bluuebunny/crossref_metadata_2025 huggingface dataset:
Therapeutic vaccination by two DerG LEAPS conjugates incorporating different PG (aggrecan) epitopes protect by different immune mechanisms in the PG G1 domain induced mouse model of rheumatoid arthritis
Authors: Zimmerman Daniel H , Mikecz Katalin , Rosenthal Kenneth S , Markovics Adrienn , Carambula Roy , Ciemielewski Jason , Glant Tibor
Published: January 2023

Background

Development of a therapeutic vaccine for rheumatoid arthritis (RA) requires modulating antigen specific inflammatory T cell responses that drive disease. RA can be initiated and driven by responses to multiple antigens but peptide vaccines usually contain few epitopes. The challenge is to elicit the appropriate responses by these vaccines.

Method

We used the proteoglycan (PG, aggrecan) G1 domain-induced arthritis model of RA. Mice were immunized with either one or two different DerG LEAPS vaccines containing PG epitopes (PG70 or PG275Cit) in Seppic ISA51vg adjuvant subcutaneously after signs of arthritis were noted. Mice were examined trice weekly for arthritis progression. At study end serum antibodies to the vaccine epitopes and splenic T-cell responses to PG G1 domain were determined.

Results

Immunization with DerGPG70 or DerGPG275Cit conjugate, alone or in combination, stopped the progression of disease. DerGPG275Cit was effective in protection but did not induce significant serum antibodies whereas DerGPG70 (alone or with DerGPG275Cit) induced both protection and antibodies. Spleen cells from DerGPG70-immunized mice had an anti-inflammatory response to PG G1 in vitro.

Conclusions

Immunization with DerG LEAPS conjugates of different epitopes of PG alone or together were effective at modulating the inflammatory response and stopping disease progression. As observed previously, the DerGPG70 conjugate modulated disease through up-regulation of Th2, and down-regulation of Th1 and Th17 responses. Results suggest mechanistically different immune responses to the two vaccines. Combination of them provided not only broader epitope coverage, but also a greater therapeutic effect than either vaccine alone.
An Accidental Discovery of a New Potential Mouse Model of the Axial Spondyloarthritis
Authors: Banda Nirmal K
Published: January 2023

Many preclinical mouse models of arthritis exists but none of the axial spondyloarthritides (axSpA) mouse model to examine the pathogenic mechanism(s) and the effect of drugs. Collagen antibody induced arthritis (CAIA) is one of the most widely used mouse model of arthritis and it is complement-dependent. We found in 100s of mice developing CAIA also developed axSpA. To induce CAIA in mice, at day 0, mice were injected intraperitoneally with anti-collagen antibodies, followed by lipopolysaccharide injection at day 3. On day 10, mice were sacrificed and assessed according to gross spinal morphology and paw synovitis scores. CAIA mice demonstrated an exaggerated kyphosis through the spine, as well as hypertrophic cartilage and osseous damage of the intravertebral joints. ImmunoHistoChemical staining of the kyphotic area revealed increased complement C3 staining and macrophage staining, with localization to the intravertebral joint margins. In vivo imaging showed that anti-collagen antibodies-conjugated with IRDye® 800CW not only localized to cartilage surface in the joints but also in the spine in arthritic mice. Thus CAIA exhibited salient features of axSpA therefore it could be a by-product of autoimmune arthritis. This new mouse model of axSpA may allow investigators to shed light on the basic aspects of axSpA development and treatment.
Reconstitution of iNKT cells enhances antitumor immunity through liver education of iNKT cells and modulation of tumor microenvironment
Authors: Zhang Hui , Zhu Zhaohui , Zhang Faya , Modrak Samantha
Published: January 2023

Invariant natural killer (iNKT) cells are a specific population of T cells that functionally are more like innateimmune regulatory cells and play important roles in antitumor immunity. It is reported that reconstitution of iNKT cells enhances antitumor immunity in human and mice, however, the underlying mechanism is not known. Using thymocytes and splenocytes to reconstitute iNKT cells in iNKT KO mice, we found that reconstitution of iNKT cells dramatically inhibits tumor development, progression and metastasis, and increases the survival of tumor-bearing mice. Most of the transferred cells accumulated in the liver. After two weeks of transfer around 70% of the transferred thymocytes in the liver differentiated into iNKT cells. Transferring CXCR6− thymocytes enhanced antitumor immunity, transferring CXCR6− splenocytes inhibited antitumor immunity. The enhanced antitumor immunity was abolished in IFN-γ and iNKT double KO mice, which suggests that this antitumor immunity is IFN-γ-dependent. Depletion of NK cells, but not CD8+ T cells or CD4+ T cells, diminishes iNKT cell reconstitution-induced antitumor immunity. Reconstitution of iNKT cells increased tumor-infiltrated lymphocytes (TIL), specifically IFN-γ-producing CD8+ T cells and NK cells, but decreased MDSC and M2 macrophages and down regulated the expression of TGFβ, Arginase and iNOS in the tumor. Taken together, liver education of iNKT cells plays a key role in iNKT cell reconstitution-induced antitumor immunity. This enhanced antitumor immunity depends on IFN-γ and NK cells of the host through modulating tumor microenvironment.
Targeted ablation of FoxP3+ T cells activates peripheral and tumor-infiltrating cytotoxic CD8+ T cells in multiple syngeneic mouse tumor models
Authors: Delfino Teresita , Wang Yingyun , Schartner Jill , Rutz Sascha
Published: January 2023

FoxP3+ regulatory T cells (Treg) are essential to prevent autoimmunity, but have also been implicated in suppressing anti-tumor immune responses. We assessed the impact of Treg depletion in 8 different syngeneic tumor models, including cancer immunotherapy-sensitive and -resistant models. Robust, rapid tumor control or regression was observed in all models tested. Analysis of both peripheral and tumor-infiltrating T cells revealed that Treg depletion resulted in robust proliferation and cytokine production by CD8+ T cells in lymph nodes, spleen, and tumor tissue. Tumor regression was CD8+ T cell-dependent, as antibody mediated-depletion of these cells abrogated tumor regression in a representative model. Interestingly, studies using FTY720 to pharmacologically block T cell egress from the lymph nodes suggest that following Treg depletion, cancer immunotherapy-sensitive models may not require new T cell infiltration into the tumor for tumor regression. This is in contrast to an immunotherapy-resistant model, which failed to regress following Treg depletion when lymph node egress was concurrently blocked. These data suggest that Tregs play a critical role in suppressing both priming of novel anti-tumor CD8+ T cells responses as well as limiting effector functionality of tumor-infiltrating CD8+ T cells within the tumor microenvironment.
Intratumoral Localization of Pattern Recognition Receptor Signaling Informs CAR T Cell Design
Authors: Johnson Lexus , June Carl H , Minn Andy J
Published: January 2023

Immune therapies have significantly improved outcomes for cancer patients with poor prognosis, but mechanisms that underlie response or resistance to therapy remain elusive. Type I and Type II interferon signaling is required for immune activation but is also known to drive programs of tumor-intrinsic resistance. We propose that activation of pattern recognition receptor (PRR) signaling in specific cellular compartments underlies this seemingly contradictory observation. We have recently identified the highly structured RNA RN7SL1 (7SL) as a novel intratumoral PAMP (pathogen associated molecular pattern) capable of activating PRR signaling. When over-expressed in tumor cells, the resulting tumors are resistant to immune checkpoint blockade (ICB) therapy. However, 7SL is stimulatory to primary human DCs, and improves T cell priming in vitro. Furthermore, direct intratumoral injection of 7SL improves immune activation and response to ICB in vivo. Thus, we propose the localization of PAMP recognition and PRR signaling may represent an important determinant of response to ICB therapy. Based on this finding, we have developed a CAR T cell capable of producing 7SL in the tumor microenvironment, as well as a fully syngeneic B16 solid tumor model for CAR T cell therapy. Murine 7SL-CAR T cells control tumors more robustly than controls and synergize with ICB. Importantly, this effect is dependent on endogenous T cells, indicating that 7SL-CAR T cells are jump-starting an endogenous polyclonal anti-tumor immune response. In total, this represents a novel insight into the role of PRR signaling in tumors and leverages the findings to create a therapy with potential to serve patients that are currently refractory to CAR-T or ICB therapies alone.
Robust antitumor effects of SNA-based T cell therapy
Authors: Dominguez Donye , Chernyak Natalia , guan Monica , Chou Yushang , Long Alan , Qin Lei , Cole Lisa , Ann Jihae , Chen Siqi , Fan Jie , Lee Andrew , Mirkin Chad , Zhang Bin
Published: January 2023

The use of toll like receptor (TLR) agonists for immunotherapy remains under intense investigation. Activation of TLRs unleash the power of dendritic cells (DCs) to induce a robust antitumor CD8+ T cell response. However, the role of TLRs in CD8+ T cells has been largely overlooked. T cells do express endosomal TLR9, but it is not readily targetable in these non-phagocytic cells. Using the novel delivery platform of spherical nucleic acids (SNAs), we have devised a way to efficiently activate endosomal TLR9 by CpG, while simultaneously delivering tumor antigens to intracellular compartments in T cells. Activation of TLR9 in CD8+ T cells endows them with special properties. First, T cells themselves express appreciable levels of co-stimulatory molecules after activation of CpG derived from SNAs, thereby functioning as antigen presenting cells to cross-prime antigen-specific CD8+ T cells. Second, SNA activated T cells act as chaperones (T chaperones) of SNA material. Through exosomal transfer, T chaperones shuttle CpG and antigen, mouse or human, to bystander DCs for further cross-presentation of antigen. Third, antigen specific T chaperones are highly cytotoxic and resist tumor induced immune suppression. These combined effects of adoptive therapy using tumor antigen specific T chaperones achieve tumor regression in aggressive melanoma models without irradiation or cytokine supplementation, which complicate T cell-based therapies. T chaperones represent a novel cell-based therapy that co-delivers adjuvant and antigen to efficiently boost antitumor CD8+ T cell activity, eliminating the need for viral manipulation and shortens the long, and costly, production time associated with other T cell therapies.
A lipid mediated paracrine signaling network stimulates tumor associated macrophage development in cancer and metastasis
Authors: Rai Vivek , Ray Rashmi
Published: January 2023

Tumor microenvironment consist of dynamic interactions between tumor cells and the surrounding non-transformed cells. Inflammatory cells constitute a major population of the non-transformed cells. Tumor associated macrophages (TAMs), the predominant population of inflammatory cells have major roles in cancer progression and metastasis but the exact stimulus and triggers for the tumor cells-macrophage interaction remains unclear. Autotaxin or lysophospholipaseD (LysoPLD) catalyses the synthesis of lyophosphatidic acid - the smallest phospholipid from lysophosphatidylcholine by its enzymatic action. Autotaxin is implicated in breast cancer, ovarian cancer and many other cancers. Lysophosphatidic acid (LPA) is involved in numerous biological processes encompassing cell growth, cell proliferation, cell migration, cancer and metastasis. LPA effects are mediated on different cell types via its cognate G-protein coupled receptors (GPCRs) or non-receptor pathways. Our recent study has shown that LPA converts monocytes into macrophages both in mice and humans and has an important role to play with immune cells. Here, we show that cancer cells in tumor growth are associated with tumor associated macrophages via a signature paracrine link. Our studies identify previously unknown signaling link between tumor cells and macrophages. Furthermore, we identify that suppression of this paracrine network can suppress tumor growth. This study suggests that inhibition of this paracrine network may act as a new therapeutic approach to control cancer and metastasis
Immune microenvironment landscape and the role of neutrophils in glioblastoma
Authors: Friedmann-Morvinski Dinorah , Magod Prerna , Agemy Lilach , Rousso-Noori Liat
Published: January 2023

Glioblastoma (GBM) is an aggressive, highly invasive primary brain tumor with near total fatality. Using a Cre-inducible lentiviral GBM mouse model we previously showed that gliomas can originate from terminally differentiated neurons and astrocytes, which can dedifferentiate to a stem cell-like state upon transformation. We believe that the tumor microenvironment (TME) may contribute to the process of tumor reprogramming. Although the majority of infiltrating cells in the tumor are peripheral macrophages and microglia, recent appreciation of the effects of neutrophils in cancer directed our efforts in understanding their role in GBM.

Flow cytometry analysis revealed differences in the brain TME of both the innate and adaptive immune populations compared to healthy brain tissue, changes were also seen in spleen and bone marrow even at early stages of GBM development. The neutrophils population varies not only at different time-points but also between tumor subtypes. We believe neutrophils switch from anti-tumor to pro-tumor phenotype. Depletion of neutrophils right before tumor initiation accelerated the onset of the disease while co-transplantation of equal ratio of glioma cells and naïve neutrophils delayed the initiation of tumors lesions. In-vitro assays showed higher migration and formation of neutrophils extracellular traps (NETs) on exposure to glioma cells condition media or glioma derived exosomes. Our findings suggest neutrophils play a role in tumor initiation and progression, and further understanding the transition of neutrophils from anti-tumor to pro-tumor phenotype will shed light into new strategies to treat GBM.
Isorhapontigenin shows strong anti-melanoma cell activity, and induces anti-tumor immunity
Authors: Song Yinhong , Yu Zouzou , Tong Chenxi , Liu Mei , Jiang Xiaoling , Zeng Yan , Chen Changrong
Published: January 2023

Background

Melanoma is one of the deadliest forms of skin cancer, and its incidence has continued to rise during the past decade. Isorhapontigenin (ISO) is a new derivative of stilbene compound, isolated from Chinese herb Gnetum Cleistostachyum with capability of antitumor.

Purpose

The potential anti-melanoma activity, impact on genes’ expression, as well as the influence of immune response against melanoma of ISO were detected in our studies in order to clarify its anti-melanoma effect and the corresponding mechanism.

Methods

Cytotoxicity of ISO on melanoma cell line B16F10 was studied by performing 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Cell cycle analysis by flow cytometry, annexin V staining, RT-qPCR and Western blot were used to elucidate the mechanism of action of ISO at the cellular level. The in vivo anti-tumor ability of ISO was measured in C57BL/6 mouse syngraft models.

Results

ISO induced dose-dependent inhibitory effect on the proliferation of B16F10 cell by arresting cells at G0/G1 phase promoting the apoptosis of the cell, which was associated with an increase in the Bax:Bcl-2 ratio and decrease in the proliferative-related genes CYCLIN D1 and SURVIVIN in vitro. In addition, ISO could inhibit the tumor immune escape by promoting the expression of MHC molecules on the membrane of tumor cells, and decreased the infiltration of Tregs and MDSCs which involved in suppressing the anti-tumor effects of immune system. On the other hand, ISO increased the infiltration of T cells, NK cells and NKT cells, which was able to enhance the anti-tumor immune response.

Conclusion

Taken together, these findings indicate that ISO may be useful as a treatment for Melanoma and thus warrants further investigation.
CD122-selective IL-2 complexes treat ovarian carcinomas and melanoma, alter Treg differentiation and induce more CD8+CXCR5+TCF-1+ stem T cells when combined with αPD-L1
Authors: Deng Yilun , Gupta Harshita B. , Drerup Justin Michael , Reyes Ryan Michael , Mendez Jenny , Zhang Xinyue , Padron Alvaro S , Garcia Myrna , Curiel Tyler J
Published: January 2023

Anti-tumor T cells (Teffs) engage IL2 largely through CD122, vs. CD25 for regulatory T cells (Tregs). In ID8agg ovarian cancer (OC), CD122-selective IL2 complexes (IL2c, made from CD122-blocking ab+IL-2), increased Teff numbers and functions as expected but unexpectedly reduced tumor Treg function, with near-complete tumor rejection in OC. IL2c reduced lineage specific molecules (e.g., Tbet) and cytokines (e.g., IFNg) in tumor Tregs, suggesting IL2c improves Treg differentiation, but not explaining IL2c-driven Treg defects. In vitro, IL-2 and IL-2c produced similar Treg numbers and Foxp3 from naïve CD4+T cells, with functional data pending. αPD-L1 improved IL2c efficacy for full tumor rejection in OC and B16 melanoma. Increased CD8+CXCR5+PD1+Tim3−TCF-1+stem-like T cells (CXCR5+) are associated with favorable αPD-1 or αPD-L1 outcome in mice and humans. IL2c + αPD-L1 markedly increased CXCR5+in B16 melanoma and to a lesser extent in OC. IL2c + αPD-L1 increased tumor CXCR5+perforin expression in B16 vs. either single agent. Injecting tumor-infiltrating CXCR5+into RagKO mice produced CXCR5+Tim3− and CXCR5−Tim3+cells confirming tumor CXCR5+stem cell properties. These data demonstrate several novel IL2c mechanisms to exploit in distinct cancers and treatment combinations. Ongoing work tests IL2c, αPD-L1 and combo treatment effects on CXCR5+ stemness and mechanisms for CXCR5+ induction, and mechanisms for IL2c-driven Treg dysfunction.
An engineered vaccine encompassing an MHC class I antigen presenting molecule induces self-reacting CTL in a setting of profound tolerance
Authors: Pease Larry R , Parks Christopher A , Kranz David , Barry Michael
Published: January 2023

Engineered antigen presenting molecules can promote allorecognition and activate CTL capable of recognizing weak or self-antigens in the context of self-MHC. Traditionally, allogeneic antigen presenting molecules differ from self MHC molecules in their interactions with the bound peptides and with TCR expressed in the immune repertoire to promote peptide dependent alloimmunity. Alloreactive T cells target their inducing alloantigens, as well as third-party alloantigens, but as a result of tolerance, generally not self-antigen presenting molecules. Certain residues located on the alpha-1/2 domains of class I antigen presenting molecules interface with TCR CDR loops and do not influence the nature or orientation of bound peptides. These residues are more conserved within and across species than are residues that determine peptide antigen binding properties. Class I variants designed with amino acid substitutions at key positions within the conserved helical structures at this MHC:TCR interface are found to provide strong activating signals to alloreactive CD8 T cells in the self tolerant immune repertoire. We demonstrate that CTL activated in this manner can break self-tolerance by reacting to self-peptides presented by native MHC. The ability to activate self-tolerant T cells capable of cross-reacting on self-peptide-MHC in vivo represents a novel approach for inducing autoimmunity, with possible application the development of cancer vaccines.
Single-cell RNA sequencing reveals different subsets of macrophage and dendritic cells in human skin
Authors: Xue Dan , Tabib Tracy , Morse Christina , Lafyatis Robert
Published: January 2023

Objective

Single-cell RNA sequencing(scRNA-seq) was performed to cells from whole human skin without pre-purifying to reveal the transcriptional landscape and heterogeneity of macrophages and dendritic cell (Mϕ/DC) subsets.

Methods

Single-cell RNA sequencing was implemented to ten enzymatically-digested human skin biopsies and analysis was performed with Seurat, then with gene ontology (GO) enrichment analysis and immunofluorescent staining.

Results

Upon scRNA-seq, 27869 cells from ten skin samples were analyzed in Seurat and 22 distinct clusters were identified using smart local moving clustering. One cluster highly specifically expressed MS4A4A, CD1C, HLA genes and other Mϕ/DC traditional markers, verifying as Mϕ/DC. Upon reanalysis of this cluster (988 cells), we identified three macrophage subsets, marked by high expression of CCR1, MARCO or TREM2; six dendritic cell subsets, marked by high expression of CXorf21, MCOLN2, CLEC9A, KIAA0101 or LAMP3 and one Langerhans cell subset. The KIAA0101 subset expressed a set of marking proliferation genes and all were in G2M phase, indicating represent a progenitor population. GO enrichment analyses indicated different putative functions of Mϕ/DC subsets. Then, the two major macrophage subsets could be defined immunofluorescently by CCR1 and MARCO double staining. And immunofluorescent staining of TREM2 and MS4A4A verified the existence of TREM2 macrophages.

Conclusion

Using scRNA-seq, we were able to uncover the transcriptional landscape and phenotypic heterogeneity of Mϕ/DC subsets in human skin, which will open up more opportunities to explore distinct immune cell populations and their roles in health and pathological conditions.
A single infection with a malaria parasite protects mice from lethal autoimmune glomerulonephritis
Authors: Amo Laura , Kole Hemanta K , Scott Bethany , Moore Ian N , Qi Cheng-Feng , Wu Juan , Wang Hongsheng , Bolland Silvia
Published: January 2023

The progression of autoimmune diseases such as Systemic Lupus Erythematous (SLE) can be altered by environmental factors, including parasitic infections. Epidemiological studies have shown a relative absence of SLE in West Africa (areas of endemic malaria), even though SLE is highly prevalent in populations of African descent living outside of Africa. We hypothesize that malaria infections might reduce the incidence of lupus. In the present work, we assess the effect of the non-lethal Plasmodium yoelii 17XNL in a well-characterized animal model of SLE (FcγRIIB−/−). Mice deficient in the inhibitory Fc receptor for IgG (FcγRIIB) develop spontaneous disease driven by the production of anti-nuclear autoantibodies, a broad immune activation, splenomegaly, anemia and lethal glomerulonephritis with a pathology similar to human SLE. We infected FcγRIIB−/− mice at 2 months of age with P. yoelii and followed the progression of autoimmune disease for 5 months. Infection with P. yoelii increased the titer of a broad range of autoantibodies, with concomitant immune-complex deposition and complement fixation in the kidney. While lupus-prone mice succumbed from renal disease detected by protein in urine, malaria-infected lupus-prone mice did not develop proteinuria or lethal disease. Infection with the parasite did not alter tolerance mechanisms leading to lupus but instead prevented end-point tissue pathology. P. yoelii infection seems to inhibit pathogenic leukocyte infiltration without affecting the autoimmune inflammation of glomeruli. These results indicate a possible route for lupus protection that specifically targets renal function, which remains a cause of the substantial morbidity and mortality in SLE patients.
Ikaros regulates chromatin landscape in mature B cells, and is critical for B cell tolerance
Authors: Rodriguez Princess D , Howell William , Amiel Eyal , Frietze Seth , Schjerven Hilde
Published: January 2023

The zinc finger transcription factor Ikaros plays central roles in the gene regulatory programs associated with proper B cell development. While mutations in the IKZF1 gene have been implicated in autoimmune disease and various B cell malignancies, the resultant epigenetic and transcriptional alterations in Ikzf1-mutated B cells remain largely unknown. We recently found in mouse models that deletion of the fourth zinc finger domain in the Ikzf1 gene (Ikzf1ΔF4/ΔF4) results in a B cell hyper-reactive (HR) phenotype. We hypothesized that Ikaros is required to establish and maintain an epigenetic program for co-stimulatory activation of B cells. To test this hypothesis, we have generated genome-wide transcriptome and chromatin-accessibility profiles of wildtype and Ikzf1ΔF4/ΔF4naïve splenic B cells following ex vivo co-stimulatory activation. We found that Ikaros mutant B cells have an overall similar accessible chromatin landscape as wildtype, but Ikzf1ΔF4/ΔF4B cells exhibit deregulated chromatin structure at inflammatory gene pathways linked to B cell growth and survival. To then explore the direct Ikaros targets we determined the Ikaros chromatin binding sites by Cleavage Under Targets & Release Using Nuclease (CUT&RUN) in wildtype B cells. Integration of gene expression and chromatin binding data indicates that Ikaros has a role in the repression of gene targets associated with cell proliferation and inflammation. Our data supports the model that Ikaros antagonizes NFkB actions at inflammatory gene pathways. Overall, our results reveal distinct mechanisms of Ikaros in the regulation of inflammatory genes in mature B cells and defines altered pathways that may be responsible for the HR phenotype of Ikaros-mutated B cells.
Multiple Myd88-dependent Toll-like receptor signaling pathways mediate inflammation in a mouse model of primary Sjögren’s syndrome
Authors: Kiripolsky Jeremy , Kasperek Eileen M , Yu Guan , Kramer Jill M
Published: January 2023

Primary Sjögren’s syndrome (pSS) is an autoimmune disease characterized by exocrine gland dysfunction and immune hyperactivity. The adaptor Myd88 is critical for immune function, as most TLRs utilize Myd88 for signal transduction. Previous work by our group found Myd88 is required for pSS disease. Our objective was to identify the specific Myd88-dependent TLR pathways that mediate salivary and systemic inflammation. We used the pSS mouse model NOD.B10Sn-H2b/J (NOD.B10), Myd88-deficient NOD.B10 mice (NOD.B10Myd88−/−), and C57BL/10 controls. For all experiments we used female mice that were at least 26 weeks old, the age that NOD.B10 females display clinical disease. We isolated spleens from NOD.B10 mice and C57BL/10 controls and performed RNA-sequencing. We then harvested salivary tissue or spleens from NOD.B10, NOD.B10Myd88−/−, and C57BL/10 mice. We performed qPCR and flow cytometry to assess expression of Myd88-dependent TLRs. We cultured salivary tissue from NOD.B10 and NOD.B10Myd88−/− mice and performed cytokine arrays on the supernatants. Finally, we treated salivary tissue with a TLR4 inhibitor and quantified spontaneous IL-6 secretion. We identified dysregulation of numerous TLR-related networks in pSS splenocytes. We found upregulation of Myd88 dependent-TLRs in both the spleen and salivary tissue from NOD.B10 mice. Salivary tissue from Myd88-sufficient NOD.B10 females exhibited spontaneous secretion of IL-6, MCP-1 and TNFa, and this was diminished in NOD.B10Myd88−/− mice. Finally, TLR4 blockade reduced IL-6 secretion in NOD.B10 salivary tissue. Our preliminary data suggest Myd88-dependent pathways contribute to the inflammatory landscape in pSS, and inhibition of such will likely have therapeutic utility.
Regulation of plasmacytoid dendritic cells function via ER stress pathway
Authors: chaudhary vidyanath , Barrat Franck J.
Published: January 2023

Systemic sclerosis is a life-threatening autoimmune disorder which causes fibrosis and chronic inflammation. One of the hallmarks of systemic sclerosis is production of high levels of type I interferon, which plays a significant role in the activation of B cells for antibody production. Plasmacytoid dendritic cells (pDCs) are the key immune cells producing high levels of interferon in autoimmune diseases such as systemic sclerosis. There is evidence suggesting that the microscopic structure of pDCs shows abundant levels of endoplasmic reticulum (ER) in their cytoplasmic compartment. Besides this, studies in patients with autoimmune diseases show that ER stress is induced in the peripheral blood mononuclear cells(PBMCs). We have observed that induction of ER stress in pDCs isolated from the PBMCs of healthy individuals markedly reduces the production of cytokines in TLR9-induced pDCs. This indicates that ER stress might be involved in the regulation of pDCs function. Hence, it is intriguing to further study the underlying mechanisms of how activation of ER stress in pDCs may be involved in the pathogenesis of autoimmune diseases.
Small numbers of T cells specific to different myelin-antigens cooperate to induce autoimmunity in a murine model of multiple sclerosis
Authors: Robinson Rachel R , Rojano Elba M , Forsthuber Thomas
Published: January 2023

T cells play a crucial role in many autoimmune diseases including multiple sclerosis (MS). Despite this, mechanisms of peripheral tolerance and dysregulation of these mechanisms during autoimmunity remain elusive. Myelin-reactive T cells are present in the blood of both healthy controls and patients with MS and T cells from patients with MS react to many different myelin peptides. The strongest genetic risk factor for developing MS is the HLA-DRB1*1501 haplotype, indicating that peptide binding to this MHC aids in disease induction. It is unlikely and not supported by current human studies that patients exhibit large populations of T cells specific for a single myelin-antigen. In contrast, a quorum of 2–5 low avidity autoreactive T cells specific for one epitope might suffice to induce autoimmunity. We hypothesized that a quorum could be established and induce autoimmunity by small numbers of antigen-specific T cells recognizing different myelin antigen peptides, particularly in genetically susceptible individuals. We utilized wild-type C57BL/6 mice and transgenic mice expressing the human MHC HLA-DRB1*1501 allele to assess the effect of inducing experimental autoimmune encephalomyelitis (EAE), the murine model of multiple sclerosis, with low numbers of different myelin peptide-specific T cells or myelin peptides at suboptimal doses. In this proof-of-principle study, we found that EAE can be initiated by the cooperation of small numbers of myelin peptide-specific T cells reactive against different myelin epitopes. The results support the hypothesis that a quorum of T cells could be reached by multiple antigen-specific T cells and could suffice to initiate autoimmune disease.
Clinicopathological and Prognostic Significance of CCR5/CCL5 axis in Hepatocellular Carcinoma
Authors: Singh Santosh Kumar , Mishra Manoj K , Lillard James W. , Singh Rajesh
Published: January 2023

Hepatocellular carcinoma (HCC) is one of the world’s most devastating malignancy which accounts for more than 90% liver cancer. The poor diagnosis of HCC lead’s scientists to search for novel treatment options to improve patient survival rates. There is accumulating evidence that chemokines and chemokine receptors (CCRs) contribute to tumor development and progression, as well as metastasis. Chemokine CCL5 (RANTES) has been reported to facilitate tumor progression and metastasis in various cancers, including liver cancer. However, the function of CCR5/CCL5 interaction in HCC proliferation and migration is poorly understood. Hence, the present study was undertaken to investigate the expression and role of the CCR5/CCL5 axis interface in the progression and invasiveness of HCC. In vitro study of liver cancer cell lines SNU387, PLC/PRF 5, and SK-HEP-1 showed higher expression of CCR5. Flow cytometry, RT-PCR, western blot, immunofluorescence techniques were used to quantify the expression of chemokine CCR5 and CCL5 in liver cancer cells. To determine the clinical significance of CCR5 expressed by HCC cell lines, a microarray of liver cancer tissues stained for CCR5 and CCL5 were analyzed. Our results show significantly (p< 0.001) higher expression of CCR5 and CCL5 in cancerous tissues compared to normal liver tissues. Furthermore, in order to delineate the role of CCR5/CCL5 interaction in HCC cell proliferation and migration, we pre-treated the HCC cell lines with the Maraviroc (CCR5 antagonist) in presence of CCL5. This data demonstrates both biological and clinical significance of CCR5 and CCL5 expression that may contribute to HCC metastasis, thus inhibition of its axis could be a promising avenue to treat liver cancer.
Transcriptomic analysis of human IL-7 receptor alphalow and higheffector memory CD8+ T cells reveals an age-associated signature linked to influenza vaccine response in older adults
Authors: Park Hong-Jai , Shin Min Sun , Kim Minhyung , Bilsborrow Joshua , Mohanty Subhasis , Montgomery Ruth R , Shaw Albert , Yoo Sungyoung , Kang Insoo
Published: January 2023

Here we investigated the relationship of the age-associated expansion of IL-7 receptor alpha low (IL-7Rαlow) effector memory (EM) CD8+ T cells with the global transcriptomic profile of peripheral blood cells in humans. We found 231 aging signature genes of IL-7Rαlow EM CD8+T cells that corresponded to 15% of the age-associated genes (231/1,497) reported by a meta-analysis study on human peripheral whole blood from approximately 15,000 individuals, having high correlation with chronological age. These aging signature genes were the target genes of several transcription factors including MYC, SATB1, and BATF, which also belonged to the 231 genes, supporting the upstream regulatory role of these transcription factors in altering the gene expression profile of peripheral blood cells with aging. We validated the differential expression of these transcription factors between IL-7Rαlow and high EM CD8+ T cells as well as in peripheral blood mononuclear cells (PBMCs) of young and older adults. Finally, we found a significant association with influenza vaccine responses in older adults, suggesting the possible biological significance of the aging signature genes of IL-7Rαlow EM CD8+T cells. The results of our study support the relationship of the expansion of IL-7Rαlow EM CD8+ T cells with the age-associated changes in the gene expression profile of peripheral blood cells and its possible biological implications.
IL-2 Signals Program the Fate of Exhausted CD8 T cells
Authors: Kalia Vandana , Xiao Hanxi , Yuzefpolskiy Yevgeniy , Sarkar Surojit
Published: January 2023

The high affinity IL-2 receptor (IL-2Ra, CD25) is a key activation marker, which is transiently and heterogeneously expressed during the priming . While IL-2 signals are deemed critical for expansion of antigen-specific CD8 T cells, the role of IL-2 in programing the fate of exhausted CD8 T cells in vivo is poorly understood. Exhausted CD8 T cells are broadly divided into 2 subsets: one with brightest levels of inhibitory receptors such as PD-1, and lower mitochondrial spare respiratory capacity (SRC) – these cells are terminally exhausted and are largely unresponsive to PD-1 checkpoint blockade immunotherapy. The other subset has intermediate levels of inhibitory receptors and retains higher mitochondrial SRC – these are the clinically relevant cells that are highly responsive to PD-1 blockade. Based on the ability of IL-2 signals to drive glycolytic burnout in T cells, we hypothesized that the stochastic heterogeneity in in vivo assimilation of IL-2 signals during priming drives distinct CD8 T cell fates late during exhaustion. We sorted in vivo primed CD8 T cells based on their heterogeneity in CD25 expression, and adoptively transferred them into infection-matched chronically infected recipients. We observed that the exhausted CD8 T cells generated from CD25Hi donor cells phenotypically and functionally resembled terminally exhausted cells. Furthermore, knocking down the expression of CD25 or Special AT-rich sequence binding protein 1 (SATB1), which is known to regulate CD25 expression, resulted in better survival and function of exhausted CD8 T cells. Our data underscores the critical role of IL-2 signals in the exhaustion of CD8 T cells and has clinical significance in current combination checkpoint blockade immunotherapies.
IL-21 produced by CD4+ T cells drives differentiation of CD8+ T cells during polyomavirus encephalitis
Authors: Ren Heather , Mockus Taryn , Shwetank  , Lauver Matthew , Netherby Colleen , Jin Ge , Lukacher Aron
Published: January 2023

The demyelinating brain disease Progressive Multifocal Leukoencephalopathy (PML), caused by JC polyomavirus (JCV), is a life-threatening complication for patients with HIV/AIDS, hematologic malignancies, rheumatologic diseases, and those receiving long-term immunomodulating therapies. Using mouse polyomavirus (MuPyV), our laboratory developed a robust model of polyomavirus-associated demyelinating leukoencephalitis, complete with viral infection and T cell infiltration. Increasing evidence supports the importance of CD4+ T cells in controlling JCV in the brain and thereby preventing PML. Recent studies in our laboratory show that CD4+ T cells are essential for differentiation and maintenance of MuPyV-specific brain-resident memory CD8+ T cells (bTRM). We find that the help CD4+ T cells are provide to virus-specific CD8+ T cells for their appropriate bTRM differentiation is IL-21. PD-1hiCXCR5hi MuPyV-specific CD4+ T cells are the primary sources of brain IL-21 and co-localize with CD8+ T cells in the brain. Brain MuPyV-specific CD8+ T cells have increased expression of the IL-21 receptor (IL21R) 15 days post-infection, which correlates to the time-point when they begin to express CD103, a canonical TRM marker. TRM maintenance is independent of circulating cells. In line with this property, we show that MuPyV-specific CD8+ T cells are not depleted by parentally administered anti-CD8. In addition, brain CD8+ T cells from IL21R−/ − mice have decreased expression of CD103 and are susceptible to depletion from peripheral anti-CD8. These data indicate that during polyomavirus infection, IL-21 by CD4+ T cells drives CD8+ T cells to differentiate into tissue resident memory in the brain.
Elucidating the role of cytokine signaling in the homeostasis of innate immune cells with JAK inhibitors
Authors: Vian Laura , Lee Mimi , Sciumè Giuseppe , Gazaniga Nathalia , Dell'Orso Stefania , Brooks Stephen , Gadina Massimo
Published: January 2023

The role of the JAK/STAT pathway in the function on innate lymphoid cells (ILCs) has been mostly investigated using genetically modified animals. While elegant, these studies present with the limitation of a complete loss of ILC populations. Therefore, pharmacological manipulation of this signaling cascade with JAK inhibitors (jakinibs) is an attractive alternative strategy. Here we investigated the effect of pan- and JAK-selective inhibitors on the development and functions of murine, IFN-γ producing, ILCs and iNKT cells after oral administration of tofacitinib (JAK1; JAK3 and JAK2), ruxolitinib (JAK1; JAK2), or PF-06651600 (JAK3). We observed a significant reduction in number and function (IFN-γ production) of both splenic and hepatic NKp46+ cells with either tofacitinib or PF- 006651600. The reduction appears to correlate with the number of proliferating Ki67+ NKp46+ cells. In addition, using an iNKT-dependent mouse model of acute liver inflammation, we observed a significant decreased production of IFN-γ by liver iNKT upon tofacitinib administration. Notably, we found that PF-06651600 selectively impairs the thymic development and the proliferation of IFN-γ-producing iNKT1 cells. Overall our data suggest that the transient inhibitory activity with jakinibs has strong impact on proliferation and activity of mouse IFN- γ producing innate cells. These preliminary findings bring us one step closer to elucidating mechanisms underlying innate cells homeostasis.
Expression of GM-CSF is regulated by transcription factor Fli-1
Authors: Zhang Xian , Richard Mara Lennard , Wang Xuan , Henry Brittany , Li Pengfei , Fan Hongkuang , Gilkeson Gary S
Published: January 2023

Granulocyte-macrophage colony-stimulating factor (GM-CSF), also known as colony-stimulating factor 2 (CSF-2) plays an important role in regulating mature myeloid cell populations under both homeostatic and inflammatory conditions. GM-CSF maintains low basal levels under homeostatic conditions, but its levels can quickly become elevated during infection or inflammation. GM-CSF is rapidly produced during inflammatory or autoimmune reactions by a variety of cells, including endothelial cells, macrophages, dendritic cells (DCs), T cells, neutrophils, eosinophils, and more. It has been reported that the expression of GM-CSF is regulated by several transcription factors including NF-kB, AP-1, and RUNX1. Fli-1 belongs to the ETS transcription factor family. We have demonstrated that the Fli-1 transcription factor is a novel regulator in modulating the expression of many inflammatory mediators, including monocyte chemotactic protein 1 (MCP-1), chemokine (C-C motif) ligand 5 (CCL5) and interleukin 6 (IL-6). In this report, we found that murine endothelial cells transfected with Fli-1 specific siRNA produced significant lower levels of GM-CSF after stimulation with Toll-like receptor 4 ligand LPS compared to the cells transfected with control siRNA. Human primary glomerular endothelial cells transfected with Fli-1 specific siRNA had reduced Fli-1 protein and produced significantly less GM-CSF following LPS stimulation compared to the cells transfected with control siRNA. Transient transfection assays show that Fli-1 drives transcription from the GM-CSF promoter in a dose-dependent manner. Together, the results indicate that Fli-1 is a novel, critical transcription factor in regulating the expression of GM-CSF.
Nrf2 selectively regulates IL-22 and IL-17A production in Th17 cells
Authors: Lin Xun , Gaudino Stephen , Kolls Jay , Kumar Pawan
Published: January 2023

Th17 cell-derived IL-17A and IL-22 are critical for mounting both inflammatory and tissue protective responses. It remains unclear whether cytokine responses can be modified differently to achieve targeted functional outcome. A therapeutic strategy which inhibits CD4+ T cell-derived inflammatory IL-17A responses but concomitantly promotes IL-22-dependent tissue protective or regenerative response is of great clinical significance.

Here, we showed that nuclear factor (erythroid-derived 2)-like 2 (Nrf2) selectively regulated IL-17A and IL-22 responses in CD4+ T cells. We found that Nrf2−/− mice had reduced IL-22 responses in Ovaalbumin (Ova) + LPS and separately concanavalin A (Con A) administered mice. Furthermore, CDDO-Im, a selective Nrf2 activator, induced IL-22 but suppressed IL-17A response in CD4+ T cells polarized under Th17 cell condition. Our qPCR data revealed that CDDO-Im-activated CD4+ T cells had lower Rorc, but increased Cybb, Sod1 and Sod3 transcript. The expression pattern of Il23r and Sod2 was unaltered. CDDO-Im-dependent IL-17A but not IL-22 response was regulated by Sod3. Interestingly, we found that CDDO-Im induced aryl hydrocarbon receptor (Ahr) and its downstream Cyp1a1 and Cyp1b1 gene expression. The luciferase reporter assay data showed that CDDO-Im regulated Ahr promoter activity in a dose-dependent manner. Additionally, CDDO-Im induced Nqo1 expression, a Nrf2-activated gene, in WT mice but not in Ahr−/− mice. Finally, we confirmed that the CDDO-Immediated induction of IL-22 production in CD4+ T cells was abrogated in Ahr−/− mice.

Collectively, our data show that Nrf2 promotes IL-22 production while it inhibits IL-17A expression in Th17 cells.
Bone marrow transplantation rescues monocyte recruitment defect and improves cystic fibrosis in mice
Authors: Fan Zhichao , Miller Jacqueline , Herro Rana , Ehinger Erik , Conrad Douglas J. , Mikulski Zbigniew , Zhu Yanfang Peipei , Marcovecchio Paola M. , Hedrick Catherine C. , Ley Klaus
Published: January 2023

Cystic fibrosis (CF) is an inherited life-threatening disease accompanied by repeated lung infections and multi-organ inflammation that affects tens of thousands of people worldwide. The causative gene, cystic fibrosis transmembrane conductance regulator (CFTR), is mutated in CF patients. Monocytes from CF patients show a deficiency in integrin activation and adhesion. Since monocytes play critical roles in controlling infections, defective monocyte function may contribute to CF progression. In this study, we demonstrate that monocytes from CFTRΔF508 mice (CF mice) show defective adhesion under flow. Transplanting CF mice with wild-type bone marrow after sublethal irradiation replaced most (60–80%) CFmonocytes with wild-type monocytes, significantly improved survival, and reduced inflammation. Wild-type/CFmixed bone marrow chimeras directly demonstrated defective CF monocyte recruitment to the bronchoalveolar lavage and the intestinal lamina propria in vivo. Our findings show that providing wild-type monocytes by bone marrow transfer rescues gastrointestinal (GI) mortality in CF mice, suggesting that wild-type bone marrow stem cells might mitigate CF inflammation.
Understanding the role of IL-33 in endometriosis associated inflammation and pathology
Authors: Miller Jessica E , Koti Madhuri , Tayade Chandrakant
Published: January 2023

Introduction

Endometriosis (EM) is a chronic inflammatory disease categorized by the growth of endometrial tissue on the ovaries, peritoneal wall and other internal organs. Despite it’s prevalence (176 million women worldwide), the etiology is unknown. Currently, there is a significant knowledge gap regarding how the immune microenvironment contributes to the progression of EM (specifically inflammation, pain and fibrosis). We have shown that interleukin(IL)-33 is produced by EM lesions and drives pathology in a mouse model of EM. We are now investigating, mechanistically, how IL-33 contributes to EM and whether IL-33 neutralization could alleviate the pathology.

Methods

Female C57BL/6 mice were induced with EM and were treated with PBS (n=10) or IL-33 (n=10) every other day. After 2 weeks, the mice were euthanized and plasma, peritoneal fluid (PF) and EM lesions were collected. Cytokines in the plasma and PF were analyzed using a multiplex array. Immune cell populations in the PF were evaluated using CyTOF. Finally, sectioned lesions were stained for markers of innervation, proliferation and fibrosis. To establish cause and effect, mice were induced with EM and treated with IL-33+anti-IL-33 antibody (n=10) or IL-33+isotype control (n=10).

Results

Mice treated with IL-33 had elevated cytokines (e.g. IL-5) in the plasma and PF and EM lesions exhibited drastic changes in morphology. Additionally, both innate and adaptive immune cells were altered in the PF of IL- 33 treated mice. Neutralizing IL-33 reduced inflammation and immune cell recruitment.

Conclusions

Overall, these results show that IL-33 drives hallmark pathologies of endometriosis and neutralization of IL-33 could provide a novel therapeutic target for EM.