Humoral immunity is important in limiting clinical disease in malaria, yet the longitudinal B cell response to infection remains unclear. We performed a 1-year prospective study in patients treated for acute P. falciparum malaria for the first time, or with previous exposure to the disease. Using an unbiased exploratory approach with mass cytometry, followed by targeted flow cytometry, we found that ~80% of mature B cells that proliferated in response to acute infection expressed CD11c. Only ~40% of CD11c+ B cells displayed an atypical B cell phenotype, with the remaining cells primarily made up of activated- and resting memory B cells. The CD11c+ B cells expanded rapidly following infection, with previous exposure to malaria resulting in a significantly larger increase compared to individuals with primary infection. This was attributed to an expansion of switched CD11c+ B cells that was absent in primary infected individuals. The rate of contraction of the CD11c+ B cell compartment was independent of previous exposure to malaria and displayed a slow decay with a half-life of ~300 days. Collectively, these results identify CD11c as a marker of B cells responding to malaria and further highlight differences in primary- and secondary B cell responses during infection.
Christopher Sundling, Caroline Rönnberg, Victor Yman, Muhammad Asghar, Peter Jahnmatz, Tadepally Lakshmikanth, Yang Chen, Jaromir Mikes, Mattias N. Forsell, Klara Sondén, Adnane Achour, Petter Brodin, Kristina E.M. Persson, Anna Färnert
Human antibody-secreting cells (ASC) triggered by immunization are globally recognized as CD19loCD38hiCD27hi. Yet, different vaccines give rise to antibody responses of different longevity, suggesting ASC populations are heterogeneous. We define circulating ASC heterogeneity in vaccine responses using multi-color flow cytometry, morphology, VH repertoire, and RNA transcriptome analysis. We also tested differential survival using a novel human cell-free system that mimics the bone-marrow (BM) microniche. In peripheral blood, we identified three CD19pos and two CD19neg ASC subsets. All subsets contributed to the vaccine-specific responses and were characterized by in vivo proliferation and activation. VH repertoire demonstrated strong oligoclonality with extensive interconnectivity among the five subsets and switched memory B cells. Transcriptome analysis showed separation of CD19pos and CD19neg subsets that included pathways such as cell cycle, hypoxia, TNFA, and unfolded protein response (UPR). They also demonstrated similar long-term in vitro survival after 48 days. In summary, vaccine-induced ASC with different surface markers (CD19 and CD138) derive from shared proliferative precursors yet express distinctive transcriptomes. Equal survival indicates that all ASC compartments are endowed with long-lived potential. Accordingly, in vivo survival of peripheral long-lived plasma cells may be determined in part by their homing and residence in the BM microniche.
Swetha Garimilla, Doan C. Nguyen, Jessica L. Halliley, Christopher Tipton, Alexander F. Rosenberg, Christopher F. Fucile, Celia L. Saney, Shuya Kyu, Denise Kaminski, Yu Qian, Richard H. Scheuermann, Greg Gibson, Inaki Sanz, F. Eun-Hyung Lee
Following injury, leukocytes are released from hematopoietic organs and migrate to the site of damage to regulate tissue inflammation and repair, however leukocytes lacking β2-adrenergic receptor (β2AR) expression have marked impairments in these processes. β-blockade is a common strategy for the treatment of many cardiovascular etiologies, therefore the objective of our study was to assess the impact of prior β-blocker treatment on baseline leukocyte parameters and their responsiveness to acute injury. In a temporal and βAR isoform-dependent manner, chronic β-blocker infusion increased splenic vascular cell adhesion molecule-1 (VCAM-1) expression and leukocyte accumulation (monocytes/macrophages, mast cells and neutrophils) and decreased chemokine receptor 2 (CCR2) expression, migration of bone marrow cells (BMC) and peripheral blood leukocytes (PBL), as well as infiltration into the heart following acute cardiac injury. Further, CCR2 expression and migratory responsiveness was significantly reduced in the PBL of patients receiving β-blocker therapy compared to β-blocker-naïve patients. These results highlight the ability of chronic β-blocker treatment to alter baseline leukocyte characteristics that decrease their responsiveness to acute injury and suggest that prior β-blockade may act to reduce the severity of innate immune responses.
Laurel A. Grisanti, Claudio de Lucia, Toby P. Thomas, Aron Stark, John T. Strony, Valerie D. Myers, Remus Berretta, Daohai Yu, Celestino Sardu, Raffaele Marfella, Erhe Gao, Steven R. Houser, Walter J. Koch, Eman A. Hamad, Douglas G. Tilley
Skeletal muscle weakness in patients suffering from rheumatoid arthritis (RA) adds to their impaired working abilities and reduced quality of life. However, little molecular insight is available on muscle weakness associated with RA. Oxidative stress has been implicated in the disease pathogenesis of RA. Here we show that oxidative post-translational modifications of the contractile machinery targeted to actin result in impaired actin polymerization and reduced force production. Using mass spectrometry, we identified the actin residues targeted by oxidative 3-nitrotyrosine (3-NT) or malondialdehyde adduct (MDA) modifications in weakened skeletal muscle from mice with arthritis and patients afflicted by RA. The residues were primarily located to three distinct regions positioned at matching surface areas of the skeletal muscle actin molecule from arthritis mice and RA patients. Moreover, molecular dynamic simulations revealed that these areas, here coined “hotspots”, are important for the stability of the actin molecule and its capacity to generate filaments and interact with myosin. Together, these data demonstrate how oxidative modifications on actin promote muscle weakness in RA patients and provide novel leads for targeted therapeutic treatment to improve muscle function.
Maarten M. Steinz, Malin Persson, Bejan Aresh, Karl Olsson, Arthur J. Cheng, Emma Ahlstrand, Mats Lilja, Tommy R. Lundberg, Eric Rullman, Kristina Ängeby Möller, Katalin Sandor, Sofia Ajeganova, Takashi Yamada, Nicole Beard, Björn C.G. Karlsson, Pasi Tavi, Ellinor Kenne, Camilla I. Svensson, Dilson E. Rassier, Roger Karlsson, Ran Friedman, Thomas Gustafsson, Johanna T. Lanner
Tregs require IL-2 signaling for signal transducer and activator of transcription 5 (STAT5)-mediated induction of Foxp3. While phosphatase 2A (PP2A) is a negative regulator of IL-2 production in effector T cells and Tregs do not produce IL-2, it is not known whether PP2A controls IL-2 signaling in Tregs. To address the role of PP2A in IL-2 signaling in Tregs we studied mice engineered to lack PP2A in all Foxp3-expressing cells. We report that PP2A is required to enable Foxp3 expression and to maintain sufficient numbers of Tregs in the thymus. We show for the first time that PP2A prevents the selective loss of surface IL-2Rβ and preserves IL-2R signaling potency in Tregs. The loss of IL-2Rβ in thymus- and spleen-derived Tregs that lack PP2A is due to increased sheddase activity. Pan-sheddase or selective A disintegrin and metalloproteinase 10 (ADAM10) inhibition, like forced expression of IL-2Rβ in PP2A-deficient Tregs restored IL-2Rβ expression and signaling. Thus, PP2A restrains the sheddase activity of ADAM10 in Treg cells to prevent the cleavage of IL-2Rβ from the cell surface to enable competent IL-2R signaling which is essential for Tregs development and homeostasis.
Amir Sharabi, Hao Li, Isaac R. Kasper, Wenliang Pan, Esra Meidan, Maria G. Tsokos, Vaishali R. Moulton, George C. Tsokos
Drug refractory epilepsy (RE) is a chronic neurological disease with varied etiology that represents a group of patients whose seizures do not respond to anti-epileptic drugs. The immune system may have a role in seizure and epilepsy development, but the specific mechanisms of inflammation that lead to epileptogenesis and contribute to RE are unknown. Here, we used mass cytometry to comprehensively study the immune system of pediatric patients with RE and compared their immune profile and function with patients with age-matched autoimmune encephalitis (AIE) and healthy controls. Patients with RE and AIE displayed similar immune profiles overall, with changes in CD4+ and CD8+ T-cell subsets and an unbalance toward pro-inflammatory IL-17 production. In addition, patients with RE uniquely showed an altered balance in natural killer cell subsets. A systems level intercellular network analysis identified rewiring of the immune system leading to loss of inhibitory/regulatory intercellular connections and emergence of pro-inflammatory pathogenic functions in neuro-inflammatory immune-cell networks in patients with AIE and RE. These data underscore the contribution of systemic inflammation to the pathogenesis of seizures and epileptogenesis and have direct translational implications in advancing diagnostics and therapeutics design.
Pavanish Kumar, Derrick Wei Shih Chan, Amanda Lim, Bhairav Paleja, Simon Ling, Lai Li Yun, Su Li Poh, Adeline Ngoh, Thaschawee Arkachaisri, Joo Guan Yeo, Salvatore Albani
Many lung diseases result from a failure of efficient regeneration of damaged alveolar epithelial cells (AECs) after lung injury. During regeneration, AEC2s proliferate to replace lost cells, after which proliferation halts and some AEC2s transdifferentiate into AEC1s to restore normal alveolar structure and function. Although the mechanisms underlying AEC2 proliferation have been studied, the mechanisms responsible for halting proliferation and inducing transdifferentiation are poorly understood. To identify candidate signaling pathways responsible for halting proliferation and inducing transdifferentiation, we performed single cell RNA sequencing on AEC2s during regeneration in a murine model of lung injury induced by intratracheal LPS. Unsupervised clustering revealed distinct subpopulations of regenerating AEC2s: proliferating, cell cycle arrest, and transdifferentiating. Gene expression analysis of these transitional subpopulations revealed that TGFβ signaling was highly upregulated in the cell cycle arrest subpopulation and relatively downregulated in transdifferentiating cells. In cultured AEC2s, TGFβ was necessary for cell cycle arrest but impeded transdifferentiation. We conclude that during regeneration after LPS-induced lung injury, TGFβ is a critical signal halting AEC2 proliferation but must be inactivated to allow transdifferentiation. This study provides insight into the molecular mechanisms regulating alveolar regeneration and the pathogenesis of diseases resulting from a failure of regeneration.
Kent A. Riemondy, Nicole L. Jansing, Peng Jiang, Elizabeth F. Redente, Austin E. Gillen, Rui Fu, Alyssa J. Miller, Jason R. Spence, Anthony N. Gerber, Jay R. Hesselberth, Rachel L. Zemans
We have previously reported that the carboxy-terminal proteolytic cleavage product of the COL6α3 chain that we refer to as “endotrophin” has potent effects on transformed mammary ductal epithelial cells in rodents. Endotrophin (ETP) is abundantly expressed in adipose tissue. It is a chemoattractant for macrophages, exerts effects on endothelial cells and through epithelial-mesenchymal transition (EMT) enhances progression of tumor cells. In a recombinant form, human endotrophin exerts similar effects on human macrophages and endothelial cells as its rodent counterpart. It enhances EMT in human breast cancer cells and upon overexpression in tumor cells, the cells become chemoresistant. Here, we report the identification of endotrophin from human plasma. It is circulating at higher levels in breast cancer patients. We have developed neutralizing monoclonal antibodies against human endotrophin and provide evidence for the effectiveness of these antibodies to curb tumor growth and enhance chemosensitivity in a nude mouse model carrying human tumor cell lesions. Combined, the data validate endotrophin as a viable target for anti-tumor therapy for human breast cancer and opens the possibility for further use of these new reagents for anti-fibrotic approaches in liver, kidney, bone marrow and adipose tissue.
Dawei Bu, Clair Crewe, Christine M. Kusminski, Ruth Gordillo, Alexandra L. Ghaben, Min Kim, Jiyoung Park, Hui Deng, Wei Xiong, Xiao-Zheng Liu, Per Eystein Lønning, Nils Halberg, Adan Rios, Yujun Chang, Anneliese Gonzalez, Ningyan Zhang, Zhiqiang An, Philipp E. Scherer
In demyelinating diseases such as Multiple Sclerosis (MS), demyelination of neuronal fibers impairs impulse conduction and causes axon degeneration. While neuronal activity stimulates oligodendrocyte production and myelination in normal conditions, it remains unclear whether the activity of demyelinated axons restores their loss-of-function in a harmful environment. To investigate this question, we established a model to induce a moderate optogenetic stimulation of demyelinated axons in the corpus callosum at the level of the motor cortex in which cortical circuit activation and locomotor effects were reduced in adult freely moving mice. We demonstrate that a moderate activation of demyelinated axons enhances the differentiation of oligodendrocyte precursor cells onto mature oligodendrocytes, but only under a repeated stimulation paradigm. This activity-dependent increase in the oligodendrocyte pool promotes an extensive remyelination and functional restoration of conduction, as revealed by ultrastructural analyses and compound action potential recordings. Our findings reveal the need of preserving an appropriate neuronal activity in the damaged tissue to promote oligodendrocyte differentiation and remyelination, likely by enhancing axon-oligodendroglia interactions. Our results provide new perspectives for translational research using neuromodulation in demyelinating diseases.
Fernando C. Ortiz, Chloé Habermacher, Mariana Graciarena, Pierre-Yves Houry, Akiko Nishiyama, Brahim Nait-Oumesmar, Maria Cecilia Angulo
Changes in neuronal activity alter blood flow to match energy demand with the supply of oxygen and nutrients. This functional hyperemia is maintained by interactions between neurons, vascular cells, and glia. However, how changing neuronal activity prevalent at the onset of neurodegenerative disease affects neurovascular elements is unclear. Here, in mice with photoreceptor degeneration, a model of neuron-specific dysfunction, we combined assessment of visual function, neurovascular unit structure, and the blood-retina barrier permeability. We found that the rod loss paralleled remodeling of the neurovascular unit, comprised of photoreceptors, retinal pigment epithelium, and Muller glia. When significant visual function was still present, blood flow became disrupted and blood-retina barrier began to fail, facilitating cone loss and vision decline. Thus, in contrast to the established view, vascular deficit in neuronal degeneration is not a late consequence of neuronal dysfunction, but is present early in the course of disease. These findings further establish the importance of vascular deficit and blood retina barrier function in neuron-specific loss, and highlight it as a target for early therapeutic intervention.
Elena Ivanova, Nazia M. Alam, Glen T. Prusky, Botir T. Sagdullaev
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