Immunology
Abstract
Epigenetic scarring of terminally dysfunctional (TDysf) CD8+ T cells hinders long-term protection and response to immune checkpoint blockade during chronic infections and cancer. We developed a faithful in vitro model for CD8+ T cell terminal dysfunction as a platform to advance T cell immunotherapy. Using TCR-transgenic CD8+ T cells, we found that 1-week peptide stimulation, mimicking conditions in previous models, failed to induce a stable exhaustion program. In contrast, prolonged stimulation for 2–3 weeks induced T cell dysfunction but triggered activation-induced cell death, precluding long-term investigation of exhaustion programs. To better mimic in vivo exhaustion, we provided post-effector, chronic TGF-β1 signals, enabling survival of chronically stimulated CD8+ T cells for over 3 weeks. These conditions induced a state of terminal dysfunction, marked by a stable loss of effector, cytotoxicity, and memory programs, along with mitochondrial stress and impaired protein translation. Importantly, transcriptomic and epigenetic analyses verified the development of terminal exhaustion-specific signatures in TDysf cells. Adoptive transfer of TDysf cells revealed their inability to recall effector functions or proliferate after acute lymphocytic choriomeningitis virus rechallenge. This tractable model system enables investigation of molecular pathways driving T cell terminal dysfunction and discovery of therapeutic targets for cancer or chronic infections.
Authors
Amir Yousif, Abbey A. Saadey, Ava Lowin, Asmaa M. Yousif, Ankita Saini, Madeline R. Allison, Kelley Ptak, Eugene M. Oltz, Hazem E. Ghoneim
Abstract
In allogeneic hematopoietic transplantation, donor αβ T cells attack recipient tissues, causing graft versus host disease (GVHD). A longstanding question has been how GVHD is maintained despite T cell exhaustion from chronic alloantigen stimulation. In other exhaustion models, CD8 responses are sustained by CD39loTim-3loToxhiTCF-1hi precursor exhausted T cells (TPEX). Here we characterize CD8+ TPEX in the B6(H-2b)→129(H-2b) GVHD model wherein responses against the minor histocompatibility antigen H60 can be tracked using MHCI-tetramers (TetH60). Early after transplant, TetH60+ CD8 cells were uniformly PD-1hiToxhi, whereas TetH60– cells also had PD-1loToxlo cells, indicative of more diverse antigen experiences. Among TetH60+ and TetH60– populations were CD39loTCF-1hi cells. Upon competitive retransplantation, TetH60+CD39loTCF-1hi cells outcompeted TetH60+CD39hiTCF-1lo cells and underwent self-renewal, whereas CD39hiTCF-1lo cells did not yield TCF-1hi cells. To test the role of TCF-1, we studied CD8 cells lacking long TCF-1 isoforms (p45–/–). P45–/– cells were outcompeted by WT cells when transplanted into 129 recipients, though they expanded similarly in syngeneic recipients. In the B6→C3H.SW(H-2b) model, p45–/– CD8 cells caused less weight loss than did WT CD8 cells; however, histopathologic GVHD was similar in both groups. P45–/– and WT CD8 cells also had similar graft versus leukemia activity. These results highlight the complex biology of TCF-1 in supporting alloreactive T cell function.
Authors
Kevin Quann, Faruk Sacirbegovic, Sarah Rosenberger, Emily R. McFerran, Kentin C. Codispot, Laura Garcia-Dieguez, Alexander M. Rowe, Wenzhong Wei, Dhanpat Jain, Jennifer M. McNiff, Warren Shlomchik
Abstract
Clinical trials have identified 2 distinct eosinophilic esophagitis (EoE) treatment phenotypes: those that show proton pump inhibitor (PPI) responsiveness (PPI-R) and those that show PPI unresponsiveness (PPI-UR). Comprehensive clinical, endoscopic, and RNA-Seq analyses of patients with EoE prior to and following PPI therapy have not previously been performed to our knowledge. We showed that clinical, endoscopic, and histologic evaluation of esophageal biopsies from pediatric PPI-R and PPI-UR individuals with EoE prior to PPI therapy (diagnosis) were indistinguishable. RNA-Seq analyses revealed common immune and inflammatory transcriptional signatures in both PPI-R EoE and PPI-UR EoE esophageal biopsy samples at diagnosis and distinct signatures enriched for processes related to neuropeptide signaling and cell cycle and division. PPI therapy induced histologic, endoscopic, and transcriptional remission in PPI-R EoE, but not in PPI-UR EoE. Persistent disease in PPI-UR EoE was associated with the presence of Th2 inflammatory and dedifferentiated esophageal epithelial transcriptomic signatures, while PPI-R EoE revealed genes enriched in cellular responses to LPS, host defense against viruses, and type I IFN signaling. In silico analyses identified common and unique EoE disease gene drivers in PPI-R and PPI-UR EoE. These studies indicate that the 2 EoE phenotypes have unique transcriptomic elements that underlie the molecular nature of PPI-R and PPI-UR EoE disease.
Authors
Somdutta Chakraborty, Ankit Sharma, Sahiti Marella, Christian F. Rizza, Patrick A. O’Brien, Varsha Ganesan, Gila Idelman, Susie Min, Mayee Chen, Talaya McCright-Gill, Nancy Gonzalez, Alexandros D. Polydorides, Paul S. Foster, Simon P. Hogan, Mirna Chehade
Abstract
In the rheumatoid arthritis (RA) synovium, resident fibroblast-like synoviocytes (FLS) express MHC class II molecules (HLA-D) but lack the co-stimulatory signals typically required for T cell activation. Here, we demonstrate that antigen presentation by FLS induces a distinct T cell activation state characterized by high CD69, yet reduced CD25 and HLA-DR expression, suppressed proliferation, and decreased effector cytokine production compared to professional antigen presenting cells (APCs), such as macrophages. FLS were also capable of suppressing macrophage-induced T cell activation, underscoring their dominant immunomodulatory role in the synovial microenvironment. Mechanistically, we identify indoleamine 2,3-dioxygenase (IDO1)-mediated tryptophan depletion as the primary driver of FLS-induced T cell hypo-responsiveness. Spatial transcriptomics revealed colocalization of IDO1 and CD69 within ectopic lymphoid structures in RA synovium, further supporting the in vivo relevance of this pathway. These findings provide the groundwork for positioning FLS as critical T cell regulators in RA and highlight the importance of preserving their immunosuppressive properties when therapeutically targeting pathogenic FLS functions.
Authors
Melissa R. Romoff, Preethi K. Periyakoil, Edward F. DiCarlo, Daniel Ramirez, Susan M. Goodman, Christina S. Leslie, Alexander Y. Rudensky, Laura T. Donlin, Melanie H. Smith
Abstract
Secreted high mobility group box protein 1 (HMGB1) regulates the adaptive immune response and acts as a biosensor for cells undergoing necrosis, stress, and inflammatory stimulation. However, its role in B cells remains enigmatic. Here, we demonstrate that HMGB1 is critical for peripheral B cell homeostasis and humoral immunity. Conditional deletion of Hmgb1 in B cells led to expanded marginal zone B cells, reduced B1a cells, and impaired antigen-specific antibody responses. Mechanistically, HMGB1 deficiency enhanced proximal and distal B cell receptor (BCR) signaling, probably via increased CD21 expression, which lowered the BCR activation threshold. This phenotype was linked to reduced lymphoid enhancer-binding factor 1 (LEF1) levels, a Wnt-responsive transcription factor, as HMGB1 directly bound the Lef1 promoter to sustain its transcription, thereby repressing Cd21. Furthermore, HMGB1 constrained actin reorganization by suppressing the MST1/DOCK8/WASP axis, which feedback-modulated BCR clustering and signalosome recruitment. Collectively, HMGB1 ensures optimal BCR signaling by transcriptionally and cytoskeletally tuning activation thresholds, highlighting its dual role as a nuclear regulator and cytoskeletal modulator in B cell immunity.
Authors
Qiuyue Chen, Ziyin Zhang, Nanshu Xiang, Li Luo, Xin Dai, Danqing Kang, Lu Yang, Yingzi Zhu, Jiang Chang, Yukai Jing, Na Li, Qianglin Chen, Panpan Jiang, Ju Liu, Yanmei Huang, Heather Miller, Xinyuan Zhou, Fang Zheng, Quan Gong, Chaohong Liu
Abstract
Immune checkpoint therapy has changed cancer treatment, including non-small cell lung cancer (NSCLC). The unresponsiveness of PD-L1lo/– tumors to anti–PD-1/PD-L1 immunotherapy is attributed to alternative immune evasion mechanisms that remain elusive. We previously reported that farnesoid X receptor (FXR) was increased in PD-L1lo/– NSCLC. Herein, we found that immune checkpoint HVEM was positively correlated with FXR but inversely correlated with PD-L1 in NSCLC. HVEM was highly expressed in FXRhiPD-L1lo NSCLC. Consistently, clinically relevant FXR antagonist dose-dependently inhibited HVEM expression in NSCLC. FXR inhibited cytokine production and cytotoxicity of cocultured CD8+ T cells in vitro, and it shaped an immunosuppressive tumor microenvironment (TME) in mouse tumors in vivo through the HVEM/BTLA pathway. Clinical investigations show that the FXR/HVEM axis was associated with immunoevasive TME and inferior survival outcomes in patients with NSCLC. Mechanistically, FXR upregulated HVEM via transcriptional activation, intracellular Akt, Erk1/2 and STAT3 signals, and G1/S cycle progression in NSCLC cells. In vivo treatment experiments demonstrated that anti-BTLA immunotherapy reinvigorated antitumor immunity in TME, resulting in enhanced tumor inhibition and survival improvement in FXRhiPD-L1lo mouse Lewis lung carcinomas. In summary, our findings establish the FXR/HVEM axis as an immune evasion mechanism in PD-L1lo/– NSCLC, providing translational implications for future immunotherapy in this subgroup of patients.
Authors
Xiaolong Xu, Bin Shang, Hancheng Wu, Xiuye Jin, Junren Wang, Jing Li, Daowei Li, Bin Liang, Xingguang Wang, Lili Su, Wenjie You, Shujuan Jiang
Abstract
Hepatic ischemia-reperfusion injury (IRI) disrupts cellular signaling pathways and contributes to early allograft dysfunction (EAD) in orthotopic liver transplantation (OLT). In this study, we found that the hepatic RNA binding protein Human Antigen R (HuR) regulated the 3′ untranslated region (UTR) of Carcinoembryonic Antigen-Related Cell Adhesion Molecule 1 (Ceacam1) following ischemic stress. Hepatocyte-specific preinjury HuR-null mice exhibited elevated LDH-5 isoenzyme activity and reduced Ceacam1-S expression, reflecting tissue-specific injury. In situ hybridization demonstrated that the stability of Ceacam1 mRNA depended on HuR. Luciferase assays identified Ceacam1 3′UTR cis-elements responsive to high oxygen tension. HuR-targeting short-activating RNAs (saRNAs) preferentially induced the alternative splicing of Ceacam1-S. Antisense oligos directed to the Ceacam1 3′UTR protected WT mice against acute liver injury. In the clinical arm, increased HuR and CEACAM1 expression were associated with reduced proinflammatory phenotype and a lower incidence of EAD in patients with OLT (n = 164). Human discarded livers with elevated ELAVL1/CEACAM1 levels correlated with improved tissue homeostasis. These findings suggest that HuR regulation of Ceacam1 represents a key determinant of donor tissue quality and offers a potential target for future therapeutic strategies in OLT recipients.
Authors
Brian Cheng, Tristan D. Tibbe, Siyuan Yao, Megan Wei, Zeriel Y. Wong, Taylor Torgerson, Richard Chiu, Aanchal S. Kasargod, Kojiro Nakamura, Monica Cappelletti, Myung Sim, Douglas G. Farmer, Fady Kaldas, Jerzy W. Kupiec-Weglinski, Kenneth J. Dery
Abstract
Palmoplantar pustulosis (PPP) is a chronic inflammatory skin disorder marked by erythematous pustules and desquamation on the palms and soles. While IL-17 pathways are implicated in PPP, IL-17 blockers have shown modest efficacy, underscoring the need for a deeper understanding of IL-17 involvement. To dissect the cellular and spatial architecture of PPP, we performed single-cell RNA-Seq (scRNA-Seq) on lesional, nonlesional, and healthy acral skin to examine cellular composition, transcriptomic profiles, and cell-cell interactions. Unbiased clustering revealed 9 major cell types, including an inflammatory keratinocyte subset enriched in IL-17A/TNF signatures and marked by high IL-36G expression. Within the lymphocyte compartment, we identified a hybrid “regTh17” population coexpressing regulatory markers (FOXP3, CTLA4, TIGIT), IL17F, and IL26. This regTh17 subset was distinguished by elevated IL1R1 and CD39, suggesting an IL-1β–driven differentiation. Spatial analyses demonstrated significant neighborhood enrichment of regTh17 cells with IL-36G+ supraspinous keratinocytes. RegTh17 cells were the predominant source of IL-17F and IL-26 signals, whereas keratinocytes were predicted as their main receivers. We further observed regTh17 coexpressing TNFRSF4 (OX40) and TNFRSF18 (GITR) specifically at sites of IL36G+ keratinocyte interactions, implicating these pathways in amplification of the IL-17/IL-36 inflammatory loop. Together, our integrated single-cell and spatial profiling uncovers Th17 plasticity in PPP, identifies a regTh17-keratinocyte interaction, and highlights IL-17F, IL-26, OX40/OX40L, and GITR/GITRL as candidate targets for precision therapies in this challenging disease.
Authors
Tran H. Do, Rachael Bogle, Haihan Zhang, Xianying Xing, Mehrnaz Gharaee-Kermani, Madalina Raducu, Jennifer Fox, Rundong Jiang, Olesya Plazyo, Paul W. Harms, Mio Nakamura, Enze Xing, Michel Gilliet, Allison C. Billi, J. Michelle Kahlenberg, Robert L. Modlin, Ozge Uluckan, Lam C. Tsoi, Johann E. Gudjonsson
Abstract
Acute lower respiratory infections are the primary cause of global mortality in post-neonatal children. Most respiratory viruses primarily involve upper airway infection and inflammation, yet nasal responses are poorly characterized. Using a mouse model of human metapneumovirus (HMPV), we found viral burden was higher in nasal airways and exhibited delayed clearance. Despite high burden, there was low nasal expression of type I and III interferon (IFN). Single-cell RNA-sequencing (scRNA-seq) from HMPV-infected mice showed lower nasal interferon-stimulated gene (ISG) expression and nasal enrichment of genes negatively regulating IFN. scRNA-seq of COVID-19 patients confirmed lower ISG expression in upper airways. HMPV infection downregulated nasal expression of interferon regulatory factor-3, suggesting a mechanism for limited response. To rescue the quiescent environment, we administered type I or III IFN to upper airways early post-infection, leading to lower nasal HMPV titer and virus-specific CD8+ T-cell upregulation. Intranasal immunization adjuvanted with type I or III IFN improved immune response, reduced clinical disease, and enhanced viral clearance in HMPV and influenza infection. IFN adjuvant increased recruitment of dendritic cells, resident-memory T-cells, and neutralizing antibodies. These findings reveal locally suppressed IFN production contributes to a quiescent nasal immune landscape that delays viral clearance and impairs mucosal vaccine responses.
Authors
Jorna Sojati, Olivia B. Parks, Taylor Eddens, Jie Lan, Monika Johnson, John V. Williams
Abstract
Glutaminolysis is enhanced in T cells of lupus patients and in follicular helper T (Tfh) cells, a critical subset of CD4+ T cells that provide help to autoreactive B cells, in lupus mice. Glutaminolysis inhibitors reduced lupus activity in association with a decreased frequency of Th17 cells in mice. Here, we thought to determine the role of glutaminolysis in murine Tfh cells. The pharmacological inhibition of glutaminolysis with DON reduced the expression of the critical costimulatory molecule ICOS on lupus Tfh cells, in association with a reduction of autoantibody production and B cell differentiation markers. Accordingly, profound transcriptomic and metabolic changes, including a reduction of glycolysis, were induced by DON in lupus Tfh cells, whereas healthy Tfh cells showed minor changes. The T cell-specific genetic inhibition of glutaminolysis largely phenocopied the effects of DON on Tfh cells and B cells in an autoimmune genetic background with minor changes in Tfh and B cells in healthy controls. Furthermore, we showed that T cell-specific glutaminolysis inhibition impaired T-dependent humoral responses in autoimmune mice as well as their Tfh response to a viral infection. Overall, these results suggest that lupus Tfh cells have a greater intrinsic requirement of glutaminolysis for their helper functions.
Authors
Seung-Chul Choi, Yong Ge, Milind V. Joshi, Damian Jimenez, Abigail Castellanos Garcia, Cassandra LaPlante, Lauren T. Padilla, Chaoyu Ma, Nu Zhang, Jeffrey C. Rathmell, Mansour Mohamadzadeh, Laurence Morel
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