Pathological angiogenesis is a major cause of irreversible blindness in individuals of all age groups with proliferative retinopathy (PR). Mononuclear phagocytes (MPs) within neovascular areas contribute to aberrant retinal angiogenesis. Due to their cellular heterogeneity, defining the roles of MP subsets in PR onset and progression has been challenging. Here, we aimed to investigate the heterogeneity of microglia associated with neovascularization and characterize the transcriptional profiles and metabolic pathways of pro-angiogenic microglia in a mouse model of oxygen-induced proliferative retinopathy (OIR). Using transcriptional single-cell sorting, we comprehensively map all microglia populations in retinas of room air (RA) and OIR mice. We unveil several unique types of PR-associated microglia (PRAM) and identify markers, signaling pathways, and regulons associated with these cells. Among these microglia subpopulations, we found a highly proliferative microglia subset with high self-renewal capacity and a hyper-metabolic microglia subset that expresses high levels of activating microglia markers, glycolytic enzymes and pro-angiogenic insulin-like growth factor 1. Immunohistochemical staining shows these PRAMs were spatially located within or around neovascular (NV) tufts. These unique microglia-types have the potential to promote retinal angiogenesis, which may have important implications for future treatment of PR and other pathological ocular angiogenesis-related diseases.
Zhiping Liu, Huidong Shi, Jiean Xu, Qiuhua Yang, Qian Ma, Xiaoxiao Mao, Zhimin Xu, Yaqi Zhou, Qingen Da, Yongfeng Cai, David J.R. Fulton, Zheng Dong, Akrit Sodhi, Ruth B. Caldwell, Yuqing Huo
We describe affected members of a two-generation family segregating a Stargardt disease-like phenotype caused by a two base pair deletion-insertion, c.1014_1015delGAinsCT;p(Trp338_Asn339delinsCysTyr), in BEST1. The variant was identified by whole exome sequencing and its pathogenicity was verified through chloride channel recording using wild-type (WT) and transfected mutant HEK293 cells. Clinical examination of both patients revealed a similar phenotype at two different disease stages that were attributable to difference in their age at presentation. Hyperautofluorescent flecks along the arcades were observed in the proband, while the affected mother exhibited more advanced retinal pigment epithelium (RPE) loss in the central macula. Full-field electroretinogram testing was unremarkable in the daughter, however, moderate attenuation of generalized cone function was detected in the mother. Electro-oculogram testing in the daughter was consistent with widespread dysfunction of the RPE characteristic of Best disease. Whole-cell patch clamp recordings revealed statistically significant decrease in chloride conductance of the mutant compared to WT cells. This report broadens the clinical spectrum of BEST1-associated retinopathy in the form of a mother and daughter with BEST1 genotype phenocopying Stargardt disease.
Masha Kolesnikova, Jin Kyun Oh, Jiali Wang, Winston Lee, Jana Zernant, Pei-Yin Su, Angela H. Kim, Laura A. Jenny, Tingting Yang, Rando Allikmets, Stephen H. Tsang
Deficiency of glycogen debranching enzyme in glycogen storage disease type III (GSD III) results in excessive glycogen accumulation in multiple tissues, primarily the liver, heart, and skeletal muscle. We recently reported that an adeno-associated virus (AAV) vector expressing a bacterial debranching enzyme (Pullulanase) driven by the ubiquitous CMV enhancer/chicken β-actin (CB) promoter cleared glycogen in major affected tissues of infant GSD IIIa mice. In this study, we developed a novel dual promoter consisting of a liver-specific promoter (LSP) and the CB promoter for gene therapy in adult GSD IIIa mice. Ten-week treatment with an AAV vector containing the LSP-CB dual promoter in adult GSD IIIa mice significantly increased Pullulanase expression and reduced glycogen contents in the liver (-60%), heart (-76%), and skeletal muscle (-63%), accompanied by the reversal of liver fibrosis, improved muscle function, and significant decrease in plasma biomarkers alanine aminotransferase, aspartate aminotransferase, and creatine kinase. Compared to the CB promoter, the dual promoter effectively decreased Pullulanase-induced cytotoxic T lymphocyte responses and enabled persistent therapeutic gene expression in adult GSD IIIa mice. Future studies are needed to determine the long-term durability of the dual promoter mediated expression of Pullulanase in adult GSD IIIa mice and in large animal models.
Jeong-A Lim, Priya S. Kishnani, Baodong Sun
Identifying host factors that contribute to pneumonia incidence and severity are of utmost importance to guiding the development of more effective therapies. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a scavenger receptor known to promote vascular injury and inflammation, but it is unknown whether and how LOX-1 functions in the lung. Here, we provide evidence of substantial accumulation of LOX-1 in the lungs of ARDS patients and in mice with pneumonia. Unlike previously described injurious contributions of LOX-1, we found that LOX-1 is uniquely protective in the pulmonary airspaces, limiting proteinaceous edema and inflammation. We also identified alveolar macrophages and recruited neutrophils as two prominent sites of LOX-1 expression in the lungs, whereby macrophages are capable of further induction during pneumonia and neutrophils exhibit a rapid, but heterogenous elevation of LOX-1 in the infected lung. Blockade of LOX-1 led to dysregulated immune signaling in alveolar macrophages, marked by alterations in activation markers and a concomitant elevation of inflammatory gene networks. However, bone marrow chimeras also suggested a prominent role for neutrophils in LOX-1-mediated lung protection, further supported by LOX-1+ neutrophils exhibiting transcriptional changes consistent with reparative processes. Taken together, this work establishes LOX-1 as a tissue-protective factor in the lungs during pneumonia, possibly mediated by its influence on immune signaling in alveolar macrophages (AMs) and LOX-1+ airspace neutrophils.
Filiz T. Korkmaz, Anukul T. Shenoy, Elise Symer, Lillia A. Baird, Christine V. Odom, Emad Arafa, Ernest L. Dimbo, Elim Na, William Molina-Arocho, Matthew Brudner, Theodore J. Standiford, Jawahar L. Mehta, Tatsuya Sawamura, Matthew R. Jones, Joseph P. Mizgerd, Katrina T. Traber, Lee J. Quinton
BACKGROUND. A patient-derived organoid (PDO) platform may serve as a promising tool for translational cancer research. In this study, we evaluated PDO’s ability to predict clinical response to gastrointestinal (GI) cancers. METHODS. We generated PDOs from primary and metastatic lesions of patients with GI cancers, including pancreatic ductal adenocarcinoma, colorectal adenocarcinoma, and cholangiocarcinoma. We compared PDO response with the observed clinical response for donor patients to the same treatments. RESULTS. We reported an approximately 80% concordance rate between PDO and donor tumor response. Importantly, we found a profound influence of culture media on PDO phenotype, where we showed significant difference in response to standard of care chemotherapies, distinct morphologies, and transcriptomes between media within the same PDOs cultures. CONCLUSION. While we demonstrate a high concordance rate between donor tumor and PDO, these studies also showed the important role of culture media when using PDOs to inform treatment selection and predict response across a spectrum of GI cancers. TRIAL REGISTRATION. Not applicable. FUNDING. This work was supported by the Joan F. & Richard A. Abdoo Family Fund in Colorectal Cancer Research, CA265050, GI Cancer program of the Mayo Clinic Cancer Center, Mayo Clinic SPORE in Pancreatic Cancer, Center of Individualized Medicine (Mayo Clinic), Department of Laboratory Medicine and Pathology (Mayo Clinic), Incyte Pharmaceuticals and Mayo Clinic Hepatobiliary SPORE, a University of Minnesota-Mayo Clinic Partnership grant, and the Early Therapeutic program (Department of Oncology, Mayo Clinic).
Tara L. Hogenson, Hao Xie, William J. Phillips, Merih D. Toruner, Jenny J. Li, Isaac P. Horn, Devin J. Kennedy, Luciana L. Almada, David L. Marks, Ryan M. Carr, Murat Toruner, Ashley N. Sigafoos, Amanda N. Koenig-Kappes, Rachel L.O. Olson, Ezequiel J. Tolosa, Cheng Zhang, Hu Li, Jason D. Doles, Jonathan Bleeker, Michael T. Barrett, James H. Boyum, Benjamin R. Kipp, Amit Mahipal, Joleen M. Hubbard, Temperance J. Scheffler Hanson, Gloria M. Petersen, Surendra Dasari, Ann L. Oberg, Mark J. Truty, Rondell P. Graham, Michael J. Levy, Mojun Zhu, Daniel D. Billadeau, Alex A. Adjei, Nelson Dusetti, Juan L. Iovanna, Tanios S. Bekaii-Saab, Wen Wee Ma, Martin E. Fernandez-Zapico
Rest has long been considered beneficial to patient healing, yet remarkably there are no evidence-based experimental models determining how it benefits disease outcomes. Here, we create a novel experimental rest model in mice that briefly extends the morning rest period. We found, in two different major cardiovascular disease conditions (cardiac hypertrophy, myocardial infarction), that imposing a short, extended period of morning rest each day limits cardiac remodeling, as compared to controls. Mechanistically, rest mitigates autonomic-mediated hemodynamic stress on the cardiovascular system, relaxes myofilament contractility, attenuates cardiac remodeling genes, consistent with the benefits on cardiac structure and function. These same rest-responsive gene pathways underlie the pathophysiology of many major human cardiovascular conditions, as demonstrated by interrogating open-source transcriptomic data, and thus patients with other conditions may also benefit from a morning rest period in a similar manner. Our findings implicate rest as a key driver of physiology, creating an entirely new field – as broad and important as diet, sleep, or exercise – and provide a strong rationale for investigation of rest-based therapy for major clinical diseases.
Cristine J. Reitz, Mina Rasouli, Faisal J. Alibhai, Tarak Nath Khatua, W. Glen Pyle, Tami A. Martino
BRD4 is a bromodomain extra-terminal domain (BET) family member and functions primarily as a chromatin reader regulating genes involved in cell fate decisions. Here we bred Brd4f/fOx40-Cre mice in which Brd4 was conditionally deleted in OX40-expressing cells to examine the role of BRD4 in regulating immune responses. We found that the Brd4f/fOx40-Cre mice developed profound alopecia and dermatitis while other organs and tissues were not affected. Surprisingly, lineage-tracing experiments using the Rosa26f/f-Yfp mice identified a subset of hair follicle stem cells (HFSCs) that constitutively express OX40 and deletion of Brd4 specifically in such HFSCs resulted in cell death and a complete loss of skin hair growth. We also found that death of HFSCs triggered massive activation of the intra-dermal γδ T cells, which induced epidermal hyperplasia and dermatitis by producing the inflammatory cytokine IL-17. Interestingly, deletion of Brd4 in Foxp3+ Tregs, which also constitutively express OX40, compromised their suppressive functions and this in turn contributed to the enhanced activation of γδ T cells as well as the severity of dermatitis and hair follicle destruction. Thus, our data demonstrate an unexpected role of BRD4 in regulating skin follicle stem cells and skin inflammation.
Mou Wen, Yuanlin Ying, Xiang Xiao, Preston R. Arnold, Guangchuan Wang, Xiufeng Chu, Rafik M. Ghobrial, Xian C. Li
Pancreatic ductal adenocarcinoma (PDA) remains resistant to immune therapies, largely due to robustly fibrotic and immunosuppressive tumor microenvironments. It has been postulated that excessive accumulation of immunosuppressive myeloid cells influences immunotherapy resistance and recent studies targeting macrophages in combination with checkpoint blockade have demonstrated promising preclinical results. Yet, our understanding of tumor-associated macrophage (TAM) function, complexity, and diversity in PDA remains limited. Here, analysis reveals significant macrophage heterogeneity, with bone marrow-derived monocytes serving as the primary source for immunosuppressive TAMs. These cells also serve as a primary source of TNF-α, which suppresses expression of the alarmin IL33 in carcinoma cells. Deletion of Ccr2 in genetically engineered mice decreases monocyte recruitment resulting in profoundly decreased TNF-α and increased IL33 expression, decreased metastasis, and increased survival. Moreover, intervention studies targeting CCR2 with a new orthosteric inhibitor (CCX598) renders PDA susceptible to checkpoint blockade resulting in reduced metastatic burden and increased survival. Our data indicate that this shift in anti-tumor immunity is influenced by increased levels of IL-33, which increases dendritic cell and cytotoxic T cell activity. These data demonstrate that interventions to disrupt infiltration of immunosuppressive macrophages, or their signaling, have the potential to overcome barriers to effective immunotherapeutics for PDA.
Ajay Dixit, Aaron L. Sarver, Jon Zettervall, Huocong Huang, Kexin Zheng, Rolf A. Brekken, Paolo Provenzano
Hevin/Sparcl1 is an astrocyte-secreted protein and regulates synapse formation. Here we show that astrocytic hevin signaling plays a critical role in maintaining chronic pain. Compared to wild-type mice, hevin-null mice exhibited normal mechanical and heat sensitivity but reduced inflammatory pain. Interestingly, hevin-null mice have faster recovery than wild-type mice from neuropathic pain after nerve injury. Intrathecal injection of wild-type hevin was sufficient to induce persistent mechanical allodynia in naïve mice. In hevin-null mice with nerve injury, AAV-mediated re-expression of hevin in GFAP-expressing spinal cord astrocytes could reinstate neuropathic pain. Mechanistically, hevin is crucial for spinal cord NMDA receptor (NMDAR) signaling. Hevin potentiated NMDA currents mediated by the GluN2B-containing NMDARs. Furthermore, intrathecal injection of a neutralizing antibody against hevin alleviated acute and persistent inflammatory pain, postoperative pain, and neuropathic pain. Secreted hevin was detected in mouse cerebrospinal fluid (CSF) and nerve injury significantly increased CSF hevin abundance. Finally, neurosurgery caused rapid and substantial increases in SPARCL1/HEVIN levels in human CSF. Collectively, our findings support a critical role of hevin and astrocytes in the maintenance of chronic pain. Neutralizing of secreted hevin with monoclonal antibody may provide a new therapeutic strategy for treating acute and chronic pain and NMDAR-medicated neurodegeneration.
Gang Chen, Jing Xu, Hao Luo, Xin Luo, Sandeep K. Singh, Juan J. Ramirez, Michael L. James, Joseph P. Mathew, Miles Berger, Cagla Eroglu, Ru-Rong Ji
TP53 mutation (TP53mut) is one of the most important driver events facilitating tumorigenesis, which could induce a series of chain reactions to promote tumor malignant transformation. However, the malignancy progression patterns under TP53 mutation still remain less known. Clarifying the molecular landscapes of TP53mut tumors will help us understand the process of tumor development and aid precise treatment. Here, we distilled genetic and epigenetic features altered in TP53mut cancers for cluster-of-cluster analysis. Using integrated classification, we derived five different subtypes of TP53mut patients. These subtypes have distinct features in genomic alteration, clinical relevance, microenvironment dysregulation and potential therapeutics. Among the five subtypes, COCA3 was identified as the subtype with worst prognosis, causing an immunosuppressive microenvironment and immunotherapeutic resistantance. Further drug efficacy research highlighted olaparib as the most promising therapeutic agents for COCA3 tumors. Importantly, the therapeutic efficacy of olaparib in COCA3 and immunotherapy in non-COCA3 tumors was validated in vivo experiment. Summarily, our study first explored the important molecular events and developed a subtype classification system with distinct targeted therapy strategies for different subtypes of TP53mut tumors. These multi-omics classification systems provided a valuable resource that significantly expands the knowledge of TP53mut tumors and might eventually benefit in clinical practice.
Xin Chen, Tianqi Liu, Wu Jianqi, Chen Zhu, Gefei Guan, Cunyi Zou, Qing Guo, Xiaolin Ren, Chen Li, Peng Cheng, Wen Cheng, Anhua Wu
Preeclampsia is a serious pregnancy disorder that lacks effective treatments other than delivery. Improper sensing of oxygen changes during placentation by prolyl hydroxylases (PHD), specifically PHD2, causes placental Hypoxia-Inducible Factor-1 (HIF1) buildup and abnormal downstream signaling in early-onset preeclampsia; yet therapeutic targeting of HIF1 has never been attempted. Here we generated a conditional (placenta-specific) knockout of Phd2 in mice (Phd2-/- cKO) to reproduce HIF1 excess and to assess anti-HIF therapy. Conditional deletion of Phd2 in the junctional zone (JZ) during pregnancy increased placental HIF1 content, resulting in abnormal placentation, impaired remodeling of the uterine spiral arteries, and fetal growth restriction. Pregnant dams developed new-onset hypertension at mid-gestation (E9.5) in addition to proteinuria and renal and cardiac pathology, hallmarks of severe preeclampsia in humans. Daily injection of acriflavine, a small-molecule inhibitor of HIF1, to pregnant Phd2-/- cKO mice from E7.5 (prior to hypertension) or E10.5 (after hypertension has been established) to E14.5 corrected placental dysmorphologies and improved fetal growth. Moreover, it reduced maternal blood pressure and reverted renal and myocardial pathology. Thus, therapeutic targeting of the HIF pathway may improve placental development and function, as well as maternal and fetal health, in preeclampsia.
Julien Sallais, Chanho Park, Sruthi Alahari, Tyler Porter, Ruizhe Liu, Merve Kurt, Abby Farrell, Martin Post, Isabella Caniggia
Acute graft-versus-host disease (aGvHD) is a life-threatening complication of allogeneic hematopoietic cell transplantation (allo-HCT) inflicted by alloreactive T cells primed in secondary lymphoid organs (SLOs) and subsequent damage to aGvHD target tissues. In recent years, regulatory T cell (Treg) transfer and/or expansion has emerged as a promising therapy to modulate aGvHD. However, cellular niches essential for fostering Tregs to prevent aGvHD have not been explored, yet. Here, we tested whether and to what extent MHC class II (MHCII) expressed on Ccl19+ fibroblastic reticular cells (FRCs) shape the donor CD4+ T cell response during aGvHD. Animals lacking MHCII expression on Ccl19-Cre-expressing FRCs (MHCIIΔCcl19) showed aberrant CD4+ T cells activation in the effector phase resulting in exacerbated aGvHD that was associated with significantly reduced expansion of Foxp3+ Tregs and invariant natural killer T (iNKT) cells. Skewed Treg maintenance in MHCIIΔCcl19 mice resulted in loss of protection from aGvHD provided by adoptively transferred donor Tregs. In contrast, although FRCs upregulated co-stimulatory surface receptors, degraded and processed exogenous antigens after myeloablative irradiation, FRCs were dispensable to activate alloreactive CD4+ T cells in two mouse models of aGvHD. In sum, these data reveal an immunoprotective, MHCII-mediated function of FRC niches in secondary lymphoid organs (SLOs) after allo-HCT and highlights a hitherto unknown framework of cellular and molecular interactions that regulate CD4+ T cell alloimmunity.
Haroon Shaikh, Joern Pezoldt, Zeinab Mokhtari, Juan Gamboa Vargas, Duc-Dung Le, Josefina Peña Mosca, Estibaliz Arellano-Viera, Michael A.G. Kern, Caroline Graf, Niklas Beyersdorf, Manfred B. Lutz, Angela Riedel, Maike Büttner-Herold, Alma Zernecke, Hermann Einsele, Antoine-Emmanuel Saliba, Burkhard Ludewig, Jochen Huehn, Andreas Beilhack
The DNA methyltransferase inhibitor decitabine has classically been used to reactivate silenced genes and as a pre-treatment for anti-cancer therapies. In a new variation of this idea, this study explores the concept of adding low-dose decitabine following administration of chemotherapy to bolster therapeutic efficacy. We find that addition of decitabine following treatment with the chemotherapy gemcitabine improves survival and slows tumor growth in a mouse model of high-grade sarcoma. Unlike prior studies in epithelial tumor models, low-dose decitabine did not induce a robust anti-tumor T cell response in sarcoma. Furthermore, low-dose decitabine synergizes with gemcitabine independently of the immune system. Mechanistic analyses demonstrate that the combination therapy induces bi-phasic cell cycle arrest and apoptosis. Therapeutic efficacy was found to be sequence dependent, with gemcitabine priming cells for treatment with decitabine through inhibition of ribonucleotide reductase. This study identifies a unique application of low-dose decitabine to augment the cytotoxic effects of conventional chemotherapy in an immune-independent manner. The concepts explored in this study represent a promising new paradigm for cancer treatment by augmenting chemotherapy through addition of low-dose decitabine to increase tolerability and improve patient response. These findings have widespread implications for the treatment of sarcomas and other aggressive malignancies.
Wade R. Gutierrez, Amanda Scherer, Jeffrey D. Rytlewski, Emily A. Laverty, Alexa P. Sheehan, Gavin R. McGivney, Qierra R. Brockman, Vickie Knepper-Adrian, Grace A. Roughton, Dawn E. Quelle, David J. Gordon, Varun Monga, Rebecca D. Dodd
The fluid covering the surface of airway epithelia represents a first barrier against pathogens. The chemical and physical properties of the airway surface fluid are controlled by the activity of ion channels and transporters. In cystic fibrosis (CF), loss of CFTR chloride channel function causes airway surface dehydration, bacterial infection, and inflammation. We investigated the effects of IL-17A plus TNF-α, two cytokines with a relevant role in CF and other chronic lung diseases. Transcriptome analysis revealed a profound change with upregulation of several genes involved in ion transport, anti-bacterial defense, and neutrophil recruitment. At the functional level, bronchial epithelia treated in vitro with the cytokine combination showed upregulation of ENaC sodium channel, ATP12A proton pump, ADRB2 beta-adrenergic receptor, and SLC26A4 anion exchanger. The overall result of IL-17A/TNF-α treatment was hyperviscosity of the airway surface as demonstrated by fluorescence recovery after photobleaching (FRAP) experiments. Importantly, stimulation with a beta-adrenergic agonist switched airway surface to a low viscosity state in non-CF but not in CF epithelia. Our study suggests that CF lung disease is sustained by a vicious cycle in which epithelia cannot exit from the hyperviscous state thus perpetuating the proinflammatory airway surface condition.
Daniela Guidone, Martina Buccirossi, Paolo Scudieri, Michele Genovese, Sergio Sarnataro, Rossella De Cegli, Federico Cresta, Vito Terlizzi, Gabrielle Planelles, Gilles Crambert, Isabelle Sermet-Gaudelus, Luis J.V. Galietta
Acquired aplastic anemia (AA) is caused by autoreactive T-cell-mediated destruction of early hematopoietic cells. Somatic loss of human leukocyte antigen (HLA) Class I alleles was identified as a mechanism of immune escape in surviving hematopoietic cells of some AA patients. However, pathogenicity, structural characteristics and clinical impact of specific HLA alleles in AA remain poorly understood. Here, we evaluated somatic HLA loss in 505 AA patients from two multi-institutional cohorts. Using a combination of HLA mutation frequencies, peptide-binding structures, and association with AA in an independent cohort of 6,323 patients from the National Marrow Donor Program, we identified 19 AA risk alleles and 12 non-risk alleles and established a novel AA HLA pathogenicity stratification. Our results define pathogenicity for the majority of common HLA-A/B alleles across diverse populations. Our study demonstrates that HLA alleles confer different risks of developing AA, but once AA develops, specific alleles are not associated with response to immunosuppression or trans-plant outcomes. However, higher pathogenicity alleles, particularly HLA-B*14:02, are associated with higher rates of clonal evolution in adult AA patients. Our study provides novel insights into the immune pathogenesis of AA, opening the door to future autoantigen identification and improved under-standing of clonal evolution in AA.
Timothy S. Olson, Benjamin F. Frost, Jamie L. Duke, Marian Dribus, Hongbo M. Xie, Zachary D. Prudowsky, Elissa Furutani, Jonas Gudera, Yash B. Shah, Deborah Ferriola, Amalia Dinou, Ioanna Pagkrati, Soyoung Kim, Yixi Xu, Meilun He, Shannon Zheng, Sally Nijim, Ping Lin, Chong Xu, Taizo Nakano, Joseph H. Oved, Beatriz M. Carreno, Yung-Tsi Bolon, Shahinaz M. Gadalla, Steven G.E. Marsh, Sophie Paczesny, Stephanie J. Lee, Dimitrios S. Monos, Akiko Shimamura, Alison A. Bertuch, Loren Gragert, Stephen Spellman, Daria V. Babushok
Rosacea is a chronic skin disorder characterized by abnormal neurovascular and inflammatory conditions on the central face. Despite increasing evidence suggests that rosacea is associated with metabolic disorders, the role of metabolism in rosacea pathogenesis remains unknown. Here, via targeted metabolomics approach, we characterized significantly altered metabolic signatures in rosacea patients, especially for amino acid-related metabolic pathways. Among these, glutamic acid and aspartic acid are highlighted and positively correlated with the disease severity in rosacea patients. We further demonstrated that glutamic acid and aspartic acid can facilitate the development of erythema and telangiectasia, typical features of rosacea, in the skin of mice. Mechanistically, glutamic acid and aspartic acid stimulate the production of vasodilation-related neuropeptides from peripheral neuron and keratinocytes, and induce the release of nitric oxide from endothelial cells and keratinocytes. Interestingly, we provided evidence showing that doxycycline can improve the symptoms of rosacea patients possibly by targeting amino acid metabolic pathway. These findings reveal that abnormal amino acid metabolism promotes neurovascular reactivity in rosacea, and raise the possibility of targeting dysregulated metabolism as a promising strategy for clinical treatment.
Tangxiele Liu, Wenqin Xiao, Mengting Chen, Rui Mao, San Xu, Qinqin Peng, Zhixiang Zhao, Qian Wang, Hongfu Xie, Zhili Deng, Ji Li
Expression of the transcription factor Interferon Regulatory Factor 4 (IRF4) is required for the development of lung conventional dendritic cells type 2 (cDC2s) that elicit Th2 responses, yet how IRF4 functions in lung cDC2s throughout the acute and memory allergic response is not clear. Here, we use a novel mouse model that loses IRF4 expression after lung cDC2 development to demonstrate that mice with IRF4-deficient DCs display impaired memory responses to allergen. This defect in the memory response is a direct result of ineffective Th2 induction and impaired recruitment of activated effector T cells to the lung after sensitization. IRF4-deficient DCs demonstrate defects in their migration to the draining lymph node and in T cell priming. Finally, T cells primed by IRF4-competent DCs mediate potent memory responses independently of IRF4-expressing DCs, demonstrating that IRF4-expressing DCs are not necessary during the memory response. Thus, IRF4 controls a program in mature DCs governing Th2 priming and effector responses, but IRF4-expressing DCs are dispensable during tissue resident-memory T cell (TRM cell)-dependent memory responses.
Daniel F. Camacho, Tania E. Velez, Maile K. Hollinger, Esther Wang, Chanie L. Howard, Eli P. Darnell, Domenick E. Kennedy, Paulette A. Krishack, Cara L. Hrusch, Marcus R. Clark, James J. Moon, Anne I. Sperling
Pseudomonas aeruginosa undergoes diversification during infection of the cystic fibrosis (CF) lung. Understanding these changes requires model systems that capture the complexity of the CF lung environment. We previously identified loss-of-function mutations in the two-component regulatory system sensor kinase gene pmrB, in P. aeruginosa from CF and from experimental infection of mice. Here, we demonstrate that whilst such mutations lower in vitro MICs for multiple antimicrobial classes, this is not reflected in increased antibiotic susceptibility in vivo. Loss of PmrB impairs aminoarabinose modification of lipopolysaccharide, increasing the negative charge of the outer membrane and promoting uptake of cationic antimicrobials. However, in vivo, this can be offset by increased membrane binding of other positively charged molecules present in lungs. The polyamine spermidine readily coats the surface of PmrB-deficient P. aeruginosa, reducing susceptibility to antibiotics that rely on charge differences to bind the outer membrane and increasing biofilm formation. Spermidine is elevated in lungs during P. aeruginosa infection in mice and during episodes of antimicrobial treatment in people with CF. These findings highlight the need to study antimicrobial resistance under clinically relevant environmental conditions. Microbial mutations carrying fitness costs in vitro may be advantageous during infection, where host resources can be utilised.
Chowdhury M. Hasan, Sian Pottenger, Angharad E. Green, Adrienne A. Cox, Jack S. White, Trevor Jones, Craig Winstanley, Aras Kadioglu, Megan H. Wright, Daniel R. Neill, Joanne L. Fothergill
The individual contribution of specific myeloid subsets such as CD1c+ conventional dendritic cells (cDC) to perpetuation of Rheumatoid Arthritis (RA) pathology remains unclear. In addition, the specific innate sensors driving pathogenic activation of CD1c+ cDCs in RA patients and their functional implications have not been characterized. Here, we assessed phenotypical, transcriptional and functional characteristics of CD1c+ and CD141+ cDCs and monocytes from the blood and synovial fluid of RA patients. Increased levels of CCR2 and the IgG receptor CD64 on circulating CD1c+ cDC associated with the presence of this DC subset in the synovial membrane in RA patients. Moreover, synovial CD1c+ cDCs are characterized by increased expression of proinflammatory cytokines and high abilities to induce pathogenic IFNγ+IL-17+ CD4+ T cells in vitro. Finally, we identified the crosstalk between Fcγ Receptors and NLRC4 as a new potential molecular mechanism mediating pathogenic activation, CD64 upregulation and functional specialization of CD1c+ cDCs in response to dsDNA-IgG in RA patients.
Cristina Delgado-Arévalo, Marta Calvet-Mirabent, Ana Triguero-Martinez, Enrique Vázquez de Luis, Alberto Benguría-Filippini, Raquel Largo, Diego Calzada-Fraile, Olga Popova, Ildefonso Sánchez-Cerrillo, Ilya Tsukalov, Roberto Moreno-Vellisca, Hortensia de la Fuente, Gabriel Herrero-Beaumont, Almudena R. Ramiro, Francisco Sánchez-Madrid, Santos Castañeda, Ana Dopazo, Isidoro González-Álvaro, Enrique Martin-Gayo
No disease-modifying drug exists for osteoarthritis (OA). Despite success in animal models, candidate drugs continue to fail in clinical trials due to the unmapped interpatient heterogeneity and disease complexity. We have utilized a single-cell cytometry-by-time-of-flight (cyTOF) based platform to precisely outline the effects of candidate drugs on human OA chondrocytes. OA chondrocytes harvested from patients undergoing total knee arthroplasty were treated with two drugs, an NF-κB pathway inhibitor, BMS-345541, and a chondroinductive small molecule, Kartogenin, that showed preclinical success in animal models for OA. cyTOF conducted with 30 metal isotope-labeled antibodies parsed the effects of the drugs on inflammatory, senescent, and chondroprogenitor populations. The NF-κB pathway inhibition decreased the expression of NF-κB, HIF2A and iNOS in multiple chondrocyte clusters and significantly depleted four p16ink4a expressing senescent populations including NOTCH1+STRO1+ chondroprogenitors. While Kartogenin also affected select p16ink4a expressing senescent clusters, there was a less discernible effect on chondroprogenitor populations. Overall, BMS-345541 elicited a uniform drug response in all patients while only a few responded to Kartogenin. These studies demonstrate that a single-cell cyTOF-based drug screening platform can provide insights into patient response assessment and their stratification.
Neety Sahu, Fiorella C. Grandi, Nidhi Bhutani