bluuebunny/crossref_metadata_2025 huggingface dataset:
Intestinal IL-17R signaling controls liver inflammation by constraining microbiome induced TLR9 signaling and IL-18 production
Authors: Castillo Patricia , Kumar Pawan , Hand Timothy W , Kolls Jay K
Published: January 2023

Th17 cells and the intestinal microbiome have been linked to liver diseases such as autoimmune and viral hepatitis. Patients exhibit intestinal dysbiosis and increased bacterial translocation. In a mouse model of T cell mediated liver inflammation, concanavalin A (Con A) hepatitis, global Il17ra deficiency and gentamicin treatment are protective. Yet, it is unknown if IL-17 contributes to microbial dysbiosis and bacterial translocation in hepatitis, and how these factors together affect disease.

To study this, we generated intestinal epithelial-specific IL-17RA deficient mice (Il17rafl/flx villin cre+ mice). Absence of enteric IL-17R signaling induced commensal dysbiosis and increased intestinal Th17 cells and Il18. These mice were more susceptible to Con A, exhibiting worsened liver damage and lethality. Single cell RNA sequencing partnered with protein-based assays revealed elevated liver IL-18, IFNγ, and FasL in Il17rafl/flx villin cre+ mice, while cohousing and antibiotic studies implicated the microbiome in disease exacerbation. Il17rafl/flx villin cre+ mice also had increased liver CpG levels, which induced IL-18. Moreover, anti-IL18 ameliorated Con A hepatitis.

We propose that disrupted enteric IL-17 signaling promotes intestinal dysbiosis and translocation of CpG to the liver, stimulating TLR9 and IL-18 production. IL-18 then induces hepatic lymphocyte activation and cell death to worsen inflammation. Understanding the mechanism underlying exacerbated disease in Il17rafl/flx villin cre+ mice will elucidate the role of enteric Th17 and the microbiome in hepatitis, with broader implications on the effects of impaired intestinal immunity and subsequent release of microbial products seen in other diseases.
Sensing Fungal Dysbiosis by Gut-Resident CX3CR1+ Mononuclear Phagocytes Aggravates Allergic Airway Disease
Authors: Li Xin , Leonardi Irina , Semon Alexa , Doron Itai , Gao Iris H , Putzel Gregory Garbe’s , Kim Youngjun , Kabata Hiroki , Artis David , Fiers William D , Ramer-Tait Amanda E , ILIEV ILIYAN
Published: January 2023

Sensing of the gut microbiota, including fungi, regulates mucosal immunity. Whether fungal sensing in the gut can influence immunity at other body sites is unknown. Here we show that fluconazole-induced gut fungal dysbiosis has persistent effects on allergic airway disease in a house dust mite challenge model. Mice with a defined community of bacteria, but lacking intestinal fungi were not susceptible to fluconazole-induced dysbiosis, while colonization with a fungal mixture recapitulated the detrimental effects. Gut-resident mononuclear phagocytes (MNPs) expressing the fractalkine receptor CX3CR1 were essential for the effect of gut fungal dysbiosis on peripheral immunity. However, how mycobiota influence immunity in gut distal sites is not well understood. We developed protocols for gut-targeted depletion of phagocytes to investigate the influence of fungi on gut-lung crosstalk. Depletion of CX3CR1+ MNPs or selective inhibition of Syk signaling downstream of fungal sensing in these cells ameliorated lung allergy. These results indicate that disruption of intestinal fungal communities can mediate gut-lung-directed immune crosstalk and aggravate disease severity through fungal sensing by gut-resident CX3CR1+ MNPs.

Highlights

Gut fungal dysbiosis persistently aggravates allergic airway disease (AAD) in mice.

Gut colonization by commensal fungi is both required and sufficient to aggravate AAD in a mycobiome-free mouse model.

Intestinal CX3CR1+ mononuclear phagocytes (MNPs) are essential for the systemic effects of gut fungal dysbiosis on AAD.

Inhibition of Syk-mediated fungal sensing in intestinal CX3CR1+ MNPs ameliorates AAD.
IL-17 receptor and IL-22 receptor signaling in Citrrobacter rodentium infection
Authors: Matsunaga Yasuka , Wanek Alanna , Song Kejin , Flemington Cathy , Clark Trevon , Bitoun Jacob , Kolls Jay
Published: January 2023

Th17 cells producing cytokines, IL-17A and IL-22 are important key player in host defense. We have reported that intestinal epithelial expression of IL-17 receptors in gut epithelial cells control the development of microbiota. To study the roles of these receptors in response to an attaching and effacing pathogen we challenged Citobacter rodenium (C. rodentium) to intestinal epithelial specific IL-17RA or IL-22RA1 KO mice. Conditional deletion of Il17ra or Il22ra1 resulted in increased bacterial growth in the colon as well as enhanced dissemination to the spleen. It suggested both cytokine receptors are required for control of this infection. In addition this was supported by unbiased RNAseq analysis of the colonic epithelium that showed diminished expression of Pigr, Duox2 and Duoxa2 in the absence of IL-17RA signaling and reduced Bcl3 expression in the absence of IL-22ra1 expression. To elucidate whether production from epithelial cells effect bacteria killing we cultured crypt and stimulated IL-17A. It resulted in increasing Pigr, Duoxa2 and Duox2 were associated with increasing bacteria killing. Pigr binds dimeric IgA and translocation into lumen from lamina propria, while Duox2 and Duoxa2 make complex and produce hydrogen peroxide into lumen from apical epithelial cell membrane. These result suggested mucosal immunity to C. rodentium requires both IL-17RA and IL-22ra1 signaling and that IL-17RA regulates transcytosis of C. rodentium specific IgA as well as luminal hydrogen peroxide concentrations to control bacterial growth. In contrast, Il-22Ra1 regulates epithelial Bcl3 expression, which restrains TNFα and NF-κB signaling in the colonic epithelium.
Maternal Immunoglobulin A protects against the development of necrotizing enterocolitis in preterm infants
Authors: Hand Timothy W , Gopalakrishna Kathyayini , Macadangdang Benjamin R , Rogers Matthew , Tometich Justin T , Ji Junyi , Firek Brian A , Baker Robyn , Burr Ansen H.P. , Ma Congrong , Good Misty , Morowitz Michael
Published: January 2023

Neonates are particularly susceptible to infection by colonizing microbes, and mammals protect their offspring via antibodies secreted into breast milk. Necrotizing Enterocolitis(NEC) is a disease of preterm infants characterized by intestinal epithelial damage and inflammation associated with the microbiota. The incidence of NEC is significantly lower in infants fed with maternal milk, though the mechanisms underlying this protective benefit are not clear. Here, we show that maternal Immunoglobulin A (IgA) is an important factor in protection against NEC. Analysis of IgA-binding within fecal samples from preterm infants indicated that breast milk was the predominant source of IgA in the first month of life and that a relative decrease in the fraction of bacteria bound by IgA is associated with the development of NEC. Sequencing of IgA-bound and unbound bacteria revealed that NEC was associated with increasing domination of the IgA unbound microbiota by Enterobacteriaceae, prior to disease onset. Further, we confirmed that IgA is critical in preventing NEC in a murine model, where we demonstrate that pups reared by IgA deficient mothers are susceptible to disease despite exposure to maternal milk. This study illustrates the importance of maternal IgA in shaping the host-microbiota relationship of preterm neonates and provides evidence that IgA is a critical factor in maternal milk necessary for the prevention of NEC.
Indole-3-carbinol prevents murine colitis development via an IL-22-dependent mechanism that regulates anti-microbial peptides and mucus production
Authors: Busbee Philip Brandon , Alrafas Haider , Dopkins Nicholas , Nagarkatti Mitzi , Nagarkatti Prakash S
Published: January 2023

Colitis is an inflammatory bowel disease characterized by acute or chronic inflammation within the colon. In our previous work, we showed that indole-3-carbinol (I3C), a naturally-occurring plant product found in a number of cruciferous vegetables, was able to prevent development of colitis in murine models. I3C prevented colitis-associated microbial dysbiosis characterized by an overabundance of potentially pathogenic gram-negative bacteria (e.g. Bacteroides acidifaciens), in addition to increasing anti-inflammatory gram-positive butyrate-producing species (e.g. Roseburia). This increase in butyrate by I3C, validated by metabolomics analysis, leads to a decrease in the pro-inflammatory T cell response (Th17) and an increase in anti-inflammatory Tregs, which was confirmed by sodium butyrate supplementation experiments. In the current study, we further defined the mechanisms by which I3C was able to prevent colitis-associated microbial dysbiosis and dysregulation of gut microbial metabolome by evaluating expression of key intestinal regulatory genes in immune cells and colonic epithelial cells (CECs). Transcriptome analysis and PCR validation of CECs showed mucus production (e.g. Muc2) and colonic anti-microbial peptides (e.g. beta-defensins) were significantly increased after I3C. I3C was also shown to, in a butyrate-independent manner, increase expression of colonic bacterial defensive interleukin-22 (IL-22) by epigenetic regulation. Lastly, in vivo IL-22 neutralization experiments negated the beneficial effects of I3C, thus proving this cytokine was crucial in I3C-mediated prevention of colitis development and regulation of key intestinal regulatory components such as mucus and AMP production.
B cells produce IL-27 to upregulate PD-L1 expression and promote breast cancer development
Authors: Yan Hui , Viswanadhapalli Suryavathi , Chupp Daniel , Fernandez Maria , Wu Shuai , Wang Jingwei , Moroney Justin , Taylor Julia , John IM , Rivera Carlos , Luo Yiliao , Liu Junhao , Sareddy Gangadhara , Casali Paolo , Vadlamudi Ratna , Xu Zhenming
Published: January 2023

The host immune response can be corrupted by cancer cells during tumor progression. Here we showed that B lymphocytes played an important role in promoting murine breast cancer (BCa) development, as B cell-deficient mMT mice displayed virtually abrogated tumor growth from transplanted syngeneic ERa+ breast adenocarcinoma cells, concomitant with reduced PD-L1 expression and increased BCa cell death. In murine BCa tumors and tumor-associated lymph nodes, B cells were the major source of IL-27, a pleotropic cytokine with both pro- and anti-inflammatory functions. Deficiency in IL-27 expression specifically in B cells abolished the BCa tumor growth, showing that the effect of B cells in BCa development was mainly through IL-27 production, as likely induced by TLR ligands in the tumor microenvironment together with CD154 and IL-21 expressed by tumor-infiltrating CD4+ T cells. IL-27 treatment of BCa cells resulted in STAT1 and STAT3 phosphorylation, upregulated PD-L1 gene expression and enhanced cell growth. In addition, IL-27 induced activated B cells to express PD-L1 in vitro, consistent with high levels of PD-L1 expression by B cells in tumors. In humans, IL-27 receptor genes were highly expressed in the breast tissue, at levels comparable to those in lymphoid organs, and high IL-27 gene expression was associated with reduced survival in BCa patients, who displayed increased circulating IL-27 levels. Finally, IL-27 supported human BCa cell growth in the presence of tamoxifen, suggesting a role of IL-27 in the drug resistance. Thus, TLR-fueled IL-27 induction in B cells reinforces PD-L1 expression in the tumor environment to drive BCa cell growth and to promote generation of PD-L1hi B regulatory cells, leading to BCa tumor progression.
Precisely timed histone deacetylase inhibition creates a highly proliferative intratumoral CD8 T cell population and sensitizes tumors to checkpoint blockade
Authors: McCaw Tyler R , Liu Mingyong , Li Mei , Starenki Dmytro , Cooper Sara , Arend Rebecca , Forero Andres , Buchsbaum Donald , Randall Troy D
Published: January 2023

We previously showed that a class I histone deacetylase (HDAC) inhibitor, entinostat (ENT), dramatically improves CD8 T cell-mediated control of TS/A murine mammary tumors, but only when given in a precise window. If given too early ENT halts initial T cell activation and expansion and has no effect if given too late— once T cells start becoming dysfunctional. This led us to hypothesize that HDAC inhibition can maintain CD8 T cells in a less differentiated state that enhances effector and proliferative capacities. To test this, we first treated TS/A tumor-bearing mice with ENT and performed single cell RNA-sequencing on sorted tumor-infiltrating CD8 T cells. Relative to vehicle, ENT treated samples clustered separately and included a distinct, highly proliferative cluster of CD8 T cells, a finding corroborated by bulk RNA-sequencing and flow cytometry. Although TS/A tumors do not respond to anti-PD1 alone, ENT-induced changes in CD8 T cell phenotype sensitized tumors to PD1 blockade, leading to tumor rejection in most mice when both agents were timed appropriately. Patient tumors, however, have often passed this window to intervene; thus, we devised a tripartite strategy to “reset the clock”. Treating first with an oncolytic virus to liberate new antigens and stimulate new tumor-specific CD8 T cells, then with ENT and anti-PD1 at the right times significantly impaired growth of late-stage tumors and even led to rejection. Our findings show that precisely timed HDAC inhibition generates highly proliferative and strongly cytotoxic CD8 T cell populations whose functions can be further elaborated with correctly timed checkpoint blockade. Resetting the clock using a tumor-lytic strategy then renders this a potent immunotherapy platform.
Differences in T cell responses to Bordetella Pertussis in adults as a function of whole cell versus acellular childhood vaccination
Authors: da Silva Antunes Ricardo Filipe , Babor Mariana , Pomaznoy Mikhail , Khalil Natalie , Cortese Mario , Mentzer Alexander J , Seumois Gregory , Petro Christopher D , Purcell Lisa A , Vijayanand Pandurangan , Crotty Shane , Pulendran Bali , Peters Bjoern , Sette Alessandro
Published: January 2023

In the mid-1990s vaccine-related side effects prompted the replacement of the whole Pertussis (wP) by a new acellular Pertussis vaccine (aP). Unexpectedly, whooping cough cases have recently increased, and the switch to the aP vaccine suspected to underlie this rise in morbidity, despite the use of routine boosters. Here we compared the immune responses to aP boosters in children who received their initial doses with either wP or aP vaccines. The use of ex vivo AIM assays highlighted a Th2 vs Th1/Th17 differential polarization of PT-specific memory CD4+ T cells as a function of childhood vaccination. The nature of the differences was further investigated for memory subset composition, activation, exhaustion marker expression, and transcriptomic profiles. Remarkably, donors originally primed with aP were defective in their ex vivo capacity to expand memory cells following a booster aP immunization and less capable to proliferate in vitro in response to PT epitopes. By contrast antibody responses are boosted in both aP and wP cohorts following IgG subclass distribution. Most recently, we have assessed early immune responses followed aP boosting for the aP and wP cohorts using longitudinal transcriptomics data from PBMC. We have identified several modules of genes that are co-expressed; using cell frequency data from CyTOF we infer modules correlated with specific cell types. Furthermore, we identified certain modules that show differential expression profiles between the two cohorts and are mostly enriched in immune related functions. In conclusion, our data suggest that the original priming after birth with aP and wP vaccines induce different T cell phenotypes associated with long-term T cell polarization.
Differential infection of dendritic cell subsets by virus-based vaccine platforms
Authors: Chiale Carolina , Robek Michael D
Published: January 2023

The use of virus-based vectors as vaccine platforms represents an attractive alternative to immunization with non-replicating antigens. However, the precise mechanism by which many virus-based vaccine vectors prime the immune response remains unknown. Specific dendritic cell (DC) subtypes play key roles in priming immune responses and controlling virus replication, but their functions in generating protective immunity following vaccination with viral vectors are not well understood. We hypothesized that highly immunogenic viral vectors based on alphaviruses and rhabdoviruses infect DCs and promote direct antigen presentation that leads to the efficient induction of antigen-specific T cells. We found that distinct DC subtypes are differentially infected with specific virus-based vaccine vectors, and display differences in activation following infection. Mouse bone marrow-derived DCs that are produced by GM-CSF stimulation and resemble moDCs rapidly succumbed to infection. In contrast, DCs that are produced by Flt3L stimulation and resemble cDCs and pDCs were efficiently infected but survived. DC activation following virus infection was more pronounced in the Flt3L-derived DCs, and showed distinctive requirements for further virus replication. Although Flt3L-derived DCs produced significant amounts of type I IFN when infected with the vaccine platforms, the mechanism of DC infection and activation was independent of type I IFN receptor signaling. These results highlight the differential survival and activation of specific DC subsets following infection with viral vaccine vectors, and indicate potentially unique roles of DC subtypes in activating the immune response following immunization.
Regulatory Cell Therapy for Sjögren’s Syndrome with E. coli Colonization Factor Antigen I (CFA/I) Fimbriae
Authors: Akgul Ali , Nelson Andrew S , Maddaloni Massimo , Hoffman Carol , Ella Bhagyaraj , Abbott Jeffrey R , Nguyen Cuong , Pascual David W
Published: January 2023

Sjögren’s syndrome (SjS) is one of the most common autoimmune diseases impacting several million people every year in the US. SjS results from autoimmune attack of exocrine (salivary and lacrimal) glands, and women are nine times more likely to be affected than men. To date, no vaccine or therapeutic exists to cure SjS, and patients must rely on lifelong therapies to treat symptoms. Oral treatment with the adhesin from enterotoxigenic E. coli CFA/I fimbriae has been shown to protect against several autoimmune diseases in an antigen (Ag)-independent manner. Lactococcus lactis was recently adapted to express CFA/I (LL-CFA/I), and found effective in suppressing inflammation by the induction of infectious tolerance via Ag-specific regulatory T cells (Tregs) producing IL-10 and TGF-β. We hypothesized that in the spontaneous, genetically defined model for SjS in C57BL/6.NOD-Aec1Aec2 mice, oral treatments with LL-CFA/I would offset the development of inflammatory T cells via Treg induction. To test this, 6 wk-old C57BL/6.NOD-Aec1Aec2 mice were orally dosed with LL-CFA/I and treated every 3 wks; control groups were given L. lactis vector or PBS. We found a significant recovery in salivary flow following treatment with LL-CFA/I by 28 wks of age. Treatment also significantly reduced the incidence of inflammatory infiltrates into the submandibular and lacrimal glands compared to PBS-treated mice. Treg analysis revealed a significant 4-fold increase in Foxp3+ Tregs expressing CD25, not CD39, and produced a 3-fold increase in IL-10. These data show that orally dosing with LL-CFA/I weakens or halts SjS progression via activation of Tregs. These studies will provide the basis for future testing in SjS patients. Work supported by NIH DE026450.
The inoculation of α-Galactosylceramide and HPV18-E1 protein promote a cytokines profile involved in the generation of central memory CD8 + T cells
Authors: Amador Alfredo , Gonzalez-Telona Ana Daniela , Lizano Marcela
Published: January 2023

CD8+ mediated immune response plays a major role in the clearance of Human Papillomavirus (HPV) infected cells. In the natural history of an HPV infection, the expression of HPV-E1 helicase during viral replication makes it a potential immunological target. Currently, there are no therapeutic vaccines that eliminate the lesions caused by HPV. Nevertheless, there are many studies on developing therapeutic vaccines against HPV. Vaccines and immunological therapies mostly use adjuvants, particularly α-galactocilceramide (α-GalCer) is an adjuvant that promotes a CD8+ T lymphocyte response mainly.

We demonstrate in previous work, that immunization of HPV type 18 E1 protein along with α-GalCer adjuvant elicited CD8 T cell-immune response, able to eliminate tumor cells expressing HPV18 E1 protein.

In this study, we analyzed the effect of α-GalCer in the generation of memory T CD8+ populations. We evaluated the serum levels of IL-15, IL-7, IL-4, IL-31, IL-2, TNF-α and the number of memory CD8+ T cells.

The serum levels of IL-15, IL-7, IL-4 and TNF- α are increase in mice inoculated with α-GalCer plus HPV18-E1 with respect to mice inoculated with α-GalCer or HPV18-E1 protein. These data correlate with the increase of central memory T CD8+ population in mice inoculated with α-GalCer plus HPV18-E1.

These data indicate that the administration of α-GalCer along with HPV18-E1 protein helps the generation of central memory CD8 + T lymphocytes. This adjuvant could be useful in the development of new therapeutic strategies to clearance HPV established infections.

INCAN: (017/076/17) (CEI/1076/17)
Transduction of retinoic acid-inducible gene 1 by Ebola virus-like particles enhances antigen-presentation
Authors: Martinez Osvaldo , Ngu Mee Kee Esther , Warneke Paul , burmeister Brock
Published: January 2023

Ebola virus (EBOV), a filovirus family member, is a highly pathogenic virus that causes Ebola hemorrhagic fever resulting in documented mortality rates in humans as high as 50%. EBOV virus-like particles (VLPs) are morphologically and biochemically similar to parental virus, yet because they lack a genome and cannot replicate, are safe enough to be used as vaccines. We hypothesize that addition of a constitutionally active retinoic acid-inducible gene 1 (RIG-I) would enhance the ability of the EBOV VLPs to induce antigen-presentation from infected antigen-presenting cells. Expression of EBOV VP40 in 293T cells induces the spontaneous production of VLPs into the media supernatant and if expressed with EBOV glycoprotein (GP), will produce VLPs studded with the attachment GP. Recombinant chimeric constitutively active (ca)RIG-I-VP40 matrix and a nonfunctional mutant L58A (mu)RIG-I-VP40 matrix genes were constructed to produce VLPs containing constitutively active and nonfunctional RIG-I, respectively. Supernatant from 293Ts transfected with caRIG-I-VP40, muRIG-I-VP40 or VP40 along with GP expression plasmids were tested for the presence of VLPs. Western blotting of purified VLPs confirmed the presence of RIG-I in caRIG-I-VP40 and muRIG-I-VP40, but not VP40 containing VLPs. Monocyte-like THP-1 reporter cells were treated with nothing, VP40+GP, caRIG-I-VP40+GP and muRIG-I-VP40+GP VLPs as well as LPS control and tested for induction of interferon signaling. CaRIG-I containing, but not muRIG-I containing VLPs induced interferon signaling as well as greater MHC I upregulation.
Daidzein and its analogs reactivate HIV replication from latently infected CD4 T cells
Authors: Liu Li , Liang Jianguo , Lu Xiaofan , Chen Zhiwei
Published: January 2023

Human immunodeficiency virus (HIV) persists in a latent form in infected individuals under antiretroviral therapy (ART) through integrating proviral DNA into the genome of host immune cells. In these latently infected cells, the transcription of HIV proviral DNA remains silent, which enables the infected cells to escape from the killing by anti-viral CD8+ T cells. One of the recent new therapeutic strategies for HIV eradication aims to reactivate HIV production in the latently infected cells, so that these cells could be detected and eliminated by the immune system. Here we show that small molecules isolated from Spatholobus suberectus Dunn., Daidzein and its analogues, reactivate HIV replication from latency. Daidzein reactivates HIV replication in latently infected CD4 T cell lines J lat and A72 dose dependently, but not monocytes. Moreover, by using cells that contain an integrated HIV LTR linked to the luciferase reporter gene, we found that Daidzein activates HIV replication at the level of transcription. This effect is through Akt signaling pathway, and do not induce T cell activation. Structure-activity relationships (SARs) analysis of Daidzein and its five analogues revealed that three analogues, Daidzin, Glycitein and Glycitin, reactivate HIV replication without obvious cell toxicity observed, and 4′-hydroxyisoflavone is their bio functional core structure. Therefore, Daidzein and its analogues have potential to be modified to improve their potency and developed into significant components for HIV therapy.
HIV-induced production of CCL2 may promote rapid seeding of the latent HIV reservoir
Authors: Packard Thomas Alexander , Herzig Eytan , Luo Xiaoyu , Egedal Johanne H , Grimmett Zachary W , Hasenkrug Kim J , Roan Nadia , Greene Warner C
Published: January 2023

Seeding of the latent HIV reservoir occurs quickly within hours to a few days after initial viral infection. It is the persistence of virus within this reservoir that thwarts an HIV cure and necessitates life-long antiretroviral therapy (ART). Central memory CD4 T cells harboring latent HIV proviruses form a major part of this reservoir. How the virus establishes latency so rapidly within this subset of CD4 T cells remains unknown.

We find that HIV infection leads to rapid production of the CCL2 chemokine in human lymphoid CD4 T cells. Cas9RNP-mediated depletion of IFI16 and STING genes significantly reduces production of CCL2 occurring in response to HIV. IFI16 was previously implicated as the key sensor triggering CD4 T cell pyroptosis. Thus, infection may also stimulate a second pathway of IFI16-mediated signaling involving STING culminating in CCL2 production. CCL2 functions a potent chemoattractant for cells expressing CCR2. Lymphoid CCR2+ CD4 T cells express the CCR5 co-receptor for HIV and markers of central memory. Additionally, these cells express many markers previously associated with the latent HIV reservoir. CCR2+ CD4 T cells are highly permissive to latent and productive infection by both R5 and X4-tropic viruses. When CCR2+ CD4 T cells are purified from HIV-infected individuals on suppressive ART, these cells are enriched up to 75-fold for HIV proviral DNA compared to naïve cells (n=10, p=0.0078).

Together, our findings support a model where HIV infection triggers the production of CCL2, leading to directed recruitment and infection of CCR2+ CD4 central memory T cells. Thus, HIV hijacks a component of the innate immune response to rapidly recruit precisely those target cells that ensure its long term persistence.
Impaired T cell activation resulting from IL-33 induced myeloid derived suppressor cells and IL-10 secreting Tregs is a major determinant of susceptibility to HSV1 infection of the central nervous system
Authors: Chandran Ramakrishna , Mendonca Stacee , Cantin E M
Published: January 2023

Resolution of viral infections of the central nervous system (CNS) is contingent on eliciting protective immunity with minimal pathology. C57BL/6 (B6) mice elucidate efficient innate and anti-viral T cell responses which render them resistant to infection with a virulent HSV 17+ virus strain. Virus replication is limited to ganglia and brainstem. However, susceptible mice such as 129S6 and BALB/c mice harbor increased virus in the CNS. These mice show increased myeloid derived suppressor cells (MDSC) and Tregs along with associated cytokines such as G-CSF, GM-CSF, IL-10 and IL-33 resulting in reduced numbers of virus specific CD8 T cells and effector T cells with increased expression of suppressive markers such as PD-1 and LAG-3. This impaired immune response resulted in disease and mortality. Neutralization of checkpoint inhibitors or immunosuppressive cytokines and depletion of myeloid cells or Tregs did not alter susceptibility rates. However, reducing expression of IL-33 in the CNS had a beneficial effect on T cell activation and the overall outcome of disease. Intriguingly, immunizing susceptible mice with a HSV thymidine kinase mutant virus incapable of infecting CNS induced memory T cells that suppressed MDSCs and protected from a subsequent challenge with HSV 17+. Interestingly IFNg, B cells or antibody were not essential but instead revealed the importance of the T cell myeloid interaction in determining disease outcomes. IL-33 plays critical roles as an immune-modulatory Th2 cytokine regulating immune-mediated diseases and tissue function especially in the CNS and as an alarmin in response to pathogens. This study shows that tempering IL-33 responses is essential to predicting improved outcomes to CNS viral infections.
Vaccinia virus E5 is a dominant inhibitor of the cytosolic DNA sensor cGAS
Authors: Yang Ning , Wang Yi , Shuman Stewart , Deng Liang
Published: January 2023

The cytosolic DNA sensor cGAS plays an important role in detecting viral nucleic acid, which leads to type I IFN production. We have previously shown that infection with conventional dendritic cells with modified vaccinia virus Ankara (MVA), a highly attenuated vaccinia strain (VACV), induces IFN production via a cGAS/STING-dependent mechanism. However, MVA or VACV infection triggers cGAS degradation and its mechanism is still unknown. VACV is a cytoplasmic DNA virus, which encodes more than 200 genes. In this study, we screened 70 vaccinia viral early genes for inhibition of cGAS/STING pathway using a dual luciferase system. We found that vaccinia E5 is a dominant inhibitor of cGAS and is the key protein mediating cGAS degradation. MVAΔE5R induces much higher levels of type I IFN than MVA in multiple cell types, including bone marrow derived dendritic cells (BMDC), bone marrow-derived macrophages (BMDM), and skin primary fibroblasts. MVAΔE5R-mediated type I IFN production is dependent on cGAS. Furthermore, MVAΔE5R gains replication capability in cGAS−/− skin fibroblasts. As a vaccine vector, skin scarification or intradermal vaccination with MVAΔE5R-OVA leads to much higher OVA-specific CD8+ T cell responses than MVA-OVA in vivo. Intratumoral injection of MVAΔE5R leads to stronger anti-tumor immune responses and better survival compared with MVA. Finally, in an intranasal infection model, VACVΔE5R is at least 100-fold attenuated compared with WT VACV. Taken together, our results provide strong evidence that E5 is a key viral virulence factor targeting the cytosolic DNA sensor cGAS and thereby inhibits type I IFN production.
Respiratory syncytial virus disease severity is associated with distinct CD8+ T cell profiles
Authors: Siefker David , Vu Luan , You Dahui , McBride Andrew , Taylor Ryleigh , Jones Tamekia L. , DeVincenzo John P. , Cormier Stephania
Published: January 2023

Most infants are infected with RSV by their second birthday. Despite known risk factors such as prematurity or congenital heart disease, most hospitalized infants are previously healthy. The mechanisms underlying the immunopathology of severe disease is incompletely understood. Our objective was to compare T cell profiles in nasal aspirates between infants with moderate and severe RSV disease hospitalized without immune deficiencies or congenital heart disease, during 2014–2017. Infants requiring intensive care were categorized as having severe disease, while infants maintained in the general ward were categorized as having moderate disease. The T cell profile in nasal aspirates collected 7 days after hospitalization (median 11 days after first symptom onset) was determined by flow cytometry. Nasal aspirates from patients with severe disease were characterized by a higher proportion of CD8+ T cells expressing IL-4 (Tc2) (P=0.024) and a lower proportion of CD8+ T cells expressing IFNγ (Tc1) (P=0.019) compared to patients with moderate disease. Patients with reduced frequencies of Tc1, Tc17, or Th17 cells had shorter durations of hospitalization (P=0.0023, P=0.0188, P=0.0437, respectively). Additionally, patients with severe disease had lower concentrations of IL-17 in nasal aspirates compared to patients with moderate disease (P=0.0133). These results suggest that RSV disease severity is associated with a specific T cell profile, and that Tc2 cells may play an underappreciated role in the development of RSV immunopathology.
Relative timing of type I interferon response and virus replication determines disease outcome during respiratory virus infection
Authors: Channappanavar Rudragouda , Fehr Anthony , Zheng Jian , Abrahante Juan , Mack Matthias , Meyerholz David , Perlman Stanley
Published: January 2023

Type 1 interferons (IFN-I) generally protect mammalian hosts from virus infections, but in some cases, IFN-I is pathogenic. Because IFN-I is protective, it is commonly used to treat virus infections for which no specific approved drug or vaccine is available. The Middle East respiratory syndrome-coronavirus (MERS-CoV) is such an infection, yet little is known about the role of IFN-I in this setting. Here we show that IFN-I signaling is protective during MERS-CoV infection. Blocking IFN-I results in delays in virus clearance and impaired MERS-CoV-specific T cell responses. Administration of IFN-I within 1 day post infection (dpi) protected mice from lethal infection. In marked contrast, delayed IFN-I enhanced the inflammatory response, converting a sublethal to a fatal infection. Together, these results suggest the relative timing of IFN-I signaling and maximal virus replication is key in determining the disease outcome and that IFNab should be used cautiously to treat virus infections.
Lung-draining lymph-node-resident memory CD8 T cells mediate local protective immunity
Authors: Anthony Scott M , Van Braeckel-Budimir Natalija , Moioffer Steven , Hartwig Stacey M , Varga Steven M , Badovinac Vladimir P , Harty John T
Published: January 2023

Recent studies indicate tissue-resident memory CD8 T cells (Trms) form at both the sites of initial infection and in lymphoid tissues upon resolution of systemic or localized viral infections. We examined if lymph node Trms were also generated in lung-draining mediastinal lymph nodes (mLN) during intranasal infections. Both transgenic P14 cells and endogenous influenza-specific CD69+/CD103+ Trms were detected in lung-draining mLN but absent in non-draining LNs after intranasal infection with recombinant influenza or vaccinia viruses. Similar to lung Trms, which wane over time, these primary memory (1M) mLN Trms had limited persistence with continual contraction throughout the memory phase. As influenza is a seasonal virus, and we previously observed that repeated antigen exposure extended the durability of lung Trms, we investigated the effects of multiple antigen exposures on the mLN Trm population. Similar to their lung Trm-counterparts, the lung-draining LN Trms derived from repeatedly stimulated (4M) memory CD8 T cells exhibited extended survival, down-regulation of Eomesodermin and the shared IL-2/15Rβ CD122 compared to 1M LN Trms. During parabiosis, both 1M and 4M mLN Trms largely maintained residence within their respective hosts. Importantly, hosts containing 4M P14 mLN Trms provided superior early protection against LCMV challenge in the mLN compared to 1M hosts, demonstrating the functional capacity for mLN Trms to contribute to protective anti-viral immune responses. Overall, these data indicate that lung-draining lymph node Trms co-express CD69 and CD103, largely maintain LN-tissue residency and can contribute to subsequent viral infections.
The role of Exchange Proteins Directly Activated by cAMP in Respiratory Syncytial Virus-induced Pulmonary Pathogenesis
Authors: Choi Eun-Jin , Ren Junping , Wu Wenzhe , Garofalo Roberto P , Bao Xiaoyong
Published: January 2023

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection in children, the elderly, and immunocompromised patients, representing a huge medical burden globally. However, a specific and effective treatment for RSV infection is not currently available, largely due to poorly understood host-RSV interaction. To facilitate the development of preventive and therapeutic approaches, continuous identification of novel host and viral molecules involved in host responses to RSV and associated pathogenesis is greatly needed. We previously have shown that modulation of cellular EPAC2 protein significantly impacts cellular responses and viral replication in both upper and lower airway epithelial cell models of RSV infection, suggesting EPAC2 could be a promising therapeutic target for RSV infection. In this study, mice deficient in EPAC2 was used to investigate the contribution of EPAC2 to pulmonary responses to RSV infection. EPAC2−/− mice showed less body weight loss and better respiratory function than wildtype mice after RSV infection, suggesting attenuated disease severity. RSV-induced cytokines/chemokines and neutrophil cells in bronchoalveolar lavage fluid (BALF) were also significantly less in EPAC2−/− mice. In addition, histopathology studies revealed less pathogenesis by EPAC2 deletion. Taken together, our data indicate that EPAC2 is a key host molecule contributing to RSV-induced inflammatory responses.
Cancer in Pacific Populations in New Zealand and ctDNA technology for early cancer detection
Authors: Sika-Paotonu Dianne , Guilford Parry
Published: January 2023

The burden of cancer amongst Pacific Peoples living in New Zealand and in the Pacific Region remains a public health concern. The utilization of a simple blood sample to enable the detection and monitoring of cancer growth is an attractive alternative to obtaining invasive biopsy samples from solid cancers in people. This can be achieved through the use of circulating tumour DNA (ctDNA) detection which refers to small fragments of tumour DNA found in the blood circulation that can be isolated and analysed to allow monitoring of tumour size and progression. This method of cancer monitoring and the potential for early cancer detection, will enhance the precision of cancer diagnostics and surveillance that will likely lead to improved cancer health outcomes over time. This work sought to investigate the potential utilization of ctDNA to support and improve the diagnosis, care and health outcomes of Pacific cancer patients in the New Zealand. A high degree of support and interest was expressed by health professionals, researchers, senior academics and lay persons engaged with for the development of ctDNA as a potential diagnostic tool for early cancer detection, particularly with regards to achieving reductions in adverse cancer statistics and addressing inequalities. As in the Pacific Region, one of the key features of interest was the potential for frail or remotely located patients to be monitored for their cancer without their need to travel or be transported to the Hospital setting which can create a significant source of stress for some. The applicability of ctDNA as an early cancer detection method was explored in depth and received with approval within the research, academic and clinical health care settings.
Human plasmablast migration toward CXCL12 requires glucose oxidation by enhanced pyruvate dehydrogenase activity via AKT
Authors: Nam Bora , Pak Hyo-Kyung , Park Chan-Sik
Published: January 2023

Migration of human plasmablast to the bone marrow is essential for the final differentiation of plasma cells and maintenance of effective humoral immunity. This migration is controlled by CXCL12/CXCR4-mediated activation of the protein kinase AKT. Herein, we show that the CXCL12-induced migration of human plasmablasts is dependent on glucose oxidation. Glucose depletion markedly inhibited plasmablast migration by 67%, and the glucose analog 2-deoxyglucose (2-DG) reduced the migration by 53%; conversely, glutamine depletion did not reduce the migration. CXCL12 boosted the oxygen consumption rate (OCR), and 2-DG treatment significantly reduced the levels of all measured tricarboxylic acid (TCA) cycle intermediates. AKT inhibitors blocked the CXCL12-mediated increase of OCR. CXCL12 enhanced the pyruvate dehydrogenase (PDH) activity by 13.5-fold in an AKT-dependent manner to promote mitochondrial oxidative phosphorylation. The knockdown and inhibition of PDH confirmed its indispensable role in CXCL12-induced migration. Cellular ATP levels fell by 91% upon exposure to 2-DG, and the mitochondrial ATP synthase inhibitor oligomycin inhibited CXCL12-induced migration by 85%. Low ATP levels inhibited the CXCL12-induced activation of AKT and phosphorylation of myosin light chains by 42%, which are required for cell migration. Thus, we have identified a mechanism that controls glucose oxidation via AKT signaling and PDH activation, which supports the migration of plasmablasts. This mechanism can provide insights into the proper development of long-lived plasma cells and is, therefore, essential for optimal humoral immunity.
Transcription factor T-bet is required for pathogenicity of GM-CSF-only Th cells in experimental autoimmune encephalomyelitis
Authors: Rasouli Javad , Ciric Bogoljub , Casella Giacomo , Yoshimura Satoshi , Zhang Guang-Xian , Rostami Abdolmohamad
Published: January 2023

Myelin-specific Th1 and Th17 cells play a pivotal role in the pathogenesis of multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). However, recent descriptions of a new Th subset, tentatively called “GM-CSF-only” Th cells, or ThG cells, raised the possibility that these cells also contribute to the pathogenesis of MS and other autoimmune diseases. ThG cells express GM-CSF, but lack expression of master transcription factors (TF) and hallmark cytokines of established Th cell lineages. Previous studies showed that Th17 cells switch to Th1 phenotype by upregulating T-bet, which is required for their encephalitogenicity. To test whether pathogenicity of ThG cells is also dependent on T-bet, we evaluated the role of T-bet expression by ThG cells in EAE. T-bet was not required for development of ThG cells in vitro or in vivo. Adoptively transferred ThG cells upregulated T-bet expression in the CNS of EAE mice, while they had low or undetectable levels of RORγt, GATA3 and Foxp3. Next, we adoptively transferred ThG cells that developed from WT and T-bet-deficient 2D2 mice to RAG−/− recipients. WT ThG cells were highly encephalitogenic, whereas T-bet-deficient ThG cells failed to induce disease. Taken as a whole, our data show that ThG cells, similar to Th17 cells, have plastic phenotype, upregulate T-bet and IFN-γ in the CNS of EAE mice, and that their encephalitogenicity is dependent on T-bet.
X-chromosome inactivation maintenance is perturbed in T cells from lupus mouse models and human lupus patients
Authors: Anguera Montserrat
Published: January 2023

Sex differences exist with immune responses, yet females are more susceptible to autoimmune and inflammatory disorders. One important biological factor underlying this sex bias is the X-chromosome, which is enriched for immunity-related genes. Female mammals use X-Chromosome Inactivation (XCI) to equalize gene expression between the sexes, which generates a transcriptionally silent inactive X-chromosome enriched with heterochromatic modifications and the long noncoding RNA Xist. We have discovered that mature naïve T cells are missing Xist RNA and heterochromatin marks on the inactive X (Xi), and these marks return to most cells with in vitro stimulation. During T cell development in the thymus, Xist RNA is not localized to the Xi and heterochromatin marks are progressively lost. Xist RNA returns to the Xi in mature T cell subsets between days 2 and 3 of in vitro activation. During lupus disease progression in the female-biased mouse model NZB/W F1, Xist RNA signals aberrantly appear in naïve T cells and Xist RNA becomes mis-localized from the Xi in activated T cells. These abnormal XIST RNA localization patterns are also observed in activated T cells from pediatric lupus patients in disease remission. Using next-generation sequencing, we found significant overexpression of X-linked genes across the X in female SLE T cells compared to healthy females. We propose that the dynamic mechanism of XCI maintenance female lymphocytes predisposes the Xi to become partially reactivated and to overexpress immunity-related genes. Our work provides the first mechanistic evidence for the female-specific enhanced immune responses and increased susceptibility for autoimmunity.
Germinal center organization mediated by T-bet dependent expression of CXCR3 and CCR6
Authors: Peel Jessica Nicole , Stone Sara L , Scharer Christopher D. , Boss Jeremy , Lund Frances E
Published: January 2023

The germinal center (GC) is composed of distinct regions with proliferation and affinity maturation of GC B cells (GCB) occurring in the dark zone (DZ) and antigen-mediated selection and commitment of the long-lived plasma cell (LL-PCs) precursors occurring in the light zone (LZ). Despite the key role for GCs in establishing enduring humoral immunity, we know little about the dynamics of the GCB cell response. We showed that the transcription factor T-bet is expressed by GCB cells and that B cell intrinsic T-bet is required for the LL-PC response following influenza (flu) infection. To test whether the loss of LL-PCs in these mice was due to a role for T-bet in regulating the GCB response, we infected 50:50 Tbx21−/−: pepboy chimeric mice with flu and enumerated GCB cells. We found that Tbx21−/− B cells were overrepresented in the GC relative to the pepboy cells and that Tbx21−/− GCB cells were enriched in the LZ. Partitioning of GCB cells in the LZ/DZ is regulated by chemokines and we showed that T-bet controls transcription of two chemokine receptors, CXCR3 and CCR6, which are expressed by GCB cells. Using 50:50 Tbx21−/−: pepboy chimeras, we tested whether T-bet regulates GCB cell positioning by inducing CXCR3 and repressing CCR6. We observed that the CCR6+ GCB cell subset, which is known to localize in the LZ, was very enriched in Tbx21−/− GCB cells. By contrast, the CXCR3+ GCB subset, which was dominated by WT cells, was enriched in the DZ. To determine the role for CXCR3 in GCB cell localization, we infected 50:50 Cxcr3−/− :pepboy chimeras and found that the Cxcr3−/− GCB cells are enriched in the LZ. Therefore, T-bet regulates GCB cell localization and potentially the dynamics of the GC response by controlling CXCR3 and CCR6 expression.