Retinoic Acid-inducible Gene Research Articles

UBR5 promotes antiviral immunity by disengaging the transcriptional brake on RIG-I like receptors

The Retinoic acid-Inducible Gene I (RIG-I) like receptors (RLRs) are the major viral RNA sensors essential for the initiation of antiviral immune responses. RLRs are subjected to stringent transcriptional and posttranslational regulations, of which ubiquitination is one of the most important. However, the role of ubiquitination in RLR transcription is unknown. Here, we screen 375 definite ubiquitin ligase knockout cell lines and identify Ubiquitin Protein Ligase E3 Component N-Recognin 5 (UBR5) as a positive regulator of RLR transcription. UBR5 deficiency reduces antiviral immune responses to RNA viruses, while increases viral replication in primary cells and mice. Ubr5 knockout mice are more susceptible to lethal RNA virus infection than wild type littermates. Mechanistically, UBR5 mediates the Lysine 63-linked ubiquitination of Tripartite Motif Protein 28 (TRIM28), an epigenetic repressor of RLRs. This modification prevents intramolecular SUMOylation of TRIM28, thus disengages the TRIM28-imposed brake on RLR transcription. In sum, UBR5 enables rapid upregulation of RLR expression to boost antiviral immune responses by ubiquitinating and de-SUMOylating TRIM28.

The Impact of Pathogens on Sepsis Prevalence and Outcome.

Sepsis, a severe global healthcare challenge, is characterized by significant morbidity and mortality. The 2016 redefinition by the Third International Consensus Definitions Task Force emphasizes its complexity as a "life-threatening organ dysfunction caused by a dysregulated host response to infection". Bacterial pathogens, historically dominant, exhibit geographic variations, influencing healthcare strategies. The intricate dynamics of bacterial immunity involve recognizing pathogen-associated molecular patterns, triggering innate immune responses and inflammatory cascades. Dysregulation leads to immunothrombosis, disseminated intravascular coagulation, and mitochondrial dysfunction, contributing to the septic state. Viral sepsis, historically less prevalent, saw a paradigm shift during the COVID-19 pandemic, underscoring the need to understand the immunological response. Retinoic acid-inducible gene I-like receptors and Toll-like receptors play pivotal roles, and the cytokine storm in COVID-19 differs from bacterial sepsis. Latent viruses like human cytomegalovirus impact sepsis by reactivating during the immunosuppressive phases. Challenges in sepsis management include rapid pathogen identification, antibiotic resistance monitoring, and balancing therapy beyond antibiotics. This review highlights the evolving sepsis landscape, emphasizing the need for pathogen-specific therapeutic developments in a dynamic and heterogeneous clinical setting.

USP13 Cooperates with MARCH8 to Inhibit Antiviral Signaling by Targeting MAVS for Autophagic Degradation in Teleost.

Mitochondrial antiviral signaling protein (MAVS), as a central adapter protein in retinoic acid-inducible gene I-like receptor signaling, is indispensable for innate antiviral immunity. Yet, the molecular mechanisms modulating the stability of MAVS are not fully understood in low vertebrates. In this study, we report that the deubiquitinase ubiquitin-specific protease 13 (USP13) acts as a negative regulator of antiviral immunity by targeting MAVS for selective autophagic degradation in teleost fish. USP13 is induced by RNA virus or polyinosinic:polycytidylic acid stimulation and acts as a negative regulator to potentiate viral replication in fish cells. Mechanistically, USP13 functions as a scaffold to enhance the interaction between MAVS and the E3 ubiquitin ligase MARCH8, thus promoting MARCH8 to catalyze MAVS through K27-linked polyubiquitination for selective autophagic degradation. Taken together, to our knowledge, our study demonstrates a novel mechanism by which viruses evade host antiviral immunity via USP13 in fish and provides a new idea for mammalian innate antiviral immunity.

The LxIS motif is the language between virus and host RLRs: From fish to mammalian viruses

Regulation of host interferon (IFN) production is a crucial strategy for viral immune escape. We propose that the LxIS motif language, a marker for exogenous viral proteins, interacts with host retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) factors. In this study, the viral proteins containing the LxIS motif of grass carp reovirus (GCRV), and cyprinid herpesvirus 3 (CyHV-3), which belong to the fish RNA and DNA viruses, were found to interact with RLRs, and the general functions of these proteins were negative regulation of IFN activation. Terrestrial animal viruses with this motif language, such as SARS-CoV-2 and so on, are also examined. Several viral proteins that were not reported previously were found to be associated with the RLR pathway. Taken together, the LxIS motif language elucidates a straightforward and efficient method for identifying viral proteins that target host RLRs to regulate IFN expression, and is conserved in both lower and higher vertebrates.

Identification of a potent and specific retinoic acid-inducible gene 1 pathway activator as a Hepatitis B Virus antiviral through a novel cell-based reporter assay

Chronic Hepatitis B Virus (HBV) infection remains a global burden. To identify small molecule RIG-I agonists as antivirals against HBV, we developed an HBV-pgRNA-based interferon-β (IFN-β) luciferase reporter assay with high level of assay sensitivity, specificity and robustness. Through HTS screening, lead compound (JJ#1) was identified to activate RIG-I signaling pathway by inducing TBK1 phosphorylation. Knockdown experiments demonstrated that JJ#1-induced retinoic acid-inducible gene 1 (RIG-I) signaling pathway activation was MAVS-dependent. Furthermore, JJ#1 exhibited HBV antiviral potency in HBV-infected cell models by reducing HBV DNA and antigens (HBsAg and HBeAg).

Histone deacetylase 8 promotes innate antiviral immunity through deacetylation of RIG-I.

Histone deacetylates family proteins have been studied for their function in regulating viral replication by deacetylating non-histone proteins. RIG-I (Retinoic acid-inducible gene I) is a critical protein in RNA virus-induced innate antiviral signaling pathways. Our previous research showed that HDAC8 (histone deacetylase 8) involved in innate antiviral immune response, but the underlying mechanism during virus infection is still unclear. In this study, we showed that HDAC8 was involved in the regulation of vesicular stomatitis virus (VSV) replication. Over-expression of HDAC8 inhibited while knockdown promoted VSV replication. Further exploration demonstrated that HDAC8 interacted with and deacetylated RIG-I, which eventually lead to enhance innate antiviral immune response. Collectively, our data clearly demonstrated that HDAC8 inhibited VSV replication by promoting RIG-I mediated interferon production and downstream signaling pathway.

Analysis of the role of dihydromyricetin derived from vine tea (Ampelopsis grossedentata) on multiple myeloma by activating STAT1/RIG-I axis.

Multiple myeloma (MM) is a plasma cell malignancy and remains incurable as it lacks effective curative approaches; thus, novel therapeutic strategies are desperately needed. The study aimed to explore the therapeutic role of dihydromyricetin (DHM) in MM and explore its mechanisms. Human MM and normal plasma samples, human MM cell lines, and normal plasma cells were used for in vitro experiments. Cell counting kit-8 (CCK-8), flow cytometry, and trans-well assays were performed for the assessment of cell viability, apoptosis, migration, and invasion, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to assess the mRNA expression of signal transducer and activator of transcription 1 (STAT1) and retinoic acid-inducible gene I (RIG-I). Western blotting was employed to assess E-cadherin, N-cadherin, signal transducer, STAT1, p-STAT1, and RIG-I protein expression. A tumor xenograft model was used for in vivo experiments. Here, dihydromyricetin (DHM) dose-dependently restrained viability, apoptosis, migration, and invasion, and facilitated apoptosis of U266 cells. After DHM treatment, the E-cadherin level was increased and the N-cadherin level was decreased in U266 and RPMI-8226 cells, suggesting the inhibitory effects of DHM on epithelial-mesenchymal transition (EMT) in MM. Besides, the levels of p-STAT1/STAT1 and RIG-I were down-regulated in MM. However, the STAT1 inhibitor fludarabine undid the suppressive effect of DMH on the malignant characteristics of U266 cells. Also, DHM inhibited MM tumor growth and EMT, and activated STAT1/RIG-I pathway in vivo. Collectively, this study first revealed that DHM can restrain EMT and tumor growth in MM by activating STAT1/RIG-I signaling, which provides a novel drug for the treatment of MM.

H5N1 avian influenza virus PB2 antagonizes duck IFN-β signaling pathway by targeting mitochondrial antiviral signaling protein1

Type I interferon (IFN)-mediated innate immune responses represent the first line of host defense against viral infection. However, the molecular mechanisms by which avian influenza virus (AIV) inhibits type I IFN production in ducks are not well understood. Here, we first found that the polymerase basic 2 (PB2) protein of H5N1 subtype AIV inhibited the type I IFN responses by targeting duck mitochondrial antiviral signaling protein (MAVS). We further demonstrated that H5N1-PB2 bound to the △transmembrane (△TM) domain of duck MAVS, and the polymerase basic 1 (PB1) binding domain (PBD) and RNA binding nuclear import domain (RND) of H5N1-PB2 interacted with MAVS to inhibit type I IFN expression in ducks. Collectively, our findings contribute to understanding the molecular mechanism by which AIV proteins regulate the retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) signaling pathway to evade host antiviral immune responses in ducks.

Orange-spotted grouper nervous necrosis virus-encoded protein A induces interferon expression via RIG-I/MDA5-MAVS-TBK1-IRF3 signaling in fish cells.

As a major pathogen, nervous necrosis virus (NNV) infects more than 120 fish species worldwide and is virulent to larvae and juvenile fish, hampering the development of the fish fry industry. Understanding virus-host interaction and underlying mechanisms is an important but largely unknown issue in fish virus studies. Here, using channel catfish ovary and fathead minnow cells as models for the study of innate immunity signaling, we found that NNV-encoded ProA activated interferon signaling via the retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) pathway which was still suppressed by the infection of wild-type NNV. This finding has important implications for the comprehension of NNV protein function and the immune response from different cells. First, RIG-I is the key node for anti-NNV innate immunity. Second, the response intensity of RLR signaling determines the degree of NNV proliferation. This study expands our knowledge regarding the overview of signal pathways affected by NNV-encoded protein and also highlights potential directions for the control of aquatic viruses.

Anemoside B4 inhibits SARS-CoV-2 replication in vitro and in vivo

ObjectiveAnemoside B4 (AB4), the most abundant triterpenoidal saponin isolated from Pulsatilla chinensis, inhibited influenza virus FM1 or Klebsiella pneumoniae-induced pneumonia. However, the anti-SARS-CoV-2 effect of AB4 has not been unraveled. Therefore, this study aimed to determine the antiviral activity and potential mechanism of AB4 in inhibiting human coronavirus SARS-CoV-2 in vivo and in vitro. MethodsThe cytotoxicity of AB4 was evaluated using the Cell Counting Kit-8 (CCK8) assay. SARS-CoV-2 infected HEK293T, HPAEpiC, and Vero E6 cells were used for in vitro assays. The antiviral effect of AB4 in vivo was evaluated by SARS-CoV-2-infected hACE2-IRES-luc transgenic mouse model. Furthermore, label-free quantitative proteomics and bioinformatic analysis were performed to explore the potential antiviral mechanism of action of AB4. Type I IFN signaling-associated proteins were assessed using Western blotting or immumohistochemical staining. ResultsThe data showed that AB4 reduced the propagation of SARS-CoV-2 along with the decreased Nucleocapsid protein (N), Spike protein (S), and 3C-like protease (3CLpro) in HEK293T cells. In vivo antiviral activity data revealed that AB4 inhibited viral replication and relieved pneumonia in a SARS-CoV-2 infected mouse model. We further disclosed that the antiviral activity of AB4 was associated with the enhanced interferon (IFN)-β response via the activation of retinoic acid-inducible gene I (RIG-1) like receptor (RLP) pathways. Additionally, label-free quantitative proteomic analyses discovered that 17 proteins were significantly altered by AB4 in the SARS-CoV-2 coronavirus infections cells. These proteins mainly clustered in RNA metabolism. ConclusionOur results indicated that AB4 inhibited SARS-CoV-2 replication through the RLR pathways and moderated the RNA metabolism, suggesting that it would be a potential lead compound for the development of anti-SARS-CoV-2 drugs.

Retinoic acid-inducible gene-1 knockdown induces immature properties in dendritic cells and prolongs the survival time of allograft mice

BackgroundThe mature state of dendritic cells (DCs) determines their ability to regulate immune responses. Retinoic acid-inducible gene-1 (RIG-1) plays a critical role in DC activation and maturation. RIG-1 activation triggers mitogen-activated protein kinase and nuclear factor-kappa B signal transduction. In this study, we aimed to investigate the effects of inhibiting RIG-1 expression in DCs and its potential in inducing immune tolerance. MethodsDCs were transduced with the recombinant lentiviral vector (Lv) to inhibit RIG-1 expression. A murine islet and skin transplantation model were constructed to find out whether DC-DDX58-RNAi could prolong allograft survival. The phenotypes of DCs and T-cells were analyzed using flow cytometry. Cytokines in serum were detected by the enzyme-linked immunosorbent assay. Protein levels were determined by Western blot. ResultsRIG-1-deficient DCs had low expression of costimulatory molecules and major histocompatibility complex and a strong phagocytic ability. DC-DDX58-RNAi induced regulatory T cell differentiation in the transplant recipient spleens. The DC-DDX58-RNAi-treated recipients showed satisfactory islet allograft function and longer survival time. ConclusionInhibition of RIG-1 with DDX58-RNAi prevented the activation and maturation of the DCs, affected T cell differentiation, protected the biological function of the allograft, and prolonged graft survival. These findings may have important therapeutic implications for new immunomodulatory regimens.

Abstract A032: Antitumor activities of different RIG-I agonists in a murine model of kidney adenocarcinoma

Abstract Purpose: Activation of retinoic acid-inducible gene I (RIG-I) signaling induces tumor cell apoptosis and enhances NK, dendritic, and effector T cell activity. We applied two distinct RIG-I agonists: purified influenza virus RNA (IV RNA) in combination with an agonistic OX40 receptor-specific monoclonal antibody (anti-OX40) and a 3p-hpRNA (5' triphosphate hairpin RNA) combined with unmethylated CpG oligonucleotide (CpG). We also investigated the CpG and anti-OX40 combination. The study aimed to establish the efficacy of these compounds in a murine model of kidney adenocarcinoma (Renca). Methods: Renca cells were subcutaneously engrafted in 40 BALB/c mice. The animals were allocated into three treatment groups and a control group. Intratumoral administration of specific adjuvant combinations was carried out thrice as follows: Group 1: CpG+anti-OX40, Group 2: CpG+3p-hpRNA, Group 3: IV RNA+anti-OX40, and Group 4: PBS. Daily monitoring encompassed assessment of primary tumor sizes and physiological parameters. Deceased mice underwent dissection, and histological evaluations were performed. Treatment efficacy was ascertained based on tumor growth, distant metastases' occurrence, and cytokine levels' alterations. Statistical analysis was conducted, with p-values<0.05 considered statistically significant. Results: On day 38 post-implantation, the survival rates for the various groups were as follows: Group 4: 20%, Group 1: 70% (sig), Group 2: 50% (ns), and Group 3: 60% (sig). Measurements of primary tumor sizes were conducted following the final treatment on day 17 and subsequently every seven days until day 38. On day 17, no significant disparities in primary tumor sizes were observed between Groups 1, 2, 3, and 4. However, on day 24, Groups 1, 2, and 3; on day 31, Groups 1 and 2 exhibited significantly smaller primary tumors than Group 4. By day 38, no significant differences in tumor sizes were evident among the groups. Notably, discernible variations in primary tumor characteristics were observed: Groups 1, 2, and 3 demonstrated incipient demarcation, whereas Group 4 exhibited diffuse primary tumors accompanied by necrosis. Concerning metastatic development, the results are currently under evaluation and will be available for presentation at the conference, alongside the outcomes of cytokine ELISA tests. Discussion: After concluding the experiment, we deem the treatments to hold considerable promise. Notably, while the treatment effects were sustained, tangible outcomes were achieved: prolonged survival of the mice, reduction in primary tumor sizes, and mitigated metastatic formation. However, following the final treatment, there was a notable decline in the animals' overall quality of life. Consequently, we decided to terminate the experiment. In light of our findings, we ascertain that the treatments serve as a solid foundation for further exploration. By modifying the composition of the treatment cocktails and adjusting the treatment frequency, there exists the potential for further enhancement of the achieved results. Citation Format: Dominik Gulyás, Márta Lőrincz, István Jankovics, Béla Dénes, Dóra Földi, Rhiannon Rodgers, Katie Commins, Gábor Andócs, Gábor Kovács. Antitumor activities of different RIG-I agonists in a murine model of kidney adenocarcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A032.

A novel protein encoded by circVPS13D attenuates antiviral innate immunity by targeting MAVS in teleost fish.

The expression of circVPS13D was upregulated with SCRV invasion, which proved that circVPS13D was involved in the regulation of the antiviral immune response. Our study revealed that the existence of circVPS13D promoted the replication of SCRV. Functionally, circVPS13D negatively regulates the antiviral responses of fish. Mechanistically, we confirmed that circVPS13D inhibited RLRs antiviral signaling pathway via the encoded protein VPS13D-170aa by targeting MAVS. Our study provided novel insights into the roles of protein-coding circRNAs and supported VPS13D-170aa as a negative regulator in the antiviral immune responses of teleost fish.

Zebrafish spop promotes ubiquitination and degradation of mavs to suppress antiviral response via the lysosomal pathway

Retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) signaling pathways are required to be tightly controlled to initiate host innate immune responses. Fish mitochondrial antiviral signaling (mavs) is a key determinant in the RLR pathway, and its ubiquitination is associated with mavs activation. Here, we identified the zebrafish E3 ubiquitin ligase Speckle-type BTB–POZ protein (spop) negatively regulates mavs-mediated the type I interferon (IFN) responses. Consistently, overexpression of zebrafish spop repressed the activity of IFN promoter and reduced host ifn transcription, whereas knockdown spop by small interfering RNA (siRNA) transfection had the opposite effects. Accordingly, overexpression of spop dampened the cellular antiviral responses triggered by spring viremia of carp virus (SVCV). A functional domain assay revealed that the N-terminal substrate-binding MATH domain regions of spop were necessary for IFN suppression. Further assays indicated that spop interacts with mavs through the C-terminal transmembrane (TM) domain of mavs. Moreover, zebrafish spop selectively promotes K48-linked polyubiquitination and degradation of mavs through the lysosomal pathway to suppress IFN expression. Our findings unearth a post-translational mechanism by which mavs is regulated and reveal a role for spop in inhibiting antiviral innate responses.

MDS-MSC Derived Exosomes Promote MDS Cells Malignant Proliferation through RIG-1/Isgs Pathway Via RN7SL1

The interactions between bone marrow stem cells (BM-MSCs) and myelodysplastic syndromes (MDS) cells perform a crucial role in MDS pathogenesis by secreting cytokines ,growth factors, and exosomes. Although BM-MSCs induced growth of MDS cells has been studied, the role of exosomes from MDS-derived MSC (MDS-MSC-EXO) in this action remains unclear. In our study, we investigate the effect of MDS-MSC-EXO on the viability, proliferation, migration, cellular apoptosis of MDS cell lines, such as MDS-L and SKM-1. Our data identified that MDS-MSC-EXO can promote proliferation, migration, and decrease cell apoptosis. Both LR-MDS-MSC and HR-MDS-MSC-derived exosomes increased MDS cell growth.Exosomes mediate local cell-to-cell communication by transferring mRNAs, miRNAs, and proteins. In our study, we found that MDS-MSC-EXO expressed higher unshielded RN7SL1,an endogenous RNA normally shielded by RNA binding proteins SRP9/14, than that from health controls (HC-MSC-EXO). MDS cloned cells expressed higher retinoic acid inducible gene-I (RIG-I) and interferon-stimulated genes (ISGs), such as ISG15, IFIT1 and IFITM1, which are the receptor and effector molecules of RN7SL1. Both MDS-MSC-EXO and exogenous RN7SL1 can promote MDS cloned cells proliferation and stimulate ISGs expression by activating RIG-1. RIG-I inhibitor can block this effect. We demonstrate that MDS-MSC may promote MDS cloned cell proliferation through RIG-ISGs signal activated by exosomes-derived RN7SL1. Overall, our studies provide new insights that MSC-EXO functions the crucial roles in mediating the progression of MDS via RIG-1/ISGs activation, and demonstrate novel targets of RN7SL1 to the pathogenesis of MDS.This study will provide new ideas for MDS treatment in the aspect of microenvironment.

Hantaan virus inhibits type I interferon response by targeting RLR signaling pathways through TRIM25

Hantaan virus (HTNV) is responsible for hemorrhagic fever with renal syndrome (HFRS), primarily due to its ability to inhibit host innate immune responses, such as type I interferon (IFN-I). In this study, we conducted a transcriptome analysis to identify host factors regulated by HTNV nucleocapsid protein (NP) and glycoprotein. Our findings demonstrate that NP and Gc proteins inhibit host IFN-I production by manipulating the retinoic acid-induced gene I (RIG-I)-like receptor (RLR) pathways. Further analysis reveals that HTNV NP and Gc proteins target upstream molecules of MAVS, such as RIG-I and MDA-5, with Gc exhibiting stronger inhibition of IFN-I responses than NP. Mechanistically, NP and Gc proteins interact with tripartite motif protein 25 (TRIM25) to competitively inhibit its interaction with RIG-I/MDA5, suppressing RLR signaling pathways. Our study unveils a cross-talk between HTNV NP/Gc proteins and host immune response, providing valuable insights into the pathogenic mechanism of HTNV.

A novel RNF125 variant associated with Tenorio syndrome alters ubiquitin chain binding.

A key signalling pathway required for clearance of viruses from host cells relies on the receptor protein, retinoic acid-inducible gene I (RIG-I). The activity of RIG-I is tightly controlled, and once bound to viral dsRNA, addition of lysine 63-linked ubiquitin chains activates signalling. Meanwhile, the addition of lysine 48-linked ubiquitin chains to RIG-I is required to terminate signalling when the infection has been resolved. Really interesting new gene (RING) finger protein 125 (RNF125) is the E3 ligase responsible for addition of the ubiquitin chains that terminate signalling, with disruption of its function associated with Tenorio syndrome. Here we describe a novel RNF125 gene variant in an individual with clinical symptoms including intellectual disability, macrocephaly and congenital heart disease, consistent with Tenorio syndrome. The newly identified Tenorio syndrome-associated variant [(NM_017831.4):c.670G>C p.Glu224Gln] is the first to be found in the ubiquitin interaction motif (UIM) of RNF125. While the E3 ligase activity of this RNF125 variant is retained, it has an impaired ability to interact with lysine 63-linked ubiquitin chains. The function of the UIM in RNF125 is uncertain; however, this study suggests that the UIM binds lysine 63-linked ubiquitin chains, and that this interaction is required for the normal function of RNF125.

Bioinformatics and in vitro study reveal the roles of microRNA-346 in high glucose-induced human retinal pigment epithelial cell damage.

To study microRNAs (miRNAs) and their potential effects in high glucose-induced human retinal pigment epithelial cell damage. We screened the GSE52233 miRNA expression dataset for differentially expressed miRNAs (DEMs). The target genes of the top 10 DEMs were predicted using miRWalk 2.0 database, followed by function enrichment and protein-protein interaction analysis. miRNA expression was determined in the human retinal pigment epithelial cell line ARPE-19 treated with high glucose (HG) by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Cell proliferation was determined using cell counting kit (CCK)-8 assay. Cell cycle, apoptosis, and reactive oxygen species (ROS) levels were determined by flow cytometry. The direct interaction between miRNA and targets was validated using dual-luciferase reporter assay. Thirty-nine DEMs were screened, and we predicted 125 miRNA-mRNA pairs for the top 10 DEMs, including 119 target genes of seven DEMs such as miR-346, which was upregulated in diabetic retinopathy (DR). miR-346 target genes were substantially enriched in the regulation of intracellular transport and retinoic acid-inducible gene I (RIG-I)-like receptor signaling pathway. Expression of three upregulated and downregulated miRNAs were verified by qRT-PCR in HG-treated ARPE-19 cells. Expression of miR-346 was elevated in HG treated ARPE-19 cells in a dose-dependent manner. HG inhibited cell proliferation and induced apoptosis, which were partly reversed by transfecting an miR-346 inhibitor, which even decreased the ROS levels elevated due to HG. Argonaute 2 (AGO2) was a target of miR-346. miR-346 is a key miRNA and plays an important role in HG-induced damage in human retinal pigment epithelial cells.

Emodin modulates the host antiviral response of fish against spring viremia of carp virus infection

As a natural small molecule monomer widely found in various traditional Chinese herbs, emodin has garnered substantial attention due to its pharmacological properties. Its widespread exploration for application such as growth enhancement, immune modulation, and antimicrobial activity in aquaculture. In the present study, our focus centers on investigate the anti-spring viremia of carp virus (SVCV) activity of emodin, thereby facilitate the development of commercially viable methods for the prevention and treatment of SVCV infection. We assessed the inhibitory effects of emodin against SVCV through both in vitro and in vivo paradigms. In the in vitro assay, cells were firstly infected with emodin mixed with SVCV to impact of emodin on viral replication, apoptosis, and mitochondrial depolarization. Notably, the results underscore that emodin, at a concentration of 16 μM, engenders a remarkable 99.99% suppression of SVCV infection. Interestedly, it distinctly influences the internalization of viral particles into the cells as well as modulate the antiviral response. Furthermore, in the in vivo assay, zebrafish (Danio rerio) fed with an emodin-supplemented commercial diet exhibited a notably elevated survival rate, surpassing that SVCV-exposed group which fed with only commercial diet (up to 22%), a significant reduction in viral load in host was also discerned. Emodin supplementation elicited substantial upregulation of host antiviral related genes, such as interferon-φ (IFN-φ), retinoic acid inducible gene I (RIG-I), and IFN-stimulated gene product 15 (ISG15), by 4.98-, 3.97-, and 7.42-fold on day 7, respectively. In summation, our results indicate that emodin has anti-SVCV activity, and the confluence of emodin's well-established extraction protocol, environmental friendliness, and multiple bioactivities collectively predict it has the potential for commercial application as an antiviral disease additive in aquaculture.

EXTH-35. UNRAVELING THE MECHANISTIC UNDERPINNINGS OF MRNA VACCINE MEDIATED INNATE IMMUNE ACTIVATION AND LYMPHOCYTE TRAFFICKING FOR HIGH-GRADE GLIOMAS

Abstract BACKGROUND High-grade gliomas are an aggressive subset of CNS tumors in need of novel treatment strategies. However, tumor heterogeneity, the immunosuppressive tumor microenvironment (TME) and the lack of immunogenic tumor specific targets have limited the translational power of many immunotherapies for high-grade gliomas. The promising application of mRNA vaccines has the potential to restore immunosurveillance through peripheral and intratumoral reprograming of the TME. METHODS Our lab has developed a lipid particle multilamellar aggregate (RNA-LPA) vaccine that encapsulates tumor specific RNA and delivers the payload to the immune system in a highly immunogenic fashion, similar to that of a virus. We sought to assess anti-tumor activity across preclinical murine models and investigate the mechanisms for response. RESULTS We demonstrated the immune activation potential of RNA-LPAs encoding for glioma associated antigens (i.e., pp65 and H3K27M) in preclinical murine models. We also showed activity of personalized RNA-LPA in canine and human studies (NCT04573140) for patients with high-grade gliomas. Exogenous RNA is recognized by pathogen recognition receptors (PRRs) which trigger a type I interferon (IFN) response and proinflammatory cytokine production. While most RNA based vaccines activate an innate immune response primarily through toll-like receptor 7 (TLR7), we reveal that RNA-LPAs activate the innate immune system through retinoic acid-inducible gene 1 protein (RIG-I). As a result of PRR signaling, the type I IFN and cytokine/chemokine response (IL-6, G-CSF, TNF-alpha, CXCL9, CXCL10, CCL2, CCL4) mediates massive recruitment of activated dendritic cell and activated T cell in lymphoreticular organs where antigen presenting cells and cytolytic T cell colocalize to initiate an adaptive immune response against glioma specific antigens. CONCLUSION RNA-LPAs are a novel platform immunotherapy that initiate anti-tumor immunity against malignant brain tumors through noncanonical activation of intracellular PRRs and widespread recruitment of peripheral blood mononuclear cells to sites of RNA-LPA localization.