Recently, by utilizing conventional and tamoxifen inducible Bmal1 (Brain and muscle Arnt-like protein 1) knockout mice, we found that delaying the loss of circadian rhythms to adulthood attenuates the impact on general integrity and survival at least under 12h light/12h dark conditions (LD). To understand further the contribution of Bmal1 in post-natal life under conditions of circadian disruption, we subjected inducible knockout mice (KO) and their littermate controls (Ctrl) to forced desynchrony protocols including cycles with non-24h periods, randomized light/dark cycles, and jet lag, and monitored their locomotor activity using radiotelemetry. Under these conditions, control mice cannot be entrained, as reflected by their maintenance of circadian behavior irrespective of schedules. By contrast, KO mice displayed higher activity levels in the dark phases of most cycles. Under a 3h light/3h dark regime, Ctrls displayed higher activity levels in the dark phases of all cycles although there were still obvious circadian rhythms, suggesting that an ultradian mechanism is also involved. Insulin sensitivity was markedly reduced by disrupted light schedules as expected in Ctrls, but not in the KOs. Thus, Bmal1 deletion in adult mice facilitates adaptation to new light/dark schedules and protects from insulin resistance induced by circadian disruption.
Guangrui Yang, Lihong Chen, Jiayang Zhang, Baoyin Ren, Garret A. FitzGerald
Biomechanical forces and endothelial-to-mesenchymal transition (EndoMT) are known to mediate valvulogenesis. However, the relative contributions of myocardial contractile and hemodynamic shear forces remain poorly understood. We integrated 4-D light-sheet imaging of transgenic zebrafish models with moving-domain computational fluid dynamics to determine effects of changes in contractile forces and fluid wall shear stress (WSS) on ventriculobulbar (VB) valve development. Augmentation of myocardial contractility with isoproterenol increased both WSS and Notch1b activity in the developing outflow tract (OFT) and resulted in VB valve hyperplasia. Increasing WSS in the OFT, achieved by increasing blood viscosity through EPO mRNA injection, also resulted in VB valve hyperplasia. Conversely, decreasing myocardial contractility by Tnnt2a morpholino oligonucleotide (MO) administration, 2,3-butanedione monoxime treatment, or Plcγ1 inhibition completely blocked VB valve formation, which could not be rescued by increasing WSS or activating Notch. Decreasing WSS in the OFT, achieved by slowing heart rate with metoprolol or reducing viscosity with Gata1a MO, did not affect VB valve formation. Immunofluorescent staining with the mesenchymal marker, DM-GRASP, revealed that biomechanical force-mediated Notch1b activity is implicated in EndoMT to modulate valve morphology. Altogether, increases in WSS result in Notch1b- EndoMT-mediated VB valve hyperplasia, whereas decreases in contractility result in reduced Notch1b activity, absence of EndoMT, and VB valve underdevelopment. Thus, we provide developmental mechanotransduction mechanisms underlying Notch1b-mediated EndoMT in the OFT.
Jeffrey J. Hsu, Vijay Vedula, Kyung In Baek, Cynthia Chen, Junjie Chen, Man In Chou, Jeffrey Lam, Shivani Subhedar, Jennifer Wang, Yichen Ding, Chih-Chiang Chang, Juhyun Lee, Linda L. Demer, Yin Tintut, Alison L. Marsden, Tzung K. Hsiai
Extracellular mRNAs (ex-mRNAs) potentially supersede extracellular miRNAs (ex-miRNAs) and other RNA classes as biomarkers. We performed conventional small-RNA-sequencing (sRNA-seq) and sRNA-seq with T4 polynucleotide kinase (PNK) end-treatment of total exRNA isolated from serum and platelet-poor EDTA, ACD, and heparin plasma to study the effect on ex-mRNA capture. Compared to conventional sRNA-seq PNK-treatment increased the detection of informative ex-mRNAs reads up to 50-fold. The exRNA pool was dominated by hematopoietic cells and platelets, with additional contribution from the liver. About 60% of the 15- to 42-nt reads originated from the coding sequences, in a pattern reminiscent of ribosome-profiling. Blood sample type had a considerable influence on the exRNA profile. On average approximately 350 to 1,100 distinct ex-mRNA transcripts were detected depending on plasma type. In serum, additional transcripts from neutrophils and hematopoietic cells increased this number to near 2,300. EDTA and ACD plasma showed a destabilizing effect on ex mRNA and non-coding RNA ribonucleoprotein complexes compared to other plasma types. In a proof-of-concept study, we investigated differences between the exRNA profiles of patients with acute coronary syndrome (ACS) and healthy controls. The improved tissue resolution of ex mRNAs after PNK-treatment enabled us to detect a neutrophil-signature in ACS that escaped detection by ex miRNA analysis.
Kemal M. Akat, Youngmin A. Lee, Arlene Hurley, Pavel Morozov, Klaas E.A. Max, Miguel Brown, Kimberly Bogardus, Anuoluwapo Sopeyin, Kai Hildner, Thomas G. Diacovo, Markus F. Neurath, Martin Borggrefe, Thomas Tuschl
Breast cancer bone metastases often cause a debilitating non-curable condition with osteolytic lesions, muscle weakness and a high mortality. Current treatment comprises chemotherapy, irradiation, surgery and anti-resorptive drugs that restrict but do not revert bone destruction. In metastatic breast cancer cells, we determined the expression of sclerostin, a soluble Wnt inhibitor that represses osteoblast differentiation and bone formation. In mice with breast cancer bone metastases, pharmacological inhibition of sclerostin using an anti-sclerostin antibody (Scl-Ab) reduced metastases without tumor cell dissemination to other distant sites. Sclerostin inhibition prevented the cancer-induced bone destruction by augmenting osteoblast-mediated bone formation and reducing osteoclast-dependent bone resorption. During advanced disease, NF-κB and p38 signaling was increased in muscles in a TGF-β1-dependent manner, causing muscle fiber atrophy, muscle weakness and tissue regeneration with an increase in Pax7-positive satellite cells. Scl-Ab treatment restored NF-κB and p38 signaling, the abundance of Pax7-positive cells and ultimately muscle function. These effects improved the overall health condition and expanded the life span of cancer-bearing mice. Together, these results demonstrate that pharmacological inhibition of sclerostin reduces bone metastatic burden and muscle weakness with a prolongation of the survival time. This might provide novel options for treating musculoskeletal complications in breast cancer patients.
Eric Hesse, Saskia Schröder, Diana Brandt, Jenny Pamperin, Hiroaki Saito, Hanna Taipaleenmäki
Imatinib (Gleevec) reverses type 1 diabetes (T1D) in NOD mice and is currently in clinical trials in individuals with recent-onset disease. While research has demonstrated that imatinib protects islet β cells from the harmful effects of ER stress, the role the immune system plays in its reversal of T1D has been less well understood, and specific cellular immune targets have not been identified. In this study, we demonstrate that B lymphocytes, an immune subset that normally drives diabetes pathology, are unexpectedly required for reversal of hyperglycemia in NOD mice treated with imatinib. In the presence of B lymphocytes, reversal was linked to an increase in serum insulin concentration, but not an increase in islet β cell mass or proliferation. However, improved β cell function was reflected by a partial recovery of MafA transcription factor expression, a sensitive marker of islet β cell stress that is important to adult β cell function. Imatinib treatment was found to increase the antioxidant capacity of B lymphocytes, improving reactive oxygen species (ROS) handling in NOD islets. This study reveals a novel mechanism through which imatinib enables B lymphocytes to orchestrate functional recovery of T1D β cells.
Christopher S. Wilson, Jason M. Spaeth, Jay Karp, Blair T. Stocks, Emilee M. Hoopes, Roland W. Stein, Daniel J. Moore
Mitogen-activated protein kinase (MAPK) signaling consists of an array of successively acting kinases. The extracellular signal-regulated kinases 1/2 (ERK1/2) are major components of the greater MAPK cascade that transduce growth factor signaling at the cell membrane. Here we investigated ERK1/2 signaling in skeletal muscle homeostasis and disease. Using mouse genetics, we observed that the muscle-specific expression of a constitutively active MEK1 mutant promotes greater ERK1/2 signaling that mediates fiber-type switching to a slow, oxidative phenotype with type I myosin heavy chain expression. Using a conditional and temporally regulated Cre strategy as well as Mapk1 (ERK2) and Mapk3 (ERK1) genetically targeted mice, MEK1-ERK2 signaling was shown to underlie this fast-to-slow fiber type switching in adult skeletal muscle as well as during development. Physiologic assessment of these activated MEK1-ERK1/2 mice showed enhanced metabolic activity and oxygen consumption with greater muscle fatigue resistance. Moreover, induction of MEK1-ERK1/2 signaling increased dystrophin and utrophin protein expression in a mouse model of limb-girdle muscle dystrophy and protected myofibers from damage. In summary, sustained MEK1-ERK1/2 activity in skeletal muscle produces a fast-to-slow fiber-type switch that protects from muscular dystrophy, suggesting a therapeutic approach to enhance the metabolic effectiveness of muscle and protect from dystrophic disease.
Justin G. Boyer, Vikram Prasad, Taejeong Song, Donghoon Lee, Xing Fu, Kelly M. Grimes, Michelle A. Sargent, Sakthivel Sadayappan, Jeffery D. Molkentin
A vast body of literature has established GRK2 as a key player in the development and progression of heart failure. Inhibition of GRK2 improves cardiac function post injury in numerous animal models. In recent years, discovery of several non-canonical GRK2 targets has expanded our view of this kinase. Here, we describe the novel and exciting finding that cardiac GRK2 activity can regulate whole body metabolism. Transgenic mice with cardiac-specific expression of a peptide inhibitor of GRK2 (TgβARKct) display an enhanced obesogenic phenotype when fed a high fat diet (HFD). In contrast, mice with cardiac-specific overexpression of GRK2 (TgGRK2) show resistance to HFD induced obesity. White adipose tissue (WAT) mass was significantly enhanced in HFD fed TgβARKct mice. Furthermore, regulators of adipose differentiation were differentially regulated in WAT from mice with gain or loss of GRK2 function. Using complex metabolomics we found that cardiac GRK2 signaling altered myocardial BCAA and endocannabinoid metabolism and modulated circulating BCAA and endocannabinoid metabolite profiles on a HFD, and one of the BCAA metabolites identified here enhances adipocyte differentiation in vitro. Taken together, these results suggest that metabolic changes in the heart due to GRK2 signaling on a HFD control whole body metabolism.
Benjamin P. Woodall, Kenneth S. Gresham, Meryl A. Woodall, Mesele-Christina Valenti, Alessandro Cannavo, Jessica Pfleger, J. Kurt Chuprun, Konstantinos Drosatos, Walter J. Koch
BACKGROUND. Subgroups of patients with relapsed or refractory (R/R) chronic lymphocytic leukemia (CLL) exhibit suboptimal outcomes after standard therapies, including oral kinase inhibitors. We and others have previously reported on safety and efficacy of autologous CD19-targeted CAR T-cells for these patients; here we report safety and long-term follow-up of CAR T-cell therapy with or without conditioning chemotherapy for patients with R/R CLL and indolent B-cell non-Hodgkin lymphoma (B-NHL). METHODS. We conducted a phase 1 clinical trial investigating CD19-targeted CAR T-cells incorporating a CD28 costimulatory domain (19-28z). Seventeen of 20 patients received conditioning chemotherapy prior to CAR T-cell infusion. Five patients with CLL received ibrutinib at the time of autologous T-cell collection and/or CAR T-cell administration. RESULTS. This analysis included 16 patients with R/R CLL and 4 patients with R/R indolent B-NHL. Cytokine release syndrome (CRS) was observed in all 20 patients but grades 3 and 4 CRS and neurological events were uncommon (10% for each). Ex vivo expansion of T-cells and proportions of CD4+/CD8+ CAR T-cells with CD62L+CD127+ immunophenotype were significantly greater in patients on ibrutinib at leukapheresis. Three of 12 evaluable CLL patients receiving conditioning chemotherapy achieved CR (two had minimal residual disease–negative CR). All patients achieving CR remained progression-free at median follow-up of 53 months. CONCLUSION. Conditioning chemotherapy and 19-28z CAR T-cells were acceptably tolerated across investigated dose levels in heavily pretreated patients with R/R CLL and indolent B-NHL, and a subgroup of patients achieved durable CR. Ibrutinib therapy may modulate autologous T-cell phenotype. TRIAL REGISTRATION. ClinicalTrials.gov NCT00466531. FUNDING. Juno Therapeutics.
Mark B. Geyer, Isabelle Rivière, Brigitte Sénéchal, Xiuyan Wang, Yongzeng Wang, Terence J. Purdon, Meier Hsu, Sean M. Devlin, M. Lia Palomba, Elizabeth Halton, Yvette Bernal, Michel Sadelain, Jae H. Park, Renier J. Brentjens
Pancreatic ductal adenocarcinoma (PDA) is characterized by an activating mutation in KRAS. Direct inhibition of KRAS through pharmacological means remains a challenge; however, targeting key KRAS effectors has therapeutic potential. We investigated the contribution of TANK-binding kinase 1 (TBK1), a critical downstream effector of mutant active KRAS, to PDA progression. We report that TBK1 supports the growth and metastasis of KRAS-mutant PDA by driving an epithelial plasticity program in tumor cells that enhances invasive and metastatic capacity. Further, we identify that the receptor tyrosine kinase Axl induces TBK1 activity in a Ras-RalB-dependent manner. These findings demonstrate that TBK1 is central to an Axl-driven epithelial-mesenchymal transition in KRAS-mutant PDA and suggest that interruption of the Axl-TBK1 signaling cascade above or below KRAS has potential therapeutic efficacy in this recalcitrant disease.
Victoria H. Cruz, Emily N. Arner, Wenting Du, Alberto E. Bremauntz, Rolf A. Brekken
The prefrontal cortex controls food reward seeking and ingestion, playing important roles in directing attention, regulating motivation towards reward pursuit, and the assignment of reward salience and value. The cell types that mediate these behavioral functions, however, are not well described. We report here that optogenetic activation of vasoactive peptide expressing (VIP) interneurons in both the infralimbic (IL) and prelimbic (PL) divisions of the medial prefrontal cortex in mice is sufficient to reduce acute, binge-like intake of high calorie palatable food in the absence of any effect on low calorie rodent chow intake in the sated animal. In addition, we discovered that the behavioral mechanisms associated with these changes in feeding differed between animals that underwent either IL or PL VIPergic stimulation. While IL VIP neurons showed the ability to reduce palatable food intake, this effect was dependent upon the novelty and relative value of the food source. In addition, IL VIP neuron activation significantly reduced novel object- and novel social investigative behavior. Activation of PL VIP neurons, however, produced a reduction in high calorie palatable food intake that was independent of food novelty. Neither IL nor PL VIP excitation changed motivation to obtain food reward. Our data show how neurochemically-defined populations of cortical interneurons can regulate specific aspects of food reward-driven behavior, resulting in a selective reduction in intake of highly valued food.
Brandon A. Newmyer, Ciarra M. Whindleton, Peter M. Klein, Mark P. Beenhakker, Marieke K. Jones, Michael M. Scott
Dysregulation of the JAK/STAT signaling pathway is associated with Multiple Sclerosis (MS) and its mouse model, Experimental Autoimmune Encephalomyelitis (EAE). Suppressors Of Cytokine Signaling (SOCS) negatively regulate the JAK/STAT pathway. We previously reported a severe, brain-targeted, atypical form of EAE in mice lacking Socs3 in myeloid cells (Socs3ΔLysM), which is associated with cerebellar neutrophil infiltration. There is emerging evidence that neutrophils are detrimental in the pathology of MS/EAE, however, their exact function is unclear. Here we demonstrate that neutrophils from the cerebellum of Socs3ΔLysM mice show a hyper-activated phenotype with excessive production of reactive oxygen species (ROS) at the peak of EAE. Neutralization of ROS in vivo delayed the onset and reduced severity of atypical EAE. Mechanistically, Socs3-deficient neutrophils exhibit enhanced STAT3 activation, a hyper-activated phenotype in response to G-CSF, and upon G-CSF priming, increased ROS production. Neutralization of G-CSF in vivo significantly reduced the incidence and severity of the atypical EAE phenotype. Overall, our work elucidates that hypersensitivity of G-CSF/STAT3 signaling in Socs3ΔLysM mice leads to atypical EAE by enhanced neutrophil activation and increased oxidative stress, which may explain the detrimental role of G-CSF in MS patients.
Zhaoqi Yan, Wei Yang, Luke Parkitny, Sara A. Gibson, Kevin S. Lee, Forrest Collins, Jessy S. Deshane, Wayne Cheng, Amy S. Weinmann, Hairong Wei, Hongwei Qin, Etty N. Benveniste
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