Immunology
Abstract
Macrophage migration inhibitory factor (MIF) is an upstream regulatory cytokine that is associated with advanced disease and poor outcomes in multiple cancer types, including melanoma. We investigated whether anti-MIF therapy could enhance the antitumor effects of the immune checkpoint inhibitor anti–programmed cell death 1 (anti–PD-1) in 2 murine tumor models. The therapeutic efficacy of anti-MIF, alone or combined with anti–PD-1, was tested in the YUMMER1.7 melanoma and MC38 colorectal cancer models. Tumor growth and survival were assessed in untreated Mif-knockout (KO) and low-expression human MIF allele (CATT5) mice and compared with wild-type (WT) or high-expression MIF allele (CATT7) mice. Tumor-bearing animals underwent cytokine profiling, tumor immunohistochemistry, flow cytometry, and scRNA-Seq. We also correlated functional variant MIF alleles with melanoma incidence and progression in patients. Our results showed that combined anti-MIF and anti–PD-1 significantly reduced tumor growth, improved survival, and promoted tumor regression, accompanied by enhanced TH1 cytokine levels, increased macrophage activation–related cytokines, and increased type 1 conventional dendritic cells. scRNA-Seq analysis revealed an expansion of intratumor Cd74/C1q/Aif1-expressing macrophages, which exhibited an antitumor phenotype, in response to anti-MIF therapy. MIF-KO and CATT5 mice exhibited reduced tumor burdens compared with WT or CATT7 mice alone and in the presence of anti–PD-1. In patients with melanoma, the high-MIF expression genotype (-173C/C) occurred at higher frequencies compared with healthy controls. These findings highlight that the addition of anti-MIF to anti–PD-1 reduces tumor growth, enhances antitumor responses, prolongs survival, and augments key intratumor immune cell populations involved in immune activation against tumors. This approach merits further consideration for clinical trial development.
Authors
Thuy T. Tran, Gabriela Athziri Sánchez-Zuno, Lais Osmani, Jasmine Caulfield, Caroline Naomi Valdez, Marta Piecychna, Lin Leng, Michelle E. Armstrong, Seamas C. Donnelly, Carlo B. Bifulco, Terri Clister, Rajan P. Kulkarni, Lin Zhang, Mario Sznol, Lucia Jilaveanu, Harriet M. Kluger, Insoo Kang, Richard Bucala
Abstract
Chronic lung allograft dysfunction (CLAD) is the leading cause of mortality after lung transplantation, yet its molecular mechanisms remain poorly understood. To elucidate the pathogenesis of CLAD, we conducted a comprehensive single-cell transcriptomic analysis of CLAD lungs, integrating our generated datasets with approximately 1.6 million cells from 15 published studies of other fibrotic lung diseases. By applying pseudo-bulk approaches to mitigate batch effects, we identified molecular signatures specific to CLAD and those shared with idiopathic pulmonary fibrosis, COVID-19, and other fibrotic conditions. Our analysis revealed CLAD-specific cellular subsets including Fibro.AT2 cells, exhausted CD8+ T cells, and superactivated macrophages while suggesting that pathogenic keratin 17–positive, keratin 5–negative (KRT17+KRT5−) cells represent a common fibrotic mechanism across fibrotic lung diseases. Additionally, we performed donor-recipient cell deconvolution in lung allografts, uncovering distinct transcriptional programs and intercellular crosstalk between donor- and recipient-derived cells that drive allograft fibrosis. Recipient-derived stromal and immune cells showed enhanced pro-fibrotic and allograft rejection pathways compared with their donor counterparts. By leveraging insights from other fibrotic diseases to elucidate CLAD-specific mechanisms, our study provides a molecular framework for understanding CLAD pathogenesis and identifies potential therapeutic targets for this treatment-refractory condition.
Authors
Yuanqing Yan, Taisuke Kaihou, Emilia Lecuona, Xin Wu, Masahiko Shigemura, Haiying Sun, Chitaru Kurihara, Ruli Gao, Felix L. Nunez-Santana, G.R. Scott Budinger, Ankit Bharat
Abstract
High-affinity antibody production depends on CD4+ T-follicular helper (Tfh) cells. In humans, peripheral blood Tfh cells are heterogenous, as evidenced by differential expression of the chemokine receptors, CXCR3 and CCR6, which to date have served to classify three subsets, pTfh1, pTfh2 and pTfh17. Although pTfh1 responses dominate during blood-stage Plasmodium infections, a clear association with protective antibody responses remains to be described. We hypothesise that pTfh cells exhibit greater phenotypic and functional heterogeneity than that described by CXCR3/CCR6 alone, and that these more nuanced pTfh subsets play distinct roles during Plasmodium infection. We map pTfh cell heterogeneity in healthy individuals prior to and during controlled human malaria infection (CHMI) using parallel scRNA-seq and VDJ-seq. We uncover two pTfh1 subsets or differential phenotypic states, distinguishable by CCR7 expression. Prior to infection, Tfh1-CCR7neg cells exhibit higher baseline expression of inflammatory cytokines and genes associated with cytotoxicity. While Tfh1-CCR7pos cells have higher GC signatures. Indeed, during CHMI, Tfh1-CCR7pos, Tfh1-CCR7neg, and Tfh2 cells, all clonally expand and become activated. However, only Tfh1-CCR7pos and Tfh2 cells positively associate with protective antibody production. Hence, our data reveal further complexity amongst human Tfh cells, and highlight two distinct subsets associated with antibody-mediated immunity to malaria.
Authors
Megan S.F. Soon, Damian A. Oyong, Nicholas L. Dooley, Reena Mukhiya, Zuleima Pava, Dean Andrew, Jessica R. Loughland, James S. McCarthy, Jo-Anne Chan, James G. Beeson, Christian Engwerda, Ashraful Haque, Michelle J. Boyle
Abstract
IgA protects the body from invaders in the mucosal sites, but its role in allergic diseases, such as hay fever, is poorly understood. We demonstrate an increased susceptibility to cedar pollen-induced hay fever associated with increasing pollen penetration into the body in IgA-deficient mice, indicating that IgA prevents pollen invasion on the mucosa. We identified Bryostatin 1, an anti-carcinogenic Protein kinase C (PKC) δ activator, as an IgA/IgE class-switching regulator in B cells. Bryostatin 1 enhanced IgA production through induction of germline transcript (GLT) α via PKCδ-MEK/ERK-RUNX1 pathway and suppressed IgE by reducing GLTε through the PKCδ-STAT5-ID2 pathway. Production of Th2 cytokines and eosinophils infiltration in the lungs was also reduced. Furthermore, hay fever alleviation by Bryostatin 1 demonstrated diminished symptoms in mice in vivo three months subsequent to nasal administration.
Authors
Naoki Morita, Kohta Yamamoto, Ryutaro Tamano, Peng Gao, Takahiro Nagatake, Takenori Inomata, Tianxiang Huang, Yasuhiro Yamada, Takahiro Adachi, Manabu Sugai, Keiichi I. Nakayama, Hirotatsu Kojima, Reiko Shinkura
Abstract
The efficacy of anti-CD20 therapies places B cells and their interaction with T cells at the center of attention for multiple sclerosis (MS) pathogenesis. Follicular T helper cells (Tfh), which guide B cell maturation in germinal centers within lymph nodes (LNs), are elevated in the circulation and cerebrospinal fluid of patients with MS (pwMS). However, the LN spatial landscape has remained largely without investigation for pwMS. Using cyclic immunofluorescence, we assessed cell abundance and spatial connections in FFPE LNs of 33 pwMS and 35 non-MS controls. The presence of EBV was analyzed through EBER immunostaining and multiplex quantitative PCR. Our analysis showed that Tfh cells were expanded in LNs of pwMS and accumulated especially in the mantle zone and B cell follicles compared with controls. The Tfh/T follicular regulator ratio was increased in pwMS, while B cell ratios were similar between the cohorts. The interaction of Tfh cells with follicular B cells was higher in pwMS. Interestingly, Tfh accumulation was also observed in 5 prediagnostic MS cases. No signs of EBV latency were detected in either group. These findings highlight LNs as a site of early and persistent immune activation in pwMS, with therapeutic implications to be further addressed.
Authors
Joona Sarkkinen, Eliisa Kekäläinen, Leo Hannolainen, Ada Junquera, Johannes Dunkel, Maria F. Perdomo, Mikko I. Mäyränpää, Sini M. Laakso
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
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