bluuebunny/crossref_metadata_2025 huggingface dataset:
Inhibition of heme oxygenase-1 activity suppresses Mycobacterium tuberculosis infection in vivo by a mechanism dependent on T lymphocytes and IFN-γ production
Authors: Costa Diego L , Namasivayam Sivaranjani , Amaral Eduardo P , Arora Kriti , Mittereder Lara R , Maiga Mamoudou , Boshoff Helena I , Goulding Celia W , Andrade Bruno B , Sher Alan
Published: January 2023

Infection with M. tuberculosis (Mtb) induces the upregulation of heme oxygenase-1 (HO-1), a powerful antioxidant enzyme that catalyzes the degradation of heme into iron, biliverdin and carbon monoxide. We observed that administration of tin protoporphyrin IX (SnPPIX), a well-characterized HO-1 activity inhibitor, to Mtb-infected mice resulted in pulmonary bacterial load reduction. Moreover, adjunct administration of SnPPIX to Mtb-infected mice receiving antibiotic treatment improved and accelerated pathogen clearance. Interestingly, both the induction of HO-1 expression in lungs of Mtb-infected mice and the efficacy of in vivo SnPPIX-mediated bacterial burden reduction were dependent on the presence of host TCR-α+ lymphocytes as well as IFN-γ suggesting a role for the immune response in the activity of the drug. Further investigation revealed that in the lungs of these immunodeficient mice the numbers of parenchymal CD11b+ myeloid cells staining positive for HO-1 were substantially reduced. SnPPIX treatment of Mtb-infected mouse bone marrow derived macrophages also resulted in diminished bacterial loads in vitro and this effect was reverted by iron supplementation and enhanced by iron chelation. Based on these observations we propose that HO-1 inhibition promotes control of Mtb by limiting the availability of nutrient iron and that the host Th1 response regulates this effect by controlling the recruitment to the lungs of a subset of myeloid cells that is both capable of HO-1 expression and permissive to bacterial infection. This work was supported by the intramural research program of the NIAID.
Non-apoptotic signaling through Fas activates Akt and promotes T cell differentiation through Caspase-8
Authors: Yi Fei , Ouyang Claudia , Yamamoto Tori , Cruz Anthony , Chan Francis K , Frazzette Nick , Thomas Stacy , Restifo Nicholas P , Siegel Richard M.
Published: January 2023

The TNF receptor Fas (CD95, TNFRSF6) is known for inducing apoptosis in primary cells via binding with Fas ligand (FasL) and activation of caspase-8 through the adapter protein FADD. In addition, recent research showed that the apoptotic signaling requires palmitoylation of Fas near its transmembrane region to localize on the lipid rafts. Fas palmitoylation-defective mutant is unable to induce apoptosis in primary cells but continues to enhance T cell differentiation into effector memory cells (CD44hiCD62Llo), indicating that Fas can also trigger non-apoptotic signaling in primary T cells. Fas non-apoptotic signaling has unexpected roles in modulating T cell tumor immunotherapy and autoimmunity (Klebanoff et al. 2016; Cruz et al. 2016). We have investigated the signaling pathways underlying non-apoptotic Fas signaling in T cells. Co-stimulation with oligomerized Fas Ligand activates Akt and ribosomal S6 protein phosphorylation, and Fas induced T cell differentiation is blocked by Akt or mTOR inhibitors. Further studies showed that T cells from RIP3−/− and Caspase-8−/− double deficient mice, but not mice deficient in RIP3, were refractory to Fas-induced differentiation, indicating that Caspase 8 is essential for Fas non-apoptotic signaling. Similar results were obtained with FADD. Our findings suggest that in addition to apoptotic functions, Fas also promotes non-apoptotic signaling via Caspase-8 and induces T cell differentiation by activating the Akt pathway. These results reveal a common role for Caspase-8 in non-apoptotic and apoptotic functions of Fas, with potentially different caspase-8 substrates mediating these diverse functions of a single receptor.
Chronic mTOR activity impaires lysosome maturation in lupus
Authors: Monteith Andrew J , Vincent Heather , Kang Sunah , Li Patrick , Claiborne Tauris , Moorman Nathaniel , Vilen Barbara
Published: January 2023

Multiple genetic and environmental factors activate lymphocytes and myeloid cells to promote systemic lupus erythematosus (SLE). This complexity has made it difficult to define underlying mechanisms. We previously demonstrated that hematopoietic cells from lupus-prone mice accumulate nuclear antigens on their cell surface due to an inability to mature the lysosome. On myeloid cells, the accumulation of nuclear antigens reflects IgG-immune complexes (IgG-ICs) bound to FcγRs. Further, lupus-prone mice lacking FcγRI fail to develop myeloid and lymphoid-associated pathologies suggesting a critical role for FcγRI in SLE. How FcγRI signaling influences lysosomal maturation is unclear. Herein, we show that lysosome acidification, a key step in the maturation process, relies on the strict regulation of mTORC2, activation of caspase-1, and cleavage of Rab39a. In macrophages from MRL/lpr mice, we find that chronic mTORC2 activation disrupts lysosomal maturation by inducing constitutive phosphorylation of cofilin. This prevents the association of cofilin with phagosomes containing IgG-ICs. As a result, inflammasome-independent activation of caspase-1 in proximity to the phagosome does not occur. Consequently, Rab39a remains intact on the phagosomal membrane, and the lysosome does not acidity. These findings reveal unique roles for mTORC2, cofilin, and pyroptototic caspases critical to an unappreciated signaling pathway promoting lysosome acidification. Further, this signaling cascade becomes disrupted in SLE, preventing the lysosome-mediated degradation of apoptotic debris upstream of multiple lupus-associated pathologies.
2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) induces tolerance by altering the gut microbiome
Authors: Neamah Wurood Hantoosh , Nagarkatti Mitzi , Nagarkatti Prakash
Published: January 2023

The compound 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) belongs to the polychlorinated dibenzodioxins (PCDDs) and is known to act as an immunosuppressant for humoral and cell-mediated responses. In the current study, we investigated TCDD-mediated mechanisms related immunosuppression. Gut microbiota serves an important role in immune function and development of disease, and it has been shown that changes in host microbiota is not only a marker of disease, but also actively contributes to disease pathogenesis. We, therefore, investigated how 3 days of TCDD exposure in mice could alter the gut microbiome compared to control groups. We performed 16S rRNA sequencing on bacterial DNA isolated from fecal samples from the experimental groups. We found separate clustering of the bacterial composition in TCDD versus the vehicle treated groups based on principal coordinate analysis. Interestingly, we found that TCDD-treated mice showed significant decrease in Sutterella, a bacterium belonging to Proteobacteria phylum, Betaproteobacteria class, Burkholderiales order and Alcaligenaceae family. Moreover, Sutterella is present normally in gut flora and elevated in some inflammatory disorders. Collectively, our data suggests that TCDD induces tolerance by inducing dysbiosis in gut microbiome that leads to impairment of proinflammatory bacteria.
Complexes assembled from myelin self-antigen and a toll-like receptor antagonist promote tolerance
Authors: Hess Krystina L , Jewell Christopher M
Published: January 2023

Myelin, a matrix that insulates the axons of neurons, is attacked by the immune system in multiple sclerosis (MS), leading to neurodegeneration. MS treatments are currently limited because they are non-curative and broadly-acting immunosuppressants that can leave patients immunocompromised. Recent reports show co-delivery of myelin peptides (MOG) along with regulatory signals can promote tolerance to myelin rather than cause inflammation. New studies reveal inflammatory toll-like receptors (TLRs) play a significant role in driving disease in both humans and mice. Further, several reports demonstrate that regular treatment with TLR9 antagonist, GpG, attenuates disease in mouse models of MS. We hypothesized that GpG DNA and cationic MOG peptide would electrostatically self-assemble into complexes, allowing co-delivery, increased cargo protection, and enhanced uptake by immune cells. These complexes might redirect the response against myelin through suppression of inflammatory TLR9 signaling during myelin presentation. Importantly, polyplexes significantly reduced inflammatory TLR9 signaling in a reporter cell line. Primary dendritic cells (DCs) internalized MOG-GpG complexes, resulting in a reduction of activation and decreased proliferation of myelin-reactive T cells. In a mouse model of MS, experimental autoimmune encephalomyelitis (EAE), complex injection reduced disease severity more than 2-fold and incidence by 30%. This data demonstrates the ability of GpG to condense self-antigens while remaining an active immunomodulator. This work also suggests a new carrier-free platform for co-delivery of antigens and immune signals that still provides the more attractive features of biomaterials.
Effect of cannabidiol (CBD) on CD4+ cells under differential immunological activation state.
Authors: Dhital Saphala , Kaplan Barbara L. F.
Published: January 2023

We previously reported that cannabidiol (CBD), one of the Cannabis sativa-derived cannabinoids, induced functional CD4+CD25+FOXP3+ Tregs in response to low level stimulation of PMA/Io. CBD increased CD25+FOXP3+ cells in CD4+, CD4+CD25+, and CD4+CD25− T cells, as well as in CD4+ T cells derived from FOXP3-GFP mice. T cell stimulation conditions were suboptimal PMA/Io (4 nM/0.05 μM, S/o) and ultrasuboptimal PMA/Io (1 nM/0.0125 μM, Us/o). We also observed that CBD + Us/o increased IL-2, IL-10 and TGF-β1 immunomodulatory cytokines. Here, we measured the effect of CBD + Us/o or S/o on CD4+ cells. Splenocytes were treated with CBD (1–10 μM) for 30 min plus S/o or Us/o activation and cultured for 1 day or 5 days. Immunofluorescence staining showed that CBD + Us/o treatment increased the percentage of CD4+ cells while CBD + S/o treatment decreased the percentage of CD4+cells. CD4+CD25+ expression by Us/o or S/o activation alone was also measured. CD25 expression in response to Us/o activation was lower than that by S/o activation in CD4+ cells. This study will provide important information on the effect of CBD on CD4 cells under differential immunological activation state by PMA/Io.
Bee venom phospholipase A2 (bvPLA2) protects against LPS-induced abortion
Authors: Bae Hyunsu , Baek Hyunjung , Shin Dasom , Hwang Deok-Sang
Published: January 2023

Implantation of the fertilized egg into maternal uterus depends on the fine balance between inflammatory and anti-inflammatory processes that prepare the uterus for this important event. Regulatory T cells (Tregs) are a key regulator for the protection of allogenic fetus against rejection by maternal immune system. In the present study, we carried out to investigate the protective effects of bvPLA2 on LPS-induced abortion mice model. LPS was injected intraperitoneally to induce spontaneous abortion at day 9.5 of pregnancy and the mice were administered bvPLA2. The results clearly revealed that bvPLA2 can prevent the fetal loss accompanied by growth restriction in remaining viable fetus. When the LPS-induced abortion mice were treated with bvPLA2, Treg cells were significantly increased compared to the no pregnant, WT and LPS group. After LPS injection, Th1 type cells (CD4+IFN-γ+) were markedly increased compared to the WT group mice, while bvPLA2 treatment showed no significant differences in CD4+IFN-γ+cell populations. These findings suggest that bvPLA2 has protective effects in LPS-induced abortion mice model by regulating Treg populations. Further studies are necessary in order to define the detailed mechanisms underlie Treg function by bvPLA2 treatment during in LPS-induced model of abortion.

Acknowledgement: This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MSIP) (No.NRF-2015R1C1A1A02037172)
Characterization of merocytic dendritic cells homeostasis
Authors: Audiger Cindy , Chabot-Roy Geneviève , Lesage Sylvie
Published: January 2023

In contrast to conventional dendritic cells (cDC), when merocytic DC (mcDC) present antigens (Ag) derived from apoptotic bodies, T cell anergy is reversed rather than induced. Although helpful to tumor clearance, reversing T cell anergy is detrimental in autoimmunity. Interestingly, mcDC are present in higher proportion in type 1 diabetes (T1D)-prone NOD mice than in B6 mice. Still, little is known about the immunological properties of mcDC that define them as a DC subset. Phenotypic characterization of mcDC revealed that they express the cDC-restricted transcription factor, Zbtb46. Moreover, we found that mcDC are potent inducers of mixed lymphocyte reactions, a key trait defining them as cDC. Investigation of other transcription factors associated with specific cDC functions, such as Irf4 and Irf8, reveal that mcDC are heterogeneous, with phenotypic and functional characteristics more closely resembling CD8α cDC. Comparative gene profiles by principal component analysis revealed that mcDC clustered more closley to CD8α cDC than other DC subsets. Still, mcDC are distinct from CD8α cDC, as mcDC are present in Batf3-deficient mice. Together, these data demonstrate that mcDC are a cDC subset with unique ability to break antigen specific tolerance. Understanding the homeostatic regulation of this new cDC subset will permit to develop new therapies for diseases, such as autoimmunity or cancer where they can be either pathogenic or beneficial.
Role of group 2 innate lymphoid cells (ILC2) in SHARPIN-deficient mice
Authors: HogenEsch Harm , Elesela Srikanth , Chien Syu-Jhe , Silva Kathleen , Kennedy Vicki , Sundberg John P.
Published: January 2023

SHARPIN is a component of the linear ubiquitination assembly complex and a key regulator of NFkB and integrin signaling. SHARPIN-deficient mice develop a phenotype known as chronic proliferative dermatitis (cpdm), characterized by progressive epidermal hyperplasia, apoptosis of keratinocytes, cutaneous and systemic eosinophilic inflammation, and hypoplasia of secondary lymphoid organs. We recently reported that the cutaneous inflammation in SHARPIN-deficient mice (Sharpincpdm) develops independently of B and T lymphocytes. We therefore sought to determine the role of innate lymphoid cells (ILCs) in the dermatitis of Sharpincpdm mice. ILCs were identified as a discrete population of CD45+ Lin−CD90.2hi cells in the skin and draining lymph nodes (LN) of wild type (WT) and Sharpincpdm mice. The number of ILCs was markedly increased in Sharpincpdm mice. The majority of cells were group 2 ILC (ILC2) as indicated by labeling for GATA3, IL5, and IL13. The skin of Sharpincpdm mice had increased expression of Il33 and Tslp mRNA. To determine the role of IL33, double mutant mice were generated in which the receptor for IL33 (Il1lr1 also known as ST2) was deleted. Loss of IL33-signaling greatly reduced the number of ILCs and reduced the severity of the dermatitis. These experiments suggest that the dermatitis in Sharpincpdm mice is driven by IL33-dependent ILC2. Supported by the NIH (AR049288).
Intestinal barrier disruption inversely regulates liver inflammation via IL-10-producing macrophages
Authors: Taniki Nobuhito , Nakamoto Nobuhiro , Mikami Yohei , Kanai Takanori
Published: January 2023

The gut-liver axis is a clinical topic as a potential therapeutic target of liver diseases. The interaction between microbial products and immune responses in the liver is still unknown. In the current study, we have demonstrated IL-10-producing CD11b+ F4/80+ macrophages contribute to immune tolerance in the inflamed liver under diminished intestinal barrier function in murine tandem model of dextran sulfate sodium (DSS)-colitis and Concanavalin A (ConA)-hepatitis. DSS-ConA mice with increased intestinal permeability developed less liver injury compared with DW-ConA mice. Of note, the severity of colitis was inversely correlated with subsequently induced liver injury. In DSS-ConA livers, increased CD11b+ macrophages showed regulatory characteristics with IL-10 production in vitro. Pretreatment with α-IL-10 antibody partially exacerbated the suppressed liver injury by DSS-ConA in vivo. Furthermore, pretreatment with CD11b+ macrophages derived from DSS-ConA livers protected mice from the subsequent liver injury. Mechanistically, we observed the contribution of gut microbiota, as the immunological tolerance induced by DSS-ConA is partially ablated by gut sterilization. Pretreatment with the portal serum derived from DSS-ConA mice partially protected mice from the subsequent liver injury. These results suggest that altered gut microbiota and the metabolites that pass through the portal vein with disrupted intestinal barrier directly gave rise to immunological permissiveness against the excessive stimulation in the liver. This study clarified the contribution of the gut-liver axis in the maintenance of immune balance in the liver and highlighted the microbiota as a potential therapeutic target for liver diseases.
Pathogenesis of Hepatic Deposited IgG-induced Liver Inflammation in SLE
Authors: Deng Guo-Min , Fang Xiang , Guo Xuanxuan
Published: January 2023

Hepatic disorders are frequent in patients with systemic lupus erythematosus ( SLE), yet the etiology and pathogenesis of liver injury in SLE remain unclear. We used lupus-prone mice and established an animal model of intrahepatic deposited lupus IgG to investigate the pathogenesis of liver damage in SLE. We found that liver damage spontaneously developed in lupus-prone mice and that there were a large number of inflammatory cell infiltrates around the portal areas of the liver, apoptosis of hepatocytes, and IgG deposition in the liver. Liver inflammation developed in mice with intrahepatic injection of immune complexes (ICs) from SLE patients. We found that ICs stimulated Kupffer cells (KCs) to secrete TNF-α and mediated hepatic apoptosis. Moreover, IL-12 secreted by KCs induced the accumulation and activation of natural killer cells (NKs) in the liver. Further, IFN-γ produced by activated NKs directly mediated liver damage and also enhanced the TNF-α-mediated apoptotic pathway. The depletion of KCs and NKs abolished apoptosis induced by ICs in vivo, suggesting that KCs and NKs have a synergic effect on liver injury. Lupus IgG induced Syk activation, and inhibitor of sky suppressed skin inflammation triggered by lupus IgG. Taken together, our findings confirmed that the hepatic disease was mediated by SLE, which may provide insight into the cellular and molecular mechanisms of lupus hepatitis; further, our results may aid in future investigations of potential therapeutic strategies for the treatment and control of lupus-mediated injury in disease settings.
VDJServer – a web-accessible analysis portal for immune repertoire sequencing analysis
Authors: Toby Inimary , Christley Scott , Scarborough Walter , Rounds William H , Fonner John , Mock Stephen , Monson Nancy , Scheuermann Richard H , Cowell Lindsay G
Published: January 2023

VDJServer is a comprehensive, web-accessible system for analysis of immune repertoire sequencing data. VDJServer provides a complete analysis workflow from pre-processing of sequence reads, to V(D)J assignment, to repertoire characterization and comparison. Recent enhancements in VDJServer include: --Automatic parallelization of analysis tools to handle very large data sets running on a high-performance supercomputer--Import and export subject and sample metadata. User-defined sample groups allows for sophisticated group analysis and comparison.--Extensive analysis functionality such as gene segment usage, CDR3 patterns, clonality, diversity measures, somatic mutation patterns, B cell lineage trees, and quantification of selection pressure. Analysis is performed for both samples and sample groups.--Interactive charting of analysis data provides exploratory visualization for ad-hoc comparison of samples and sample groups. Charts can be downloaded as image files for use in presentations and publications. All analysis data can be downloaded in standard TSV format for use with external tools.--Novel process workflow metadata that is automatically captured by VDJServer. Hiding the complexities of command line tools and their parameters, yet providing complete transparency of the analysis workflow for reproducibility.

VDJServer allows users to upload antigen receptor repertoire sequences and execute a customizable workflow for all steps in the analysis. Data and analysis results can be privately shared with other users for collaborative projects. VDJServer is funded by the NIAID and is freely available.
Tumor-experienced human NK cells express PD-L1 and display immunoregulatory functions
Authors: Fuertes Mercedes B , Iraolagoitia Ximena L Raffo , Ziblat Andrea , Nuñez Sol Y , Torres Nicolas I , Secchiari Florencia , Domaica Carolina I , Zwirner Norberto W , Sierra Jessica M
Published: January 2023

While NK cells play a key role in the elimination of infected cells and tumors, regulatory functions of NK cells are emerging in several models of viral infections and autoimmunity. In the tumor context, we have recently described an immunoregulatory population of PD-L1-expressing NK cells arising in tumor-bearing mice that inhibits CD8+ T cells priming. Since little is known about the expression of PD-L1 on human NK cells, the aim of this work was to study its expression on human NK cells after tumor recognition and the underlying mechanisms. After culture of human PBMCs from healthy donors with K562 tumor cells, PD-L1 expression was induced on NK cells. Transwell and receptor blockade experiments showed a dependence on cell-to-cell contact that involved NKG2D engagement. PD-L1 expression on isolated NK cells stimulated with K562 tumor cells could be further increased by soluble factors derived from tumor-stimulated PBMCs or by the addition of monocytes. Neutralization of different cytokines identified IL-18 as the main factor responsible for PD-L1 up-regulation, which can be produced by monocytes. These PD-L1hi NK cells were immunoregulatory, as they inhibited DC maturation in vitro. This population may be clinically relevant, as we detected PD-L1hi NK cells in the tumor microenvironment and in peripheral blood of human patients with kidney cancer, making PD-L1 expression on NK cells an attractive candidate as prognostic/diagnostic biomarker. Our results indicate that PD-L1 is expressed in tumor-experienced human NK cells which subsequently acquire immunoregulatory functions. Thus, rational manipulation of these regulatory cells could lead to improved anti-tumor immunity in vivo
Relevance of host and tumor PD-L1 expression in PD-L1 and PD-1 blockade
Authors: LIN HENG , Wei Shuang , Vatan Linda , Kryczek Ilona , Zou Weiping
Published: January 2023

Therapeutic blockade of PD-1 or PD-L1 with monoclonal antibodies (mAb) leads to durable tumor regressions in patients across cancer types. Clinical responses to PD-L1 and PD-1 blockade may be associated with increased PD-L1 expression in the tumor tissues, tumor-infiltrating T cells, and tumor neoantigens. However, it is unclear which patient population(s) benefits from these treatments. Furthermore, why do some patients with PD-L1 negative tumor respond to PD-L1 and PD-1 blockade therapy? To dissect the molecular and cellular mechanisms that account for the clinical efficacy of PD-L1 and PD-1 blockade, we used Rag1tm1Mom(Rag1−/−), NOD.SCID gc-deficient (NSG), PD-L1 genetic deficient (PD-L1−/−), and PD-1 genetic deficient (PD-1−/−) mice, and studied PD-L1 blockade in MC38, ID8, B16-F10, and LLC models. We found a loss of therapeutic effect of anti-PD-L1 mAb treatment in Rag1−/−, NSG, PD-L1−/−, and PD-1−/−mice. We demonstrated the importance of PD-L1 expression on antigen presenting cells (APCs), particularly dendritic cells (DCs) in the tumor microenvironment and draining lymph nodes, in anti-PD-L1 treatment efficacy. Thus, the host immune system is indispensable for anti-PD-L1 therapy, and the host, rather than cancer cell-intrinsic PD-L1 may account for the therapeutic efficacy of PD-L1 signaling blockade.
Trichomonas associated protection against Salmonella infection
Authors: Lu Chunchi , Zhao Xiaodan , Lo Chia-Ta , Lyu Sin-Yi , Smith Kelly D
Published: January 2023

Colonization resistance (CR), defined as the ability of intestinal microbiota and host defenses to protect against pathogens, is the first line defense against intestinal infection. In our studies of innate immune recognition of intestinal Salmonella, we discovered Naip5−/− and Rag1−/− mice harbor microbiota that protect mice against Salmonella infection, which is associated with intestinal colonization by Tritrichomonas muris (Tm). Tm is a protozoon commonly found in laboratory, pet shop and wild mice, which has no definitive disease association. Recent studies have demonstrated Tm and other intestinal parasites are sensed by tastant receptors expressed by specialized intestinal tuft cells, which enhances intestinal barrier by tuft cell and goblet cell hyperlasia. Helminths and protozoa have been associated with changes in the intestinal microbiota, suggesting that intestinal protozoa may produce biologically significant alterations in the composition of gut bacteria. We demonstrate that Tm colonization of mice is associated with decreased inflammation in the intestine during Salmonella infection of streptomycin pretreated mice. Tm colonization is also associated with increased resistance to Salmonella infection and CR. Finally, we demonstrate that Tm colonized mice have an altered gut microbiome, suggesting that Tm cultivates a protective microbiome that helps prevent Salmonella colonization and infection. Our studies indicate that intestinal protozoa may influence the complex environment within the gut to help limit Salmonella invasion of the gut ecosystem and Salmonella induced intestinal inflammation, and provide insight into novel strategies that may help prevent Salmonella infection and transmission.
A Novel Population of TCRαβ-expressing CD11bhighCD14+F4/80+ Macrophages induced by a Pathogenic Murine Malaria enhances phagocytosis of parasites
Authors: Oakley Miranda S , Chorazeczewski Joanna , Majam Victoria , Chawla Bhavna , Aleshnick Maya , Takeda Kazuyo , Akue Adovi , KuKuruga Mark , Kumar Sanjai
Published: January 2023

Macrophages, a component of the innate immune system, are equipped with a wide array of invariant receptors that facilitate the host defense against invading pathogens via phagocytosis and the release of inflammatory cytokines, and help maintain tissue homeostasis by initiating the repair and remodeling processes. In malaria, macrophages perform both immune protective (parasite clearance) and pathogenic (severe malaria anemia and cerebral malaria) functions. Utilizing the virulent asexual stage Plasmodium berghei ANKA (Pb-A) parasite, which causes experimental cerebral malaria (ECM) in mice, we have identified a novel population of TCRαβ-expressing macrophages that expands during Pb-A infection and preferentially migrates to and sequesters in the brain during ECM. This novel population of macrophages is CD3−CD4−CD8− and has a Vβ TCR repertoire that is distinctly different from conventional T cells. We also find that optimal expression of combinatorial TCRαβ on CD11bhigh cells requires coexpression of CD14 and F4/80. In depth immunological studies demonstrate that macrophage TCRαβ induces proliferation, alters IL-4 and MIP1β production, and enhances phagocytosis of infected erythrocytes. Expansion and activation of TCRαβ+CD11bhighCD14+F4/80+ macrophages during malaria infection point towards a convergence of the innate and adaptive immune responses where both arms of the immune system cooperate to modulate the host response to invading pathogens. Our results suggest that targeting macrophage effector functions through vaccines and drugs should be explored as an approach to reduced malaria mortality and morbidity.
Effects of pH, Charybdotoxin, and PC12 cells on Jurkat T-cell proliferation
Authors: Snitsarev Vladislav , Petroff Elena
Published: January 2023

Neuroimmunological diseases involve T-cell attacking neurons. We showed previously that acid-sensing ion channels (ASICs) and voltage-gated channels (Kv) from different cells can interact leading to inhibition of Kv. We hypothesized that this interaction may exist between Kv of Jurkat T-cells (JTL) and ASICs of PC12 cells (neuronal model), and that this interaction is pH- and Charybdotoxin (ChTX)-dependent.

JTL were seeded on top of confluent PC12, or without PC12. The effects of 200 nm ChTX and pH 7.2 on JTL doubling time (DT) were tested.

Without PC12, DT was 12.2±0.5h at pH 7.4, 25.6±1.1h with ChTX, and 17.0±1.0h at pH 7.2; in ChTX at pH 7.2, DT was 40.8±5.0h.

With PC12, JTL DT was 34.1±2.9h at pH 7.4, 30.1±2.3h with ChTX, and 20.7±1.8h at pH 7.2; in ChTX at pH 7.2, DT was 32.2±1.8h.

Paired comparisons without PC12 show that DT was significantly longer at pH 7.2 vs pH 7.4 and in ChTX at pH 7.4; pH 7.2 and ChTX synergistically increased DT vs pH 7.2 and vs ChTX at pH 7.4.

With PC12, all corresponding DTs were significantly longer. Paired comparisons showed that at pH 7.2 DT was significantly shorter than at pH 7.4, opposite to without PC12. This can be potentially explained by lower pH relieving inhibition of Kv on JTL by the ASICs on PC12. At pH 7.4, ChTX did not have any effect possibility due to JTL’s Kv already blocked by PC12’s ASICs, and at pH 7.2 ChTX inhibited the free Kv.

Thus, both acidic pH and ChTX independently slow down JTL proliferation, and in the presence of PC12, effects of low pH and ChTX are consistent with ASIC-Kv interaction.

We speculate that ASICs expressed in APCs and Kv in lymphocytes may be a new unrecognized interaction in the immunological synapse, and disruption of neuronal ASIC-lymphocytic Kv interaction may play a role in autoimmune disease.
Blunting age-associated chronic inflammation preserves hematopoiesis and immunity and reduces leukemogenesis
Authors: Henry Curtis J , Casas-Selves Matias , Kim Jihye , Zaberezhnyy Vadym , Aghili Leila , Daniel Ashley E. , Jimenez Linda , Azam Tania , McNamee Eoin N. , Clambey Eric T. , Klawitter Jelena , Serkova Natalie J. , Tan Aik-Choon , Dinarello Charles A. , DeGregori James
Published: January 2023

Aging is the single most important prognostic factor for the development of many cancers. While mutational accumulation may increase the risk of cancer in aged individuals, declining immunity due to chronic inflammation is also a likely contributing factor.

We have demonstrated that reducing aging-associated chronic inflammation abrogates fitness declines in B-progenitor cells, and significantly reduces leukemogenesis in aged mice (Henry et al., JCI, 2015). In addition to preserving the function of B-progenitor cells, subsequent studies have revealed that reducing chronic inflammation in aged mice augments immune responses. Specifically, reducing inflammation in aged mice (≥22 months) results in a two-fold reduction in the number of splenic M2 macrophage and T-regulatory cell populations, as well as, a three-fold reduction in the surface expression of the inhibitory protein PD-L1 on innate immune cells. Reducing chronic inflammation in aged mice also results in a three-fold increase in the percentage of interferon-gamma producing CD4+ and CD8+ T-cell lymphocytes when stimulated. These phenotypes were observed in aged, anti-inflammatory transgenic mice (alpha-1-anti-trypsin and interleukin-37) and in geriatric mice (≥27 months) treated with recombinant interleukin-37. Overall, these findings suggest that reducing chronic inflammation in aged populations can rejuvenate hematopoiesis and immunity, while also creating a less permissive environment for leukemogenesis.
Absence of estrogen receptor alpha in males does not impact disease severity during nephrotoxic serum-induced nephritis
Authors: Corradetti Chelsea , Cesaroni Matteo , Madaio Michael , Caricchio Roberto
Published: January 2023

Systemic lupus erythematosus (SLE) exhibits a strong sex bias in which 90% of patients are female. Interestingly, male SLE patients experience greater progression and severity of disease. Glomerulonephritis (GN) remains one of the most serious manifestations of SLE and accounts for most morbidity and mortality among lupus patients. Due to the sex bias, our lab is interested in studying how estrogens may play a role in the pathogenesis of GN and in recent studies we revealed that estrogens exacerbate GN in females in an estrogen receptor alpha- (ERa)-dependent manner by altering activity of metabolic pathways. Although estrogen concentrations are lower, we were interested in investigating whether these sex hormones can also influence disease in males. We found that absence of ERa did not significantly affect the severity of NTS-induced nephritis (NTN) in male mice. Both immune cells and renal cells were involved in NTN progression, and chimera experiments revealed absence of ERa in either of these compartments does not alter the severity of disease. Although absence of ERa in males did not alter NTN disease phenotype, interestingly, analysis of the glomeruli of these mice revealed large differences in gene expression profiles between WT and ERaKO males even if the progression of NTN was similar. WT males exhibited increased expression of genes related to immune activation and cell adhesion, followed by downregulation of metabolic pathways, while ERaKO males undergo very few genetic changes during NTN. From these results, we conclude that ERa does not play a role in GN pathogenesis in males; however, the potential impacts of the gene expression changes induced by its absence warrants further investigation on other aspect of lupus disease.
Emergence of T Regulatory Cells Following Neural Precursor Cell Transplantation in Mouse Models of Multiple Sclerosis
Authors: McIntyre Laura Lynn , Plaisted Warren Christopher , Nguyen Quy , Kessenbrock Kai , Lane Thomas , Walsh Craig Michael
Published: January 2023

Multiple Sclerosis (MS) is a chronic, autoimmune disease of the central nervous system for which there is no cure. Defects in neurological function are a result of demyelination and axonal loss caused by infiltrating immune cells. Transplantation of neural precursor cells (NPCs) is a promising therapeutic strategy to treat neurological diseases. However, studies evaluating NPC transplantation often employ a syngeneic donor or immune suppressed subjects. Therefore, it is important to consider how transplanted cells elicit an immune response. We report that human neural precursor cells (hNPCs) induce antigen specific Tregs which influence endogenous repair pathways to promote remyelination in murine models of MS. We observed remyelination and decreased neuroinflammation in mice receiving an intra-spinal transplant of syngeneic NPCs, which replaced damaged cells. Alternatively, mice receiving transplants of xenogeneic human NPCs displayed remyelination, a decrease in neuroinflammation, and an increase in CD4+CD25+FoxP3+ Tregs. Recovery was not a result of direct cell replacement as hNPCs underwent xenograft rejection. Importantly, ablation of Tregs abrogated histopathological improvement. Furthermore, hNPCs promoted generation of Tregs in vitro. These findings support an emerging role for Tregs in promoting tissue regeneration, distinct from immune modulation.
Altered flow triggers activated T cell accumulation and inflammation in early atherosclerosis
Authors: Xu Maria Mei , Menoret Antoine , Zhou Beiyan , Murphy Patrick A. , Vella Anthony T.
Published: January 2023

Atherosclerosis is linked to systemic inflammation. Yet, the inflamed plaques predicating cardiovascular morbidity and mortality preferentially develop in areas of low and disturbed flow (LDF). Partial carotid artery ligation (PCAL) of the left carotid exposes vascular endothelium to LDF without disturbing flow contralaterally and when combined with high fat diet, LDF regions develop plaques within 2 wks. We show that just 48 hr after PCAL, LDF triggers significant upregulation of IL-36, a new member of the proinflammatory IL-1 family, and Cathepsin S, its enzymatic activator. The IL-1 family and IFNγ are intimately tied to atherogenesis and our past work establishes IL-36 as a robust stimulator of IFNγ from effector T cells. Thus, we hypothesize that LDF triggers an IL-36-IFNγ loop, promoting early disease independent of plaque-specific antigen. To test this, we assessed the homing of adoptively transferred CD8 TCR transgenic T cells that were antigen-primed and CD137-costimulated within recipient mice. Comparison of physiologic LDF regions (aortic arch) with experimentally induced LDF (PCAL) reveals that effector CD8 T cells substantially and preferentially accumulate within LDF areas, even without TCR-specific stimuli in the vessel wall. Notably, cells isolated from these regions produce IFNγ in response to IL-36. Mass cytometry (CYTOF) and single cell RNA-seq analyses are underway to build mechanistic understandings of how LDF triggers immune cell recruitment and signaling cascades. Overall, our studies propose a novel role for IL-36 within the vascular system and provide insight into the early pathogenesis of atherosclerosis, especially in the context of autoimmunity and antigen-independent T cell activation.
Loss of the von-Hippel Lindau gene in osteocytes alters B lymphocyte development and immune response
Authors: Chicana Betsabel J. , Genetos Damian , Yellowley Clare , Murguesh Deepa , Loots Gabriela G. , Manilay Jennifer O.
Published: January 2023

The von-Hippel Lindau protein (VHL) regulates hypoxia-inducible factor (HIF) degradation. Conditional-deletion of VHL in osteoblasts and in hematopoietic progenitors have demonstrated a role for VHL in these cell types, but how changes in bone homeostasis affects immune development are not fully understood. Osteoblasts (OBs) support B cell development. OBs mature into osteocytes (OCYs) and to test if OCYs also support B cell development, we generated OCY-specific (Dmp-Cre) conditional VHL knockout (cKO) mice. cKO mice display dysregulated bone growth, high bone density, smaller bone marrow (BM) cavity volume, and overall decrease in BM cellularity compared to controls. In line with this, the frequencies and numbers of B cell precursors (B220+ IgM−), immature B (B220+ IgMint) and recirculating B (B220+ IgMhigh) cells in the bone marrow are significantly decreased. These data suggest that the change in bone homeostasis adversely affects B cell development in a cell-extrinsic manner. Although the absolute numbers of B cells in the spleens of cKOs were normal, we hypothesized that loss of VHL in OCYs could permanently and adversely affect B cell functional response. To test this, VHL cKO mice were immunized with a T-independent antigen (NP-Ficoll). IgM and IgG3 antigen-specific titers to NP-Ficoll in VHL cKOs were similar to WT mice. Testing of B cell responses to T-dependent antigens (NP-OVA) is ongoing. Taken together, our data suggest that OCY-specific VHL deletion might be helpful to increase bone density, but at the expense of B cell numbers. Further investigation is required to understand the molecular mechanisms by which VHL in OCYs contribute to its putative role as a B cell niche cell.
Long-lived plasma cells regulate patterns of hematopoiesis in aging.
Authors: Pioli Peter D. , Casero David , Montecino-Rodriguez Encarnacion , Morrison Sherie L. , Dorshkind Kenneth A.
Published: January 2023

In both humans and mice, aging results in significant alterations to the patterns of bone marrow hematopoiesis. This is highlighted by a precipitous reduction in B lymphopoiesis while myelopoiesis is enhanced. Treatment of young mice with factors such as GM-CSF or IL-1 can skew hematopoiesis towards myeloid development. This suggests that imbalances in the relative concentrations of various cytokines such as inflammatory mediators, whose production is thought to increase with age, could contribute to suppressed lymphoid and enhanced myeloid development in aging. In this case, the resulting stimulation of myelopoiesis would dominate the limited number of niches in the bone marrow environment at the expense of lymphopoiesis. However, whether the age-related decline in lymphopoiesis and increase in myelopoiesis are linked or represent distinct processes is not known. We observed that long-lived plasma cells from old mice acquire an inflammatory gene signature and accumulate in the bone marrow. Addition of long-lived plasma cells to long-term in vitro cultures demonstrated the ability of these cells to promote granulopoiesis from hematopoietic stem cells as well as myeloid progenitors. However, while depletion of long-lived plasma cells in old mice by injection of anti-CD138 antibodies resulted in a significant reduction in myeloid development, B lymphopoiesis remained suppressed. These data demonstrate a previously unappreciated ability of inflammatory plasma cells to stimulate myelopoiesis and provide evidence that, in the context of aging, enhanced myelopoiesis and depressed lymphopoiesis are distinct, independently regulated processes.
Regulation of B Lymphocyte Development by Extracellular ST6Gal-I Sialyltransferase
Authors: Irons Eric Edward , Lau Joseph
Published: January 2023

Post-translational glycosylation of membrane proteins is increasingly appreciated as a means by which cell signaling and transcriptional networks are regulated. Although canonical glycosylation occurs intracellularly, the presence of active glycosyltransferases in the blood has been recognized for some time. We have recently reported that these extracellular enzymes, long thought to be functionless, can in fact remodel glycans by the extrinsic pathway of glycosylation. This axis enables glycosyltransferases to systemically regulate immunologically-relevant processes such as leukocyte production and IgG sialylation. In this study, we report that B cell development is both intrinsically and extrinsically regulated by the α2,6-sialyltransferase ST6Gal-I. We observed that endogenous expression of ST6Gal-I within bone marrow and splenic B cells could not fully account for cell surface sialylation. Bone marrow transplantation experiments between wild-type and ST6Gal-1 KO mice revealed an unexpected contribution of extrinsic ST6Gal-I towards maintenance of type 1 transitional B cells, the target of negative selection in the periphery, whereas intrinsic ST6Gal-I was necessary for marginal zone B cell development. The deficit in transitional B cells upon loss of host ST6Gal-1 was not due to impaired migration from the bone marrow, but a developmental block between the immature and transitional stages. Extrinsic sialylation of immature B cells enhanced BAFF-dependent signaling, CD23 expression, and elevated serum anti-histone auto antibodies in vivo. Collectively, these results highlight a novel glycan-dependent axis of regulation of B lymphopoiesis by extrinsic ST6Gal-I, with implications for autoimmunity.
IL-27 induces autophagy during monocyte-to-macrophage differentiation through a novel LC3-independent pathway
Authors: Laverdure Sylvain , Wang Ziqiu , Nagashima Kunio , Lane H. Clifford , Imamichi Tomozumi
Published: January 2023

IL-27, a member of the IL-12 cytokine family composed of IL-12, IL-23 and IL-35, has previously been shown to be an antiviral cytokine inhibiting infection of several viruses, among which HIV-1, HIV-2, HSV, HCV, HBV and Influenza virus. We have already reported that IL-27 induces HIV-1 inhibition in macrophages and dendritic cells in an IFN-independent manner, and that the cytokine enhances potential reactive oxygen species induction from cells. In this study, we demonstrated for the first time that IL-27 triggers autophagy induction in human AB serum-induced macrophages, while other IL-12 cytokine family members did not have such effect on these cells; IL-27 was able to induce autophagy both in terminally differentiated macrophages, causing a 2 to 5-fold increase in autophagosome accumulation, as well as during monocyte-to-macrophage differentiation, where IL-27 effect was even stronger (generating a 20-fold increase in autophagosome formation). Furthermore, we also determined that two key autophagy proteins, mTOR and LC3, were not involved in this phenomenon: while we were able to assess autophagosome accumulation in IL-27-treated cells via electron microscopy and immunofluorescence, this was not linked to either mTOR dephosphorylation nor LC3 lipidation, implying that IL-27 induces autophagy through a novel non-canonical pathway. Finally, we established that blocking IL-27 signaling during macrophage differentiation using both Ruxoliotinib or Tofacitinib, respectively JAK1/2 and JAK3 inhibitors, successfully inhibited autophagy triggering together with STAT3 phosphorylation, which suggests that the JAK-STAT pathway is responsible for regulating IL-27-induced autophagy.
Identification of a conserved multi-lineage Il9 enhancer
Authors: Koh Byunghee , Qayum Amina Abdul , Fu Yongyao , Kaplan Mark H.
Published: January 2023

The immune system provides resistance to the myriad of pathogens in the environment, but can also respond inappropriately causing allergic inflammation and autoimmune disease. CD4+ T cells, which play a crucial role in adaptive immune system, can be divided into subsets based on their effector functions. T helper 9 (Th9) cells, derived by the IL-4/STAT6 and TGFβ signaling pathways, produce IL-9 as a hallmark cytokine. Through IL-9 production, Th9 cells protect against parasite infection but are also involved in allergic inflammation and autoimmune diseases. In this study, we demonstrate that a conserved noncoding sequence (CNS) located 25kb upstream of the Il9 transcription start site, termed Il9 CNS-25, is critical for regulating Il9 expression in Th cell subsets and IL-9 producing CD8+ T cells (Tc9). Th9 cells derived from Il9 CNS-25 mutant (Il9ΔCNS-25) mice produce significantly less IL-9. Il9 CNS-25 was bound by factors that promote Il9 gene expression including STATs, IRF4 and BATF, promoted chromatin modifications and transcription factor binding at the promoter, and mediated accessibility of the locus. Il9ΔCNS-25 mice showed attenuated airway inflammation compared to control mice. The Il9 CNS-25 region in mice is conserved with an IL9 CNS-18 region in the human genome. We deleted CNS-18 in primary human Th9 cells and observed diminished IL-9 production. Thus, we have demonstrated that the Il9 CNS-25/IL9 CNS-18 elements are respectively critical for Il9/IL9 gene expression in Th9 cells and other IL-9 producing T cell subsets.