IL-2 Signaling Research Articles

Optimizing Early Clinical Investigations in Cancer Immunotherapy: The Translational Journey of RG6292, a Novel, Selective Treg-Depleting Antibody.

The multifaceted IL-2/IL-2R biology and its modulation by promising therapeutic agents are highly relevant topics in the cancer immunotherapy field. A novel CD25-Treg-depleting antibody (Vopikitug, RG6292) has been engineered to preserve IL-2 signaling on effector T cells to enhance effector activation and antitumor immunity, and is currently being evaluated in the clinic. The Entry into Human-enabling framework described here investigated the characteristics of RG6292, from in vitro quantification of CD25 and RG6292 pharmacology using human tissues to in vivo assessment of PK/PD/safety relationships in cynomolgus monkeys as non-human primate species (NHP). Fundamental knowledge on CD25 and Treg biology in healthy and diseased tissues across NHP and human highlighted the commonalities between these species in regard to the target biology and demonstrated the conservation of RG6292 properties between NHP and human. The integration of in vitro and in vivo PK/PD/safety data from these species enabled the identification of human relevant safety risks, the selection of the most appropriate safe starting dose and the projection of the pharmacologically-relevant dose range. The first clinical data obtained for RG6292 in patients verified the appropriateness of the described approaches as well as validated the full clinical relevance of the projected safety, PK, and PD profiles from animal to man. This work shows how the integration of mechanistic non-clinical data increases the predictive value for human, allowing efficient transition of drug candidates and optimizations of early clinical investigations.

Dietary Protein Optimization for Growth and Immune Enhancement in Juvenile Hybrid Sturgeon (Acipenser baerii × A. schrenckii): Balancing Growth Performance, Serum Biochemistry, and Expression of Immune-Related Genes.

This study aimed to evaluate the effects of dietary protein levels on growth performance, serum indices, body amino acid composition, and intestinal gene expression in juvenile hybrid sturgeon (Acipenser baerii × A. schrenckii). Hybrid sturgeons (initial weight 29.21 ± 2.04 g) were fed isolipidic diets containing 30%, 33%, 36%, 39%, 42% or 45% crude protein for 12 weeks (n = 18 tanks, 30 fish/tank). Results showed significant differences between treatments, where weight gain and protein efficiency ratio peaked optimally between 35.9% and 38.3% dietary protein. Serum parameters such as glucose, alanine aminotransferase, aspartate aminotransferase, superoxide dismutase, and lipid peroxidation levels varied significantly with changes in dietary protein levels. Specifically, the highest enzymatic activities and growth parameters were observed in groups fed with 33% to 39% protein, enhancing whole-body concentrations of lysine, leucine, phenylalanine, proline, and glutamic acid. Immune parameters such as immunoglobulin M and lysozyme activity also showed peak levels at higher protein concentrations, particularly notable at 42% for lysozyme and 36% for both component 3 and immunoglobulin M. Gene expression related to immune and growth pathways, including MyD88, TLR1, IL-8, IL-6, NF-κB, and IL1β, was significantly upregulated at protein levels of 33% to 36%, with a noted peak in expression at 39% for TLR1, IL-10, and TOR signaling genes, before diminishing at higher protein levels. Overall, the dietary protein requirement for juvenile hybrid sturgeon ranges from 35.9% to 38.3% crude protein.

185 Maternal live yeast supplementation protects newly weaned piglets against lipopolysaccharide challenge by regulating immune-associated milk proteins

Abstract Sow milk serves as a crucial source of nutritional, immunological, and growth-promoting components for the piglets. Our previous studies established that maternal live yeast (LY) supplementation altered the expression of antioxidant and immune-regulatory genes in the intestinal mucosa of suckling piglets. The current study aimed to investigate the effects of maternal LY supplementation during late gestation and lactation on milk proteomic profile in dams and health of offspring under lipopolysaccharide (LPS) challenge. On d 77 of gestation, 40 sows were allocated to 2 dietary treatments: without (CON) or with LY supplementation at 0.05% of diet during gestation and 0.1% during lactation. Milk samples were collected on d 0, 10, and 18 post-partum (n = 6), respectively. Within 24 h postweaning, piglets from the CON and LY groups were intraperitoneally injected with sterile saline or LPS [25 μg/kg body weight (BW)] and divided into four groups (n = 8): 1) saline + CON (CS), 2) saline + LY (YS), 3) LPS + CON (CLPS), and 4) LPS + LY (YLPS). Data were analyzed in a 2 × 2 factorial arrangement with main effects of maternal diet and immunological challenge. Rectal temperature was measured hourly for 4 h post-injection after which piglets were euthanized. Results showed that LPS increased concentrations of serum tumor necrosis factor (TNF) -α and interlekin (IL) -10 (P < 0.05) and tended to increase IL-8 in the ileal mucosa of piglets (P < 0.10). At 3 h post-injection, YLPS piglets tended to have a lower rectal temperature than CLPS piglets (P < 0.10). YLPS piglets also had decreased protein levels of TNF-α in the ileal mucosa (P < 0.05), with a tendency for greater protein abundance of E-cadherin but decreased mRNA abundance of catalase (P < 0.10) in the jejunal mucosa at 4 h post-injection (P < 0.05). In addition, YLPS piglets had less mRNA abundance of TLR2 and TLR4 in the liver than CLPS piglets (P < 0.05). With shotgun proteomics analysis, we found that milk of LY sows had more abundant proteins associated with immunity, including immunoglobulin and complement protein (Ig-like and C1q domain-containing proteins) on d 0 and d 10. Insulin-like growth factor binding protein (IGFBP) 5 was also more abundant in the d 0 milk of LY sows, whereas prostaglandin synthase was greater in d 10 and d 18 milk of CON sows. Additionally, pathway analysis showed that proteins identified in the milk of CON sows have roles in neutrophil degranulation and IL-12 signaling and production, whereas pathways associated with regulation of complement system, IGF and DHCR24 signaling, were enriched in the milk of LY sows. These results suggest that maternal LY supplementation could protect newly weaned piglets from bacteria endotoxin effect by suppressing inflammatory response, improving expression of tight junction proteins in offspring, and altering immune-associated proteins in sow milk.

Unpaired cysteine insertions favor transmembrane dimerization and induce ligand-independent constitutive cytokine receptor signaling.

Naturally occurring gain-of-function (GOF) mutants have been identified in patients for a variety of cytokine receptors. Although this constitutive activation ofcytokine receptors is strongly associated with malignant disorders, ligand-independent receptor activation is also a useful tool in synthetic biology e.g. to improve adoptive cellular therapies with genetically modified T-cells. Balanced Interleukin (IL-)7 signaling via a heterodimer of IL-7 receptor (IL-7Rα) and the common γ-chain (γc) controls T- and B-cell development and expansion, whereas uncontrolled IL-7 signaling can drive acute lymphoid leukemia (ALL) development. The ALL-driver mutation PPCL in the transmembrane domain of IL-7Rα is a mutational insertion of the four amino acids proline-proline-cysteine-leucine and leads to ligand-independent receptor dimerization and constitutive activation. We showed here in the cytokine-dependent pre-B-cell line Ba/F3 that the PPCL-insertion in a synthetic version of the IL-7Rα induced γc-independent STAT5 and ERKphosphorylation and also proliferation of the cells and that booster-stimulation by arteficial ligands additionally generated non-canonical STAT3 phosphorylation via the synthetic IL-7Rα-PPCL-receptors. Transfer of theIL-7Rα transmembrane domain with the PPCL insertion intonatural and synthetic cytokine receptor chains of the IL-6, IL-12 and Interferon families also resulted in constitutive receptor signaling. In conclusion, our data suggested that the insertion of the mutated PPCL IL-7Rα transmembrane domain is an universal approach to generate ligand-independent, constitutively active cytokine receptors.

The Possible Roles of IL-4/IL-13 in the Development of Eosinophil-Predominant Severe Asthma.

Bronchial asthma is characterized by airway inflammation, airway hyperresponsiveness, and reversible airway obstruction. Eosinophils contribute to the pathogenesis of airway disease mainly by releasing eosinophil-specific granules, lipid mediators, superoxide anions, and their DNA. Type-2 cytokines such as interleukin (IL)-4 and IL-13 also play roles in the development of bronchial asthma. Among these cytokines, IL-4 is involved in T-cell differentiation, B-cell activation, B-cell differentiation into plasma cells, and the production of immunoglobulin E. Although IL-13 has similar effects to IL-4, IL-13 mainly affects structural cells, such as epithelial cells, smooth muscle cells, and fibroblasts. IL-13 induces the differentiation of goblet cells that produce mucus and induces the airway remodeling, including smooth muscle hypertrophy. IL-4 and IL-13 do not directly activate the effector functions of eosinophils; however, they can induce eosinophilic airway inflammation by upregulating the expression of vascular cell adhesion molecule-1 (for adhesion) and CC chemokine receptor 3 ligands (for migration). Dupilumab, a human anti-IL-4 receptor α monoclonal antibody that inhibits IL-4 and IL-13 signaling, decreases asthma exacerbations and mucus plugs and increases lung function in moderate to severe asthma. In addition, dupilumab is effective for chronic rhinosinusitis with nasal polyps and for atopic dermatitis, and IL-4/IL-13 blocking is expected to suppress allergen sensitization, including transcutaneous sensitization and atopic march.

B cell-intrinsic IL-27 and IFNγ receptor signaling drive PD-L1 expression, IgG2a autoantibody production, VEGF-dependent tumor angiogenesis and cancer progression

Abstract B effector cells produce Abs and autoantibodies, while B regulatory cells use cytokines and transmembrane proteins to fine tune immune functions. Our lab recently revealed that tumor-infiltrating B cells produce IL-27 to promote immune evasion, partly by upregulating tumoral PD-L1, including that expressed by B cells per se. To investigate the autocrine activity of IL-27 in tumor progression, we generated μMT:Il27ra–/– bone marrow chimera mice lacking the IL-27 receptor specifically in B cells. These mice displayed substantially reduced tumor growth from orthotopically engrafted ER+ E0771 breast cancer cells, as associated with only moderately decreased PD-L1 levels but, unexpectedly, severely compromised blood vessel integrity. The residual PD-L1 levels and tumor growth were virtually abolished by additional B cell-specific deficiency in IFNγ receptor signaling, as occurring in double KO μMT:Il27ra–/–Ifngr1–/– mice, in which the microvessel permeability was also worsened. Mechanistically, this aligned with our findings that IL-27-producing B cells expressed high levels of VEGFA, whose transcription in activated B cells could be further enhanced by IL-27 and IFNγ. Double KO mice also lacked anti-dsDNA IgG2a autoantibodies, as elicited by tumor engraftment. Thus, B cell-intrinsic IL-27 and IFNγ receptor signaling synergize to promote B-cell PD-L1 expression, IgG2a autoantibody production, and vascular integrity, all of which may contribute to cancer development.

Induction of CXCL3 in airway epithelial cells by a novel IL-9 receptor complex

Abstract Cytokine signaling is critical for cell-to-cell communication in development, homeostasis and disease. Interleukin (IL)-9 signaling promotes allergic diseases by acting on both immune and structural cells. In hematopoietic cells, IL-9 receptor (IL-9R) consists of two subunits – the α-chain (IL-9Rα) and a common γ (γc) chain. However, IL-9-responsive epithelial cells express IL-9Rα but not the γc chain raising a question on how these cells respond to IL-9 in the absence of the established IL-9R complex. Here, we discovered that in airway epithelial cells, IL-9Rα pairs with IL-13Rα1, forming a novel type II IL-9R. Using proximity ligation assay, we show that the IL-9Rα /IL-13Rα1 chains are in IL-9-dependent proximity in both mouse tracheal epithelial cells (mTEC) and human small airway epithelial cells (SAEC). IL-9-induced STAT3 activation in SAEC was blocked by anti-IL-13Rα1 but not by anti-γc. Using RNA-seq, in vitro IL-9 treatment of SAEC induced the expression of genes including CXCL3, a key chemokine implicated in asthma. Importantly, intranasal administration of IL-9 increased Cxcl3 expression in mTECs and CXCL3 protein in the lung, that corresponds with an expansion in the number of CXCR2+ immune cells in the lung. Thus, we identify a novel type II IL-9R which is composed of IL-9Rα and IL-13Rα1 and show that IL-9 signaling via type II IL-9R induces CXCL3 expression in epithelial cells to contribute to airway inflammation.

Abstract 1028: Cytotoxic CD4+ T Cells: Unraveling The Residual Risk In Lipid Lowering For Type 2 Diabetes Patients

Type 2 Diabetes (T2D) significantly increases the risk of cardiovascular diseases (CVDs), marked by severe plaque pathology resistant to standard cholesterol-lowering treatments. While the roles of the innate immune system and macrophages in CVDs are well-established, T cell alterations, particularly involving cytotoxic CD4 T cells (CD4 CTLs), remain unexplored in human atherosclerotic plaques. In our study, we aimed to identify the phenotype and function of CD4 CTLs in blood & plaque tissue, in the context of T2D. Using unbiased single-cell RNA sequencing (scRNA-seq) analysis, we examined over 70,000 immune cells from 22 plaques of patients undergoing carotid endarterectomy (T2D: n=12, Non-Diabetic (ND): n=10), & peripheral blood mononuclear cells (PBMCs) from the CHORD study (NCT04369664) (T2D: n=12, ND: n=4). scRNA-seq analysis revealed a diverse population of CD4 T cells, like naïve, central memory (CM), effector memory (EM), tissue resident memory (Trm), T regulatory (Tregs), & two clusters of cytotoxic (CTL) T cells expressing canonical cytotoxicity-related genes. Cluster 5 expressed GZMB, GNLY, PRF1 , whereas Cluster 10 expressed GZMA, GZMK . Cell-cell communication analysis demonstrated that both CTL clusters received signaling from dendritic cells and macrophages, including TREM2+ macrophages. CTL T cells reciprocally communicated with clusters of EM and EMRA CD8 T cells, underscoring their central role in plaque cell-cell communication. Notably, Cluster 5, one of these two CTL clusters, was expanded in T2D plaque samples (p=0.032) and expressed genes involved in chemotaxis, IL-2 signaling, & transcription factors (e.g., RUNX3, TBX21 ) involved in T cell activation. The equivalent of CD4 CTL phenotype was found to be expanded in T2D individuals compared to controls, & these elevated levels persisted in T2D patients following intensive lipid-lowering therapy, indicating resistance to the treatment regimen. In summary, our study emphasizes the crucial role of CD4 CTLs in the immune dynamics of atherosclerotic plaques. The observed resistance of CTLs to lipid-lowering interventions underscores the necessity for targeted therapeutic strategies to address the distinctive immune changes in T2D-associated cardiovascular diseases.

Regulatory T cells tonically inhibit spontaneous activation of naturally arising memory-phenotype CD4+ T lymphocytes

Abstract Foxp3- CD4+ T lymphocytes consist of naïve (CD44lo), foreign antigen-specific memory, and self antigen-driven memory-phenotype (MP; both CD44hi) cells at homeostasis. We recently showed that self-reactive MP cells spontaneously proliferate and differentiate into the T-bet+ subset. How excess proliferation and Th1 differentiation of MP cells are inhibited in steady state remains unclear. Regulatory T cells (Tregs) can inhibit several immune responses, posing a possibility that the same lymphocytes may suppress spontaneous MP cell activation. Here we show using Foxp3-diphtheria toxin receptor-transgenic mice that both preexisting MP and naïve CD4+ T cells spontaneously proliferate and differentiate into Th1 cells upon Treg depletion. MP as compared to naïve CD4+ T cells more rapidly proliferate and differentiate three to five days post Treg depletion whereas these responses are mainly attributed to by the latter T cells at a later time point. Moreover, we demonstrate that these early MP proliferation and differentiation are driven in a manner dependent on CD28 and IL-2 signals, respectively. Furthermore, our data suggest that such activation of MP and naïve CD4+ T cells contribute to development of multi-organ inflammation at early and later time points, respectively. Together our results argue that Tregs tonically inhibit early, spontaneous proliferation and Th1 differentiation of MP CD4+ T lymphocytes, thereby contributing to maintenance of T cell homeostasis.

Siglec-H ablation in plasmacytoid dendritic cells alleviates acute liver injury by promoting IL-21+ CD4 T cells

Abstract Siglec-H is an immune receptor specifically expressed in plasmacytoid dendritic cells (pDCs) that inhibits IFN-α production and regulates pDC maturation. Although, the role of pDCs in autoimmune disorders is well-known, the effect of Siglec-H expression in pDCs in acute liver injury (ALI) remains elusive. To investigate Siglec-H/pDCs axis in ALI, BDCA2 transgenic and Siglec-H knockout (SH KO) mice were administrated with concanavalin A (ConA) to induce a T cell-mediated ALI in combination of either pDCs depletion with anti-BDCA2 antibody or anti-IL-21R and Stat3 blockade. Here, we report that specific depletion of pDCs in BDCA2 transgenic mice exacerbated ALI. Surprisingly, SH KO mice were resistant to ALI (lower ALT, hyaluronic acid, IL-6 and TNF-α). Th1/IFN-γ response and Stat1 signaling were both inhibited in the liver of SH KO mice while intrahepatic IL-21 and Stat3 signaling pathways were elevated. Mechanistically, we identified that Siglec-H-null pDCs exhibited immunosuppressive phenotypes (CD40LowCCR9Low), resulting in a defective CD4 T cell activation and a promotion of IL-21-producing CD4 T cells in the liver. Finally, IL-21R and Stat3 inhibitions in Siglec-H deficient mice restored ConA-induced ALI. We conclude that, Siglec-H-null pDCs mediate hepatic tolerance via induction of IL-21+CD4 T cells in the liver during ALI. These findings indicate that targeting Siglec-H/pDCs axis may provide a novel approach to modulate CD4 T cell polarization and liver disease.

Understanding molecular differences in endocrine and exocrine pancreatic tissue in healthy and unhealthy metabolic states

Type II diabetes (T2D) is a metabolic disorder characterized by hyperglycemia largely due the development of insulin-resistance, and in some cases, due to insuffcient insulin secretion. The endocrine portion of the pancreas, the islets of Langerhans, is composed of a diverse population of cells (beta, alpha, delta) that play vital roles in glucose homeostasis. Intercellular signaling events that occur within this diverse cell population play an active role in T2D progression. Therefore, to properly understand the progression of T2D it is critical to understand the how both the cellular neighborhoods and cell-cell communications within the islet of Langerhans and the surrounding exocrine (non-Islet) tissue contribute to the development of T2D. As the pancreas is a heterogenous organ, techniques allowing for the identification of cellular subpopulations within the tissue, such as single-cell RNA sequencing (scRNAseq), have been critical to answering questions about the pathogenesis of T2D. However, these techniques provide no insight into the anatomical and spatial context of cells, and therefore lack the ability to determine the influence of cellular neighborhoods in T2D pathogenesis. To begin to fill the gap in our understanding of how cellular neighborhoods and cell-cell communication impacts the progression of T2D, we utilized cutting edge technology that allows for the identification of RNA and protein within tissue at cellular and subcellular level, the Nanostring CosMx Spatial Molecular Imager. In this study, we aimed to determine if the location of individual cells within the islet (exocrine adjacent versus endocrine adjacent) contributes to gene expression changes that direct disease progression. To answer this question, we collected 200 human donor pancreases and analyzed them for 1000 RNA targets using the CosMX Spatial Molecular Imager. Ig. We then performed an analysis of RNA expression of exocrine cells bordering the islets of Langerhans and endocrine cells surrounding pancreatic vasculature was compared between healthy, metabolically heathy obese (MHO), and obese T2D groups. Preliminary analysis of these data yield novel insights into the transcriptomic profile of metabolically unhealthy tissues that may play a role in the progression of T2D. In particular, we observed an upregulation of mRNA transcripts associated with pro-inflammatory IL-21 and IL-22 signaling in the alpha cells from MHO donor pancreases. Furthermore, this upregulation is primarily observed in alpha cells in close proximity to acinar cells. Overall, these data will help pave the way to addressing previously unanswered questions in the context of T2D pathogenesis and unexplored cell-cell interactions that occur between the endocrine and exocrine spaces of the pancreas. Funding: Saint Louis University. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Mapping the cellular landscape of the maternal-fetal interface in women with preterm birth and chronic chorioamnionitis

Abstract Preterm birth (PTB) affects over 15 million infants yearly. In up to 40% of late PTBs, placental examination reveals chronic chorioamnionitis (CCA), maternal mononuclear cell infiltration in the chorioamniotic membranes; yet, the cellular landscape remains fully unknown. Herein, we first used imaging mass cytometry (IMC) to map the phenotype and spatial distribution of immune cells at the maternal-fetal interface of women with PTB and CCA and non-CCA controls (n = 4 each). Cases exhibited a 5-fold increase in T cells, including effector memory CD4+ (p=0.02) and proliferative CD8+ T cells (p=0.04), and a 7-fold rise in CD163+CD206+ macrophages (p=0.05) compared to controls. NK cells were also present in both groups. Neighborhood analysis revealed that CD8+ T cells are in proximity to macrophages and CD4+Tbet+ T cells, and distanced from NK cells. Next, we performed spatial transcriptomics (STx) in each study group (n = 8 PTB with CCA, n = 7 PTB no CCA). STx highlighted diversity in several clusters of T cells, macrophages, and other immune cells found to be transcriptionally active in CCA. Gene ontology analysis indicated enrichment for CD8+ T cell activation, as well as IL-12 and IFNγ signaling in macrophages in CCA. Ongoing efforts are focused on validation and functional testing of such immune cells in order to unveil the complex interactions at the maternal-fetal interface in CCA, a placenta pathology associated with adverse outcomes in the mother and offspring.

Metabolic pathways in T cells are modulated by CCR7 and IL-7R to facilitate T cell motility.

Abstract T cells play an essential role in eliminating infections by migrating through tissues and interacting with antigen-presenting dendritic cells within lymph nodes, leading to activation. A continuous supply of energy is required for T cell movement to enable scanning of dendritic cells. Many studies have established that signaling through chemokine receptor CCR7 facilitates rapid movement, however, how CCR7 signaling induces T cell metabolism remains poorly understood. IL-7 signaling regulates glycolysis to promote T cell survival, but how signaling through the IL-7R affects T cell movement is unexplored. This study investigates the roles of CCR7 and IL-7R in metabolism to drive naïve T cell movement. Utilizing Seahorse analysis, we observe that CCL21 signaling to CCR7 has no effect on glycolysis, yet decreases cellular respiration. Conversely, IL-7 has no effect on respiration but promotes glycolysis. We then determined whether oxidative phosphorylation and glycolysis play a role in CCL21 and IL-7 driven T cell movement. We employ inhibitors of oxidative phosphorylation such as rotenone to examine the influence of different metabolic pathways on T cell mobility. Our findings provide new insights into the intersection of metabolism and motility, suggesting potential differential mechanisms of ATP production that drive interactions.

Opposing roles of SWI/SNF chromatin remodeling complex BAF and PBAF in Effector Differentiation during Chronic Viral Infection

Abstract During chronic viral infection, CD4+ T cells help in the form of IL-21 is critical for the development of highly cytolytic effector CD8+ T cells. However, the exact signaling pathways from IL-21 that regulate CD8+ T cell function remain unclear. The current dogma states that IL-21 predominantly signals through STAT3, to generate effector CD8+ T cells during chronic viral infection. Here, we reveal a novel mechanism of IL-21 signaling via STAT4 for effector CD8+ T cell development following chronic LCMV infection. Furthermore, STAT4-mediated effector differentiation is epigenetically regulated by BAF, a SWI/SNF chromatin remodeling complex. In the absence of either STAT4 or BAF, virus specific CD8+ T cells fail to differentiate into highly cytolytic effector CD8+ T cells capable of controlling viral infection. The effector differentiation phenotype is balanced by PBAF, another component of the SWI/SNF complex, which promotes CD8+ T cell exhaustion during chronic LCMV infection. Conditional knockout of PBAF from CD8+ T cells impairs the development of T cell exhaustion, resulting in immunopathology and mortality of chronic LCMV-infected mice. Interestingly, this phenotype is dependent on the STAT4 and BAF-mediated transcriptional and chromatin changes. Overall, our findings underscore the delicate balance between effector and exhaustion pathways, governed by BAF and PBAF complex, to dictate the outcomes of immunopathology during chronic viral infections.

Function of tissue-resident helper T cells in chronic lung sequelae following viral pneumonia

Abstract Respiratory viral infections such as influenza and SARS-CoV-2 can lead to the development of chronic lung disease and impaired lung function long-term after the recovery from acute morbidity. However, the mechanisms underlying the development of chronic diseases post-acute respiratory viral infection are largely elusive. Using a model of age-related development of chronic lung disease after acute influenza infection, we found that a population of lung resident PD1HiCD4+ T helper cells (TRH), which exhibit characteristics of both follicular helper T cells and resident memory cells, was greatly increased after primary influenza pneumonia. Furthermore, TRH cells were located near the dysplastic/inflammatory lung parenchyma after the resolution of primary viral pneumonia. Strikingly, genetic ablation of TRH or the blockade of TRH-derived IL-21 signaling diminished chronic lung pathology and improved lung function following viral pneumonia in aged animals. Moreover, TRH cell presence was associated with worse lung function and increased lung pathology in COVID-19 convalescents. Our data suggest that exuberant TRH responses contribute to the development of chronic lung sequelae following viral pneumonia. Additionally, our findings may pave the way to develop novel preventive and/or therapeutic options treating chronic lung diseases following acute viral pneumonia including long COVID.

Abstract 132: Immunosenescence-related Biomarkers And Potential Therapeutic Targets For Coronary Artery Disease Identified Via Plasma Proteome And Immune Cell Traits

There is a pressing need to identify novel biomarkers and therapeutic targets to address the epidemiological burden of coronary artery disease (CAD). Mendelian Randomization analysis (MR) leverages Genome-Wide Association Study (GWAS) data to infer the causal mechanism underlying the phenotype of interest. The proteome is the central mediator of the etiology and pathophysiology of multifactorial diseases such as CAD. Here, we screened for potential biomarkers and therapeutic targets for CAD through MR by using genetically predicted levels of plasma proteome followed by immune cell traits and transcriptomic analysis. The main workflow of our study is outlined in Figure 1. Briefly, MR analysis was performed with protein quantitative trait loci (pQTL) and meta-analysis of four independent GWASs. Sensitivity analysis was performed to ensure the robustness of identified proteins. Pathway Enrichment analysis on whole-blood transcriptome was performed with GSEA. Secondary MR explored the causal effect of 731 immune cell traits on CAD risk. In total, 11 overlapping candidate proteins (P<0.05, OR>1.0) were identified from the plasma proteome meta-analysis, including LPA, PLA2G7, and PCSK9, substantiating the validity of our MR analysis. Functional annotation of candidate proteins identified LCAT and CA11 as promising therapeutic targets and biomarkers. LCAT suppression is protective against atherosclerosis in mice models, while the functional significance of CA11 remains to be elucidated. Transcriptomic analysis shows enrichment in IL-2 signaling pathways reflective of the exhaustion of CD28 on cytotoxic T cells identified via secondary MR using immune-cell traits. Together, these results are indicative of immunosenescence as a risk factor for CAD. Overall, our results revealed that an altered immune landscape increased the risk for CAD and unraveled an encouraging avenue for early diagnosis and treatment of CAD.

Resistance-Exercise-Induced Circulating Factors on Damaged Skeletal Muscle Tissue Proteome in Young Untrained Men

Recovery from muscle damage is tightly orchestrated by multiple physiological processes that can be altered by circulating factors (e.g., cytokines, growth factors) and intracellular signaling pathways (e.g., autophagy). Our previous studies have demonstrated that resistance exercise (RE)-induced circulating factors alter the expression of intramuscular cytokine mRNA and autophagic and myogenic markers in exercise-induced skeletal muscle damage (EIMD). However, the global effects of these circulating factors on damaged muscle are largely understudied. Purpose: To identify global changes in the proteome and functional pathways in skeletal muscle when EIMD is followed by an upper body RE-induced circulating milieu. Hypothesis: RE-induced circulating factors alter protein expression profile in damaged muscle. Methods: Vastus lateralis samples from 3 men included in the parent study were utilized for this report. Untrained men (age: 23 ± 5yr; height: 181.6 ± 5.9cm; weight: 86.0 ± 15.8kg) completed 2 identical bouts of maximal unilateral eccentric knee extensions for 8 sets of 10 repetitions, which were immediately followed by either heavy upper body RE to induce changes in circulating factors (EX) or 20 min seated rest (CON). Muscle samples from the damaged leg at 24h were used to compare between conditions. The total number of proteins was measured using LC-MS/MS. Bioinformatic analyses were performed via Qiagen Ingenuity Pathway Analysis software. For identifying enriched biological processes and functions, proteins had to have at least 1.0-fold differential expression with an FDR-adjusted significance level < 0.05. Z scores were calculated to predict changes in biological functions. Results: A total of 900 proteins were identified. For EX, 79 proteins were downregulated, and 15 were upregulated compared to CON at 24h. Pathway analyses demonstrated that for EX, there was decreased activation in acute phase response signaling, liver X receptor/retinoid X receptor activation, nitric oxide production and reactive oxygen species in macrophages, and neutrophil extracellular trap signaling pathways compared to CON. In addition, for EX, there was an increase in macrophage IL-12 signaling and production than CON. Conclusion: RE-induced changes in circulating factors modulated pathways associated with the immune response and cytokine signaling in untrained men. Given that immune response is critical for recovery from muscle damage, these results highlight the potential role of RE-induced circulating factors (e.g., hormones, growth factors, etc.) which could play a role in mediating muscle recovery. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

IL10 signal of lung interstitial macrophages prevents progressive lung fibrosis by suppressing commensal dysbiosis

IL10 signal of lung interstitial macrophages prevents progressive lung fibrosis by suppressing commensal dysbiosis

IL-10 promotes mast cell homeostasis and differentiation towards a mucosal mast cell phenotype

Abstract We recently demonstrated an unexpected role for IL-10 in driving mast cell (MC) activation and proliferation. IL-10 stimulation significantly enhanced IgE-mediated MC activation, upregulated FceRI expression, and promoted MC-mediated food allergy. Surprisingly, IL-10 also amplified cytokine production in un-activated MCs and enhanced their proliferation and survival, suggesting a role for IL-10 in MC homeostasis. Herein, we demonstrate that IL-10 can significantly expand MC proliferation and activity during homeostatic conditions. Treatment with IL-10 enhanced the proliferation and survival of both IL-3 and IL-3+SCF cultured bone marrow-derived MCs (BMMCs). This corresponded with a dramatic increase in cytokine production in IL-10-treated cells, along with upregulation of FceRI expression. While SCF was dispensable for the increased FceRI, it significantly enhanced cytokine production compared to cells cultured in IL-3 alone. Further analysis revealed a distinctive mucosal MC phenotype in IL-10-treated cells, characterized by beta7-integrin expression and enhanced secretion of murine MC protease-1. Lastly, autocrine IL-10 also supported MC expansion and function, as blocking IL-10 signaling attenuated MC expansion, inducing apoptosis, and decreasing cytokine production. Taken together, these data suggest an important role for IL-10 in maintaining MC responses during physiological conditions.

Construction and evaluation of a metabolic correlation diagnostic model for diabetes based on machine learning algorithms.

Diabetes mellitus (DM) is a prevalent chronic disease marked by significant metabolic dysfunctions. Understanding its molecular mechanisms is vital for early diagnosis and treatment strategies. We used datasets GSE7014, GSE25724, and GSE156248 from the GEO database to build a diagnostic model for DM using Random Forest (RF) and LASSO regression models. GSE20966 served as a validation cohort. DM patients were classified into two subtypes for functional enrichment analysis. Expression levels of key diagnostic genes were validated using quantitative real-time PCR (qRT-PCR) on Peripheral Blood Mononuclear Cells (PBMCs) from DM patients and healthy controls, focusing on CXCL12 and PPP1R12B with GAPDH as the internal control. After de-batching the datasets, we identified 131 differentially expressed genes (DEGs) between DM and control groups, with 70 up-regulated and 61 down-regulated. Enrichment analysis revealed significant down-regulation in the IL-12 signaling pathway, JAK signaling post-IL-12 stimulation, and the ferroptosis pathway in DM. Five genes (CXCL12, MXRA5, UCHL1, PPP1R12B, and C7) were identified as having diagnostic value. The diagnostic model showed high accuracy in both the training and validation cohorts. The gene set also enabled the subclassification of DM patients into groups with distinct functional traits. qRT-PCR results confirmed the bioinformatics findings, particularly the up-regulation of CXCL12 and PPP1R12B in DM patients. Our study pinpointed seven energy metabolism-related genes differentially expressed in DM and controls, with five holding diagnostic value. Our model accurately diagnosed DM and facilitated patient subclassification, offering new insights into DM pathogenesis.