reduced inflammation and microglia activation to alleviate PND in aged mice undergoing PH.

Gene expression profiles in the brain of phenylketonuria mouse model reversed by the low phenylalanine diet therapy.

To gain insight into the potential protective mechanisms of low phenylalanine diet (LPD) in phenylketonuria (PKU), gene expression profiles were studied in the cerebral cortex and hippocampus of a PKU mouse model (BTBR-Pahenu2). PKU mice were fed with low Phe diet (LPD-PKU group) and normal diet (PKU group). Wild-type mice were treated with normal diet (WT group) as control. After 12 weeks, we detected gene expression in the cerebral cortex and hippocampus of the three groups by RNA-sequencing, and then screened the differentially-expressed genes (DEGs) among the groups by bioinformatics analyses. We found that the transcriptional profiles of both cerebral cortex and hippocampus changed markedly between PKU and WT mice. Furthermore, LPD changed the transcriptional profiles of the cerebral cortex and the hippocampus of PKU mice significantly, especially in the cerebral cortex, with overlaps of genes that changed with the disease and altered by LPD treatment. In the cerebral cortex, hundreds of DEGs enriched in a wide spectrum of biological processes, molecular function, and cellular component, including nervous system development, axon development and guidance, calcium ion binding, modulation of chemical synaptic transmission, and regulation of protein kinase activity. In the hippocampus, the overlapping genes were enriched in positive regulation of long term synaptic, negative regulation of excitatory postsynaptic potential, positive regulation of synapse assembly. Our results showed that genes impaired in PKU and then rescued by LPD might indicate the potential protective capability of LPD in the PKU brain.

Ring-shaped twisted Gaussian Schell-model array beams.

A new partially coherent source which can generate a beam field with a ring-shaped twisted array profile is presented, and the distribution characteristics of spectral density and degree of coherence of the field are discussed. It is shown that both the spectral density and degree of coherence will rotate along the propagating direction, but in opposite rotating directions. Furthermore, we find that the distribution properties of the ring-shaped array of the spectral density, including the number of the rings, the number of the lobes of each ring, and the distance of all adjacent lobes, can be properly controlled by adjusting structural parameters of the source.

Hollow rectangular multi-Gaussian Schell-model source.

We introduced a new class of a partially coherent source that can generate a field with a hollow rectangular profile, named the hollow rectangular multi-Gaussian Schell-mode source. The dependence of distribution of far-zone spectral density on the properties of the proposed source was analyzed. The results show that one can control the distribution properties of the far-zone intensity profile, including the thickness of the hollow edge, the shape of the hollow, the size of the hollow, and the orientation of the hollow, by adjusting the corresponding structural parameters of the source.

YouTube and food allergy: An appraisal of the educational quality of information.

BACKGROUND: Food allergy affects an estimated 8% of children and 3% of adults in the United States. Food-allergic individuals increasingly use the web for medical information. We sought to determine the educational quality of food allergy YouTube videos. METHODS: We performed a YouTube search using keywords "food allergy" and "food allergies". The 300 most viewed videos were included and analyzed for characteristics, source, and content. Source was further classified as healthcare provider, alternative medicine provider, patient, company, media, and professional society. A scoring system (FA-DQS) was created to evaluate quality (-10 to +34 points). Negative points were assigned for misleading information. Eight reviewers scored each video independently. RESULTS: Three hundred videos were analyzed, with a median of 6351.50 views, 19 likes, and 1 dislike. More video presenters were female (54.3%). The most common type of video source was alternative medicine provider (26.3%). Alternative treatments included the following: water fast, juicing, Ayurveda, apple cider, yoga, visualization, and sea moss. Controversial diagnostics included kinesiology, IgG testing, and pulse test. Almost half of the videos depicted a non-IgE-mediated reaction (49.0%).Videos by professional societies had the highest FA-DQS (7.27). Scores for videos by professional societies were significantly different from other sources (P < .001). There was a high degree of agreement among reviewers (ICC = 0.820; P < .001). CONCLUSION: YouTube videos on food allergy frequently recommend controversial diagnostics and commonly depict non-IgE-mediated reactions. There is a need for high-quality, evidence-based, educational videos on food allergy.

Transferring knowledge and skills from an international collaborative nursing educational program to clinical practice: A qualitative study.

BACKGROUND: The effective transfer of knowledge and skills is the primary goal of nursing education. Despite this, little is known about how knowledge and skills gained through international collaborative nursing educational programs is applied by nurses. OBJECTIVES: To describe the experiences of knowledge and skills transfer among nursing graduates from an international collaborative educational program to their clinical practice and ongoing study. METHODS: A qualitative design was employed for this study. In 2023, fifteen interviews were conducted with nursing graduates from a Chinese-Swedish collaborative nursing educational program at Lishui University in eastern China. Directed content analysis was utilized to analyze the interview data. RESULTS: Nursing graduates gained both knowledge and skills that surpassed the educational goals of the international collaborative program. Throughout the application and transfer of knowledge and skills, participants reported both positive and negative experiences. Notably, a gap persisted between basic nursing theoretical knowledge and clinical practice. Additionally, deficiencies were identified in the transfer of nursing research knowledge, indicating areas for improvement in future nursing education. CONCLUSION: The findings suggest that knowledge and skills from an international collaborative nursing educational program can be successfully transferred to clinical nursing practice and postgraduate study. However, addressing the gap between theoretical knowledge and practice, particularly in updated basic nursing knowledge and practice, is essential. Furthermore, there is a need to enhance awareness and attitudes towards nursing research among nurses, emphasizing the importance of continuous learning.

Syringaresinol attenuates Tau phosphorylation and ameliorates cognitive dysfunction induced by sevoflurane in aged rats.

Cognitive dysfunction following anesthesia with agents such as sevoflurane is a significant clinical problem, particularly in elderly patients. This study aimed to explore the protective effects of the phytochemical syringaresinol (SYR) against sevoflurane-induced cognitive deficits in aged Sprague-Dawley rats and to determine the underlying mechanisms involved. We assessed the impact of SYR on sevoflurane-induced cognitive impairment, glial activation, and neuronal apoptosis through behavioral tests (Morris water maze), immunofluorescence, Western blotting for key proteins involved in apoptosis and inflammation, and enzyme-linked immunosorbent assays for interleukin-1ß, tumor necrosis factor-α, and interleukin-6. SYR treatment mitigated sevoflurane-induced cognitive decline, reduced microglial and astrocyte activation (decreased Iba-1 and GFAP expression), and countered neuronal apoptosis (reduced Bax, cleaved-caspase3, and cleaved-PARP expression). SYR also enhanced Sirtuin-1 (SIRT1) expression and reduced p-Tau phosphorylation; these effects were reversed by the SIRT1 inhibitor EX527. SYR exerts neuroprotective effects on sevoflurane-induced cognitive dysfunction by modulating glial activity, apoptotic signaling, and Tau phosphorylation through the SIRT1 pathway. These findings could inform clinical strategies to safeguard cognitive function in patients undergoing anesthesia.

Demineralized Frozen-dried Allogeneic Bone Blocks with Suture Fixation Technique for reconstruction of maxillary alveolar bone deficiency: A Case Series.

PURPOSE: This study aims to evaluate the clinical outcomes of using demineralized freeze-dried allogeneic bone blocks (DFDABB) combined with the periosteal vertical mattress suture (PVMS) technique for the reconstruction of severe horizontal alveolar bone deficiencies in the maxilla. METHOD: In continuous horizontal maxillary defects cases, bone augmentation was performed using DFDABB and deproteinized bovine bone matrix (DBBM) filling the interstice. Subsequently, a resorbable collagen membrane was carefully placed over the graft surface, and both the membrane and bone graft were firmly secured using the periosteal vertical mattress suture technique (PVMS). Linear changes were assessed through superimposed cone-beam computed tomography (CBCT) scans obtained before the operation and after a healing period of 6-10 months. RESULTS: A total of 7 female patients with ten bone blocks and 13 implants were included in this study. One of the wounds was slightly ruptured postoperatively without infection, and all implants showed successful osseointegration. The average alveolar ridge width at a point 5 mm below the crest was 4.52 ± 2.03 mm before bone graft and 9.79 ± 1.57 mm after implantation, with an average increase of 5.26 ± 1.97 mm. Similarly, at a point 10 mm below the crest, the pre-graft alveolar ridge width measured 7.23 ± 3.60 mm, and post-implantation, it expanded to 11.81 ± 2.90 mm, showing an average gain of 4.58 ± 2.01 mm. CONCLUSION: This case series demonstrates the successful application of DFDABB combined with the PVMS technique to achieve adequate bone width for implantation at severe continuous horizontal bone deficiency of the maxilla. DFDABB with the PVMS technique resulted in superior horizontal bone gain during maxillary bone augmentation with horizontal continuity deficiency. However, further studies are necessary to validate these findings.

A Germinal Center Checkpoint of AIRE in B Cells Limits Antibody Diversification.

In response to antigens, B cells undergo affinity maturation and class switching mediated by activation-induced cytidine deaminase (AID) in germinal centers (GCs) of secondary lymphoid organs, but uncontrolled AID activity can precipitate autoimmunity and cancer. The regulation of GC antibody diversification is of fundamental importance but not well understood. We found that autoimmune regulator (AIRE), the molecule essential for T cell tolerance, is expressed in GC B cells in a CD40-dependent manner, interacts with AID and negatively regulates antibody affinity maturation and class switching by inhibiting AID function. AIRE deficiency in B cells caused altered antibody repertoire, increased somatic hypermutations, elevated autoantibodies to T helper 17 effector cytokines and defective control of skin Candida albicans. These results define a GC B cell checkpoint of humoral immunity and illuminate new approaches of generating high-affinity neutralizing antibodies for immunotherapy.

Antifungal Activity and Action Mechanisms of 2,4-Di-tert-butylphenol against Ustilaginoidea virens.

Ustilaginoidea virens is a destructive phytopathogenic fungus that causes false smut disease in rice. In this study, the natural product 2,4-di-tert-butylphenol (2,4-DTBP) was found to be an environmentally friendly and effective agent for the first time, which exhibited strong antifungal activity against U. virens, with an EC50 value of 0.087 mmol/L. The scanning electron microscopy, fluorescence staining, and biochemical assays indicated that 2,4-DTBP could destroy the cell wall, cell membrane, and cellular redox homeostasis of U. virens, ultimately resulting in fungal cell death. Through the transcriptomic analysis, a total of 353 genes were significantly upregulated and 367 genes were significantly downregulated, focusing on the spindle microtubule assembly, cell wall and membrane, redox homeostasis, mycotoxin biosynthesis, and intracellular metabolism. These results enhanced the understanding of the antifungal activity and action mechanisms of 2,4-DTBP against U. virens, supporting it to be a potential antifungal agent for the control of false smut disease.

Generation of host-directed and virus-specific antivirals using targeted protein degradation promoted by small molecules and viral RNA mimics.

Targeted protein degradation (TPD), as exemplified by proteolysis-targeting chimera (PROTAC), is an emerging drug discovery platform. PROTAC molecules, which typically contain a target protein ligand linked to an E3 ligase ligand, recruit a target protein to the E3 ligase to induce its ubiquitination and degradation. Here, we applied PROTAC approaches to develop broad-spectrum antivirals targeting key host factors for many viruses and virus-specific antivirals targeting unique viral proteins. For host-directed antivirals, we identified a small-molecule degrader, FM-74-103, that elicits selective degradation of human GSPT1, a translation termination factor. FM-74-103-mediated GSPT1 degradation inhibits both RNA and DNA viruses. Among virus-specific antivirals, we developed viral RNA oligonucleotide-based bifunctional molecules (Destroyers). As a proof of principle, RNA mimics of viral promoter sequences were used as heterobifunctional molecules to recruit and target influenza viral polymerase for degradation. This work highlights the broad utility of TPD to rationally design and develop next-generation antivirals.

RASopathies due to de novo pathogenic variants: clinical features, genetic findings and outcomes in nine neonates born with congenital heart defects.

BACKGROUND: There are limited information available related to neonatal characteristics of RASopathies, a group of autosomal dominant syndromes with considerable phenotypic overlap. METHODS: The retrospective review revealed 9 neonates born with congenital heart defects (CHDs) and diagnosed as RASopathies due to de novo mutations (DNMs) by trio-based exome sequencing (ES) between January 2017 and December 2020. We report in details of the neonatal course, molecular analysis and 180-days of age follow-up in affected individuals. RESULTS: The early clinical spectrum included various types of CHDs, less noticeable multiple extracardiac anomalies and unspecific symptoms like poor feeding. Of the 8 variants identified from 6 genes, 2 in RASA1 were novel: (NM_002890.2: c.2828 T > C (p.Leu943Pro)) and (NM_002890.2: c.2001del (p.Pro668Leufs*10)), which functionally impaired the protein structure. There was a relatively high mortality rate of 33.33% (3/9) for all the defects combined. A RAF1-deficient male and a RASA1-deficient male survived from severe heart failure by surgical interventions in early life. CONCLUSIONS: Our results revealed that family-based ES was useful in identifying DNMs and causal genes for sporadic diseases and screening Rasopathies shortly after birth. We recommended a family-based ES and a full phenotypic evaluation including echocardiogram, magnetic resonance imaging, ultrasonography and coagulation screening in neonates with CHDs and a suspected genetic etiology.