The gastrointestinal tract of livestock and poultry is prone to challenge by feedborne antigens, pathogens, and other stress factors in the farm environment. Excessive physiological inflammation and oxidative stress that arises firstly disrupts the intestinal epithelial barrier followed by other components of the gastrointestinal tract. In the present review, the interrelationship between intestinal barrier inflammation and oxidative stress that contributes to the pathogenesis of inflammatory bowel disease was described. Further, the role of naturally existing immunomodulatory nutrients such as the omega-3 polyunsaturated fatty acids, citrus pectin, and milk-derived exosomes in preventing intestinal barrier inflammation was discussed. Based on the existing evidence, the possible molecular mechanism of these bioactive nutrients in the intestinal barrier was outlined for application in animal diets. Apart from nutrient absorption, the intestinal epithelial layer (IEL) acts as the first line of host defense against various environmental stress factors. Excessive physiological inflammation and oxidative stress primarily disrupts the inner lining of the gastrointestinal tract such as the IEL. Impairment of IEL can further contribute to the progression of inflammatory bowel diseases (IBD), metabolic disorders, and even mortality in animals –3. TradiGastrointestinal tract is secured by a series of protective layers collectively described as the intestinal mucosa. It mainly encompasses the mucus layer, gut microbiota, IEL, immune cells dispersed in lamina propria and lamina. The IEL physically separates the circulatory system from the external milieu Fig. . BesidesGastrointestinal tract is prone to inflammatory and oxidative damages owing to continuous contact with environmental stress factors. IECs exhibit specialized pattern-recognizing receptors (PRRs) such as the toll-like receptors (TLRs) and nucleotide oligomerization domain (NOD)-like receptors for identification of pathogen-associated molecular patterns (PAMPs) or damage-associated molecular patterns (DAMPs) . SubsequThe pursuance of reducing the antibiotics and non-steroidal anti-inflammatory drugs (NSAIDs) has raised the interest in utilizing naturally existing bioactive compounds. These bioactive compounds are constituents of functional food, similar to conventional foods but provide additional therapeutic benefits. Thus, functional foods are described as the food prepared using ‘scientific intelligence’ vital for optimal health . The EurThe pectins from citrus fruit peels are increasingly recognized for its multiple health-beneficial properties. Although the application of CPn as a feed additive is emerging, preliminary studies have reported to exhibit potential anti-inflammatory, antimicrobial and prebiotic properties –33. BesiAlthough ω-3 PUFAs are widely known feed additive, its mechanism in intestinal barrier is poorly established, which limits the assessment of its efficacy. Based on previous studies, its possible modes of action are described presently. Generally, fatty acids are carbon chain structures of varying length, synthesized in the cytoplasm from Acetyl-CoA . The two2O2), dextran sulfate sodium (DSS), and 2,4,6-trinitrobenzene sulfonic acid (TNBS) [ZO]-1, OCLN, CLDN-2), while decreased the barrier permeability to HRP flux compared to DHA ) or proteins  that involved in the pro-inflammatory response of Caco-2 cells and instead activated the protein expression of inhibitor of nuclear factor kappa B (IκB) and peroxisome proliferator-activated receptor(PPAR)γ 1) and the downstream pro-inflammatory response  [[GPR]120), and tissue repair were significantly enhanced. The gene expression of anti-inflammatory cytokine IL-10, MUC2, and its regulatory gene, free fatty acid receptor-2 (FFAR-2) were also substantially increased -1) -1) . In the 1, OCLN) . In a si1, OCLN) .TNF-α, IL-1β, IL-6, IL-17a, IL-12, MCP-1 or CXCL2) , protein kinase B [Akt]) and tight junction proteins  were significantly recovered by the treatment of porcine MDEs. Additionally, DON-activated genes or proteins that involved in apoptosis  were suppressed by the porcine MDEs. Besides, the miRNAs targeting the p53 pathway were highly expressed in the IPEC-J2 cells post-incubation with porcine MDEs [ and morphological damage. Besides, the human MDEs stimulated the expression of markers specific for epithelial proliferation (Ki67+ cells) and regeneration (LGR5 gene) in the organoids. Amongst the three MDE-treatment groups, colostrum-MDEs exhibited enhanced cytoprotective effects [Further, co-administration of porcine MDEs or MDE-miRNAs with LPS decreased the impact of LPS-induced inflammation and apoptosis of IPEC-J2 cells. Particularly, damage on cell viability, genes or proteins activated in TLR4/NF-κB pathway  of IEC-6 cells under hypoxic stress. Especially, the yak MDEs or MDE-miRNAs enhanced hypoxia resistance by decreasing the protein expression of hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor-A (VEGFA), and different apoptotic markers , while upregulated prolylhydroxylases-1 (PHD-1) [IL-6, while enhanced the goblet cell abundance and MUC2 expression in the hypoxia and LPS-injured neonatal mice intestinal organoids [Several recent studies assessed the capability of MDEs in controlling the hypoxic stress in intestinal barrier. Accordingly, pre-treatment of cow MDEs, yak MDEs or yak MDE-miRNAs enhanced the survival and proliferation (Ki67 (PHD-1) , 151. Ad (PHD-1) , 151. Li (PHD-1) . Similarrganoids .p53 [MUC2, MyD88, regenerating islet-derived protein 3 gamma (RegIIIγ), GATA binding protein-4 (GATA4), immunoglobulin A (IgA), and secretory IgA were significantly increased [. The DON-activated genes and proteins that involved in apoptosis  were also significantly inhibited [ in the DSS-colitis mice. Additionally, the protein level of TGF-β1 and specific miRNAs were upregulated in the mice colon, which subsequently downregulated its target genes as the DNA methyltransferase-1/-3 (DNMT1/DNMT3) [Previously, administration of porcine MDEs in mice significantly improved the small intestinal morphology  and expression of genes and proteins specific for mucosal proliferation , while supressed the p53  (Table 4p53 . In addincreased . Furtherncreased . Similarnhibited . Further1/DNMT3) . Similar1/DNMT3) .+/GRP94+ cells) and decreased the MPO expression in the NEC-neonatal mice [+ cells), while attenuated the distal ileal injury, MPO level and, the gene expression of pro-inflammatory IL-6 [Moreover, MDEs are able to attenuate the severity of NEC induced by a combination of hypoxia, formula feeding, or LPS challenge in neonatal murine. Accordingly, in NEC-rat pups, supplementation of human pre-term MDEs accumulated in the small intestine, which subsequently increased the ileal villus integrity and enterocyte proliferation. For the first time, peptidomic profiling of MDEs revealed the presence of numerous peptides involved in epithelial proliferation, migration, regeneration, and immunomodulation . Similartal mice . Likewistal mice . In anotory IL-6 . A schemBased on European Food Safety Authority, the main target of an immunomodulatory feed additive is to stop the local inflammation and prevent further damage to the immune system. According to existing data, it can be concluded that the dietary omega-3 polyunsaturated fatty acids, citrus pectin, and milk-derived exosomes are able to terminate inflammation at the level of the intestinal barrier. The molecular mechanisms of these nutrients in the intestinal barrier are mainly associated with improving the expression of tight junction proteins, epithelial proliferation, enrichment of mucus layer, immunomodulation and prevention of inflammatory cell infiltration. Further, the nutrients support the maintenance of intestinal equilibrium even under stress conditions by enhancing epithelial proliferation and regeneration. Moreover, these nutrients have demonstrated considerable bioaccessibility and bioavailability across the intestinal epithelium, which is a major challenging factor in feed formulations. Although omega-3 polyunsaturated fatty acids are a well-known anti-inflammatory nutrient over decades, till-date its application in controlling IBD is well established in humans, unlike animals. The lack of specific molecular studies at the intestinal level of livestock and poultry is a major setback for achieving desired health outcomes in farm animals. On the other hand, existing data on citrus pectin and milk-derived exosomes are insufficient for harnessing their application as an immunomodulatory feed additive. However, in the future, the novel property of citrus pectin to form gel matrix with mucin and selectively support the intestinal adhesion of probiotics can be utilized to reconstitute the mucosal epithelium that is damaged during infections, antibiotic therapy, or IBD. Additionally, the barrier penetrating property of low molecular weight citrus pectin can be used to target the pro-inflammatory mediators such as Galectin-3 at the local and systemic level. Intriguingly, both omega-3 polyunsaturated fatty acids and miRNA/peptide cargoes of milk-derived exosomes prevent the incidence of necrotizing enterocolitis by suppressing hypoxic stress. They can be excellent nutritional supplements especially in newborn mammals to preventing undesired inflammatory stress, hypoxia, and mortality during the pre- and post-weaning periods. In summary, all these nutrients pose a promising opportunity for controlling chronic inflammatory diseases and promote gut health in farm animals. However, it is noteworthy to mention that the molecular mechanism of these natural bioactive compounds can be diverse and poor understanding can limit its practical application. Therefore, further studies, especially using high-throughput omics technologies are necessary in order to enumerate their precise molecular mechanisms for efficient utilization in animal diets.


Whole genome sequencing from multiple mink was consistent with published SARS-CoV-2 genomes with few polymorphisms. The Utah mink SARS-CoV-2 strains fell into Clade GH, which is unique among mink and other animal strains sequenced to date. While sharing the N501T mutation which is common in mink, the Utah strains did not share other spike RBD mutations Y453F and F486L found in nearly all mink from the United States. Mink in the outbreak reported herein had high levels of SARS-CoV-2 in the upper respiratory tract associated with symptomatic respiratory disease and death.The breadth of animal hosts that are susceptible to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and may serve as reservoirs for continued viral transmission are not known entirely. In August 2020, an outbreak of SARS-CoV-2 occurred on five mink farms in Utah and was associated with high mink mortality (35–55% of adult mink) and rapid viral transmission between animals. The premise and clinical disease information, pathology, molecular characterization, and tissue distribution of virus within infected mink during the early phase of the outbreak are provided. Infection spread rapidly between independently housed animals and farms, and caused severe respiratory disease and death. Disease indicators were most notably sudden death, anorexia, and increased respiratory effort. Gross pathology examination revealed severe pulmonary congestion and edema. Microscopically there was pulmonary edema with moderate vasculitis, perivasculitis, and fibrinous interstitial pneumonia. Reverse transcriptase polymerase chain reaction (RT-PCR) of tissues collected at necropsy demonstrated the presence of SARS-CoV-2 viral RNA in multiple organs including nasal turbinates, lung, tracheobronchial lymph node, epithelial surfaces, and others. Localization of viral RNA by  The recent emergence and spread of the novel coronavirus has resulted in worldwide disease and economic hardship. The virus, known as SARS-CoV-2 is believed to have originated in bats and has spread worldwide through human-to-human virus transmission. It remains unclear which animal species, other than humans, may also be susceptible to viral infection and could naturally transmit the virus to susceptible hosts. In this study, we describe the early phases of an outbreak of disease and death due to SARS-CoV-2 infection in farmed mink in Utah, United States. The investigation reveals that mink can spread the virus rapidly between animals and that the disease in mink is associated with viral infection and damage to tissues of the upper and lower respiratory system. The determination that mink are susceptible to SARS-CoV-2 indicates the need for strict biosecurity measures on mink farms to remediate mink-to-mink and human-to-mink transmission for the protection of mink, as well as prevent potential transmission from mink to humans. It may be prudent for humans working with SARS-CoV-2 mink to wear appropriate PPE until more is understood. Rhinolopphus affinis) to human beings, possibly via an undetermined intermediate host  using MagMAX-96 Viral RNA Isolation Kit, per the manufacturer’s instructions.RNA was extracted from fresh tissue samples in TRIzol Reagent and formalin fixed paraffin embedded tissues using TRIzol reagent , per the manufacturer’s instructions. FFPE tissues were cut in 10um sections and heated at 65°C for 10 minutes in TRIzol prior to RNA extraction using MagMAX-96 Viral RNA Isolation Kit, per the manufacturer’s instructions.Reverse transcriptase (RT) real-time PCR to the SARS-CoV-2 RNA-dependent RNA polymerase gene (RDRp) was performed as previously described using primers SARS-CoV-2 primers RdRp_SARSr-F2 5’-GTGARATGGTCATGTGTGGCGG-3’ and COVID-410R 5’-CCAACATTTTGCTTCAGACATAAAAAC-3’ , using Thttps://www.gisaid.org/epiflu-applications/covsurver-mutations-app.Libraries for the whole genome sequencing were generated using the Ion AmpliSeq Kit for Chef DL8 and Ion AmpliSeq SARS-CoV-2 Research Panel . Libraries were sequenced using an Ion 520 chip on the Ion S5 system using the Ion 510 & Ion 520 & Ion 530 Kit. Sequences were assembled using IRMA v. 0.6.7 and visually verified using DNAStar SeqMan NGen v. 14. Mutational analysis was performed using the GISAID EpiFlu Database CoVsurver: Mutation Analysis of hCoV-19 at In situ hybridization utilized RNAscope  technology to visualize the presence and location viral RNA in tissues harvested from infected mink. A set of anti-sense SARS-CoV-2 specific RNA probes comprised of 20 Z pairs targeting nucleotides 21,631–23,303 of the spike viral glycoprotein gene (Genbank accession number NC_045512.2) was developed by Advanced Cell Diagnostics (ACD) and performed as previously described [In parallel with the collection of tissues collected in TRIzol for RT-PCR investigation, duplicate sections of those tissues were collected in formalin to be used for ISH from mink 1–4. escribed . This asThere was detection of viral RNA in inflamed splenic tissue from an FIPV infected cat, which served as a positive assay control for performance and presence of RNA in FFPE handled similar to the mink tissues in this study. No viral RNA was detected (no cross-reactivity) in the negative control slides handled similarly to the mink tissues. These negative controls included applying the SARS-CoV-2 probe to tissues  from one healthy adult mink that died of crush injuries prior to the emergence of SARS-CoV-2, spleen from an FIPV-infected cat, intestines and trachea from a bovine calf infected with bovine coronavirus, and nasal turbinates from a cat with suppurative rhinitis collect prior to the emergence of SARS-CoV-2. Additionally, there was no FIPV detection when this probe was applied to the SARS-CoV-2 positive mink nasal turbinates and lungs .


All eukaryotic cells are composed of the cytoskeleton, which plays crucial roles in coordinating diverse cellular functions such as cell division, morphology, migration, macromolecular stabilization, and protein trafficking. The cytoskeleton consists of microtubules, intermediate filaments, and actin filaments. Cofilin, an actin-depolymerizing protein, is indispensable for regulating actin dynamics in the central nervous system (CNS) development and function. Cofilin activities are spatiotemporally orchestrated by numerous extra- and intra-cellular factors. Phosphorylation at Ser-3 by kinases attenuate cofilin’s actin-binding activity. In contrast, dephosphorylation at Ser-3 enhances cofilin-induced actin depolymerization. Cofilin functions are also modulated by various binding partners or reactive oxygen species. Although the mechanism of cofilin-mediated actin dynamics has been known for decades, recent research works are unveiling the profound impacts of cofilin dysregulation in neurodegenerative pathophysiology. For instance, oxidative stress-induced increase in cofilin dephosphorylation is linked to the accumulation of tau tangles and amyloid-beta plaques in Alzheimer’s disease. In Parkinson’s disease, cofilin activation by silencing its upstream kinases increases α-synuclein-fibril entry into the cell. This review describes the molecular mechanism of cofilin-mediated actin dynamics and provides an overview of cofilin’s importance in CNS physiology and pathophysiology. Neurons contain a cytoskeleton consisting of microtubules, neurofilaments, and actin filaments. Microtubules are composed of tubulin proteins and other polypeptides and provide the essential organization of organelles. Neurofilaments, which are class IV intermediate filaments, offer structural support to axons and influence nerve conduction velocity. Actin filaments (F-actin) are composed of globular actin monomers (G-actin). Dynamic transition can occur between G-actin and F-actin, and the polymerization-depolymerization events are spatiotemporally regulated in response to numerous extracellular and intracellular stimuli. Actin is an ATPase, and both G-actin and F-actin can bind to ATP. When ATP-bound G-actin monomers assemble into a polymer, ATP hydrolyzes rapidly to generate ADP-Pi actin, slowly releasing the inorganic phosphate producing ADP-actin subunits. Consequently, actin filaments exhibit considerable asymmetry with a plus end (barbed end or growing end) and a minus end (pointed end or shrinking end) dominated by ATP- and ADP-actins, respectively. ATP-G-actin monomers are preferentially polymerized to the barbed end whereas, ADP-actin subunits are depolymerized from the pointed end. The actin dynamics is precisely controlled by various actin-binding proteins (ABPs) such as actin-related protein 2/3 complex (Arp2/3), cortactin, formin, profilin, and actin-depolymerizing factor (ADF)/cofilin (reviewed in ).DSTN in humans, was initially identified in the chick brain )ophrenia . OPCML wturation . This reAnother study reported that functional inhibition of 14-3-3 protein in neurons in the mouse nervous system lead to behavioral deficits corresponding to the core symptoms of schizophrenia . A low l2+, increased excitotoxicity, lack of oxygen, and glucose. Overload of Ca2+ activates NADPH-oxidase (NOX) through PKC and nitric oxide (NO), leading to a high level of glutamate accumulation in extracellular space . Glfunction . Severalfunction ,120,121.hondria. , promotihondria. . Anotherhondria. .Moreover, cofilin oxidation may lead to oxidant-induced apoptosis . IschemiUsing a rat stroke model, Shu et al. (2019) suggested that rod formation disrupts dendritic mitochondrial trafficking during ischemic conditions . AccumulPharmacological induction of the 70 kDa heat shock protein (HSP70) has been suggested as a potential therapeutic intervention for stroke . The HSPPD is another age-related neurodegenerative disorder characterized by the dopaminergic neuron loss in the substantia nigra and by the presence of Lewy neurites (LNs) and Lewy bodies (LBs) in cortical and subcortical neurons . A signiExtracellular α-synucleins upregulate surface-exposed glucose-related protein of 78kDa  that becomes clustered into microdomains of the neuronal plasma membrane . InteracThe pathological α-synuclein-fibrils are transmitted from the fibril-generating cells to recipient cells and spread throughout the brain. Cofilin was found to facilitate the α-synuclein-fibril uptake into recipient cells and the expansion of α-synuclein pathology . The comPARK2) gene have been linked to autosomal recessive juvenile parkinsonism and early-onset parkinsonism [Mutations of the parkin (insonism ,136. Liminsonism . FurtherThe findings generated from a plethora of studies implicate that proper balance in cofilin activity is a prerequisite for actin turnover and CNS functions. Among various regulatory pathways already discovered for cofilin, phosphoregulation through SSHs and LIMKs is the most critical mechanism, where the Ser-3 residue of cofilin is the specific target. Roles of cofilin Cys-39 and Cys-147 residues are also becoming apparent in the context of ROS-induced rod formation in many neurodegenerative disorders. Newer studies report different molecular pathways of cofilin through which cofilin dysregulation and translocation in subcellular regions might be associated with various CNS disorders.Recent advances in experimental techniques will significantly facilitate the understanding of the signaling pathways of cofilin function in development and disease conditions. Drugs or peptides targeting the critical amino acid residues of cofilin might be a new potential therapeutic strategy for neurodegenerative disorders.


In this study we employed Hidden Markov Model-based searches to identify ~1059 Caenorhabditis elegans neuropeptide G-protein coupled receptor (Ce-NP-GPCR) encoding gene homologs in the predicted protein datasets of 10 key parasitic nematodes that span several phylogenetic clades and lifestyles. We show that, whilst parasitic nematodes possess a reduced complement of Ce-NP-GPCRs, several receptors are broadly conserved across nematode species. To prioritize the most appealing parasitic nematode NP-GPCR anthelmintic targets, we developed a novel in silico nematode parasite drug target prioritization pipeline that incorporates pan-phylum NP-GPCR conservation, C. elegans-derived reverse genetics phenotype, and parasite life-stage specific expression datasets. Several NP-GPCRs emerge as the most attractive anthelmintic targets for broad spectrum nematode parasite control. Our analyses have also identified the most appropriate targets for species- and life stage- directed chemotherapies; in this context we have identified several NP-GPCRs with macrofilaricidal potential. These data focus functional validation efforts towards the most appealing NP-GPCR targets and, in addition, the prioritization strategy employed here provides a blueprint for parasitic nematode target selection beyond NP-GPCRs.Nematode parasites undermine human health and global food security. The frontline anthelmintic portfolio used to treat parasitic nematodes is threatened by the escalation of anthelmintic resistance, resulting in a demand for new drug targets for parasite control. Nematode neuropeptide signalling pathways represent an attractive source of novel drug targets which currently remain unexploited. The complexity of the nematode neuropeptidergic system challenges the discovery of new targets for parasite control, however recent advances in parasite ‘omics’ offers an opportunity for the  Nematode parasites continue to have a global impact on human health and agricultural productivity such that novel mode-of-action anthelmintics are critical for sustained parasite control, especially in light of the escalation in anthelmintic resistance –3. WhilsWithin the neuromuscular signalling system, neuropeptide GPCRs (NP-GPCRs) have been identified as highly ‘druggable’ targets . Indeed,in silico data for parasitic nematodes  characterise the NP-GPCR complements of 10 key parasitic nematode species; (ii) develop a novel nematode drug target prioritization pipeline that incorporates NP-GPCR conservation, expression and functional data, and (iii) identify NP-GPCRs that represent putative, novel, broad spectrum parasitic nematode control targets. Integration of these multi-omics-derived datasets provides a springboard for functional biology that will improve our understanding of fundamental nematode neurosignalling and support future anthelmintic discovery efforts.In this study we employed NP-GPCRs were identified via multiple sequence alignment derived Hidden Markov Models (HMMs), using methods based on those previously described  was employed to identify potential NP-GPCRs within the predicted protein datasets of 10 phylogenetically dispersed nematode parasites  to identify the most similar sequences in C. elegans. Queries that failed to return a putative NP-GPCR as the highest scoring pair/top hit were excluded from downstream analyses. Putative NP-GPCR sequences were then filtered based on the number of transmembrane (TM) domains, as predicted by hmmtop (NP-GPCR datasets), but still included in the drug target prioritization pipeline  was used to identify convex clusters within the NP-GPCR datasets (NP-GPCR hits (761 putative parasite NP-GPCR sequences with ≥4 TM domains) were analysed alongside all putative NP-GPCRs from C. elegans . Individual clusters were used in Maximum-likelihood phylogenetic tree construction using MEGA 7 or MEGA CC ’ option to determine the most appropriate model of evolution for tree construction. All trees were constructed using: the bootstrap method (500 replicates); the LG model of evolution (G+I) with 5 discrete Gamma categories; a partial deletion of gaps (80% site coverage cut-off); and the nearest-neighbour interchange algorithm with no branch swap filter. Trees were rooted using a selection of C. elegans biogenic amine receptor sequences , predicted proteins with <4 TM regions were included as homologs of the highest scoring C. elegans BLAST hit.A drug target prioritization pipeline based on: (i) NP-GPCR-encoding gene conservation (generated in this study); (ii) nal data , and , where no recorded phenotype scored 0, reproductive scored 2, sterility scored 3, motility scored 4, and lethality scored 5. Any other recorded phenotype scored 1. Many Ce-NP-GPCR encoding genes had multiple phenotypes recorded; in this scenario, phenotype scores were combined to provide an overall total phenotype score for each NP-GPCR. Where multiple phenotypes within the same category were recorded, the category was only scored once.Phenotype data associated with T. muris  were generated following an established RNASeq pipeline. Raw sequences reads . Several key points emerge from this study:In this study we identified 1059 putative ites see Figure 1Ce-NP-GPCRs  profiles characterised previously  also display the largest and smallest complements of parasite neuropeptide encoding genes respectively.All 10 parasitic nematodes examined in this study exhibited restricted profiles of the 152 eviously , 37, wheGPCRs in addition to the NP-GPCRs reported here (data not shown); this provides confidence that all putative NP-GPCRs were identified in the available parasitic nematode datasets. Although a small number of divergent NP-GPCR sequences without an obvious C. elegans ortholog were identified in specific parasite species .The NP-GPCR encoding gene profiles of parasitic nematodes representing five nematode clades  were exNP-GPCR complement was evident; for example, whilst the clade 2 species, T. muris and T. spiralis, displayed a highly similar, reduced, NP-GPCR complement (21%), an additional clade 2 species, R. culicivorax (entomopathogenic nematode), possessed 49% of Ce-NP-GPCR homologs . These data suggest multiple distinct gene loss events in the lineages that led to present day Trichuris/Trichinella and filarid spp. It is also likely that some of the 152 Ce-NP-GPCRs arose from gene duplication events that occurred in the lineages that led to the crown clades , and so the NP-GPCRs absent from clade 2 species may not have been present in the last common ancestor of all nematodes. In contrast, both of the plant parasitic nematodes examined, B. xylophilus (clade 10) and G. pallida (clade 12), display relatively similar NP-GPCR profiles despite their distinct clade designations  and Drosophila FMRF-amide (frpr-11-13) GPCR families  indicating their general importance to nematode biology , T. muris (L2), G. pallida (J2), D. immitis (microfilariae). This indicates that, whilst NP-GPCRs have an important role across the nematode lifecycle, there may be an enhancement of NP-GPCR signaling in the larval stages that could reflect a significant need for movement/migration and development at this stage. Whether these patterns of NP-GPCR expression can be directly tied to variation in gene function between lifecycle stages remains to be investigated.The majority of NP-GPCR profiles outlined above challenge the ability to prioritise the most attractive NP-GPCRs for validation as novel drug targets. In silico approaches offer a novel route to exploit available datasets and integrate information to direct drug target selection (in silico nematode parasite NP-GPCR drug target prioritization pipeline that incorporates pan-phylum NP-GPCR conservation (generated in this study), parasite life-stage specific expression, and C. elegans-derived phenotype data to assess the target appeal of NP-GPCRs for nematode control  show highly restricted patterns of conservation.Seven of the 152 Caenorhabditis elegans functional data may inform NP-GPCR target appeal through the collation and consideration of phenotype information. In this study we collated phenotype data from C. elegans null mutant/RNAi experiments for the 152 NP-GPCRs [see WormBase . Often, multiple phenotypes were attributed to individual NP-GPCRs, therefore scores were added to yield an overall phenotype score for each receptor (see Ce-NP-GPCRs had no associated null mutant/RNAi phenotype(s) which may reflect a combination of: (a) lack of functional analyses data for Ce-NP-GPCRs, (b) use of an unsuitable C. elegans post-functional genomics bioassay and/or, (c) functional redundancy in nematode neuropeptidergic signalling systems; (ii) 16 Ce-NP-GPCRs had a lethal phenotype reported in at least one study  include npr-1, -2, -8, -9, -10, -13, -25, and F59D12.1.NP-GPCRs that are associated with lethal phenotypes in C. elegans three  are present in the most important APN/HPN species in this study  are present in the most important APN/HPN species in this study  the reported differences between phenotypes recorded for multiple distinct mutations associated with the same gene, as well as RNAi animals , (ii) the variable and often specific nature of the phenotype screens employed, and (iii) the bias in the volume/quality of functional data for specific NP-GPCRs or GPCR families. Whilst we have attempted to incorporate all of the observed C. elegans phenotypes recorded on WormBase regardless of experimental approach , the major caveats outlined above somewhat limit the utility of these data, and emphasise the need for functional analysis of all highly conserved and highly expressed NP-GPCRs in parasitic nematodes. Despite this, the approach offers a route to prioritising drug target candidates for functional validation in low throughput parasite platforms.In the context of this study, there are several important caveats to the extrapolation of the WormBase derived C. elegans phenotype. This enables for the segregation or integration of prioritisation criteria as required and for the addition of phenotype data as they become available.The format of our prioritisation pipeline allows for the distinct prioritisation of NP-GPCRs based on conservation, expression and/or NP-GPCRs are broadly expressed across the species examined in this study , the majority are also expressed in therapeutically relevant lifecycle stages ; of these, npr-1, fshr-1, F59D12.1 and pdfr-1 also display defective phenotypes in C. elegans .The available nematode RNASeq data suggest that parasite NP-GPCRs as the most appealing broad spectrum drug target candidates  have been linked to several peptides. NPR-11 has been functionally linked to NLP-1 and FLP-34 and, heterologously matched with FLP-21, -18, -34, -15 and -27  peptides, which modulate locomotion (C. elegans (pdf-1 and pdfr-1 were present together in 96% of 134 nematode genomes (unpublished observations).PDFR-1 is related to arthropod Pigment Dispersing Factor Receptor and, more distantly, to vertebrate Calcitonin and Vasoactive Intestinal Peptide receptors . These rcomotion , 59. PDF elegans , 60, 61.via RNAi experiments which resulted in slow and paralyzed worms (Microbacterium nematophilum infection in C. elegans (fshr-1 indicate that mutant C. elegans (tm3954) and RNAi worms display lethal phenotypes , has been associated with locomotion ed worms . PCDR-1  elegans . The funenotypes , 64.NP-GPCRs  were conserved in all species examined in this study, the limited functional information available for these receptors has precluded their prioritisation as the most appealing targets at this point  highly variable and often gene dependent and, (ii) focus almost exclusively on loss of function screens (lack of over-expression data), such that this results in a degree of bias within the NP-GPCR prioritisation pipeline whereby highly conserved receptors that simply lack phenotype data are not emerging among the prioritised subset. Indeed, this is supported by a lack of correlation between parasitic nematode NP-GPCR conservation and the C. elegans derived phenotype data reported here ; this suggests potential functional importance and should form the focus of future functional analyses in parasitic nematodes.Whilst several additional oint see Figure 4NP-GPCRs  emerge from our pipeline as NP-GPCRs that are expressed in both adult male and female B. malayi and D. immitis  however, an additional two NP-GPCRs (ckr-1 and Y40C5A.4) emerge that are also appealing as microfilaricides; this underscores the utility of the NP-GPCR prioritisation pipeline in teasing out species specific therapeutic targets (see Narrow spectrum anthelmintics have proven utility in nematode parasite control strategies . The drugets see 4.NP-GPCR conservation and the application of a drug target prioritisation pipeline that highlights the most attractive parasitic nematode NP-GPCRs for parasite control at this time. These data: (i) provide a comprehensive library of NP-GPCRs in key nematode parasites; (ii) enable the selection of both broad and narrow spectrum control targets; (iii) inform future validation efforts for NP-GPCRs in key parasitic nematode systems which are currently significantly lacking and, (iv) will expedite the anthelmintic development pipeline via informed target selection.Recently improved parasite ‘omics’ data have driven a paradigm-shift towards mechanism-directed drug target screening approaches, providing an opportunity to identify the most attractive nematode parasite targets. Our focus on NP-GPCRs as therapeutic targets is driven by their importance to nematode biology , howeverThe original contributions presented in the study are included in the article/AM, LA, CM, AGM, and NM designed the research. LA, CM, BC, PM, DM, AI, FM, and BAR performed the research. LA, CM, and BC analysed the data with assistance from BAR and MM. AM, LA, CM, AGM, NM, and JH wrote the manuscript. All authors contributed to the article and approved the submitted version.This work was supported by: the Biotechnology and Biological Sciences Research Council (BB/H019472/1 to AM); the Biotechnology and Biological Sciences Research Council/Merial Animal Health grant ; the Biotechnology and Biological Sciences Research Council/Boehringer Ingelheim ; the Department of Education and Learning for Northern Ireland (studentships awarded to FM and BC); the Department of Agriculture, Environment and Rural Affairs for Northern Ireland (studentships awarded to DM and AI); the National Institutes of Health-National Institute of Allergy and Infectious Diseases grant (AI159450 to MM).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The evolutionarily conserved NOTCH signaling displays pleotropic functions in almost every organ system with a simple signaling axis. Different from many other signaling pathways that can be amplified via kinase cascades, NOTCH signaling does not contain any intermediate to amplify signal. Thus, NOTCH signaling can be activated at distinct signaling strength levels, disruption of which leads to various developmental disorders. Here, we reviewed mechanisms establishing different NOTCH signaling strengths, developmental processes sensitive to NOTCH signaling strength perturbation, and transcriptional regulations influenced by NOTCH signaling strength changes. We hope this could add a new layer of diversity to explain the pleotropic functions of NOTCH signaling pathway. Drosophila and subsequent discovery of notch gene one century ago  together with other transcription co-factors to activate gene transcription  and four NOTCH receptors , all of which contain extracellular epidermal growth factor (EGF)-like domains executing ligand–receptor binding. Subsequently after binding, NOTCH receptor undergoes two successive enzymatic cleavages mediated by ADAM10 and γ-secretase, releasing the NICD into cell nucleus where it binds with NOTCH signaling transcription factor CSL together with other co-factors to activate gene transcription . The mosUnlike many other signaling pathways, NOTCH signaling can be activated at distinct strength levels due to following three reasons: (1) one NOTCH receptor can only release one NICD after ligand–receptor binding, (2) no signal intermediate or kinase cascade is involved to amplify the initial signal, and (3) NICD is subjected to proteasome-mediated degradation after transcriptional activation . Since CThe frequency of NOTCH ligand and receptor binding determines the amount of NICD generated. In order to achieve a successful binding-induced NOTCH receptor cleavage, both ligand and receptor need to be presented on cell membrane in close proximity. Since all active NOTCH ligands and receptors are trans-membrane proteins that are subjected to consistent endocytosis, recycling, and degradation, the amount of NICD that could be potentially generated is affected by endocytic regulations of the ligands and receptors . MeanwhiThe stability of NICD affects the duration of NOTCH signaling. NICD is generated following ligand–receptor binding and shuttles into cell nucleus, where it binds to transcription factor CSL together with co-factor of mastermind-like protein (MamL) and other chromatin modifiers to activate gene transcription. In addition to transcriptional regulations, CSL and MamL also recruit kinase CDK8 to phosphorylate NICD, which triggers protein ubiquitination on PEST  domain of NICD and proteasome-mediated NICD degradation . Thus, NNotch-1 haploinsufficiency has been discovered in AOS patients with variable levels of cardiovascular anomalies, such as ventricular septal defects, aortic stenosis, regurgitation, and coarctation  is also found to be haploinsufficient in AOS patients  characterized by congenital defects of limbs and scalp. rctation . In linepatients , implyinmutation . CollectJag-1  that affects multiple organs including the liver, heart, eye, kidney, and bone. AGS patients are reported to carry heterozygous mutation of Jag-1  or Notch specification and artery branching (vs. elongation). During early embryonic development, progenitor cells resident in aorta-gonad-mesonephros (AGM) region can give rise to either arterial endothelial cell or hematopoietic stem cell (HSC) depending on NOTCH signaling strength. fication . In addiranching .Jag-1-mutated mice lowered Dll-4-activated NOTCH signaling and rescued vascular branching defects  into  defects . Therefo defects . Collect-SMA and SM-22α and the final specification of VSMC fate  differentiation. VSMCs are recruited to vascular endothelial cells during vasculogenesis and play crucial roles on maintaining normal vascular tone in response to hemodynamic changes, especially in arteries where multiple layers of VSMCs are attached to endothelial cells . During SMC fate . Meanwhiof VSMCs . Therefoin vitro study also showed that over-expression of NICD in VSMCs can increase cell proliferation  with increasing concentrations gradually switched γδ T-cell into αβ T-cell  specification in T-cell lineage. Postnatal development of T immune cell in the thymus requires activation of NOTCH signaling in the hematopoietic progenitor cells (HPCs) that migrated from the bone marrow. NOTCH signaling activation inhibits non-T-cell cells including myeloid lineage during early stages and B-cell during late stages . Howeverβ T-cell , documenThe strength changes of NOTCH signaling in T-cell lineage is caused by binding with different NOTCH ligands that hold distinct receptor binding affinities. Still in human HPCs, Jag-2 exhibited strong NOTCH activation potential and directed HPCs predominantly into γδ T-cell , 2013; DNotch-2 completely eliminated MZB cells. Interestingly, heterozygous mutation of Notch-2, in which 50% of Notch-2 mRNA still expressed, resulted in partial reduction of MZB cells  showed a similar dose-dependent regulation of MZB cell differentiation. Wild-type, heterozygous mutation of MamL-1 and homozygous mutation of MamL-1 showed sequential reduction of MZB cell number  cell, an immune cell developed in marginal zone of the spleen, relies on NOTCH signaling dosage. Notch-2 is preferentially expressed in B-cell and is prominent in splenic marginal zone, suggesting its potential function on MZB cell differentiation. Homozygous mutation of ZB cells . Consistl number , suggestDll-1 similarly showed a sequential reduction of MZB cell among wild-type, heterozygous mutation of Dll-1 and homozygous mutation of Dll-1  to the developing pancreas promoted progenitor cell proliferation and consequently expanded pancreatic endocrine progenitor pool. However, saturated DAPT led to differentiation of pancreatic secreting cells accompanied by drastically reduced progenitor pool (Stepwise downregulation of NOTCH signaling strength in pancreatic endocrine progenitor cells drives the transition from quiescence to proliferation and from proliferation to differentiation. NOTCH signaling is well-known for its function on maintaining pancreatic progenitor cells, and suppressing NOTCH signaling triggered the progenitor cells to differentiate into pancreatic secreting cells . Interestor pool , confirmConclusively, high NOTCH signaling strength maintains quiescent state of pancreatic endocrine progenitor cell, moderate NOTCH signaling strength triggers its proliferation, and low NOTCH signaling strength leads to its final differentiation. Combining NOTCH haploinsufficiency-related disorders, precise strength requirements in artery endothelial cell, VSMC, T-cell, and MZB cell, developmental processes can be sensitive to NOTCH signaling strength perturbations (summarized in Hes1 and Hey1/L responded differently to the level changes of NICD, the active component of NOTCH signaling. Interestingly, Hes1 and Hey1/L did not simply adjust their expression levels in proportion to the amount of NICD generated. High and sustained NOTCH activation induced by Dll-4 initiated Hey1/L transcription compared to pulsatile NOTCH activation induced by Dll-1, instead of proportionally increasing Hes1 that was already transcribed under pulsatile NOTCH activation (Different duration and dynamics of NOTCH signaling strengths yield distinct transcriptional responses, which resulted in distinct developmental consequences. A new study reported by tivation . ConsideDosage change of NICD can influence CSL-DNA binding kinetics, NICD dimerization, and chromatin opening. A recent study using live cell imaging changed the classical model that CSL stays on chromatin to repress gene transcription until the arrival of NICD that switches CSL into a transcriptional activator . InsteadThe on/off switch of NOTCH signaling is well-recognized for its function on regulating cell differentiation, proliferation, and apoptosis. The pleotropic function of NOTCH signaling seems contradictory to the simple setting of NOTCH signaling axis. Genetic studies discovered heterozygous mutations of NOTCH signaling components in various developmental disorders and diseases, suggesting that organ development is also sensitive to NOTCH signaling dosage. More importantly, simple changes of NOTCH signaling strength influences the binary cell fate determinations and cell proliferation and differentiation during artery, postnatal T-cell, MZB cell, and pancreas development, suggesting that NOTCH signaling strength changes can be as important as on/off switch of the signaling. Underlying NOTCH signaling strength changes and NICD–CSL complex-mediated gene transcriptions are changed both quantitatively and qualitatively to direct distinct cellular responses. Therefore, NOTCH signaling strength can add a new layer of diversity to explain the pleotropic functions of NOTCH.WS and YW conceived the study. All authors contributed their expertise in NOTCH signaling, epigenetic regulation, and cardiovascular development, respectively and contributed to the finalization of this manuscript significantly.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


However, what is currently unknown is how these immunostimulatory DAMPs are counteracted to avoid immune-overactivation. Our recent work builds on these fundamentals and introduces prostaglandin E2 (PGE2) as an ‘inhibitory' DAMP—a new variable to the ICD equation. Prostaglandin E2 functions as an immunosuppressive counterpoise of adjuvant DAMPs; and thus, mechanistically precludes ICD. Furthermore, the long-debated immunogenicity of gemcitabine chemotherapy was revealed to be contingent on inhibitory DAMP blockade and not due to its inability to promote DAMP expression  as previously reported. These findings were intriguing. First, despite the presence of gemcitabine-induced hallmark DAMPs, the inhibitory DAMP  was sufficient to hinder the ICD-induced antitumoral immune response (Dying (or dead) cells are increasingly recognized to impose significant biological influence within their tissues of residence—exerting paracrine effects through proteins and metabolites that are expressed or secreted during cellular demise. For example, certain molecules function as potent mitogens, promoting the repopulation of neighboring epithelial cells. And other myriad of factors—classified as damage-associated molecular patterns (DAMPs)—function as “find me” (attractant), “eat me” (engulfment), or “danger” (activation) signals for recruiting and activating effector immune cells  to initiate inflammation. Since the discovery of immunogenic cell death (ICD), the current dogma posits DAMPs as immunological adjuvants for innate immune cell mobilization and activation, which ultimately leads to the antitumoral cross-priming of CD8response . And secresponse . In this Such finding begs the conceptual question, are there additional iDAMPs to fine-tune inflammation? Rather than a system that is solely dependent on the hallmark, adjuvant DAMP signaling, it is rational to think of ICD as a tug-of-war  between both immunostimulatory and inhibitory DAMPs. Here, we provocatively hypothesize that newer iDAMPs could be discovered within the next decade, and these newer findings will have a profound impact on the conceptualization of ICD and other physiological conditions.How are immunostimulatory DAMPs shut off? Under normal physiological conditions, the co-stimulatory molecules, CD80/86, on antigen presenting cells bind to the co-activating signal, CD28, on T cells to bolster their activation. In contrast, CD80/86 can also bind to the corresponding co-inhibitory signal, CTLA4, which functions as a competitive buffer to prevent uncontrolled T cell over-activation . Thus, it seems plausible that there are co-inhibitory mechanism(s) set in place to prevent the overt DAMP-driven immune activation. Currently, only a few DAMPs have known antagonists; of such, interleukin-1 (IL-1) and its receptor antagonist (IL-1Ra). The discovery of PGEi) the cell surface translocation of CRT from the endoplasmic reticulum and ii) the extracellular release of HMGB1 and ATP, has paved the path to systematically identify novel ICD-inducing drug candidates. How would iDAMP(s) conceptually impact ICD-drug discovery? Currently, amongst a panel of candidate drugs that molecularly promote hallmark DAMPs, only a few are capable of inducing ICD in vivo. Under these circumstances, are iDAMP(s) a cause for the low ICD-drug turnout? The incorporation of iDAMP(s) into the drug discovery pipeline may better assess a drug's ICD-inducing potential. Anecdotally, PGE2 release was approximately three-folds lower in mitoxantrone-treated (a known ICD-inducer) murine bladder cancer cells, when compared to the gemcitabine-treated cells in vitro. Such findings beg the question of whether bonafide ICD-inducers promote less iDAMP release in general? If so, what are the mechanisms? It is possible that anthracyclines  preclude COX-2 protein synthesis by preventing chromatin decondensation; thus, mitigating RNA translation. RT-qPCR-based assays can easily address this question. Can other ICD-inducers also halt COX-2 mRNA translation by different mechanisms? Further studies are required to dissect these underlying mechanism(s) that regulate iDAMPs during drug-induced cell death. In the meantime, novel drugs that promote hallmark DAMPs, but can also concurrently prevent iDAMP biosynthesis may pose as a lucrative class of ICD-inducers for clinical translation.Currently, only a handful of drugs are considered as bonafide ICD-inducers. The elegant integration of DAMP biosensor cell lines that measures fluorescence to distinguish The clinical prognostic value of DAMPs remains unresolved. On one hand, calreticulin expression—in the context of chemotherapeutic treatment—in acute myeloid leukemia (n=20), colorectal cancer (n=68), neuroblastoma (n=68), and ovarian cancer (n=22) is favorable. While in bladder cancer (n=30), mantle cell lymphoma (n=92), and neuroblastoma (n=251), calreticulin levels correlate with poor overall survival. Similarly, HMGB1 levels in breast cancer (n=232) positively correlate with favorable prognosis, while in pancreatic cancer (n=78), high HMGB1 is unfavorable. Do these mixed results imply additional iDAMP to be considered as biomarkers? Perhaps, a panel of both stimulatory and inhibitory DAMPs would be appropriate for future clinical studies to resolve their prognostic capacity.+ T cells were polarized towards a type-I phenotype. However, this strategy was in the context of a vaccination assay. To advance our findings into human clinical trials, we are currently implementing pharmacological iDAMP blockade with gemcitabine-based chemotherapy treatment in preclinical model(s). To generalize the applicability of this strategy, we are evaluating other tumor types  using the same therapeutic strategy. We hypothesize that concurrent iDAMP blockade during gemcitabine-based chemotherapy treatment will promote a type-I T cell polarizing tumor immune microenvironment, resulting in the selective expansion of tumor-targeting cytotoxic CD8+ T cells. Furthermore, we speculate the sequential administration of iDAMP blockade plus chemotherapy followed by immune-checkpoint blockade therapy (ICBT) will prime a durable T cell response, which could increase the spectrum of patient response toward ICBT . Our rationalization is supported by the works of others, where the combination of ICD-inducing chemotherapy plus ICBT displayed better therapeutic efficacy than chemotherapy or ICBT alone in both pre-clinical and clinical settings. Our finding is timely, since a recent Phase 3 clinical trial combining chemotherapy with ICBT in metastatic bladder cancer (IMvigor130) failed to meet its primary endpoint. Our results, together with those from others, provide a strong rationale to evaluate celecoxib/EP antagonists in combination with chemotherapy and ICBT. We highly anticipate future preclinical and clinical studies to evaluate the efficacy of iDAMP blockade in combination with ICB-based therapies and foresee these discoveries to benefit patients across all cancer types.Conventional strategies utilize adjuvant drugs that substantiate immunostimulatory DAMP expression/release to convert non-ICD-inducers into ICD-inducing drugs. Our current study revealed an alternative approach; inhibitory DAMP blockade converted gemcitabine (a non-ICD-inducer) into an ICD-inducing chemotherapy. By mitigating iDAMP release from dying cancer cells, bone marrow-derived dendritic cells displayed an immunogenic maturation phenotype and vaccine-draining lymph node CD8In conclusion, we hope that our study will spark the future identification of novel iDAMPs. And that these newer findings will initiate the integration of iDAMPs for assessing ICD in both basic research and clinical settings.


Age and disease prevalence are the two biggest risk factors for COVID-19 symptom severity and death. We therefore hypothesized that increased biological age, beyond chronological age, may be driving disease-related trends in COVID-19 severity. Using the UK Biobank England data, we tested whether a biological age estimate (PhenoAge) measured more than a decade prior to the COVID-19 pandemic was predictive of two COVID-19 severity outcomes . Logistic regression models were used with adjustment for age at the pandemic, sex, ethnicity, baseline assessment centers, and pre-existing diseases/conditions. 613 participants tested positive at inpatient settings between March 16 and April 27, 2020, 154 of whom succumbed to COVID-19. PhenoAge was associated with increased risks of inpatient test positivity and COVID-19 related mortality  adjusting for demographics including age at the pandemic. Further adjustment for pre-existing disease s/conditions at baseline  and at the early pandemic  decreased the association. PhenoAge measured in 2006-2010 was associated with COVID-19 severity outcomes more than 10 years later. These associations were partly accounted for by prevalent chronic diseases proximate to COVID-19 infection. Overall, our results suggest that aging biomarkers, like PhenoAge may capture long-term vulnerability to diseases like COVID-19, even before the accumulation of age-related comorbid conditions.


Anticoagulation in hospitalized COVID-19 patients has been associated with survival benefit; however, the optimal anticoagulant strategy has not yet been defined. The objective of this meta-analysis was to investigate the effect of intermediate-to-therapeutic versus prophylactic anticoagulation for thromboprophylaxis on the primary outcome of in-hospital mortality and other patient-centered secondary outcomes in COVID-19 patients.MEDLINE, EMBASE, and Cochrane databases were searched from inception to August 10th 2021. Cohort studies and randomized clinical trials that assessed the efficacy and safety of intermediate-to-therapeutic versus prophylactic anticoagulation in hospitalized COVID-19 patients were included. Baseline characteristics and relevant data of each study were extracted in a pre-designed standardized data-collection form. The primary outcome was all-cause in-hospital mortality and the secondary outcomes were incidence of thrombotic events and incidence of any bleeding and major bleeding. Pooled analysis with random effects models yielded relative risk with 95 % CIs.2=77 %) and lower incidence of thrombotic events , but increased the risk of any bleeding events  and major bleeding events significantly  versus prophylactic anticoagulation. Moreover, intermediate-to-therapeutic anticoagulation decreased the incidence of thrombotic events  among critically ill COVID-19 patients admitted to intensive care units (ICU), with increased bleeding risk  and unchanged in-hospital mortality  in such patients. The Grading of Recommendation, Assessment, Development, and Evaluation certainty of evidence ranged from very low to moderate.This meta-analysis included 42 studies with 28,055 in-hospital COVID-19 patients totally. Our pooled analysis demonstrated that intermediate-to-therapeutic anticoagulation was not associated with lower in-hospital mortality .The protocol was registered at PROSPERO on August 17th 2021 (The online version contains supplementary material available at 10.1186/s12959-021-00343-1. Coronavirus disease 2019 (COVID-19), provoked by severe acute respiratory syndrome coronavirus-2 infection, is currently the most serious public health crisis worldwide . In a reHowever, increasing evidence demonstrated that some COVID-19 patients, especially those with critical illness, still developed severe thrombotic complications despite the use of prophylactic anticoagulant –18. EnhaPrevious meta-analyses that investigated the optimal anticoagulation strategy only included limited studies and reported inconsistent conclusions –25. As mThis review was conducted following the Preferred Reporting Items for Systemic Reviews and Meta-Analyses (PRISMA) Statement  and regiLiterature search was conducted strictly and comprehensively in MEDLINE, EMBASE, and Cochrane databases. Two independent investigators (ZSR and LYP) searched studies available from database inception to August 10th 2021 without language limitation. The following key words and/or medical subject heading terms were used: ‘COVID’, ‘COVID-19’, ‘2019 novel coronavirus infection’, ‘SARS-CoV-2’, ‘2019-nCoV disease’, ‘anticoagulation’, ‘anticoagulants’, ‘anticoagulant’, ‘unfractionated heparin’, ‘UFH’, ‘fondaparinux’, ‘enoxaparin’, ‘low-molecular-weight heparin’, ‘LMWH’, ‘heparin, low molecular weight’, ‘heparin’, ‘antithrombotic’, and ‘anti-thrombosis’, ‘thromboprophylaxis’ [see Additional file Two independent investigators (ZSR and LYP) performed the initial screening of titles and abstracts. Full-length articles of identified studies were retrieved to assess eligibility. Any discrepancies were adjudicated by a third reviewer (SBH). The inclusion criteria of our meta-analysis were as follows: (1) cohort studies and RCTs, (2) studies that enrolled COVID-19 patients admitted to general wards or ICUs, and (3) studies that compared the efficacy or safety of intermediate-to-therapeutic versus prophylactic anticoagulation. The exclusion criteria were as follows: (1) studies enrolling out-hospital COVID-19 patients, or (2) animal experiments, case reports, reviews, comments, editorial comments, or (3) no relevant and sufficient data on preferred outcomes. In case a same patient population was enrolled in a few articles, we only selected the most comprehensive study.After study selection, the following data were extracted: author, publication year, study type, study location, patient characteristics, numbers of patients in different groups, in-hospital mortality, incidence of any bleeding events, incidence of major bleeding events, and incidence of thrombotic events using a predesigned standardized data-collection form by two researchers (ZSR and LYP) independently.Quality assessment of studies were conducted by two independent reviewers (ZSR and LYP). The Cochrane Collaboration tool was used to assess the risk of bias in randomized controlled trials . The quaOur primary outcome was all-cause mortality during hospitalization for COVID-19. Secondary patient-centered outcomes included the incidence of bleeding events and thrombotic complications during hospitalization for COVID-19. We also investigated these outcomes in critically ill COVID-19 patients who were admitted to ICU exclusively. Definitions of bleeding events depended on the variable definitions of each study, mostly in accordance with Myocardial Infarction (TIMI) bleeding  or the W2 index . Sensitivity analysis was conducted by omitting each study at a time to determine whether each study affected the overall estimate and to identify studies that potentially drove the results. Publication bias was assessed with funnel plots and Egger test. A two-side P-value <0.05 was considered to be of statistical significance. All statistical analyses were carried out using the meta package in R 3.6.2 and Review Manager software (version 5.4).Risk ratios (RR) with 95 % confidence interval (CI) were calculated for dichotomous outcomes such as mortality, bleeding events and thrombotic complications. A random effects model was applied for meta-analyses due to the heterogeneity of study populations and design. Heterogeneity among studies was assessed by the chi-square test and shown as an IFrom 6,002 yielded studies, we retained 74 studies after removing duplications and screening titles and abstracts. Forty-two studies involving 28,055 subjects met the inclusion criteria and were finally included in this meta-analysis after full-text review. The PRISMA flow diagram demonstrates the process of study screening and selection in detail, as shown in Fig. Of the forty-two included studies, 6 studies –39 were We included a pooled population of 23,579 hospitalized COVID-19 patients , 70–75 wThe score by NOS for the included cohort studies ranged from 6 to 9, while 10 studies were of high quality scoring 9. A full assessment is shown in Additional file 2=77 %). The sensitivity analysis by leave-one-out approach indicated that the result was relatively stable because the risk ratio remained unchangeable though p value fluctuated around 0.05, as shown in Additional file Data regarding in-hospital mortality of COVID-19 patients receiving prophylactic anticoagulation or intermediate-to-therapeutic anticoagulation were available from 39 studies , 70–75 . The predefined sensitivity analyses did not change the overall effect and the results remained stable .A total of 27 studies , 71, 73 2=0 %, Fig. Subgroup analysis of bleeding risk related to thromboprophylaxis in ICU settings included 10 studies , 64, 70 2=88 %). The predefined sensitivity analysis conformed the stability of the results in this meta-analysis, as shown in Additional file Incidence of thrombotic events was reported in 17 studies , 64, 66 2=0 %), as shown in Fig. Subgroup analysis of studies exclusively enrolling critically ill COVID-19 patients admitted to ICU indicated that patients receiving intermediate-to-therapeutic anticoagulation were associated with reduced risk of thrombotic events compared with prophylactic anticoagulation (RR=0.71, 95 %CI 0.56-0.89, p =0.03, IIn this systemic review and meta-analysis, we included 42 studies with 28,055 COVID-19 patients admitted to hospital in total. By analyzing data of 23,579 hospitalized COVID -19 patients from 33 cohort studies and 6 RCTs reporting in-hospital mortality, intermediate-to-therapeutic anticoagulation did not improve the primary outcome of in-hospital mortality in unselected hospitalized COVID-19 patients when compared with prophylactic anticoagulation. Additionally, a subgroup analysis of critically ill COVID-19 patients admitted to ICU also showed no significant survival benefit of intermediate-to-therapeutic anticoagulation against standard thromboprophylaxis. These results were inconsistent with earlier studies which reported a significant association between enhanced-dose anticoagulation and improved outcomes , 65. It Additionally, meta-analysis of data from 11 cohort studies and 6 RCTs reporting thrombotic events indicated that intermediate-to-therapeutic anticoagulation did not reduce relative risk of thrombotic events either versus prophylactic anticoagulation in general COVID-19 patients. However, a subgroup analysis of critically ill patients admitted to ICU found intermediate-to-therapeutic anticoagulation significantly reduced the incidence of thrombotic events by 30 %. These results should be interpreted with caution considering that possible insufficient thrombosis screening in patients admitted to general wards might inevitably influence the reliability of this evidence .Recent studies have also reported high incidences of hemorrhage among inpatients with COVID-19. In this meta-analysis, we observed a significant increase of incidence of any bleeding events and major bleeding events in both general and critically ill patients receiving intermediate-to-therapeutic anticoagulation when compared with prophylactic anticoagulation. Especially, in critically ill COVID-19 patients, use of intermediate-to-therapeutic dose anticoagulants significantly reduced incidence of thrombotic events by 30 % but increased incidence of bleeding events by nearly 70 %, without a beneficial effect on patient survival. Therefore, the potential profit of elevated dose anticoagulation of thromboprophylaxis on thrombosis prevention are associated with an increase in bleeding events and should thus be weighed against the risk of bleeding.Growing evidences have arisen to determine the optimal anticoagulation strategy that weighs thrombotic events and consequent bleeding risk in hospitalized patients with COVID-19. In the updated guidelines on the use of anticoagulants for thromboprophylaxis in COVID-19 patients, the American Society of Hematology recommended prophylactic anticoagulation over intermediate anticoagulation in COVID-19 patients with no confirmed or suspected thromboembolism based on low certainty of the evidence . The latCOVID-19 patients may exhibit complex coagulopathy states. Thrombotic and bleeding events are staggered in time and hypercoagulable states are not always consistent in the course of disease. A recent literature review by Tacquard, et al. demonstrated that thrombotic events occurred primarily in the first ten days after admission while bleeding events occurred most often late . Hardy eImportantly, the choices of different anticoagulation doses mainly follow local institutional protocols and were decided by the physicians in most studies, which resulted in the inconsistent anticoagulation dosage within the same group leading to potential bias . For insThis study has several limitations. Firstly, we included 36 cohort studies and only 6 RCTs. In those cohort studies, selection bias exists. Patients with higher disease severity and risk of thrombotic events were more likely to be treated by intermediate-to-therapeutic anticoagulation, which might lead to undervaluation of potential benefit. Secondly, most patients enrolled in the included studies were over 60 years old and it will be more reasonable to include patients with a wider age scope considering mortality is significantly higher in the elderly inpatients with COVID-19. Thirdly, the criteria for admission to hospital, definition of in-hospital mortality, bleeding and thrombotic events, follow-up periods and exact dosage of two groups might vary from region to region during different stages of the pandemic, affecting risk of both thrombotic and fatal outcomes. Results of the current meta-analysis should therefore be interpreted with caution considering these limitations.Among unselected in-hospital patients with COVID-19, the current meta-analysis showed that intermediate-to-therapeutic anticoagulation was not associated with lower in-hospital mortality and incidence of thrombotic events, but increased the risk of bleeding events versus prophylactic anticoagulation. In contrast, intermediate-to-therapeutic anticoagulation reduced the risk of thrombotic events with increased incidence of bleeding events in critically COVID-19 patients admitted to ICU. We recommended the use of prophylactic anticoagulation against empirical intermediate-to-therapeutic anticoagulation among unselected hospitalized COVID-19 patients based on low certainty in the evidence. For those with severer illness, the benefits to reduce thrombotic events of intermediate-to-therapeutic anticoagulation should also be weighed cautiously by clinicians because of the significantly increased risk of bleeding associated with higher dose of anticoagulant use. Furthermore, the optimal timing of anticoagulation initiation and treatment duration remains unexplored in this field. More well-designed RCTs that effectively stratify included patients by disease severity, timing of anticoagulation initiation, and treatment duration are highly encouraged to provide high-quality evidence regarding the safety and efficacy of higher dose anticoagulation in different clinical settings.Additional file 1. Detailed search strategy.Additional file 2. Baseline characteristics of each study.Additional file 3. Details of anticoagulation administration in each study.Additional file 4. Quality assessment of cohort studies by NOS score.Additional file 5. Quality assessment of RCTs by Cochrane Collaboration tool.Additional file 6. Sensitivity analysis of in-hospital mortality outcome.Additional file 7. Funnel plot for the assessment risk of publication bias of in-hospital mortality outcome.Additional file 8. The quality of evidence for in-hospital mortality assessed by GRADE framework.Additional file 9. Subgroup analysis of in-hospital mortality stratified by study regions.Additional file 10. Sensitivity analysis of bleeding events outcome.Additional file 11. Funnel plot for the assessment risk of publication bias of bleeding events outcome.Additional file 12. The quality of evidence for bleeding events assessed by GRADE framework.Additional file 13. Sensitivity analysis of thrombotic complication events outcome.Additional file 14. Funnel plot for the assessment risk of publication bias of thrombotic complication events outcome.Additional file 15. The quality of evidence for thrombotic complication events assessed by GRADE framework.


This interest is closely linked to the MSC-marked tropism for tumors, suggesting them as a rational and effective vehicle for drug delivery for both hematological and solid malignancies. Nonetheless, the therapeutic application of the MSCs in human tumors is still controversial because of the induction of several signaling pathways largely contributing to tumor progression and metastasis. In spite of some evidence supporting that MSCs may sustain cancer pathogenesis, increasing proofs have indicated the suppressive influences of MSCs on tumor cells. During the last years, a myriad of preclinical and some clinical studies have been carried out or are ongoing to address the safety and efficacy of the MSC-based delivery of therapeutic agents in diverse types of malignancies. A large number of studies have focused on the MSC application as delivery vehicles for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), chemotherapeutic drug such as gemcitabine (GCB), paclitaxel (PTX), and doxorubicin (DOX), prodrugs such as 5-fluorocytosine (5-FC) and ganciclovir (GCV), and immune cell-activating cytokines along with oncolytic virus. In the current review, we evaluate the latest findings rendering the potential of MSCs to be employed as potent gene/drug delivery vehicle for inducing tumor regression with a special focus on the  In this section, we have reviewed the recent reports respecting MSC-based cytokine delivery for cancer therapy .in vivo could migrate into tumor area in 9-L glioma-bearing murine models through the corpus callosum and finally infiltrate into the tumor bed  have represented more effective antitumor effects. Studies in breast tumor xenografts revealed that intravenously injected MSC could migrate to the breast tumor area and secrete high rates of IFN-β into tumor stroma . MoleculVirotherapy as an innovative strategy for cancer treatment has some advantages such as the potential absence of cross-resistance with conventional therapies and also the capacity to induce tumor eradication by several mechanisms. Regardless of their unique mechanisms of functions distinguishing them from other treatment options, the self-perpetuating nature of oncolytic virus (OVs) serves as a supreme strategy for therapeutic transgenic insertion. The use of MSCs as a non-systemic carrier of OVs has been investigated with remarkable achievement for the treatment of varied types of pathological conditions . In thisin vitro  using the semipermissive cotton rat (CR) model verified that CR-MSCs could sustain the replication of oncolytic adenovirus (oAd) in vitro . Furthereactions . Furthereactions . In addiprofiles . Similarprofiles .in vitro, MSC-oAd demonstrated tumor tropism in prostate cancer mice models concomitant with suppression of tumor growth in experimental models. This result arose from continued viral replication which enabled cell lysis  in a SCID mouse xenograft metastases model of breast tumors resulted in effective recruitment of injected cells into tumor area in experimental models and consequently improved mouse survival in comparison with mice treated with CRAds alone apparently by viral amplification in the MSCs . On the ll lysis .In 2010, evaluation of the safety and the efficacy of injection of autologous MSCs transduced with ICOVIR-5, a new oAd, in four children with metastatic neuroblastoma demonstrated that the injection was well tolerated; on the other hand, a complete clinical response was verified in one participant, describing MSCs as eligible vehicles for oncolytic virus delivery into metastatic tumors with very low systemic toxicity concomitant with reliable antitumor effects .in vitro and in vivo presumably by a dynamic procedure of oHSV infection and tumor eradication (It has been found that MSCs loaded with oncolytic herpes simplex virus (oHSV) (MSC-oHSV) could potently generate oHSV progeny leading to the lysis of glioblastoma (GBM) cells dication . Moreovedication . Furtherdication . In addidication . Observadication .in vitro and pancreatic tumor-bearing murine models demonstrated that combination therapy led to the highest cytotoxicity toward human pancreatic cell line BxPC-3, while combined use of MSC-HF10 and erlotinib had no significant cytotoxicity against human pancreatic cell line PANC-1 cells  (MSC-oMV) can lead to the desired therapeutic outcomes in a variety of human tumors  . In thisThe partial resistance of MSC to cytostatic and cytotoxic chemotherapeutic agents along with their suitable tropism to tumors offers a novel platform for direct targeted delivery of therapeutic agents, such as paclitaxel (PTX), gemcitabine (GCB), and doxorubicin (DOX) to tumor tissue . Regardlin vitro -compliant MSCs loaded with the chemotherapeutic drug paclitaxel (PTX) (MSC-PTX). In addition to allowing large-scale cultivation of the MSC-PTX, these modified cells show common characteristics of MSCs concerning viability, adhesion capacity, and also phenotype . Moreovein vitro . In vivoed genes . Further results . Besides results . Further results . Moreove results . Corresp results . Similarby DPSCs . The anaby DPSCs .in vitro  (PLGA) nanoparticles (NPs) in glioma xenograft rat models clarified superiority of the PTX-PLGA NP-primed MSCs over PTX-primed MSCs in terms of secretion of PTX in the form of free PTX as well as PTX-NPs, supporting glioma cells apoptosis in vitro . Moreovein vitro .in vitro. Particularly, GCB-primed GinPa-MSCs offer a reliable cell-mediated drug delivery policy for upcoming possible application in oral oncology , BM-MSC-DOX elicited remarkable cytotoxicity against tumor cells -primed human BM-MSCs (BM-MSC-DOX) could induce antitumor effects against breast cancer cells (MDA-MB-231) and anaplastic thyroid cancer cells (CAL62) . In vivoor cells .in vitro and in vivo. Compared with DOX-SPIO, MSC membrane-camouflaged nanodrug (DOX-SPIO@MSCs) showed greater cellular uptake efficiency, augmented antitumor influences, and mitigated the immune system reactions in vitro. In addition, DOX-SPIO@MSCs exerted more powerful antitumor effects, while dwindled systemic side effects in MC38 tumor-bearing C57BL/6 mice  or cytosine deaminase (CD), converting inactive intravenously injected prodrugs, including ganciclovir (GCV) and 5-fluorocytosine (5-FC), into active cytotoxic agents, which in turn, can decrease systemic toxicity by easing tumor-localized chemotherapeutic activity . Altogetin vitro and a mouse orthotopic GBM model showed that TMZ and MSC-CD with 5-FC synergistically could hinder U87 glioma cell proliferation concurrently triggering cell cycle arrest and DNA breakage (The 5-FC has been presented as a prodrug combined with the CD suicide gene. MSC-expressing CD (MSC-CD) have been formerly confirmed to suppress the development of subcutaneous prostate cancer xenografts upon converting the non-toxic 5-FC into the antineoplastic 5-fluorouracil (5-FU) . Moreovebreakage . On the breakage . Also, ibreakage . Nonethebreakage . Anotherbreakage . Furtherbreakage .Combination therapy with AT-MSCs expressing CD and lysomustine chemotherapy, a nitrosourea derivative of lysine, followed by systemic injection of 5-FC in mice bearing late-stage Lewis lung carcinoma (LLC) led to the significant attenuation of tumor growth, as evidence by decreased tumor volume . Moreovein vitro and also in U87-bearing murine models  could phosphorylate GCV to its toxic metabolites and consequently induce tumor regression in human tumors . In thise models .in vitro, in particular, on 8-MG-BA cells - and red fluorescent protein (RFP)-reporter genes, in C57/Bl6 models , is a member of TNF cytokine superfamily with attractive potent anticancer function owing to its unique competence to target cancerous cells without any harm to adjacent normal cells. This is because the expression of TRAIL-specific receptors, termed as death receptors (DR), are significantly higher in cancer cells compared with normal cells .89Zr-oxine labeling and positron emission tomography (PET)/computed tomography (CT) imaging for tracking MSC-TRAIL biodistribution in metastatic lung adenocarcinoma xenograft models evidenced the effective delivery of MSC-TRAIL to the lungs in a murine model up to 1-week postinjection, verified through in vivo bioluminescence imaging, autoradiography, and fluorescence imaging on tissue sections  or as a modified shape generally termed as soluble TRAIL (sTRAIL)  Table 5Table 5. sections . MSC-sTRsections . Accordisections . Converssections . Besidessections . On the c effect .Tumor necrosis factor-related apoptosis-inducing ligand delivery by MSC-derived extracellular vesicles (EVs) is considered an effective antitumor treatment option. In this regard, EVs derived from MSC-TRAIL showed cytotoxicity against lung cancer lines , malignant pleural mesothelioma lines , renal cancer lines (RCC10 and HA7-RCC), human breast adenocarcinoma line , and neuroblastoma line (SHEP-TET) without any cytotoxicity against primary human bronchial epithelial cells . Importain vitro largely by activation of the apoptosis intrinsic pathway in CSCs  concomitant with enrichments of the CSC niche . MSC-secAll authors contributed to the conception and the main idea of the work and read and approved the final version of the work to be published. AH, AA, HR, MS, DB, WA, FM, MZ, YY, and MY drafted the main text, figures, and tables. MJ supervised the work and provided the comments and additional scientific information. MC and YP also reviewed and revised the text.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The COVID-19 pandemic obligated academic medical programs to substantially alter the traditional Internal Medicine (IM) rounding model to decrease risk of inpatient nosocomial viral transmission. Our study aimed to describe how IM rounding practices changed during the COVID-19 pandemic and to understand the impacts of these changes on medical education. We conducted a two-phase, mixed-methods study of inpatient IM rounding team practices at a large academic hospital in Houston, TX. In the first phase (January-February 2021), we organized and audio-recorded 4 virtual (Zoom) focus groups. Each included 5-6 rounding team members, divided by: attendings; senior residents; interns; and medical and physician assistant students. In the second phase (March-May 2021), we performed 6 direct observations of IM teams during rounds. Two observers systematically recorded variables such as time spent on non-bedside versus bedside rounds, number of each team member type entering patient rooms for bedside teaching, and types of personal protective equipment (PPE) worn. Topics discussed during focus groups included comparisons of rounding team size, rounding duration, physical distancing and PPE use, bedside education, communication methods, and patient safety before and after March 2020. Perceptions of changes in each topic were generally consistent across groups (Table 1). Direct observation data showed that team rounding styles remained diverse in the proportion of rounding time spent in an office versus on the wards, and in the number and types of team members entering patient rooms. IM team members uniformly wore respiratory PPE when entering all patient rooms; use of eye protection varied. Teams spent more total time discussing patients with or suspected to have COVID-19 compared to patients without COVID-19 .Our results suggest that the COVID-19 pandemic adversely impacted bedside medical education, even into Spring of 2021. Conclusions from this study can be used to 1) address educational gaps related to COVID-19 pandemic-associated rounding changes and 2) create innovative methods of providing high-quality clinical education that will be minimally impacted by future respiratory virus pandemics.Prathit A. Kulkarni, M.D., Vessel Health, Inc. (Grant/Research Support)


Current state-of-the-art quantum dot light-emitting diodes have reached close to unity internal quantum efficiency. Further improvement in external quantum efficiency requires more efficient photon out-coupling. Improving the directivity of the photon emission is considered to be the most feasible approach. Here, we report improved emission directivity from colloidal quantum dot films. By growing an asymmetric compressive shell, we are able to lift their band-edge state degeneracy, which leads to an overwhelming population of exciton with in-plane dipole moment, as desired for high-efficiency photon out-coupling. The in-plane dipole proportion determined by back-focal plane imaging method is 88%, remarkably higher than 70% obtained from conventional hydrostatically strained colloidal quantum dots. Enhanced emission directivity obtained here opens a path to increasing the external quantum efficiencies notably. Asymmetric strain lifts the dipole degeneracy and enables directional photon emission in colloidal quantum dots. Colloidal quantum dots (CQDs) are appealing active emitting layer candidate for light-emitting diodes (LEDs) thanks to their narrow emission linewidth and wide wavelength tenability .Here, we report that a pronounced dipole orientation can be achieved in dedicatedly strained CdSe-CdS core-shell CQDs that have similar aspect ratio to CQDs that performed well in LED devices. The cores of the CQDs are strained perpendicular to the Excitons with dipoles parallel to the CQD film plane, namely, in-plane dipoles, are desired for light out-coupling because they emit photons perpendicular to the film substrate, facilitating their escape into air. Those with dipoles perpendicular to the film plane  emit photon parallel to the film plane, making light extraction difficult , because the strain effect along this direction is superimposed with the effect of crystal field. Our previous work demonstrated that this kind of strain can be achieved in drum-like wurtzite CdSe/CdS nanocrystals  computations including spin-orbit coupling for 3-nm wurtzite CdSe nanoparticles, hydrostatically and biaxially strained . Calculaxy versus pz) but remain nearly degenerate in the hydrostatic strain case  patterns of the spin-coated thin film on nondiffraction silicon wafer and ground powder -co-4,4-(N-(4-s-butylphenyl)diphenyl-amine)] (TFB) (fig. S13), which is one of the mostly used hole transport layers for high-performance QLED  images of two types of QDs. These two types of QDs represent a different cross-sectional shape while observing from the [1¯0] axis . While tAfter confirming the CQD orientation in thin films, we sought to test the transition dipole moment orientation of band-edge excitons by measuring the absorption/emission polarization direction. Since the band-edge absorbance of thin-film samples is low, a polarization-dependent PL excitation (PLE) measurement was adopted , of which the experimental principle is shown in To further quantify the polarization and the directivity of emission, CQD films are further characterized by a back-focal plane (BFP) imaging method , which iD = Pmax/Pmean  facets allow us to spin-coat films with CQDs uniformly orientated. DFT computations have explicated the origin of biaxial strain–induced directivity. The bond lengths along different crystalline axes have been changed due to biaxial strain, affecting the overlap of atomic orbitals from neighboring atoms and thus the energy of orbitals, lifting the band-edge degeneracy, and setting the in-plane polarized exciton as the ground state. These findings demonstrate a deliberate control over the band-edge states in II-VI CQDs, paving the way for ultrahigh EQE LED devices.The structures were first optimized using Perdew–Burke–Ernzerhof , sulfur powder , selenium powder , oleylamine (>98% primary amine), octadecene (90%), oleic acid (90%), trioctylphosphine (90%), trioctylphosphine oxide (99%) and octadecylphosphonic acid (97%), and 1-octanethiol (>98.5%) were purchased from Sigma-Aldrich and used without further purification; hexane  and acetone (99.5%) were purchased from Greagent. The biaxially strained CQDs were synthesized using a modified protocol of our previous work  and imaged using a JEOL JEM 2100F HRTEM operating at 200 kV. The x-ray diffraction data refinement is based on the Rietveld method  was used to collect real-space and k-space emission patterns of the CQD films. To image k-space (BFP), two coupled 200-mm focal plano-convex lenses were placed on the back optical path of the objective lens and projected the BFP image of the objective lens onto a sCMOS  camera (TUCSEN Dhyana 400BSI V2). A polarizer (Thorlabs CCM1-PBS251) was used to generate the polarization image of the BFP, and a band-pass filter (Semrock FF02-641/75-25) was used to filter out the excitation and noise light. See text S2 for the BFP image fitting details.The CQD submonolayer films were made by spin-coating the CQD in The optical efficiency of LED device is calculated by the transfer matrix method (


COVID-19 vaccination remains the most effective means to achieve control of the pandemic.In the United States, COVID-19 cases and deaths have markedly declined since their peakin early January 2021, due in part to increased vaccination coverage (The principal mode by which persons are infected with SARS-CoV-2 is through exposure torespiratory fluids carrying infectious virus.A person’s risk for SARS-CoV-2 infection is directly related to the risk forexposure to infectious persons, which is largely determined by the extent ofSARS-CoV-2 circulation in the surrounding community.Data on usage of clinical care resources to manage patients with COVID-19 reflectunderlying community disease incidence and can signal when urgent implementation oflayered prevention strategies might be necessary to prevent overloading the healthcare system. Strains on critical care capacity can increase COVID-19 mortality(Monitoring vaccination coverage in communities and organizations is recommended byCDC to gauge progress, focus vaccination efforts on populations whose coverage islow, and inform the need for additional prevention strategies. As of July 23, 2021,the proportion of the total U.S. population who is fully vaccinated is 48.9%.Certain populations are at high risk for exposure to, and thereby infection with,SARS-CoV-2. Such populations are especially well-suited for sentinel monitoringefforts to detect the early introduction and spread of COVID-19, particularly inareas with low vaccination coverage or where layered prevention strategies are notin use. CDC considers the capacity to monitor COVID-19 incidence in the followingpopulations particularly useful due to their high risk of exposure or severeillness: students and staff members of kindergarten–grade 12 schools andinstitutions of higher education, health care workers, residents and staff membersof long-term care facilities, incarcerated persons, homeless persons, and workers inhigh-density work sites (Serial screening testing is an effective method to monitor for the early introductionand spread of COVID-19 (CDC recommends additional prevention strategies to safeguard populations at highestrisk for severe outcomes from COVID-19, particularly in the context of the highlytransmissible Delta variant. Unvaccinated persons remain at risk for infection,severe illness, and death. Advanced age, pregnancy, and an increasingly well-definedset of underlying medical conditions increase the risk for serious outcomes fromCOVID-19 among unvaccinated persons.The most important public health action to end the pandemic remains increasingvaccination coverage, which saves lives, prevents illness, and reduces the spread ofCOVID-19. Effective COVID-19 prevention strategies are well documented and can helpreduce community transmission until high vaccination coverage is achieved (The widespread availability and administration of COVID-19 vaccines has changed thetrajectory of the pandemic in the United States and significantly reducedhospitalization and mortality among vaccinated persons (COVID-19 vaccines authorized in the United States are effective againstsevere illness and death from SARS-CoV-2 infection; however, current U.S.coverage is uneven. Implementation of layered prevention strategies reducesSARS-CoV-2 transmission.Given the spread of the highly transmissible Delta variant, localdecision-makers should assess the following factors to inform the need forlayered prevention strategies across a range of settings: level ofSARS-CoV-2 community transmission, health system capacity, vaccinationcoverage, capacity for early detection of increases in COVID-19 cases, andpopulations at risk for severe outcomes from COVID-19.Although increasing COVID-19 vaccination coverage remains the most effectivemeans to achieve control of the pandemic, additional layered preventionstrategies will be needed in the short-term to minimize preventablemorbidity and mortality.