Porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), imposes substantial economic losses on the global swine industry due to its profound impact on pig health and production efficiency. The main clinical manifestations are reproductive disorders such as miscarriage, stillbirth or weak foetuses in sows, as well as severe respiratory diseases in pigs of all ages. Currently, due to the high genomic diversity, antigen heterogeneity, and antibody-dependent enhancement effects of PRRSV, vaccination strategies cannot effectively control PRRSV infection. It is also difficult to control these viral infectious diseases with antiviral drugs. In this study, we demonstrate that neferine (Nef), a natural compound derived from lotus seeds, inhibits NF-κB activation during PRRSV infection. Consequently, Nef effectively suppresses PRRSV gene expression and virion replication in the PRRSV-infected Marc-145 cell line. Further studies demonstrated that Nef attenuates the nuclear translocation of p65 triggered by PRRSV infection, consequently mitigating the excessive production of inflammatory cytokines in infected cells. This anti-inflammatory mechanism may contribute to its observed antiviral efficacy against PRRSV. As a result, we conclude that Nef may be an effective clinical treatment for the acute occurrence and transmission of PRRSV infections. Furthermore, PRRSV is thought to be responsible for an increase in proinflammatory cytokines.

mRNA therapies have emerged as a transformative class of medicines, offering immense potential across a diverse array of applications. This progress has been particularly evident in the wake of the success of lipid nanoparticle (LNP)-based mRNA vaccines during the COVID-19 pandemic. As these applications expand, the demand for sustained protein production has become increasingly critical. However, conventional mRNA therapies face significant challenges, including inherent RNA instability and suboptimal expression efficiency, often requiring repeated dosing to maintain therapeutic efficacy over time. This review highlights recent advances in strategies to prolong the therapeutic efficacy of LNP-mRNA systems. We focus on preclinical and emerging approaches aimed at extending the period of protein translation by engineering both the mRNA molecule and the LNP delivery system. Sustained protein expression is a cornerstone of mRNA-based therapeutics, and addressing this challenge is vital for unlocking their therapeutic potential. We hope this review provides valuable insights to guide the development of optimized delivery platforms for LNP-mRNA therapeutics.

SARS-CoV-2, the third major coronavirus of the 21st century, causing COVID-19 disease, profoundly impacts public health and workforces worldwide. Identifying individuals at heightened risk of SARS-CoV-2 infection is crucial for targeted interventions and preparedness. This study investigated 35 SNVs within viral infection-associated genes in SARS-CoV-2 patients and uninfected controls from the Basque Country (March 2020-July 2021). Its primary aim was to uncover genetic markers indicative of SARS-CoV-2 susceptibility and explore genetic predispositions to infection. Association analyses revealed previously unreported associations between SNVs and susceptibility. Haplotype analyses uncovered novel links between haplotypes and susceptibility, surpassing individual SNV associations. Descriptive modelling identified key susceptibility factors, with rs11246068-CC (IFITM3), rs5742933-GG (ORMDL1), rs35337543-CG (IFIH1), and GGGCT (rs2070788, rs2298659, rs17854725, rs12329760, rs3787950) variation in TMPRSS2 emerging as main infection-susceptibility indicators for a COVID-19 pandemic situation. These findings underscore the importance of integrated SNV and haplotype analyses in delineating susceptibility to SARS-CoV-2 and informing proactive prevention strategies. The genetic markers profiled in this study offer valuable insights for future pandemic preparedness.

Neuroinflammation is a pivotal factor in the progression of both age-related and acute neurodegenerative disorders, including Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, multiple sclerosis, and stroke. Mitochondria, essential for neuronal health due to their roles in energy production, calcium buffering, and oxidative stress regulation, become increasingly susceptible to dysfunction under conditions of metabolic stress, aging, or injury. Impaired mitophagy in aged or injured neurons leads to the accumulation of dysfunctional mitochondria, which release mitochondrial-derived damage-associated molecular patterns (mtDAMPs). These mtDAMPs act as immune checkpoints, activating pattern recognition receptors (PRRs) and triggering innate immune signaling pathways. This activation initiates inflammatory responses in neurons and brain-resident immune cells, releasing cytokines and chemokines that damage adjacent healthy neurons and recruit peripheral immune cells, further amplifying neuroinflammation and neurodegeneration. Long-term mitochondrial dysfunction perpetuates a chronic inflammatory state, exacerbating neuronal injury and contributing additional immunogenic components to the extracellular environment. Emerging evidence highlights the critical role of mtDAMPs in initiating and sustaining neuroinflammation, with circulating levels of these molecules potentially serving as biomarkers for disease progression. This review explores the mechanisms of mtDAMP release due to mitochondrial dysfunction, their interaction with PRRs, and the subsequent activation of inflammatory pathways. We also discuss the role of mtDAMP-triggered innate immune responses in exacerbating both acute and chronic neuroinflammation and neurodegeneration. Targeting dysfunctional mitochondria and mtDAMPs with pharmacological agents presents a promising strategy for mitigating the initiation and progression of neuropathological conditions.

Type I and III interferons critically protect the host against viral infection. Previous studies showed that IFN responses are suppressed in cells infected by hepatitis E virus (HEV). Here, we studied the anti-HEV function of IFN secreted by plasmacytoid dendritic cells (pDCs), specialized producers of IFNs. We showed that pDCs co-cultured with HEV-replicating cells secreted IFN in a cell contact-dependent manner. This pDC response required the endosomal nucleic acid sensor TLR7 and adhesion molecules. IFNs secreted by pDCs reduced viral spread. Intriguingly, ORF2, the capsid protein of HEV, can be produced in various forms by the infected cells, and we wanted to study their role in anti-HEV immune response. During infection, a fraction of ORF2 localizes into the nucleus, and glycosylated forms of ORF2 are massively secreted by infected cells. We showed that glycosylated ORF2 potentiates the recognition of infected cells by pDCs, by regulating cell contacts. On the other hand, nuclear ORF2 triggers immune response by IRF3 activation. Together, our results suggest that pDCs may be essential to control HEV replication.

The innate immune signaling network follows a canonical format for signal transmission. The innate immune pathway is crucial for defense against pathogens, yet its mechanistic crosstalk with aging processes remains largely unexplored. Retinoic acid-inducible gene-I (RIG-I), a key mediator of antiviral immunity within this pathway, has an enigmatic role in stem cell senescence. Our study reveals that RIG-I levels increase in human genetic and physiological cellular aging models, and its accumulation drives cellular senescence. Conversely, CRISPR/Cas9-mediated RIG-I deletion or pharmacological inhibition in human mesenchymal stem cells (hMSCs) confers resistance to senescence. Mechanistically, RIG-I binds to endogenous mRNAs, with CDKN1A mRNA being a prominent target. Specifically, RIG-I stabilizes CDKN1A mRNA, resulting in elevated CDKN1A transcript levels and increased p21Cip1 protein expression, which precipitates senescence. Collectively, our findings establish RIG-I as a post-transcriptional regulator of senescence and suggest potential targets for the mitigation of aging-related diseases.

Traumatic brain injury (TBI) is a major global health concern and a leading cause of morbidity and mortality. Neuroinflammation is a pivotal driver of both the acute and chronic phases of TBI, with pattern recognition receptors (PRRs) playing a central role in detecting damage-associated molecular patterns (DAMPs) and initiating immune responses. Key PRR subclasses, including Toll-like receptors (TLRs), NOD-like receptors (NLRs), and cGAS-like receptors (cGLRs), are abundantly expressed in central nervous system (CNS) cells and infiltrating immune cells, where they mediate immune activation, amplify neuroinflammatory cascades, and exacerbate secondary injury mechanisms. This review provides a comprehensive analysis of these PRR subclasses, detailing their distinct structural characteristics, expression patterns, and roles in post-TBI immune responses. We critically examine the molecular mechanisms underlying PRR-mediated signaling and explore their contributions to neuroinflammatory pathways and secondary injury processes. Additionally, preclinical and clinical evidence supporting the therapeutic potential of targeting PRRs to mitigate neuroinflammation and improve neurological outcomes is discussed. By integrating recent advancements, this review offers an in-depth understanding of the role of PRRs in TBI pathobiology and underscores the potential of PRR-targeted therapies in mitigating TBI-associated neurological deficits.

Radiation pneumonitis (RP) is a common and severe complication of radiotherapy, whose pathogenesis involves complex inflammatory responses and cellular damage. Despite its clinical significance, effective treatments remain limited. This study investigates the role of radiation-induced PINK1/PRKN-mediated mitophagy and type I interferon responses in RP and evaluates the therapeutic potential of Urolithin A (UA) in regulating inflammation through mitophagy activation.

We established RP mouse models (20 Gy thoracic irradiation) and radiation-induced BEAS-2B cell models (6 Gy). We systematically investigated mitochondrial damage, mtRNA release, RIG-I/MDA5-MAVS pathway activation, and PINK1/PRKN-mediated mitophagy changes. Moreover, the effects of UA and the mitophagy inhibitor Mdivi-1 on inflammation and lung injury were analyzed.

Radiation significantly caused mitochondrial damage in lung tissues, inducing mtRNA release and RIG-I/MDA5-MAVS-mediated type I interferon response. PINK1/PRKN-mediated mitophagy was significantly enhanced, clearing damaged mitochondria and reducing cytosolic mtRNA release, thereby suppressing inflammation. Pharmacological activation of mitophagy with UA markedly improved lung pathology, reduced inflammatory cytokine levels, and inhibited excessive activation of the RIG-I/MDA5-MAVS pathway. Conversely, the knockdown of PINK1 or PRKN weakened the protective effects of UA. Both in vitro and in vivo, UA reduced radiation-induced inflammation and improved lung tissue structure and function through mitophagy.

Radiation-induced mtRNA release activates the RIG-I/MDA5-MAVS-mediated type I interferon response, driving inflammation in RP. PINK1/PRKN-mediated mitophagy significantly alleviates inflammation by reducing cytosolic mtRNA release. As a mitophagy inducer, UA demonstrates therapeutic potential for RP, providing a new direction for the development of anti-inflammatory strategies.

The immune response to SARS-CoV-2 has been implicated in the onset of multiple, seemingly unrelated, autoimmune diseases. The immune response to SARS-CoV-2 has also been implicated in the unmasking and/or production of multiple autoantibodies, even in the absence of clinical disease. Despite such data, it remains unclear whether antibodies targeting antiviral signaling proteins and mitochondrial antigens reflect bystander activation or alternatively contribute to de novo viral immune escape mechanisms. With these comments in mind, a variety of professional antibody presenting cells and including lung resident macrophages of COVID-19 infected patients are impacted and dependent on the uptake of antibody-opsonized virus by Fcγ receptors; yet infection is aborted via antibody-dependent effector mechanisms or pyroptosis, possibly leading to autoantibody production, and autoinflammatory manifestations, respectively. TRIM21/Ro52, a cytosolic E3-ubiquitin ligase with an Fc-gamma receptor domain, functions as an intracytoplasmic antibody receptor, directs immune complexes binding virions but also autoantigens to autophagy. During autophagy, Ig-virions-TRIM21/Ro52-autoantigens complexes bind directly to class II human leukocyte antigen in lysosomal compartment, leading to subsequent presentation on the cell surface. This process favors the development of a specific humoral immune response but has the potential to lead to loss of tolerance. Interestingly, TRIM21/Ro52 can also contribute to pyroptosis. We propose that TRIM21/Ro52 is well-placed at the crossroad between the inflammatory response and clinical autoimmunity.

Cutaneous Lupus is an inflammatory skin disease causing highly morbid inflamed skin and hair loss. In order to investigate the pathophysiology of cutaneous lupus, we performed single-cell RNA and spatial sequencing of lesional and non-lesional cutaneous lupus skin compared to healthy controls. Pathway enrichment analyses of lesional keratinocytes revealed elevated responses to type I interferon, type II interferon, tumor necrosis factor, and apoptotic signaling. Detailed clustering demonstrated unique fibroblasts specific to lupus skin with likely roles in inflammatory cell recruitment and fibrosis. We also evaluated the association of retroelement expression with type I interferons in the skin. We observed increased retroelement expression which correlated with interferon-stimulated genes across multiple cell types. Moreover, we saw elevated expression of genes involved in RIG-I and cGAS-STING pathways, which transduce elevated nucleic acid signals. Treatment of active cutaneous lupus with Anifrolumab reduced RIG-I and cGAS-STING pathways in addition to the most abundant retroelement family, L2b. Our studies better define type I interferon-mediated immunopathology in cutaneous lupus and identify an association between retroelement expression and interferon signatures in cutaneous lupus.

Pseudouridine synthases (PUSs) catalyze the isomerization of uridine (U)-to-pseudouridine (Ψ) and have emerging roles in development and disease. How PUSs adapt gene expression under stress remains mostly unexplored. We identify an unconventional role for the Ψ "writer" PUS10 impacting intracellular innate immunity. Using Pus10 knockout mice, we uncover cell-intrinsic upregulation of interferon (IFN) signaling, conferring resistance to inflammation in vivo. Pus10 loss alters tRNA-derived small RNAs (tdRs) abundance, perturbing translation and endogenous retroelements expression. These alterations promote proinflammatory RNA-DNA hybrids accumulation, potentially activating cyclic GMP-AMP synthase (cGAS)-stimulator of interferon gene (STING). Supplementation with selected tdR pools partly rescues these effects through interactions with RNA processing factors that modulate immune responses, revealing a regulatory circuit that counteracts cell-intrinsic inflammation. By extension, we define a PUS10-specific molecular fingerprint linking its dysregulation to human autoimmune disorders, including inflammatory bowel diseases. Collectively, these findings establish PUS10 as a viral mimicry modulator, with broad implications for innate immune homeostasis and autoimmunity.

Activation of the type I interferon (IFN-I) signaling pathway is crucial for protecting host cells against viral infections. IFN-I production requires the transcription factors IFN regulatory factor 3 (IRF3) and IRF7, and its regulation must be finely tuned to both combat infection effectively and prevent excessive immunopathology. Here, we report that selective autophagy mediated by zebrafish FK506-binding protein 5 (Fkbp5), a PPIase (peptidyl-prolyl isomerase) promotes the degradation of Irf7 and Irf3, thereby inhibiting virus-induced type I IFN production. Quantitative real-time reverse-transcription polymerase chain reaction experiments indicate that zebrafish fkbp5 is induced by viral infection. Moreover, disrupting fkbp5 in AB-line zebrafish using CRISPR/Cas9 enhances survival rates and reduces viral messenger RNA levels compared with wild-type zebrafish. In cell culture, using promoter analysis and quantitative real-time reverse-transcription polymerase chain reaction, we found fkbp5 overexpression significantly attenuates cellular antiviral capacity and facilitates viral proliferation. Mechanistically, we found that fkbp5 inhibits Irf3/7-induced IFN activation, which depends on the binding of Fkbp5 to the Irf3 or IRF association domain of Irf7 via co-immunoprecipitation and Western blot assays. Furthermore, Fkbp5 induces the autophagic degradation of Irf3 and Irf7 independent of its PPIase activity. Blocking autophagy in vivo and in vitro restores the regulation of the RLR (RIG-I-like receptor) pathway by fkbp5. These findings reveal a critical role for zebrafish fkbp5 in suppressing the activation of Irf7 and Irf3 for IFN signaling and antiviral immune responses.

Viruses modulate various aspects of host physiology, including carbon metabolism, redox balance, and mitochondrial bioenergetics to acquire the building blocks for replication and regulation of the immune response. Understanding how SARS-CoV-2 alters the host metabolism may lead to treatments for COVID-19. We report that a ubiquitous gaseous molecule, hydrogen sulfide (H2S), regulates redox, metabolism, and mitochondrial bioenergetics to control SARS-CoV-2. Virus replication is associated with down-regulation of the H2S-producing enzymes cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CTH), and 3-mercaptopyruvate sulfurtransferase (3-MST) in multiple cell lines and nasopharyngeal swabs of symptomatic COVID-19 patients. Consequently, SARS-CoV-2-infected cells showed diminished endogenous H2S levels and a protein modification (S-sulfhydration) caused by H2S. Genetic silencing or chemical inhibition of CTH resulted in SARS-CoV-2 proliferation. Chemical supplementation of H2S using a slow-releasing H2S donor, GYY4137, diminished virus replication. Using a redox biosensor, metabolomics, transcriptomics, and XF-flux analyzer, we showed that GYY4137 blocked SARS-CoV-2 replication by inducing the Nrf2/Keap1 pathway, restoring redox balance and carbon metabolites and potentiating mitochondrial oxidative phosphorylation. Treatment of SARS-CoV-2-infected mice or hamsters with GYY4137 suppressed viral replication and ameliorated lung pathology. GYY4137 treatment reduced the expression of inflammatory cytokines and re-established the expression of Nrf2-dependent antioxidant genes in the lungs of SARS-CoV-2-infected mice. Notably, non-invasive measurement of respiratory functions using unrestrained whole-body plethysmography (uWBP) of SARS-CoV-2-infected mice showed improved pulmonary function variables, including pulmonary obstruction (Penh), end-expiratory pause (EEP), and relaxation time (RT) upon GYY4137 treatment. Together, our findings significantly extend our understanding of H2S-mediated regulation of viral infections and open new avenues for investigating the pathogenic mechanisms and therapeutic opportunities for coronavirus-associated disorders.

Research into the role of chronic sterile inflammation (i.e. a prolonged inflammatory state not caused by an infectious agent), in vascular disease progression has continued to grow over the last few decades. DAMPs have a critical role in this research due to their ability to link stress-causing cardiovascular risk factors to inflammatory phenotypes seen in vascular disease. In this mini-review, we will briefly summarize the DAMPs and receptor signaling pathways that have been extensively studied in the context of vascular disease, including TLRs, RAGE, cGAS-STING, and the NLRP3 inflammasome. In particular, we will discuss how these pathways can promote the release of pro-inflammatory cytokines and chemokines as well as vascular remodeling. Next, we will summarize the results of studies which have linked the various pro-inflammatory effects of DAMPs with the phenotypes in the context of vascular diseases including atherosclerosis, fibrosis, aneurysm, ischemia, and hypertension. Finally, we will discuss some pre-clinical and clinical trials that have targeted DAMPs, their receptors, or the products of their signaling pathways, and discuss the outlook and future directions for the field at large.

Mesenchymal stromal cells (MSCs) represent a promising therapeutic approach in viral infection management. However, their interaction with viruses remains poorly understood. MSCs can support antiviral immune responses and act as viral reservoirs, potentially compromising their therapeutic potential. Innate immune system recognition of viral pathogens involves pattern recognition receptors (PRRs), including RIG-I-like receptors (RLRs), which activate mitochondrial antiviral signaling protein (MAVS). MAVS triggers antiviral pathways like IRF3 and NF-κB, leading to interferon (IFN) production and pro-inflammatory responses. This study explores the antiviral response in umbilical cord-derived MSCs (UC-MSCs) through targeted stimulation with influenza A virus-derived 5'triphosphate-RNA (3p-hpRNA), a RIG-I agonist. By investigating MAVS activation, we provide mechanistic insights into the immune response at the molecular level. Our findings reveal that 3p-hpRNA stimulation triggers immune activation of the IRF3 and NF-κB pathways through MAVS. Subsequently, this leads to the induction of type I and III IFNs, IFN-stimulated genes (ISGs), and pro-inflammatory cytokines. Critically, this immune activation occurs without compromising mitochondrial integrity. UC-MSCs retain their capacity for mitochondrial transfer to recipient cells. These results highlight the adaptability of UC-MSCs, offering a nuanced understanding of immune responses balancing activation with metabolic integrity. Finally, our research provides mechanistic evidence for MSC-based interventions against viral infections.

A balanced IFN response, tightly regulated at multiple levels, is essential for host defense against viral infection. Tripartite motif-containing (TRIM) proteins are a large group of E3 ubiquitin ligases, and have been shown to be involved in the regulation of IFN response. However, the regulatory functions of individual TRIM proteins remain controversial. Here, we show that a virus-inducible TRIM2 homolog acts as a negative regulator for IFN production in zebrafish. Zebrafish Trim2a was upregulated in response to spring viremia of carp virus (SVCV) infection, and knockout of Trim2a significantly increased the expression of antiviral genes, leading to enhanced resistance to SVCV. Overexpression of Trim2a resulted in pronounced ubiquitination of IFN regulatory factor 3 (IRF3) via K11, K27, K29, and K48, promoting IRF3 degradation and stability of SVCV phosphoprotein to favor viral replication. Moreover, TRIM2a induced ubiquitination of autophagic cargo receptor p62, which then interacted with IRF3, instigating IRF3 degradation. Further, the inhibitory effects of TRIM2a on IFN production were also observed in human HEK293 cells, suggesting that the regulatory functions of TRIM2 are likely to be conserved during evolution. Collectively, our findings demonstrate that TRIM2a is a negative regulator of IFN production, and could serve as a potential target to dampen exacerbated IFN response triggered by aberrant activation of retinoic acid-inducible gene 1 (RIG-I)-like receptors. Our study provides insights into a previously uncharacterized role of TRIM2 in the regulation of IFN signaling.

Cancer is one of the leading causes of mortality worldwide. Nanomedicines have significantly improved life expectancy and survival rates for cancer patients in current standard care. However, recurrence of cancer due to metastasis remains a significant challenge. Vaccines can provide long-term protection and are ideal for preventing bacterial and viral infections. Cancer vaccines, however, have shown limited therapeutic efficacy and raised safety concerns despite extensive research. Cancer vaccines target and stimulate responses against tumor-specific antigens and have demonstrated great potential for cancer treatment in preclinical studies. However, tumor-associated immunosuppression and immune tolerance driven by immunoediting pose significant challenges for vaccine design. In situ vaccination represents an alternative approach to traditional cancer vaccines. This strategy involves the intratumoral administration of immunostimulants to modulate the growth and differentiation of innate immune cells, such as dendritic cells, macrophages, and neutrophils, and restore T-cell activity. Currently approved in situ vaccines, such as T-VEC, have demonstrated clinical promise, while ongoing clinical trials continue to explore novel strategies for broader efficacy. Despite these advancements, failures in vaccine research highlight the need to address tumor-associated immune suppression and immune escape mechanisms. In situ vaccination strategies combine innate and adaptive immune stimulation, leveraging tumor-associated antigens to activate dendritic cells and cross-prime CD8+ T cells. Various vaccine modalities, such as nucleotide-based vaccines (e.g., RNA and DNA vaccines), peptide-based vaccines, and cell-based vaccines (including dendritic, T-cell, and B-cell approaches), show significant potential. Plant-based viral approaches, including cowpea mosaic virus and Newcastle disease virus, further expand the toolkit for in situ vaccination. Therapeutic modalities such as chemotherapy, radiation, photodynamic therapy, photothermal therapy, and Checkpoint blockade inhibitors contribute to enhanced antigen presentation and immune activation. Adjuvants like CpG-ODN and PRR agonists further enhance immune modulation and vaccine efficacy. The advantages of in situ vaccination include patient specificity, personalization, minimized antigen immune escape, and reduced logistical costs. However, significant barriers such as tumor heterogeneity, immune evasion, and logistical challenges remain. This review explores strategies for developing potent cancer vaccines, examines ongoing clinical trials, evaluates immune stimulation methods, and discusses prospects for advancing in situ cancer vaccination.

African swine fever virus (ASFV) causes a frequently fatal hemorrhagic disease in domestic pigs and wild boar. The spread from Africa to Georgia in 2007 initiated a pandemic affecting many European and most Asian countries. This has had a very high socio-economic impact and threatens global food security. The virus is a large, complex, cytoplasmic DNA virus, the only member of the Asfarviridae family and codes for 170-190 proteins. Many of these have unknown functions and do not resemble other viruses or host proteins. This complexity has hindered the development of vaccines and other tools for control. The intensity of research has increased since the spread of ASFV in Europe and Asia, leading to rapid advances in knowledge. This review summarizes recent research, including the determination by cryogenic electron microscopy of the virus capsid structure and virion proteome. Novel information on the virus replication cycle, including mechanisms of virus entry into cells and the identification of host endosomal proteins important for entry, is summarized. Multiple, novel virus immune evasion proteins and their targets in the type I interferon response and inflammation pathways have been identified. The potential for the application of this knowledge to developing novel control tools, including modified live vaccines and other interventions targeting critical virus processes or host interactions, is discussed.

Podocytes are key components of the glomerular filtration barrier - a specialized structure that is responsible for the filtration of blood by the kidneys. They therefore exist in a unique microenvironment exposed to mechanical force and the myriad molecules that cross the filtration barrier. To survive and thrive, podocytes must continually sense and respond to their ever-changing microenvironment. Sensing is achieved by interactions with the surrounding extracellular matrix and neighbouring cells, through a variety of pathways, to sense changes in environmental factors such as nutrient levels including glucose and lipids, oxygen levels, pH and pressure. The response mechanisms similarly involve a range of processes, including signalling pathways and the actions of specific organelles that initiate and regulate appropriate responses, including alterations in cell metabolism, immune regulation and changes in podocyte structure and cognate functions. These functions ultimately affect glomerular and kidney health. Imbalances in these processes can lead to inflammation, podocyte loss and glomerular disease.

Antisense oligonucleotides (ASOs) are short, synthetic sequences designed to specifically target RNA target molecules to modulate their translation and protein expression. To enhance their stability, chemically modified ASOs have been developed. Over the decades, peptide nucleic acids (PNAs) have emerged as powerful tools in molecular biology, proving their ability in gene knockdown, as well as therapeutic or diagnostic applications. In this study, we sought to explore the potential of PNAs to inhibit RNA translation through a novel mechanism of action. To enhance translation inhibition, we designed a new class of PNAs able to mediate target RNA circularization. These PNAs bind the 5' end of the RNA, interacting with CAP and 5'UTR structures, while simultaneously binding to the poly-A tail at the opposite end of the same RNA molecule. Our findings suggest target RNA circularization and inhibition of protein translation across multiple models. We established a dose-dependent effect of the circularizing PNAs on target RNA translation. Overall, our results reveal that this dual-targeting approach strongly inhibits RNA translation compared to binding solely to the 5'UTR, paving the way to the potential development and therapeutic application of this novel class of ASO PNAs.

Photochemical control of oligonucleotides bears great potential for the spatio-temporal control of therapeutic targets, such as immune sensing receptors. Retinoic acid-inducible gene I (RIG-I) is a cytoplasmic receptor of the innate immune system that triggers antiviral responses upon detection of viral RNA. RIG-I can be specifically activated by short double-stranded (ds) RNA with a blunt 5' end bearing a triphosphate, mimicking nascent viral transcripts. Tumor cells are specifically sensitive to RIG-I-induced cell death. Here we developed a potent oligonucleotide ligand for spatiotemporally controlled activation of RIG-I by light exposure. Through structural considerations and functional studies we identified a combination of two nucleoside positions in a RIG-I oligonucleotide ligand for which the substitution of both respective 2'-hydroxy groups of the ribose by photolabile protecting groups (2'-photocages) resulted in a complete loss of RIG-I ligand activity, whereas photocaging the individual positions was not sufficient to turn off RIG-I. Light exposure fully restored RIG-I activation by the photocaged RIG-I ligand, enabling light-controlled RIG-I-mediated cell death of human cancer cells which had internalized the photocaged RIG-I ligand prior to light exposure. This novel photoactivatable RIG-I oligonucleotide ligand may be applicable for precise light-controlled induction of tumor cell death in superficial cancer such as melanoma.

As one of the most primitive vertebrates, fish have evolved a distinct repertoire of viral recognition receptors. However, the existence of nuclear viral recognition receptors in fish remains uncertain. In this study, we identified a scaffold attachment factor A (SAFA), CiSAFA, as a potential nuclear recognition receptor for grass carp reovirus (GCRV) in grass carp (Ctenopharyngodon idella). CiSAFA shares high amino acid sequence similarity (70.3-99.6 %) and identity (56.1-98.6 %) with its counterparts from other vertebrates and contains three conserved domains, namely, SAP, SPRY, and AAA_33. On the basis of the subcellular location analysis, CiSAFA was found to localize to the nucleus. In vitro, CiSAFA can bind to poly(I:C) and induce interferon (IFN) expression. The expression data revealed that CiSAFA exhibited ubiquitous mRNA expression across all the tissues of grass carp. After GCRV infection, CiSAFA showed significantly upregulated mRNA expression levels and exhibited an expression pattern similar to that of IFN1 in the spleen and head kidney. The results of RNA immunoprecipitation indicated that CiSAFA might interact with dsRNA segment 6 of GCRV. The overexpression of CiSAFA significantly increased the expression levels of several representative antiviral genes, including interferon regulatory factor 3 (IRF3), IRF7, IFN1, and virus-induced gene 1 (GIG1), and inhibited GCRV replication. To our knowledge, this study represents the first discovery of a potential nuclear recognition receptor, CiSAFA, for GCRV in grass carp and reveals its antiviral immune mechanism against GCRV infection, which may provide new insight into host immune recognition system-virus interactions in fish.

Maintaining innate immune homeostasis is critical for preventing infections and autoimmune diseases but effective interventions are lacking. Here we identified C864-C869-mediated intermolecular disulfide-linkage formation as a critical step for human RIG-I activation that can be bidirectionally regulated to control innate immune homeostasis. The viral-stimulated C864-C869 disulfide linkage mediates conjugation of an SDS-resistant RIG-I oligomer, which prevents RIG-I degradation by E3 ubiquitin-ligase MIB2 and is necessary for RIG-I to perform liquid-liquid phase separation to compartmentalize downstream signalsome, thereby stimulating type I interferon signalling. The corresponding C865S 'knock-in' caused an oligomerization defect and liquid-liquid phase separation in mouse RIG-I, which inhibited innate immunity, resulting in increased viral load and mortality in mice. Using unnatural amino acids to generate covalent C864-C869 linkage and the development of an interfering peptide to block C864-C869 residues, we bidirectionally regulated RIG-I activities in human diseases. These findings provide in-depth insights on mechanism of RIG-I activation, allowing for the development of methodologies that hold promising implications in clinics.

Tembusu virus (TMUV), an emerging avian orthoflavivirus, causes severe economic losses due to egg-drop syndrome and fatal encephalitis in domestic waterfowl. To combat this threat, the host immune system plays a crucial role in controlling and eliminating TMUV infection. Understanding the mechanisms of this immune response is thus vital for developing effective strategies against the virus. In this study, we investigated the antiviral activities of duck TRIM family proteins (duTRIM) against TMUV, focusing particularly on duTRIM14 as a potent host restriction factor. We showed that overexpression of duTRIM14 significantly inhibits TMUV replication, while its deficiency leads to increased viral titers. We elucidate a novel mechanism by which duTRIM14 interacts with the TMUV NS1 protein, facilitating its K27/K29-linked polyubiquitination and subsequent proteasomal degradation. The Lys141 residue on NS1 was identified as critical for this process, with its removal significantly enhancing TMUV replication both in vitro and in vivo. Furthermore, we showed that duTRIM14 interacts with duck TBK1 (duTBK1), promoting its K63-linked polyubiquitination on Lys30 and Lys401, which substantially augments IFN-β production during TMUV infection. Taken together, these results provide a novel dual-action antiviral mechanism in which duTRIM14 suppresses TMUV replication by simultaneously promoting proteasomal degradation of NS1 and enhancing the host antiviral response by modulating duTBK1 activity.

Cytoplasmic DNA-RNA hybrids are emerging as important immunogenic nucleic acids, that were previously underappreciated. DNA-RNA hybrids, formed during cellular processes like transcription and replication, or by exogenous pathogens, are recognized by pattern recognition receptors (PRRs), including cGAS, DDX41, and TLR9, which trigger immune responses. Post-translational modifications (PTMs) including ubiquitination, phosphorylation, acetylation, and palmitoylation regulate the activity of PRRs and downstream signaling molecules, fine-tuning the immune response. Targeting enzymes involved in DNA-RNA hybrid metabolism and PTMs regulation offers therapeutic potential for inflammatory diseases. Herein, we discuss the sources, immune response, and therapeutic implications of DNA-RNA hybrids in inflammation, highlighting the significance of DNA-RNA hybrids as potential targets for the treatment of inflammation.

Hepatitis B virus (HBV), the prototypical member of the Hepadnaviridae family, is a DNA virus that replicates its genome through reverse transcription of a pregenomic RNA (pgRNA) precursor. The selective packaging of pgRNA and viral polymerase (Pol) into assembling capsids formed by the viral core protein-a process known as nucleocapsid assembly-is an essential step in the HBV lifecycle. Advances in cellular and cell-free systems have provided significant insights into the mechanisms underlying capsid assembly, Pol binding to pgRNA, Pol-pgRNA packaging, and initiation of genome replication. However, the absence of a cell-free system capable of reconstituting selective HBV Pol-pgRNA packaging into fully assembled capsids leaves fundamental questions about nucleocapsid assembly unanswered. This review summarizes the current knowledge of HBV nucleocapsid assembly, focusing on the interplay between Pol-pgRNA interactions, capsid formation, and regulation by host factors. It also highlights the contribution of cellular and cell-free systems to these discoveries and underscores the need for new approaches that reconstitute the complete HBV nucleocapsid assembly process. With the growing interest in developing nucleocapsid assembly inhibitors, some of which are currently in clinical trials, targeting Pol-pgRNA interactions and nucleocapsid assembly represents a promising therapeutic strategy for curing chronic hepatitis B.

Persistent intestinal inflammation is a major contributor to various diseases, including digestive disorders, immune dysregulation, and cancer. The NF-κB signaling pathway is pivotal in the inflammatory response of intestinal cells, regulating the secretion of inflammatory factors, mediating signal transduction, and activating receptors. In colitis, NF-κB signaling and its effector molecules are excessively activated by various stimuli, leading to overexpression of inflammatory mediators and immune regulators. Colitis, an inflammation of the intestinal mucosa, underlies many intestinal diseases, with increasing incidence. Traditional treatments such as glucocorticoids and nonsteroidal antiinflammatory drugs have significant limitations and side effects. Pogostemon cablin, a traditional Chinese medicine and food, is widely used in food, spices, and pharmaceuticals. Studies have demonstrated its positive therapeutic effects on intestinal inflammation, primarily through regulation of the NF-κB signaling pathway. Moreover, P. cablin and its active components exhibit pharmacological activities such as antiapoptotic, antioxidant, and antitumor effects. This review summarizes the original research on treating intestinal mucosal inflammation via NF-κB signaling regulation using P. cablin and its active components, providing new insights for colitis treatment.

As an evolutionarily conserved and ubiquitous mechanism of host defense, non-immune cells in vertebrates possess the intrinsic ability to autonomously detect and combat intracellular pathogens. This process, termed cell-autonomous immunity, is distinct from classical innate immunity. In this review, we comprehensively examine the defense mechanisms employed by non-immune cells in response to intracellular pathogen invasion. We provide a detailed analysis of the cytosolic sensors that recognize aberrant nucleic acids, lipopolysaccharide (LPS), and other pathogen-associated molecular patterns (PAMPs). Specifically, we elucidate the molecular mechanisms underlying key signaling pathways, including the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway, the retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs)-mitochondrial antiviral signaling (MAVS) axis, and the guanylate-binding proteins (GBPs)-mediated pathway. Furthermore, we critically evaluate the involvement of these pathways in the pathogenesis of various diseases, including autoimmune disorders, inflammatory conditions, and malignancies, while highlighting their potential as therapeutic targets.

Mucosal-mediated immune deficiency is associated with immune evasion and poor clinical outcomes in acute myeloid leukemia (AML). Here, we describe the elicitation of mucosal and systemic immune response by oral delivery of MDP-modified PEG-lipid (MDP-PEG-DSPE) and polylactic acid-polyhistidine (PLA-PHis) copolymer constructed nanosystem (mPOD) into Peyer's patches. To protect against gastrointestinal degradation, enteric-soluble capsules are utilized for encapsulating mPOD to promote penetration across intestinal mucus and engender robust Peyer's patch targeting initiated by MDP-PEG-DSPE. Compared with intravenous and intramuscular administration, the oral delivery of MDP-PEG-DSPE and 5'-triphosphate-modified RNA (ppp-RNA) into gut-associated lymphoid tissues reinforces dendritic cell maturation and migration, amplifies mucosal immune response, and boosts the production of secretory immunoglobulin A via retinoic acid-inducible gene I/nucleotide-binding oligomerization domain 2 (RIG-I/NOD2) signaling activation. In the AML murine model, the provoked mucosal immunity positively regulates the systemic cytotoxic immune reactions, which, in turn, eradicate disseminated malignant leukemic cells and provide defense against leukemia attacks.

Bilirubin metabolism crucially maintains normal liver function, but whether it contributes to antiviral immunity remains unknown. Here, we reveal that the liver bilirubin metabolic pathway facilitates antiviral innate immunity of the body. We discovered that viral infection upregulates uridine diphosphate glucuronosyltransferase 1A1 (UGT1A1) expression in the liver, which in turn stabilizes IRF3 proteins to promote type I interferon (IFN-I) production. Moreover, we found that serum unconjugated bilirubin (UCB), a unique physiological substrate of UGT1A1, can competitively inhibit the binding of IFN-I to IFN-I receptor 2 (IFNAR2), thus attenuating IFN-I-induced antiviral signaling of the body. Accordingly, effective bilirubin metabolism in the liver promotes antiviral immunity of the body by specifically employing liver UGT1A1-mediated enhancement of IFN-I production and reducing serum bilirubin-mediated inhibition of IFN-I signaling. This study uncovers the significance of bilirubin metabolism in antiviral innate immunity and demonstrates that conventional IFN-I therapy is less efficient for patients with hepatitis B virus (HBV) with high levels of bilirubin.

Orf virus (ORFV) is the type species of Parapoxvirus of the Poxviridae family that induces cutaneous pustular skin lesions in sheep and goats, and causes zoonotic infections in humans. Pattern recognition receptors (PRRs) sense pathogen-associated molecular patterns (PAMPs), leading to the triggering of the innate immune response through multiple signalling pathways involving type I interferons (IFNs). The major PAMPs generated during viral infection are nucleic acids, which are the most important molecules that are recognized by the host. The induction of type l IFNs leads to activation of the Janus kinase (JAK)-signal transducer activator of transcription (STAT) pathway, which results in the induction of hundreds of interferon-stimulated genes (ISGs), many of which encode proteins that have antiviral roles in eliminating virus infection and create an antiviral state. Genetic and functional analyses have revealed that ORFV, as found for other poxviruses, has evolved multiple immunomodulatory genes and strategies that manipulate the innate immune sensing response.

Inflammation is one defense mechanism of the body that has multiple origins, ranging from physical agents to infectious agents including viruses and bacteria. The resolution of inflammation has emerged as a critical endogenous process that protects host tissues from prolonged or excessive inflammation, which can become chronic. Failure of the inflammation resolution is a key pathological mechanism that drives the progression of numerous inflammatory diseases. Owing to the various side effects of currently available drugs to control inflammation, novel therapeutic agents that can prevent or suppress inflammation are needed. Supersulfides are highly reactive and biologically potent molecules that function as antioxidants, redox regulators, and modulators of cell signaling. The catenation state of individual sulfur atoms endows supersulfides with unique biological activities. Great strides have recently been made in achieving a molecular understanding of these sulfur species, which participate in various physiological and pathological pathways. This review mainly focuses on the anti-inflammatory effects of supersulfides. The review starts with an overview of supersulfide biology and highlights the roles of supersulfides in both immune and inflammatory responses. The various donors used to generate supersulfides are assessed as research tools and potential therapeutic agents. Deeper understanding of the molecular and cellular bases of supersulfide-driven biology can help guide the development of innovative therapeutic strategies to prevent and treat diseases associated with various immune and inflammatory responses.

Tumor cells often employ many ways to restrain type I IFN signaling to evade immune surveillance. However, whether cellular amino acid metabolism regulates this process remains unclear, and its effects on antitumor immunity are relatively unexplored. Here, we found that asparagine inhibited IFN-I signaling and promoted immune escape in bladder cancer. Depletion of asparagine synthetase (ASNS) strongly limited in vivo tumor growth in a CD8+ T cell-dependent manner and boosted immunotherapy efficacy. Moreover, clinically approved L-asparaginase (ASNase),synergized with anti-PD-1 therapy in suppressing tumor growth. Mechanistically, asparagine can directly bind to RIG-I and facilitate CBL-mediated RIG-I degradation, thereby suppressing IFN signaling and antitumor immune responses. Clinically, tumors with higher ASNS expression show decreased responsiveness to immune checkpoint inhibitor therapy. Together, our findings uncover asparagine as a natural metabolite to modulate RIG-I-mediated IFN-I signaling, providing the basis for developing the combinatorial use of ASNase and anti-PD-1 for bladder cancer.

Toxoplasma gondii stands as one of the most successful pathogens, capable of infecting nearly all warm-blooded species. It is estimated that up to 50% of human population might harbor Toxoplasmosis infections. One of the primary transmission routes is the consumption of tissue cysts from infected farm animals used for food production. Thus, controlling Toxoplasmosis in farm animals is of vital importance for human health and food safety. Selective breeding in farm animals, where available, could complement classical control measures like biosecurity measures, vaccination, and test-and-cull methods. This multidisciplinary approach will make the eradication of Toxoplasmosis more effective. For this purpose, we conducted four multi-locus genome-wide association (GWA) approaches to identify the polygenic factors underlying innate resistance to Toxoplasma gondii in naturally infected sheep. Our findings indicate that 16 single nucleotide polymorphisms (SNPs), exhibiting varying degrees of statistical power, play a significant role in host immunity against T. gondii infection. We propose the genes containing these SNPs or located within 100 ± Kb of them (PLSCR5, EPHA3, DGKB, IL12B, CGA, WDR64, TMEM158, CLMP, and SIAE) as potential candidate genes. This study represents the first exploration of host genetic factors against Toxoplasmosis in livestock, utilizing the ovine paradigm as its foundation.

Circular RNAs (circRNAs) have gained significant attention in recent years due to their diverse biological functions and potential as novel therapeutic modality. Two general mechanisms for generating circRNAs involve the utilization of group 1 and group 2 introns, which are self-splicing ribozymes found in many organisms. Although group 1 intron has been demonstrated to be highly effective in circularization, the reaction requires high temperature plus cofactors such as GTP. Consequently, undesired byproducts such as nicked RNA were generated, which requires complex purification steps before downstream applications. In this study, we have strategically designed structural elements and incorporated sequence features to enhance the efficacy of RNA circularization by group 2 introns. This innovative approach occurred at a reduced temperature and resulted in notably fewer nicks compared with group 1 introns. Furthermore, to streamline the purification process of circRNA, we incorporated two halves of streptavidin aptamer tags into the enzymatic sites of the group 2 intron. This strategic architecture guarantees the creation of a fully operational streptavidin aptamer tag solely at the circRNA junction site during in vitro circularization. This unique attribute facilitates a one-step purification process. The resulting "nickless one-step purification of engineered circular messenger RNA" system emerges as both straightforward and efficient in generating therapeutic circular messenger RNAs using simple, scalable in vitro systems.

Early detection of viral infection and rapid activation of host antiviral defenses through transcriptional upregulation of interferons (IFNs) and IFN-stimulated genes (ISGs) are critical for controlling infection. However, aberrant production of IFN in the absence of viral infection leads to auto-inflammation and can be detrimental to the host. Here, we show that the DNA-binding transcriptional repressor complex composed of Capicua (CIC) and Ataxin-1 like (ATXN1L) binds to an 8-nucleotide motif near IFN and ISG promoters and prevents erroneous expression of inflammatory genes under homeostasis in humans and mice. By contrast, during respiratory viral infection, activation of the mitogen-activated protein kinase (MAPK) pathway results in rapid degradation of the CIC-ATXN1L complex, thereby relieving repression and allowing for robust induction of IFN and ISGs. Together, our studies define a new paradigm for host regulation of IFN and ISGs through the evolutionarily conserved CIC-ATXN1L transcriptional repressor complex during homeostasis and viral infection.