bluuebunny/crossref_metadata_2025 huggingface dataset:
Bead-based Multiplex Assay Panels for Quantifying COVID-19 Vaccine-induced Serological Response, Neutralizing Antibodies and Cytokines
Authors: Zhao Amy , Zhao Jessica , Webster Edvelyn , Wheat Cassandra , Sun Binggang , Ni Jessie
Published: January 2023

The COVID-19 pandemic, caused by SARS-CoV-2 virus infection, has resulted in more than 5 million deaths and triggered severe social and economic disruptions worldwide. The emergence of multiple variants of the virus (Alpha, Beta, Gamma, Delta, and Omicron) has further made the pandemic to be one of the most dangerous in history. In order to enable biomedical researchers to understand the disease and develop advanced vaccines and treatments, BioLegend has developed a series of bead-based multiplex panels including SARS-CoV-2 serological IgA, IgM and IgG Panel, Variants Neutralization Antibody, and COVID-19 Cytokine Storm Panels. These panels can quantify simultaneously either various isotype of antibodies, functional antibodies or cytokines in one sample using commonly available flow cytometers. Highly specific antibodies were used in order to ensure assay specificities and robust performance. In addition, these multiplex assay panels are validated in biological samples (serum and plasma) obtained from donors before and after vaccination. Our preliminary data indicate that three types of vaccines induced different antibody responses and cytokine profiles. The experimental data demonstrated that these panels can be reliable, convenient and cost-effective tools for vaccine developers and COVID-19 research.
Proteomic analysis of plasma exosomes as biomarkers of response to MVP-S based immunotherapy
Authors: Smith Pascal , Teh Evelyn , Chisholm Ken , Penny Susanne , Pinto Devanand , Patterson Kelcey , Dirk Brennan , Bydoun Moamen , Fiset Stephan , Bramhecha Yogesh , Hirsch Heather , Hrytsenko Olga , Graff Jeremy
Published: January 2023

Ovarian cancer is a deadly gynaecological disease owing to its late-stage diagnosis. Recurrence and chemo-resistance develop despite initial treatment success. Maveropepimut-S (MVP-S), formerly DPX-Survivac, is a DPX-based immune educating therapy that elicits robust, targeted and, sustained specific anti-tumor T- and B-cell responses in advanced ovarian cancer patients.

Exosomes are a subtype of extracellular vesicles (EVs) secreted by cells involved in intercellular communication. They contain bioactive molecules that can influence the tumor, the immune system, and the extracellular environment. Hence, circulating exosomes could provide real time assessment of disease evolution making them ideal minimally invasive biomarkers for monitoring response to immunotherapies.

Plasma exosomes were isolated from advanced ovarian cancer patients (n=22) at pre- and on-treatment with MVP-S based therapy (DeCidE1 trial, NCT02785250). Subjects with clinical benefit (>10% tumor shrinkage) to therapy displayed a lower overall total EV protein concentration. Untargeted proteomic profiling by liquid chromatography tandem mass spectrometry identified 227 proteins in exosomes. Of these, 95 (42%) were likely from non-tumor exosomes and involved in immune modulation such as regulation of humoral response and B-cell immunity. When compared to the plasma proteome, 128 (56%) proteins were found to be exclusive to exosomes. Functional annotations of these proteins suggest their role in modulating immune pathways related to Fc receptor mediated signaling and T-cell migration.

The results highlight the potential value of plasma exosome assays for monitoring disease and as a potential surrogate response biomarker.

Supported by -IMV inc. -NRC-IRAP R&D Certificate Program.
Harnessing the unique immune biology of cytomegalovirus for cancer immunotherapy
Authors: Iyer L Ravi F , Verweij Marieke C , Nair Sujit , Morrow David , Mansouri Mandana , Beechwood Teresa , Meyer Christine , Chakravarty Dimple , Uebelhoer Luke , Ventura Abigail , Lauron Elvin J , Selseth Andrea , Axthelm Michael , Lind Evan F , Saultz Jennifer , Douglas Janet , Korman Alan , Bhardwaj Nina , Tewari Ashutosh K , Hansen Scott G , Malouli Daniel , Picker Louis J , Fruh Klaus
Published: January 2023

Pre-clinical models in non-human primates demonstrate that cytomegalovirus (CMV)-vectored vaccines are unique in their ability to elicit and indefinitely maintain high frequencies of polyfunctional effector memory T cells to heterologous pathogen antigens, including in animals that are already chronically CMV infected. By introducing defined genetic modifications into the CMV backbone it is possible to program CD8+ T cell responses that are either directed to MHC-I, MHC-II or the non-classical MHC-I molecule MHC-E. In progress clinical studies are evaluating CMV-based vaccine immunity in humans as potential vaccines against HIV. To see if the pre-clinical findings could be extended to cancer antigens we inserted known tumor associated antigens (TAA) or viral TAA into genetically modified rhesus CMV (RhCMV) and characterized the T cell response in rhesus macaques. We found T cell responses to all TAA that were comparable to pathogen antigen-specific responses in frequency, duration, phenotype, epitope density and MHC-restriction. Since many of these TAA are expressed in healthy tissue, this suggests that CMV-vectored cancer vaccines are well-suited to overcome immunological tolerance. As such, CMV-based vectors expressing TAAs could be a powerful new tool for cancer immunotherapy. We show that TAA-specific, MHC-E restricted CD8+ T cells from RhCMV/TAA-immunized RM are stimulated by TAA-expressing cancer tissues and cell lines, indicating that cancer cells can present TAA-derived peptides via MHC-E. Since MHC-E is often upregulated in cancer cells to engage the inhibitory NKG2A receptor on tumor-infiltrating T and NK cells, these results suggest that MHC-E could be used as a novel target for T cell-based immunotherapies.
Combination of NKG2A and PD-1 Blockade Improves Radiotherapy Response in Radioresistant Tumors
Authors: Battaglia Nicholas G , Murphy Joseph D , Uccello Taylor P , Hughson Angela , Gavras Nicholas W , Caldon Johnathan J , Gerber Scott A , Lord Edith M
Published: January 2023

Radiotherapy (RT) is commonly employed to treat solid tumors. Immune checkpoint blockade of programmed cell death protein 1 (PD-1) and CTLA-4 improves survival in RT patients, yet many fail to respond to combination therapy. Natural killer group 2 (NKG2) family receptors, particularly inhibitory NKG2A and activating NKG2D, have emerged as promising therapeutic targets to improve antitumor T cell responses; thus, we examined how these receptors and their ligands (Qa-1b and retinoic acid early inducible 1 [Rae-1], respectively) regulate the RT response in C57BL/6 mice bearing syngeneic B16F10 melanoma and MC38 colorectal adenocarcinoma tumors. RT (15 Gy) transiently reduced B16F10 tumor burden, whereas MC38 tumors exhibited durable response to RT. Intratumoral NK and CD8 T cells expressed NKG2A and NKG2D in both models, which was unaltered by RT. In vitro/in vivo RT increased tumor/stromal cell Qa-1b and Rae-1 expression in both models, especially B16F10 tumors, but IFN-γ stimulation induced both Qa-1b and Rae-1 only in B16F10 tumors. NKG2A/Qa-1b inhibition alone did not improve RT response in either model, but combined RT and NKG2A/PD-1 blockade improved survival in the B16F10 model. Depletion experiments indicate that the triple therapy efficacy is CD8 T cell–dependent with negligible NK cell contribution. RNA sequencing of CD8 T cells from triple therapy–treated B16F10 tumors showed increased proliferative capacity compared with RT and PD-1 blockade alone. Our work demonstrates that RT modulates NKG2A ligand expression, which inhibits RT-induced T cell responses in tumors that fail to respond to combined RT and PD-1 blockade. These results provide a rationale for combining NKG2A blockade with immune checkpoint blockade therapies and RT to improve clinical response.
A Rapid Translational Immune Response Program in CD8 Memory T Lymphocytes
Authors: Salloum Darin , Singh Kamini , Davidson Natalie R , Cao Linlin , Kuo David , Sanghvi Viraj R , Jiang Man , Lafoz Maria Tello , Viale Agnes , Ratsch Gunnar , Wendel Hans-Guido
Published: January 2023

The activation of memory T cells is a very rapid and concerted cellular response that requires coordination between cellular processes in different compartments and on different time scales. In this study, we use ribosome profiling and deep RNA sequencing to define the acute mRNA translation changes in CD8 memory T cells following initial activation events. We find that initial translation enables subsequent events of human and mouse T cell activation and expansion. Briefly, early events in the activation of Ag-experienced CD8 T cells are insensitive to transcriptional blockade with actinomycin D, and instead depend on the translation of pre-existing mRNAs and are blocked by cycloheximide. Ribosome profiling identifies ∼92 mRNAs that are recruited into ribosomes following CD8 T cell stimulation. These mRNAs typically have structured GC and pyrimidine-rich 5′ untranslated regions and they encode key regulators of T cell activation and proliferation such as Notch1, Ifngr1, Il2rb, and serine metabolism enzymes Psat1 and Shmt2 (serine hydroxymethyltransferase 2), as well as translation factors eEF1a1 (eukaryotic elongation factor α1) and eEF2 (eukaryotic elongation factor 2). The increased production of receptors of IL-2 and IFN-γ precedes the activation of gene expression and augments cellular signals and T cell activation. Taken together, we identify an early RNA translation program that acts in a feed-forward manner to enable the rapid and dramatic process of CD8 memory T cell expansion and activation.
A Baseline Cellular Antiviral State Is Maintained by cGAS and Its Most Frequent Naturally Occurring Variant rs610913
Authors: Kazmierski Julia , Elsner Carina , Döhner Katinka , Xu Shuting , Ducroux Aurélie , Pott Fabian , Jansen Jenny , Thorball Christian W , Zeymer Ole , Zhou Xiaoyi , Fedorov Roman , Fellay Jacques , Löffler Markus W , Weber Alexander N R , Sodeik Beate , Goffinet Christine
Published: January 2023

Upon recognition of aberrantly located DNA, the innate immune sensor cyclic GMP-AMP synthase (cGAS) activates stimulator of IFN genes (STING)/IFN regulatory factor (IRF)3–driven antiviral responses. In this study, we characterized the ability of a specific variant of the human cGAS-encoding gene MB21D1, rs610913, to alter cGAS-mediated DNA sensing and viral infection. rs610913 is a frequent G>T polymorphism resulting in a P261H exchange in the cGAS protein. Data from the International Collaboration for the Genomics of HIV suggested that rs610913 nominally associates with HIV-1 acquisition in vivo. Molecular modeling of cGAS(P261H) hinted toward the possibility for an additional binding site for a potential cellular cofactor in cGAS dimers. However, cGAS(wild-type [WT]) or cGAS(P261H)-reconstituted THP-1 cGAS knockout cells shared steady-state expression of IFN-stimulated genes, as opposed to cells expressing the enzymatically inactive cGAS(G212A/S213A). Accordingly, cGAS(WT) and cGAS(P261H) cells were less susceptible to lentiviral transduction and infection with HIV-1, HSV-1, and Chikungunya virus as compared with cGAS knockout or cGAS(G212A/S213A) cells. Upon DNA challenge, innate immune activation appeared to be mildly reduced upon expression of cGAS(P261H) compared with cGAS(WT). Finally, DNA challenge of PBMCs from donors homozygously expressing rs610913 provoked a trend toward a slightly reduced type I IFN response as compared with PBMCs from GG donors. Taken together, the steady-state activity of cGAS maintains a baseline antiviral state rendering cells more refractory to IFN-stimulated gene–sensitive viral infections. rs610913 failed to grossly differ phenotypically from the WT gene, suggesting that cGAS(P261H) and WT cGAS share a similar ability to sense viral infections in vivo.
Ornithine Decarboxylase in Gastric Epithelial Cells Promotes the Immunopathogenesis of Helicobacter pylori Infection
Authors: Latour Yvonne L , Sierra Johanna C , McNamara Kara M , Smith Thaddeus M , Luis Paula B , Schneider Claus , Delgado Alberto G , Barry Daniel P , Allaman Margaret M , Calcutt M Wade , Schey Kevin L , Piazuelo M Blanca , Gobert Alain P , Wilson Keith T
Published: January 2023

Colonization by Helicobacter pylori is associated with gastric diseases, ranging from superficial gastritis to more severe pathologies, including intestinal metaplasia and adenocarcinoma. The interplay of the host response and the pathogen affect the outcome of disease. One major component of the mucosal response to H. pylori is the activation of a strong but inefficient immune response that fails to control the infection and frequently causes tissue damage. We have shown that polyamines can regulate H. pylori–induced inflammation. Chemical inhibition of ornithine decarboxylase (ODC), which generates the polyamine putrescine from l-ornithine, reduces gastritis in mice and adenocarcinoma incidence in gerbils infected with H. pylori. However, we have also demonstrated that Odc deletion in myeloid cells enhances M1 macrophage activation and gastritis. Here we used a genetic approach to assess the specific role of gastric epithelial ODC during H. pylori infection. Specific deletion of the gene encoding for ODC in gastric epithelial cells reduces gastritis, attenuates epithelial proliferation, alters the metabolome, and downregulates the expression of immune mediators induced by H. pylori. Inhibition of ODC activity or ODC knockdown in human gastric epithelial cells dampens H. pylori–induced NF-κB activation, CXCL8 mRNA expression, and IL-8 production. Chronic inflammation is a major risk factor for the progression to more severe pathologies associated with H. pylori infection, and we now show that epithelial ODC plays an important role in mediating this inflammatory response.
Heat Shock Protein 70 Is a Damage-Associated Molecular Pattern That by Binding to Lipopolysaccharide and β-1,3-Glucan–Binding Protein Activates the Prophenoloxidase System in Shrimp
Authors: Junprung Wisarut , Supungul Premruethai , Sangklai Nutthapon , Tassanakajon Anchalee
Published: January 2023

Recent studies have initiated a paradigm shift in understanding heat shock protein 70 (HSP70) functions in the shrimp immune system. However, the mechanism by which Litopenaeus vannamei (Lv)HSP70 modulates the innate immune response remains unclear. This study shows that LvHSP70 binds to the pattern recognition receptor LPS and β-1,3-glucan–binding protein (LvLGBP), and subsequently leads to the activation of the prophenoloxidase system. Injection of shrimp with rLvHSP70 significantly (p < 0.05) upregulated the gene and protein expression of the key pattern recognition receptor LvLGBP. A coimmunoprecipitation and ELISA-based binding assay strongly confirmed the binding of LvHSP70 to LvLGBP at polysaccharide recognition motifs (PLS motifs) with a Kd of 4.44 μM and its competitive binding with LPS (IC50) is 8.036 μM. Conversely, LPS efficiently competed with LvHSP70 for binding to LvLGBP in a concentration-dependent manner with an IC50 of 7.662 μM, indicating that both are ligands of LvLGBP and likely bind at the same site. Binding of LvHSP70 to LvLGBP highly activated phenoloxidase activity in shrimp hemocyte lysate supernatants. Gene silencing of LvLGBP impaired the activation of phenoloxidase activity in shrimp by rLvHSP70, indicating that LvHSP70–LvLGBP interaction was essential for stimulating the immune cascade. Taken together, these results demonstrated that LvHSP70 is a ligand of LvLGBP similar to LPS and acts as a damage-associated molecular pattern to modulate the shrimp immune system via the prophenoloxidase system, eventually leading to the production of melanin and toxic reactive intermediates against invading pathogens.
Disruption of Nurse-like Cell Differentiation as a Therapeutic Strategy for Chronic Lymphocytic Leukemia
Authors: Merchand-Reyes Giovanna , Santhanam Ramasamy , Robledo-Avila Frank H , Weigel Christoph , Ruiz-Rosado Juan de Dios , Mo Xiaokui , Partida-Sánchez Santiago , Woyach Jennifer A , Oakes Christopher C , Tridandapani Susheela , Butchar Jonathan P
Published: January 2023

Chronic lymphocytic leukemia (CLL) is the most common adult leukemia, but, despite advances in treatment, many patients still experience relapse. CLL cells depend on interactions with supportive cells, and nurse-like cells (NLCs) are the major such cell type. However, little is known about how NLCs develop. Here, we performed DNA methylation analysis of CLL patient–derived NLCs using the 850K Illumina array, comparing CD14+ cells at day 1 (monocytes) versus day 14 (NLCs). We found a strong loss of methylation in AP-1 transcription factor binding sites, which may be driven by MAPK signaling. Testing of individual MAPK pathways (MEK, p38, and JNK) revealed a strong dependence on MEK/ERK for NLC development, because treatment of patient samples with the MEK inhibitor trametinib dramatically reduced NLC development in vitro. Using the adoptive transfer Eµ-TCL1 mouse model of CLL, we found that MEK inhibition slowed CLL progression, leading to lower WBC counts and to significantly longer survival time. There were also lower numbers of mouse macrophages, particularly within the M2-like population. In summary, NLC development depends on MEK signaling, and inhibition of MEK leads to increased survival time in vivo. Hence, targeting the MEK/ERK pathway may be an effective treatment strategy for CLL.
The Chicken A and E Blood Systems Arise from Genetic Variation in and around the Regulators of Complement Activation Region
Authors: Fulton Janet E , Drobik-Czwarno Wiola , Wolc Anna , McCarron Amy M , Lund Ashlee R , Schmidt Carl J , Taylor Robert L
Published: January 2023

The tightly linked A and E blood alloantigen systems are 2 of 13 blood systems identified in chickens. Reported herein are studies showing that the genes encoding A and E alloantigens map within or near to the chicken regulator of complement activation (RCA) gene cluster, a region syntenic with the human RCA. Genome-wide association studies, sequence analysis, and sequence-derived single-nucleotide polymorphism information for known A and/or E system alleles show that the most likely candidate gene for the A blood system is C4BPM gene (complement component 4 binding protein, membrane). Cosegregation of single-nucleotide polymorphism–defined C4BPM haplotypes and blood system A alleles defined by alloantisera provide a link between chicken blood system A and C4BPM. The best match for the E blood system is the avian equivalent of FCAMR (Fc fragment of IgA and IgM receptor). C4BPM is located within the chicken RCA on chicken microchromosome 26 and is separated from FCAMR by 89 kbp. The genetic variation observed at C4BPM and FCAMR could affect the chicken complement system and differentially guide immune responses to infectious diseases.
An In Vivo Model of Human Macrophages in Metastatic Melanoma
Authors: Voillet Valentin , Berger Trisha R , McKenna Kelly M , Paulson Kelly G , Tan Wei Hong , Smythe Kimberly S , Hunter Daniel S , Valente William J , Weaver Stephanie , Campbell Jean S , Kim Teresa S , Byrd David R , Bielas Jason H , Pierce Robert H , Chapuis Aude G , Gottardo Raphaël , Rongvaux Anthony
Published: January 2023

Despite recent therapeutic progress, advanced melanoma remains lethal for many patients. The composition of the immune tumor microenvironment (TME) has decisive impacts on therapy response and disease outcome, and high-dimensional analyses of patient samples reveal the heterogeneity of the immune TME. Macrophages infiltrate TMEs and generally associate with tumor progression, but the underlying mechanisms are incompletely understood. Because experimental systems are needed to elucidate the functional properties of these cells, we developed a humanized mouse model reconstituted with human immune cells and human melanoma. We used two strains of recipient mice, supporting or not supporting the development of human myeloid cells. We found that human myeloid cells favored metastatic spread of the primary tumor, thereby recapitulating the cancer-supportive role of macrophages. We next analyzed the transcriptome of human immune cells infiltrating tumors versus other tissues. This analysis identified a cluster of myeloid cells present in the TME, but not in other tissues, which do not correspond to canonical M2 cells. The transcriptome of these cells is characterized by high expression of glycolytic enzymes and multiple chemokines and by low expression of gene sets associated with inflammation and adaptive immunity. Compared with humanized mouse results, we found transcriptionally similar myeloid cells in patient-derived samples of melanoma and other cancer types. The humanized mouse model described here thus complements patient sample analyses, enabling further elucidation of fundamental principles in melanoma biology beyond M1/M2 macrophage polarization. The model can also support the development and evaluation of candidate antitumor therapies.
Carma3 Protects from Liver Injury by Preserving Mitochondrial Integrity in Liver Sinusoidal Endothelial Cells
Authors: Cheng Liqing , Wei Zhanqi , Yang Zaopeng , Lu Renlin , Yang Ming , Yu Muchun , Yang Naixue , Li Shulin , Gao Mingyi , Zhao Xueqiang , Lin Xin
Published: January 2023

Carma3 is an intracellular scaffolding protein that can form complex with Bcl10 and Malt1 to mediate G protein–coupled receptor– or growth factor receptor–induced NF-κB activation. However, the in vivo function of Carma3 has remained elusive. Here, by establishing a Con A–induced autoimmune hepatitis model, we show that liver injury is exacerbated in Carma3−/− mice. Surprisingly, we find that the Carma3 expression level is higher in liver sinusoidal endothelial cells (LSECs) than in hepatocytes in the liver. In Carma3−/− mice, Con A treatment induces more LSEC damage, accompanied by severer coagulation. In vitro we find that Carma3 localizes at mitochondria and Con A treatment can trigger more mitochondrial damage and cell death in Carma3-deficient LSECs. Taken together, our data uncover an unrecognized role of Carma3 in maintaining LSEC integrity, and these results may extend novel strategies to prevent liver injury from toxic insults.
The Brucella Effector BspI Suppresses Inflammation via Inhibition of IRE1 Kinase Activity during Brucella Infection
Authors: Li Chen , Wang Jingyu , Sun Wanchun , Liu Xiaofeng , Wang Jun , Peng Qisheng
Published: January 2023

Mammalian GTPase-activating proteins (GAPs) can inhibit innate immunity signaling in a spatiotemporal fashion; however, the role of bacterial GAPs in mediating innate immunity remains unknown. In this study, we show that BspI, a Brucella type IV secretion system (T4SS) effector protein, containing a GAP domain at the C terminus, negatively regulates proinflammatory responses and host protection to Brucella abotus infection in a mouse model. In macrophages, BspI inhibits the activation of inositol-requiring enzyme 1 (IRE1) kinase, but it does not inhibit activation of ATF6 and PERK. BspI suppresses induction of proinflammatory cytokines via inhibiting the activity of IRE1 kinase caused by VceC, a type IV secretion system effector protein that localizes to the endoplasmic reticulum. Ectopically expressed BspI interacts with IRE1 in HeLa cells. The inhibitory function of BspI depends on its GAP domain but not on interaction with small GTPase Ras-associated binding protein 1B (RAB1B). Collectively, these data support a model where BspI, in a GAP domain–dependent manner, inhibits activation of IRE1 to prevent proinflammatory cytokine responses.
An AhR–Caspase Axis Mediated Antiviral Apoptosis in an Arthropod
Authors: Fu Qihui , Li Haoyang , Wang Sheng , Liao Xuzheng , Chen Shihan , Xiao Bang , Chen Rongjian , Jiang Xiewu , He Jianguo , Li Chaozheng
Published: January 2023

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates immune modulation following exposure of animals to many environmental xenobiotics. However, its role in innate immune responses during viral infection is not fully understood, especially in invertebrates. In this study, a cDNA encoding an AhR homolog was cloned from an arthropod Litopenaeus vannamei (LvAhR). The expression of LvAhR was strongly upregulated in response to the challenge of white spot syndrome virus, a pathogen of highly contagious and fatal infectious disease of shrimp. The relevance of LvAhR to host defense was underlined by heightened susceptibility and elevated virus loads after AhR-silenced shrimp exposure to white spot syndrome virus. LvAhR could induce an apoptosis response through regulating the expression of L. vannamei caspase-1 (homologous to human caspase-3) by directly targeting its promoter that was required to couple with AhR nuclear translocator. Additionally, knockdown of L. vannamei caspase-1 resulted in elevated virus titers and a lower cell apoptotic rate. Thus, we demonstrate that an AhR–caspase axis restrains virus replication by promoting antiviral apoptosis, supporting a previously unidentified direct link between AhR signaling and caspase-mediated apoptosis signaling and, furthermore, suggests that the AhR–caspase axis could be a potential therapeutic target for enhancing antiviral responses in arthropods.
CgDM9CP-5-Integrin-MAPK Pathway Regulates the Production of CgIL-17s and Cgdefensins in the Pacific Oyster, Crassostrea gigas
Authors: Liu Yu , Wang Weilin , Sun Jiejie , Li Yinan , Wu Shasha , Li Qing , Dong Miren , Wang Lingling , Song Linsheng
Published: January 2023

DM9 domain containing protein (DM9CP) is a family of newly identified recognition receptors exiting in most organisms except plants and mammals. In the current study, to our knowledge, a novel DM9CP-5 (CgDM9CP-5) with two tandem DM9 repeats and high expression level in gill was identified from the Pacific oyster, Crassostrea gigas. The deduced amino acid sequence of CgDM9CP-5 shared 62.1% identity with CgDM9CP-1 from C. gigas, and 47.8% identity with OeFAMeT from Ostrea edulis. The recombinant CgDM9CP-5 (rCgDM9CP-5) was able to bind d-mannose, LPS, peptidoglycan, and polyinosinic-polycytidylic acid, as well as fungi Pichia pastoris, Gram-negative bacteria Escherichia coli and Vibrio splendidus, and Gram-positive bacteria Staphylococcus aureus. The mRNA transcript of CgDM9CP-5 was highly expressed in gill, and its protein was mainly distributed in gill mucus. After the stimulations with V. splendidus and mannose, mRNA expression of CgDM9CP-5 in oyster gill was significantly upregulated and reached the peak level at 6 and 24 h, which was 13.58-fold (p < 0.05) and 14.01-fold (p < 0.05) of that in the control group, respectively. CgDM9CP-5 was able to bind CgIntegrin both in vivo and in vitro. After CgDM9CP-5 or CgIntegrin was knocked down by RNA interference, the phosphorylation levels of JNK and P38 in the MAPK pathway decreased, and the expression levels of CgIL-17s (CgIL-17-3, -4, -5, and -6), Cg-Defh1, Cg-Defh2, and CgMolluscidin were significantly downregulated. These results suggested that there was a pathway of DM9CP-5-Integrin-MAPK mediated by CgDM9CP-5 to regulate the release of proinflammatory factors and defensins in C. gigas.
Cutting Edge: Hyperinflammatory Monocytes Expressing CD56 Abound in Severe COVID-19 Patients
Authors: Campana Stefania , De Pasquale Claudia , Sidoti Migliore Giacomo , Pezzino Gaetana , Cavaliere Riccardo , Venanzi Rullo Emmanuele , Nunnari Giuseppe , Caramori Gaetano , David Antonio , Bonaccorsi Irene , Pollicino Teresa , Carrega Paolo , Ferlazzo Guido
Published: January 2023

Proinflammatory monocytes play a preponderant role in the development of a cytokine storm causing fatal consequences in coronavirus disease 2019 (COVID-19) patients, highlighting the importance of analyzing in more detail monocyte distribution in these patients. In this study, we identified an atypical monocyte subpopulation expressing CD56 molecules that showed a low level of HLA-DR and high level of l-selectin. They released higher amounts of TNF-α and IL-6 and expressed genes associated with an excessive inflammatory process. Remarkably, the frequency of CD56+ monocytes inversely correlated with that of CD16+ monocytes and a high CD56+/CD16+monocyte ratio was associated with both disease severity and mortality, as well as with serum concentration of type I IFN, a factor able to induce the appearance of CD56+ monocytes. In conclusion, severe COVID-19 is characterized by the abundance of hyperinflammatory CD56+ monocytes, which could represent a novel marker with prognostic significance and, possibly, a therapeutic target for controlling the inflammatory process occurring during COVID-19.
MicroRNA miR-722 Inhibits Cyprinid Herpesvirus 3 Replication via Targeting the Viral Immune Evasion Protein ORF89, Which Negatively Regulates IFN by Degrading IRF3
Authors: Zhang Chi , Liu An-Qi , Zhang Chu , Liu Lan-Hao , Su Jianguo , Zhang Yong-An , Tu Jiagang
Published: January 2023

Cyprinid herpesvirus 3 (CyHV-3) has caused severe economic losses to carp culture, but its pathogenicity is far from clear. Our previous study has revealed that microRNA (miR)-722 was upregulated during CyHV-3 infection, indicating that miR-722 might play an important role in CyHV-3 replication. In this study, we found that overexpression of miR-722 inhibited CyHV-3 replication and promoted IFN expression. The putative target gene of miR-722 was searched over the CyHV-3 genome, and ORF89 was identified and validated as a target gene of miR-722. Overexpression of ORF89 markedly reduced the expression of IFN and IFN-stimulated genes. Mechanistically, ORF89 interacted with and degraded IFN regulatory factor 3 (IRF3), and inhibited the entry of IRF3 into the nucleus by suppressing the dimerization of IRF3. Moreover, ORF89-mediated suppression of IFN expression could be restored by adding miR-722. To our knowledge, our findings confirm a novel virus–host combat, in which CyHV-3 evades host antiviral immunity by its ORF89 protein, whereas host miR-722, upregulated on CyHV-3 infection, targets ORF89 to impede CyHV-3 replication.
ICAM-1 Abundance Is Increased in Pancreatic Islets of Hyperglycemic Female NOD Mice and Is Rapidly Upregulated by NF-κB in Pancreatic β-Cells
Authors: Martin Thomas M , Burke Susan J , Batdorf Heidi M , Burk David H , Ghosh Sujoy , Dupuy Samuel D , Karlstad Michael D , Collier J Jason
Published: January 2023

Type 1 diabetes (T1D) is classified as an autoimmune disease where pancreatic β-cells are specifically targeted by cells of the immune system. The molecular mechanisms underlying this process are not completely understood. Herein, we identified that the Icam1 gene and ICAM-1 protein were selectively elevated in female NOD mice relative to male mice, fitting with the sexual dimorphism of diabetes onset in this key mouse model of T1D. In addition, ICAM-1 abundance was greater in hyperglycemic female NOD mice than in age-matched normoglycemic female NOD mice. Moreover, we discovered that the Icam1 gene was rapidly upregulated in response to IL-1β in mouse, rat, and human islets and in 832/13 rat insulinoma cells. This early temporal genetic regulation requires key components of the NF-κB pathway and was associated with rapid recruitment of the p65 transcriptional subunit of NF-κB to corresponding κB elements within the Icam1 gene promoter. In addition, RNA polymerase II recruitment to the Icam1 gene promoter in response to IL-1β was consistent with p65 occupancy at κB elements, histone chemical modifications, and increased mRNA abundance. Thus, we conclude that β-cells undergo rapid genetic reprogramming by IL-1β to enhance expression of the Icam1 gene and that elevations in ICAM-1 are associated with hyperglycemia in NOD mice. These findings are highly relevant to, and highlight the importance of, pancreatic β-cell communication with the immune system. Collectively, these observations reveal a portion of the complex molecular events associated with onset and progression of T1D.
Mass Spectrometry Defines Lysophospholipids as Ligands for Chicken MHCY Class I Molecules
Authors: Gugiu Gabriel B , Goto Ronald M , Bhattacharya Supriyo , Delgado Melissa K , Dalton Jennifer , Balendiran Vaishnavi , Miller Marcia M
Published: January 2023

Chicken (Gallus gallus) MHCY class I molecules are highly polymorphic yet substantially different from polymorphic MHC class I molecules that bind peptide Ags. The binding grooves in MHCY class I molecules are hydrophobic and too narrow to accommodate peptides. An earlier structural study suggested that ligands for MHCY class I might be lipids, but the contents of the groove were not clearly identified. In this study, lysophospholipids have been identified by mass spectrometry as bound in two MHCY class I isoforms that differ substantially in sequence. The two isoforms, YF1*7.1 and YF1*RJF34, differ by 35 aa in the α1 and α2 domains that form the MHC class I ligand binding groove. Lyso-phosphatidylethanolamine (lyso-PE) 18:1 was the dominant lipid identified in YF1*7.1 and YF1*RJF34 expressed as recombinant molecules and renatured with β2-microglobulin in the presence of a total lipid extract from Escherichia coli. Less frequently detected were lyso-PE 17:1, lyso-PE 16:1, and lysophosphatidylglycerols 17:1 and 16:0. These data provide evidence that lysophospholipids are candidate ligands for MHCY class I molecules. Finding that MHCY class I isoforms differing substantially in sequence bind the same array of lysophospholipids indicates that the amino acid polymorphism that distinguishes MHCY class I molecules is not key in defining ligand specificity. The polymorphic positions lie mostly away from the binding groove and might define specificity in interactions of MHCY class I molecules with receptors that are presently unidentified. MHCY class I molecules are distinctive in bound ligand and in display of polymorphic residues.
Toward Understanding How Staphylococcal Protein A Inhibits IgG-Mediated Phagocytosis
Authors: Cruz Ana Rita , Bentlage Arthur E H , Blonk Robin , de Haas Carla J C , Aerts Piet C , Scheepmaker Lisette M , Bouwmeester Inge G , Lux Anja , van Strijp Jos A G , Nimmerjahn Falk , van Kessel Kok P M , Vidarsson Gestur , Rooijakkers Suzan H M
Published: January 2023

IgG molecules are crucial for the human immune response against bacterial infections. IgGs can trigger phagocytosis by innate immune cells, like neutrophils. To do so, IgGs should bind to the bacterial surface via their variable Fab regions and interact with Fcγ receptors and complement C1 via the constant Fc domain. C1 binding to IgG-labeled bacteria activates the complement cascade, which results in bacterial decoration with C3-derived molecules that are recognized by complement receptors on neutrophils. Next to FcγRs and complement receptors on the membrane, neutrophils also express the intracellular neonatal Fc receptor (FcRn). We previously reported that staphylococcal protein A (SpA), a key immune-evasion protein of Staphylococcus aureus, potently blocks IgG-mediated complement activation and killing of S. aureus by interfering with IgG hexamer formation. SpA is also known to block IgG-mediated phagocytosis in absence of complement, but the mechanism behind it remains unclear. In this study, we demonstrate that SpA blocks IgG-mediated phagocytosis and killing of S. aureus and that it inhibits the interaction of IgGs with FcγRs (FcγRIIa and FcγRIIIb, but not FcγRI) and FcRn. Furthermore, our data show that multiple SpA domains are needed to effectively block IgG1-mediated phagocytosis. This provides a rationale for the fact that SpA from S. aureus contains four to five repeats. Taken together, our study elucidates the molecular mechanism by which SpA blocks IgG-mediated phagocytosis and supports the idea that in addition to FcγRs, the intracellular FcRn is also prevented from binding IgG by SpA.
The Absence of IL-12Rβ2 Expression on Recipient Nonhematopoietic Cells Diminishes Acute Graft-versus-Host Disease in the Gastrointestinal Tract
Authors: Bastian David , Sui Xiaohui , Choi Hee-Jin , Wu Yongxia , Tian Linlu , Yang Kaipo , Liu Chen , Liu Yuejun , Yu Xue-Zhong
Published: January 2023

The gastrointestinal (GI) tract is a frequent target organ in acute graft-versus-host disease (aGVHD), which can determine the morbidity and nonrelapse mortality after allogeneic hematopoietic cell transplantation (allo-HCT). Donor T cells recognize allogeneic Ags presented by host APCs, proliferate, and differentiate into Th1 and Th17 cells that drive GVHD pathogenesis. IL-12 has been shown to play an important role in amplifying the allogeneic response in preclinical and clinical studies. This study demonstrates that IL-12Rβ2 expression on recipient nonhematopoietic cells is required for optimal development of aGVHD in murine models of allo-HCT. aGVHD attenuation by genetic depletion of IL-12R signaling is associated with reduced MHC class II expression by intestinal epithelial cells and maintenance of intestinal integrity. We verified IL-12Rβ2 expression on activated T cells and in the GI tract. This study, to our knowledge, reveals a novel function of IL-12Rβ2 in GVHD pathogenesis and suggests that selectively targeting IL-12Rβ2 on host nonhematopoietic cells may preserve the GI tract after allo-HCT.
Arid5a Mediates an IL-17–Dependent Pathway That Drives Autoimmunity but Not Antifungal Host Defense
Authors: Taylor Tiffany C , Li Yang , Li De-Dong , Majumder Saikat , McGeachy Mandy J , Biswas Partha S , Gingras Sebastien , Gaffen Sarah L
Published: January 2023

IL-17 contributes to the pathogenesis of certain autoimmune diseases, but conversely is essential for host defense against fungi. Ab-based biologic drugs that neutralize IL-17 are effective in autoimmunity but can be accompanied by adverse side effects. Candida albicans is a commensal fungus that is the primary causative agent of oropharyngeal and disseminated candidiasis. Defects in IL-17 signaling cause susceptibility to candidiasis in mice and humans. A key facet of IL-17 receptor signaling involves RNA-binding proteins, which orchestrate the fate of target mRNA transcripts. In tissue culture models we showed that the RNA-binding protein AT-rich interaction domain 5A (Arid5a) promotes the stability and/or translation of multiple IL-17–dependent mRNAs. Moreover, during oropharyngeal candidiasis, Arid5a is elevated within the oral mucosa in an IL-17–dependent manner. However, the contribution of Arid5a to IL-17–driven events in vivo is poorly defined. In this study, we used CRISPR-Cas9 to generate mice lacking Arid5a. Arid5a−/− mice were fully resistant to experimental autoimmune encephalomyelitis, an autoimmune setting in which IL-17 signaling drives pathology. Surprisingly, Arid5a−/− mice were resistant to oropharyngeal candidiasis and systemic candidiasis, similar to immunocompetent wild-type mice and contrasting with mice defective in IL-17 signaling. Therefore, Arid5a-dependent signals mediate pathology in autoimmunity and yet are not required for immunity to candidiasis, indicating that selective targeting of IL-17 signaling pathway components may be a viable strategy for development of therapeutics that spare IL-17–driven host defense.
RNA-Editing Enzyme ADAR1 p150 Isoform Is Critical for Germinal Center B Cell Response
Authors: Li Yuxing , Ruan Gui-Xin , Chen Wenjing , Huang Hengjun , Zhang Rui , Wang Jing , Li Yan , Xu Shengli , Ou Xijun
Published: January 2023

Adenosine deaminase acting on RNA (ADAR)1 is the principal enzyme for adenosine-to-inosine editing, an RNA modification–avoiding cytosolic nucleic acid sensor’s activation triggered by endogenous dsRNAs. Two ADAR1 isoforms exist in mammals, a longer IFN-inducible and mainly cytoplasm-localized p150 isoform and a shorter constitutively expressed and primarily nucleus-localized p110 isoform. Studies of ADAR1 mutant mice have demonstrated that ADAR1 is essential for multiple physiological processes, including embryonic development, innate immune response, and B and T lymphocyte development. However, it remained unknown whether ADAR1 plays a role in the humoral immune response. In this study, we conditionally delete Adar1 in activated B cells and show that ADAR1-deficient mice have a defective T cell–dependent Ab response and diminished germinal center (GC) B cells. Using various double mutant mice concurrently deficient in ADAR1 and different downstream dsRNA sensors, we demonstrate that ADAR1 regulates the GC response by preventing hyperactivation of the melanoma differentiation-associated protein 5 (MDA5) but not the protein kinase R or RNase L pathway. We also show that p150 is exclusively responsible for ADAR1’s function in the GC response, and the p110 isoform cannot substitute for the p150’s role, even when p110 is constitutively expressed in the cytoplasm. We further demonstrated that the dsRNA-binding but not the RNA-editing activity is required for ADAR1’s function in the GC response. Thus, our data suggest that the ADAR1 p150 isoform plays a crucial role in regulating the GC B cell response.
TRIM21 Regulates Virus-Induced Cell Pyroptosis through Polyubiquitination of ISG12a
Authors: Guo Mengmeng , Cao Wenyan , Chen Shengwen , Tian Renyun , Xue Binbin , Wang Luoling , Liu Qian , Deng Rilin , Wang Xintao , Wang Zhenghao , Zhang Yingdan , Yang Di , Zuo Chaohui , Li Guangdi , Tang Songqing , Zhu Haizhen
Published: January 2023

Pyroptosis is a form of regulated cell death mediated by the gasdermin protein family. During virus infection, cell pyroptosis restricts viral replication. The mechanisms of the tripartite motif (TRIM) protein family and IFN-stimulated genes (ISGs) against viruses have been studied. The role of TRIMs and ISGs in pyroptosis remains unclear. In this study, we show that TRIM21 interacts with ISG12a in viral infection and facilitates its translocation into the mitochondria by promoting its ubiquitination, thereby causing caspase 3 activation. Gasdermin E (GSDME) is specifically cleaved by caspase 3 upon viral infection, releasing the GSDME N-terminal domain, perforating the cell membrane, and causing cell pyroptosis. Our study uncovers a new mechanism of TRIM21 and ISG12a in regulating virus-induced cell pyroptosis.
FABP5 Deficiency Impaired Macrophage Inflammation by Regulating AMPK/NF-κB Signaling Pathway
Authors: Hou Yangxiao , Wei Dong , Bossila Elhusseny A , Zhang Zhaoqi , Li Sihong , Bao Jiaming , Xu Huawen , Zhang Lianfeng , Zhao Yong
Published: January 2023

Fatty acid binding protein 5 (FABP5) is mainly involved in the uptake, transport, and metabolism of fatty acid in the cytoplasm, and its role in immune cells has been recognized in recent years. However, the role of FABP5 in macrophage inflammation and its underlying mechanisms were not fully addressed. In our study, the acute liver injury and sepsis mouse models were induced by i.p. injection of LPS and cecal contents, respectively. Oleic acid (0.6 g/kg) was injected four times by intragastric administration every week, and this lasted for 1 wk before the LPS or cecal content challenge. We found that myeloid-specific deletion of FABP5 mitigated LPS-induced acute liver injury with reduced mortality of mice, histological liver damage, alanine aminotransferase, and proinflammatory factor levels. Metabolic analysis showed that FABP5 deletion increased the intracellular unsaturated fatty acids, especially oleic acid, in LPS-induced macrophages. The addition of oleic acid also decreased LPS-stimulated macrophage inflammation in vitro and reduced acute liver injury in LPS-induced or cecal content–induced sepsis mice. RNA-sequencing and molecular mechanism studies showed that FABP5 deletion or oleic acid supplementation increased the AMP/ATP ratio and AMP-activated protein kinase (AMPK) activation and inhibited the NF-κB pathway during the inflammatory response to LPS stimulation of macrophages. Inhibiting AMPK activation or expression by chemical or genetic approaches significantly rescued the decreased NF-κB signaling pathway and inflammatory response in LPS-treated FABP5-knockout macrophages. Our present study indicated that inhibiting FABP5 or supplementation of oleic acid might be used for the treatment of sepsis-caused acute liver injury.
Molecular and Structural Basis of Receptor Binding and Signaling of a Fish Type I IFN with Three Disulfide Bonds
Authors: Chen Jingjie , Guan Yanyun , Guan Hongxin , Mu Yinnan , Ding Yang , Zou Jun , Ouyang Songying , Chen Xinhua
Published: January 2023

In mammals, type I IFNs, which commonly contain one or two disulfide bonds, activate the JAK-STAT signaling pathway through binding to the common cell surface receptor formed by IFN-α/β receptor (IFNAR)1 and IFNAR2 subunits. Although type I IFNs are also known to be essential for antiviral defense in teleost fish, very little is known about mechanisms underlying the recognition of fish type I IFNs by associated receptors. In this study, we demonstrate that a type I IFN of large yellow croaker Larimichthys crocea (LcIFNi), belonging to a new subgroup of fish type I IFNs, triggers antiviral response via the conserved JAK-STAT pathway through stable binding with a heterodimeric receptor comprising subunits LcCRFB5 and LcCRFB2. LcIFNi binds to LcCRFB5 with a much higher affinity than to LcCRFB2. Furthermore, we determined the crystal structure of LcIFNi at a 1.39 Å resolution. The high-resolution structure is, to our knowledge, the first reported structure of a type I IFN with three disulfide bonds, all of which were found to be indispensable for folding and stability of LcIFNi. Using structural analysis, mutagenesis, and biochemical assays, we identified key LcIFNi residues involved in receptor interaction and proposed a structural model of LcIFNi bound to the LcCRFB2–LcCRFB5 receptor. The results show that LcIFNi–LcCRFB2 exhibits a similar binding pattern to human IFN-ω–IFNAR2, whereas the binding pattern of LcIFNi–LcCRFB5 is quite different from that of IFN-ω–IFNAR1. Altogether, our findings reveal the structural basis for receptor interaction and signaling of a type I IFN with three disulfide bonds and provide new insights into the mechanisms underlying type I IFN recognition in teleosts.
Dietary Cholesterol Metabolite Regulation of Tissue Immune Cell Development and Function
Authors: Frascoli Michela , Reboldi Andrea , Kang Joonsoo
Published: January 2023

Obesity is considered the primary environmental factor associated with morbidity and severity of wide-ranging inflammatory disorders. The molecular mechanism linking high-fat or cholesterol diet to imbalances in immune responses, beyond the increased production of generic inflammatory factors, is just beginning to emerge. Diet cholesterol by-products are now known to regulate function and migration of diverse immune cell subsets in tissues. The hydroxylated metabolites of cholesterol oxysterols as central regulators of immune cell positioning in lymphoid and mucocutaneous tissues is the focus of this review. Dedicated immunocyte cell surface receptors sense spatially distributed oxysterol tissue depots to tune cell metabolism and function, to achieve the “right place at the right time” axiom of efficient tissue immunity.