Tregs play vital roles in suppressing atherogenesis. Pathological conditions reshape Tregs and increase Treg-weakening plasticity. It remains unclear how Tregs preserve their function and how Tregs switch into alternative phenotypes in the environment of atherosclerosis. In this study, we observed a great induction of CD4+Foxp3+ Tregs in the spleen and aorta of ApoE–/– mice, accompanied by a significant increase of plasma IL-35 levels. To determine if IL-35 devotes its role in the rise of Tregs, we generated IL-35 subunit P35–deficient (IL-35P35–deficient) mice on an ApoE–/– background and found Treg reduction in the spleen and aorta compared with ApoE–/– controls. In addition, our RNA sequencing data show the elevation of a set of chemokine receptor transcripts in the ApoE–/– Tregs, and we have validated higher CCR5 expression in ApoE–/– Tregs in the presence of IL-35 than in the absence of IL-35. Furthermore, we observed that CCR5+ Tregs in ApoE–/– have lower Treg-weakening AKT-mTOR signaling, higher expression of inhibitory checkpoint receptors TIGIT and PD-1, and higher expression of IL-10 compared with WT CCR5+ Tregs. In conclusion, IL-35 counteracts hyperlipidemia in maintaining Treg-suppressive function by increasing 3 CCR5-amplified mechanisms, including Treg migration, inhibition of Treg weakening AKT-mTOR signaling, and promotion of TIGIT and PD-1 signaling.
Ying Shao, William Y. Yang, Fatma Saaoud, Charles Drummer IV, Yu Sun, Keman Xu, Yifan Lu, Huimin Shan, Ethan M. Shevach, Xiaohua Jiang, Hong Wang, Xiaofeng Yang
Common variable immunodeficiency (CVID) is characterized by profound primary antibody defects and frequent infections, yet autoimmune/inflammatory complications of unclear origin occur in 50% of individuals and lead to increased mortality. Here, we show that circulating bacterial 16S rDNA belonging to gut commensals was significantly increased in CVID serum (P < 0.0001), especially in patients with inflammatory manifestations (P = 0.0007). Levels of serum bacterial DNA were associated with parameters of systemic immune activation, increased serum IFN-γ, and the lowest numbers of isotype-switched memory B cells. Bacterial DNA was bioactive in vitro and induced robust host IFN-γ responses, especially among patients with CVID with inflammatory manifestations. Patients with X-linked agammaglobulinemia (Bruton tyrosine kinase [BTK] deficiency) also had increased circulating bacterial 16S rDNA but did not exhibit prominent immune activation, suggesting that BTK may be a host modifier, dampening immune responses to microbial translocation. These data reveal a mechanism for chronic immune activation in CVID and potential therapeutic strategies to modify the clinical outcomes of this disease.
Hsi-en Ho, Lin Radigan, Gerold Bongers, Ahmed El-Shamy, Charlotte Cunningham-Rundles
Experimental Autoimmune Encephalomyelitis (EAE) is a well-characterized animal model of Multiple Sclerosis. During the early phase of EAE, the infiltrating monocyte and monocyte-derived macrophages and activated resident microglia contribute to T cell recruitment, especially CD4+ T cells, into the CNS resulting in neuronal demyelination, however, in later stages they promote remyelination and recovery by removal of myelin debris by phagocytosis. SIRPα and CD47 are abundantly expressed in the CNS and deletion of either molecule is protective in myelin oligodendrocyte glycoprotein (MOG)-induced EAE due to failed effector T cell expansion and trafficking. Here we report that treatment with the function blocking CD47 antibody (Ab), Miap410 significantly reduced the infiltration of pathogenic immune cells, but impaired recovery from paresis. The underlying mechanism was by blocking the emergence of CD11chigh MHCIIhigh microglia at peak disease that expressed receptors for phagocytosis, scavenging, and lipid catabolism, which mediated clearance of myelin debris, and the transition of monocytes to macrophages in the CNS. In the recovery phase of EAE, Miap410 Ab treated mice had worsening paresis with sustained inflammation and limited remyelination as compared to control Ab treated mice. In summary, Ab blockade of CD47 impaired resolution of CNS inflammation, thus worsening EAE.
Huan Wang, Gail Newton, Liguo Wu, Lih-Ling Lin, Amy S. Miracco, Sridaran Natesan, Francis W. Luscinskas
Interleukin-33 (IL-33), a nuclear alarmin released during cell death, exerts context-specific effects on adaptive and innate immune cells eliciting potent inflammatory responses. We screened blood, skin and kidney tissues from patients with Systemic Lupus Erythematosus (SLE), a systemic autoimmune disease driven by unabated type I interferon (IFN) production, and found increased amounts of extracellular IL-33 complexed with Neutrophil Extracellular Traps (NETs), correlating with severe, active disease. Using a combination of molecular, imaging and proteomic approaches, we show that SLE neutrophils -activated by disease immunocomplexes- release IL-33-decorated NETs that stimulate robust IFNα synthesis by plasmacytoid dendritic cells (pDCs) in an IL-33-receptor (ST2L)-dependent manner. IL33-silenced neutrophil-like cells cultured under lupus-inducing conditions generated NETs with diminished interferogenic effect. Importantly, SLE patient-derived NETs are enriched in mature bioactive isoforms of IL-33 processed by the neutrophil proteases elastase and cathepsin G. Pharmacological inhibition of these proteases neutralized IL-33-dependent IFNα production elicited by NETs. These data demonstrate a novel role for cleaved IL-33 alarmin decorating NETs in human SLE, linking neutrophil activation, type I IFN production and end-organ inflammation with skin pathology mirroring that observed in the kidneys.
Spiros Georgakis, Katerina Gkirtzimanaki, Garyfalia Papadaki, Hariklia Gakiopoulou, Elias Drakos, Maija-Leena Eloranta, Manousos Makridakis, Georgia Kontostathi, Jerome Zoidakis, Eirini Baira, Lars Rönnblom, Dimitrios T. Boumpas, Prodromos Sidiropoulos, Panayotis Verginis, George Bertsias
Infection is a common complication of major trauma that causes significantly increased morbidity and mortality. The mechanisms however, linking tissue injury to increased susceptibility to infection remain poorly understood. To study this relationship, we present a novel murine model where a major liver crush injury is followed by bacterial inoculation into the lung. We find that such tissue trauma both impaired bacterial clearance and was associated with significant elevations in plasma heme levels. While neutrophil (PMN) recruitment to the lung in response to Staphylococcus aureus was unchanged after trauma, PMN cleared bacteria poorly. Moreover, PMN show >50% less expression of TLR2, which is responsible, in part, for bacterial recognition. Administration of heme effectively substituted for trauma. Last, day 1 trauma patients (n=9) showed similar elevations in free heme to that seen after murine liver injury and circulating PMN showed similar TLR2 reduction compared to volunteers (n=6). These findings correlate to high infection rates.
Ghee Rye Lee, David Gallo, Rodrigo W. Alves de Souza, Shilpa Tiwari-Heckler, Eva Csizmadia, James D. Harbison, Sidharth Shankar, Valerie Banner-Goodspeed, Michael B. Yaffe, Maria Serena Longhi, Carl J. Hauser, Leo E. Otterbein
Pulmonary hypertension (PH) is a severe cardiopulmonary disease characterized by complement-dependent, fibroblast-induced perivascular accumulation and pro-inflammatory activation of macrophages. We hypothesized that, in PH, nanoscale-sized small extracellular vesicles (sEVs), released by perivascular/adventitial fibroblasts, are critical mediators of complement-dependent pro-inflammatory activation of macrophages. Pulmonary adventitial fibroblasts were isolated from calves with severe PH (PH-Fibs) and age-matched controls (CO-Fibs). PH-Fibs exhibited increased secretion of sEVs, compared to CO-Fibs, and sEV biological activity was tested on mouse and bovine bone marrow-derived macrophages (BMDMs) and showed similar responses. PH-Fib-sEVs induced augmented expression of pro-inflammatory cytokines/chemokines and metabolic genes in BMDMs, compared to CO-Fib-sEVs. Pharmacological blockade of exosome release from PH-Fibs resulted in significant attenuation of pro-inflammatory activation of BMDMs. “Bottom-up” proteomic analyses revealed significant enrichment of complement and Coagulation cascades in PH-Fib-sEVs, including augmented expression of complement component C3. We therefore examined whether PH-Fib-sEVs-mediated pro-inflammatory activation of BMDMs was complement C3-dependent. Treatment of PH-Fibs with siC3-RNA significantly attenuated the capacity of PH-Fib-sEVs for pro-inflammatory activation of BMDMs. PH-Fib-sEVs mediated pro-glycolytic alterations and complement-dependent activation of macrophages toward a pro-inflammatory phenotype, as confirmed by metabolomic studies. Thus, fibroblast-released sEVs can serve as critical mediators of complement-induced perivascular/microenvironmental inflammation in PH.
Sushil Kumar, Maria G. Frid, Hui Zhang, Min Li, Suzette Riddle, R. Dale Brown, Subhash Chandra Yadav, Micaela K. Roy, Monika E. Dzieciatkowska, Angelo D’Alessandro, Kirk C. Hansen, Kurt R. Stenmark
Polarization of low-grade inflammatory monocytes facilitates the pathogenesis of atherosclerosis. However, underlying mechanisms as well as approaches for resolving monocyte polarization conducive for the regression of atherosclerosis are not well established. In this report, we demonstrate that TRAM mediates monocyte polarization in vivo and in vitro. TRAM controls monocyte polarization through activating SFK, which not only induces STAT1/STAT5-regulated inflammatory mediators CCR2 and SIRPα, but also suppresses PPARγ-regulated resolving mediator CD200R. Enhanced PPARγ and Pex5 due to TRAM deficiency facilitates peroxisome homeostasis and reduction of cellular reactive oxygen species (ROS), further contributing to the establishment of resolving monocyte phenotype. TRAM deficient monocytes can propagate the resolving phenotype to neighboring monocytes through CD200R mediated inter-cellular communication. At the translational level, we show that TRAM deficient mice are resistant to high-fat-diet induced pathogenesis of atherosclerosis. We further document that intravenous transfusion of TRAM deficient resolving monocytes into atherosclerotic mice can potently reduce the progression of atherosclerosis. Together, our data reveal that targeting TRAM may facilitate the effective generation of resolving monocytes conducive for the treatment of atherosclerosis.
Shuo Geng, Yao Zhang, Ziyue Yi, Ran Lu, Liwu Li
Asthma is a chronic inflammatory disease of the airways associated with excess production of Th2 cytokines and lung eosinophil accumulation. This inflammatory response persists in spite of steroid administration that blocks autocrine/paracrine loops of inflammatory cytokines, and the detailed mechanisms underlying asthma exacerbation remain unclear. Here, we show that asthma exacerbation is triggered by airway macrophages through a prion-like cell-to-cell transmission of extracellular particulates, including ASC protein, that assemble inflammasomes and mediate IL-1β production. OVA-induced allergic asthma and associated IL-1β production were alleviated in mice with small GTPase Arf6-deficient macrophages. The extracellular ASC specks were slightly engulfed by Arf6–/– macrophages, and the IL-1β production was reduced in Arf6–/– macrophages compared with that in WT macrophages. Furthermore, pharmacological inhibition of the Arf6 guanine nucleotide exchange factor suppressed asthma-like allergic inflammation in OVA-challenged WT mice. Collectively, the Arf6-dependent intercellular transmission of extracellular ASC specks contributes to the amplification of allergic inflammation and subsequent asthma exacerbation.
SangJoon Lee, Akari Ishitsuka, Takahiro Kuroki, Yu-Hsien Lin, Akira Shibuya, Tsunaki Hongu, Yuji Funakoshi, Yasunori Kanaho, Kyosuke Nagata, Atsushi Kawaguchi
Kawasaki disease (KD) is the leading cause of acquired heart disease among children. Murine and human data suggest that the NLRP3-IL-1β pathway is the main driver of KD pathophysiology. NLRP3 can be activated during defective autophagy/mitophagy. We used the Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis, to examine the role of autophagy/mitophagy on cardiovascular lesion development. LCWE-injected mice had impaired autophagy/mitophagy and increased levels of ROS in cardiovascular lesions, together with increased systemic 8-OHdG release. Enhanced autophagic flux significantly reduced cardiovascular lesions in LCWE-injected mice, whereas autophagy blockade increased inflammation. Vascular smooth muscle cell specific deletion of Atg16l1 and global Parkin-/- significantly increased disease formation, supporting the importance of autophagy/mitophagy in this model. Ogg1-/- mice had significantly increased lesions with increased NLRP3 activity, whereas treatment with MitoQ, reduced vascular tissue inflammation, ROS production and systemic 8-OHdG release. Treatment with MN58b or Metformin (increasing AMPK and reducing ROS), resulted in decreased disease formation. Our results demonstrate that impaired autophagy/mitophagy and ROS-dependent damage exacerbate the development of murine KD vasculitis. This pathway can be efficiently targeted to reduce disease severity. These findings enhance our understanding of KD pathogenesis and identify novel therapeutic avenues for KD treatment.
Stefanie Marek-Iannucci, A. Beyza Ozdemir, Debbie Moreira, Angela C. Gomez, Malcolm Lane, Rebecca A Porritt, Youngho Lee, Kenichi Shimada, Masanori Abe, Aleksandr Stotland, David Zemmour, Sarah Parker, Elsa Sanchez-Lopez, Jennifer Van Eyk, Roberta A. Gottlieb, Michael Fishbein, Michael Karin, Timothy R Crother, Magali Noval Rivas, Moshe Arditi
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the pandemic Coronavirus Disease 2019 (COVID-19) and now many face the burden of prolonged symptoms—long-lasting COVID-19 symptoms or “long-COVID”. Long-COVID is thought to be linked to immune dysregulation due to harmful inflammation, with the exact causes being unknown. Given the role of the microbiome in mediating inflammation, we aimed to examine the relationship between the oral microbiome and the duration of long-COVID symptoms. Tongue swabs were collected from patients presenting with symptoms concerning for COVID-19. Confirmed infections were followed until resolution of all symptoms. Bacterial composition was determined by metagenomic sequencing. We used random forest modeling to identify microbiota and clinical covariates that associated with long-COVID symptoms. Of the patients followed, 63% (17/27) developed ongoing symptomatic COVID-19 and 37% (10/27) went on to long-COVID. Patients with prolonged symptoms had significantly higher abundances of microbiota that induce inflammation, such as members of the genera Prevotella and Veillonella. Of note are species that produce lipopolysaccharides and the similarity of long-COVID patients’ oral microbiome to those of patients with chronic fatigue syndrome. All together, we our findings suggest an association with the oral microbiome and long-COVID revealing the possibility that dysfunction of the oral microbiome may contribute to this draining disease.
John P. Haran, Evan Bradley, Abigail L. Zeamer, Lindsey Cincotta, Marie-Claire Salive, Protiva Dutta, Shafik Mutaawe, Otuwe Anya, Mario Meza-Segura, Ann M. Moormann, Doyle V. Ward, Beth A. McCormick, Vanni Bucci
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