Aberrant fibroblast function plays a key role in the pathogenesis of idiopathic pulmonary fibrosis, a devastating disease of unrelenting extracellular matrix deposition in response to lung injury. Platelet-derived growth factor α–positive (Pdgfra+) lipofibroblasts (LipoFBs) are essential for lung injury response and maintenance of a functional alveolar stem cell niche. Little is known about the effects of lung injury on LipoFB function. Here, we used single-cell RNA-Seq (scRNA-Seq) technology and PdgfraGFP lineage tracing to generate a transcriptomic profile of Pdgfra+ fibroblasts in normal and injured mouse lungs 14 days after bleomycin exposure, generating 11 unique transcriptomic clusters that segregated according to treatment. While normal and injured LipoFBs shared a common gene signature, injured LipoFBs acquired fibrogenic pathway activity with an attenuation of lipogenic pathways. In a 3D organoid model, injured Pdgfra+ fibroblast–supported organoids were morphologically distinct from those cultured with normal fibroblasts, and scRNA-Seq analysis suggested distinct transcriptomic changes in alveolar epithelia supported by injured Pdgfra+ fibroblasts. In summary, while LipoFBs in injured lung have not migrated from their niche and retain their lipogenic identity, they acquire a potentially reversible fibrogenic profile, which may alter the kinetics of epithelial regeneration and potentially contribute to dysregulated repair, leading to fibrosis.
Carol S. Trempus, Brian N. Papas, Maria I. Sifre, Carl D. Bortner, Erica Scappini, Charles J. Tucker, Xin Xu, Katina L. Johnson, Leesa J. Deterding, Jason G. Williams, Dylan J. Johnson, Jian-Liang Li, Deloris Sutton, Charan Ganta, Debabrata Mahapatra, Muhammad Arif, Abhishek Basu, Lenny Pommerolle, Resat Cinar, Anne K. Perl, Stavros Garantziotis
Although SARS-CoV-2 evolution seeds a continuous stream of antibody-evasive viral variants, COVID-19 mRNA vaccines provide robust protection against severe disease and hospitalization. Here, we asked whether mRNA vaccine–induced memory T cells limit lung SARS-CoV-2 replication and severe disease. We show that mice and humans receiving booster BioNTech mRNA vaccine developed potent CD8 T cell responses and showed similar kinetics of expansion and contraction of granzyme B/perforin-expressing effector CD8 T cells. Both monovalent and bivalent mRNA vaccines elicited strong expansion of a heterogeneous pool of terminal effectors and memory precursor effector CD8 T cells in spleen, inguinal and mediastinal lymph nodes, pulmonary vasculature, and most surprisingly in the airways, suggestive of systemic and regional surveillance. Furthermore, we document that: (a) CD8 T cell memory persists in multiple tissues for > 200 days; (b) following challenge with pathogenic SARS-CoV-2, circulating memory CD8 T cells rapidly extravasate to the lungs and promote expeditious viral clearance, by mechanisms that require CD4 T cell help; and (c) adoptively transferred splenic memory CD8 T cells traffic to the airways and promote lung SARS-CoV-2 clearance. These findings provide insights into the critical role of memory T cells in preventing severe lung disease following breakthrough infections with antibody-evasive SARS-CoV-2 variants.
Brock Kingstad-Bakke, Thomas Cleven, Hailey Bussan, Boyd L. Yount Jr., Ryuta Uraki, Kiyoko Iwatsuki-Horimoto, Michiko Koga, Shinya Yamamoto, Hiroshi Yotsuyanagi, Hongtae Park, Jay S. Mishra, Sathish Kumar, Ralph S. Baric, Peter J. Halfmann, Yoshihiro Kawaoka, M. Suresh
Malaria can quickly progress from an uncomplicated infection into a life-threatening severe disease. However, the unspecificity of early symptoms often makes it difficult to identify patients at high risk of developing severe disease. Additionally, one of the most feared malaria complications — cerebral malaria — is challenging to diagnose, often resulting in treatment delays that can lead to adverse outcomes. To identify candidate biomarkers for the prognosis and/or diagnosis of severe and cerebral malaria, we have analyzed the transcriptomic response of human brain microvascular endothelial cells to erythrocytes infected with Plasmodium falciparum. Candidates were validated in plasma samples from a cohort of pediatric patients with malaria from Mozambique, resulting in the identification of several markers with capacity to distinguish uncomplicated from severe malaria, the most potent being the metallopeptidase ADAMTS18. Two other biomarkers, Angiopoietin-like-4 and Inhibin-βE were able to differentiate children with cerebral malaria within the severe malaria group, showing increased sensitivity after combination in a biomarker signature. The validation of the predicted candidate biomarkers in plasma of children with severe and cerebral malaria underscores the power of this transcriptomic approach and indicates that a specific endothelial response to P. falciparum–infected erythrocytes is linked to the pathophysiology of severe malaria.
Cláudia Gomes, Rosauro Varo, Miquel Duran-Frigola, Antonio Sitoe, Rubão Bila, Sonia Machevo, Alfredo Mayor, Quique Bassat, Ana Rodriguez
Overactive fibroblast growth factor receptor 3 (FGFR3) signaling drives pathogenesis in a variety of cancers and a spectrum of short-limbed bone dysplasias, including the most common form of human dwarfism, achondroplasia (ACH). Targeting FGFR3 activity holds great promise as a therapeutic approach for treatment of these diseases. Here, we established a receptor/adaptor translocation assay system that can specifically monitor FGFR3 activation, and we applied it to identify FGFR3 modulators from complex natural mixtures. An FGFR3-suppressing plant extract of Amaranthus viridis was identified from the screen, and 2 bioactive porphyrins, pheophorbide a (Pa) and pyropheophorbide a, were sequentially isolated from the extract and functionally characterized. Further analysis showed that Pa reduced excessive FGFR3 signaling by decreasing its half-life in FGFR3-overactivated multiple myeloma cells and chondrocytes. In an ex vivo culture system, Pa alleviated defective long bone growth in humanized ACH mice (FGFR3ACH mice). Overall, our study presents an approach to discovery and validation of plant extracts or drug candidates that target FGFR3 activation. The compounds identified by this approach may have applications as therapeutics for FGFR3-associated cancers and skeletal dysplasias.
Yun-Wen Lin, Hsiao-Jung Kao, Wei-Ting Chen, Cheng-Fu Kao, Jer-Yuarn Wu, Yuan-Tsong Chen, Yi-Ching Lee
Approximately 30% of breast cancer survivors deemed free of disease will experience locoregional or metastatic recurrence even up to 30 years after initial diagnosis, yet how residual/dormant tumor cells escape immunity elicited by the primary tumor remains unclear. We demonstrate that intrinsically dormant tumor cells are indeed recognized and lysed by antigen-specific T cells in vitro and elicit robust immune responses in vivo. However, despite close proximity to CD8+ killer T cells, dormant tumor cells themselves support early accumulation of protective FoxP3+ T regulatory cells (Tregs), which can be targeted to reduce tumor burden. These intrinsically dormant tumor cells maintain a hybrid epithelial/mesenchymal state that is associated with immune dysfunction, and we find that the tumor-derived, stem cell/basal cell protein Dickkopf WNT signaling pathway inhibitor 3 (DKK3) is critical for Treg inhibition of CD8+ T cells. We also demonstrate that DKK3 promotes immune-mediated progression of proliferative tumors and is significantly associated with poor survival and immunosuppression in human breast cancers. Together, these findings reveal that latent tumors can use fundamental mechanisms of tolerance to alter the T cell microenvironment and subvert immune detection. Thus, targeting these pathways, such as DKK3, may help render dormant tumors susceptible to immunotherapies.
Timothy N. Trotter, Carina E. Dagotto, Delila Serra, Tao Wang, Xiao Yang, Chaitanya R. Acharya, Junping Wei, Gangjun Lei, H. Kim Lyerly, Zachary C. Hartman
In youths with obesity, the gut hormone potentiation of insulin secretion — the incretin effect — is blunted. We explored the longitudinal impact of the incretin effect during pubertal transition on β cell function and insulin sensitivity. Youths with obesity and 2-hour glucose level ≥ 120 mg/dL underwent a 3-hour oral glucose-tolerance test (OGTT) and an isoglycemic i.v. glucose infusion to quantify the incretin effect. After 2 years, 30 of 39 participants had a repeated OGTT and were stratified into 3 tertiles according to the baseline incretin effect. The high–incretin effect group demonstrated a longitudinal increase in β cell function (disposition index, minimal model [DIMM]), with greater insulin sensitivity at follow-up and stable insulin secretion (φtotal). A lower incretin effect at baseline was associated with higher 1-hour and 2-hour glucose level at follow-up. The high–incretin effect group displayed a greater increase of GLP-17–36 than the moderate- and low-incretin group at baseline, while such a difference did not persist after 2 years. Glucagon suppression was reduced at follow-up in those with low-baseline incretin in respect to the high-incretin group. The incretin effect during pubertal transition affected the longitudinal trajectory of β cell function and weight in youths with obesity.
Alfonso Galderisi, Domenico Tricò, Jessica Lat, Stephanie Samuels, Ram Weiss, Michelle Van Name, Bridget Pierpont, Nicola Santoro, Sonia Caprio
The development of human prenatal adaptive immunity progresses faster than previously appreciated, with the emergence of memory CD4+ T cells alongside regulatory T cells by midgestation. We previously identified a prenatal specific population of promyelocytic leukemia zinc finger–positive (PLZF+) CD4+ T cells with heightened effector potential that were enriched in the developing intestine and accumulated in the cord blood of infants exposed to prenatal inflammation. However, the signals that drive their tissue distribution and effector maturation are unknown. Here, we define the transcriptional and functional heterogeneity of human prenatal PLZF+CD4+ T cells and identify the compartmentalization of T helper–like (Th-like) effector function across the small intestine (SI) and mesenteric lymph nodes (MLNs). IL-7 was more abundant in the SI relative to the MLNs and drove the preferential expansion of naive PLZF+CD4+ T cells via enhanced STAT5 and MEK/ERK signaling. Exposure to IL-7 was sufficient to induce the acquisition of CD45RO expression and rapid effector function in a subset of PLZF+CD4+ T cells, identifying a human analog of memory phenotype CD4+ T cells. Further, IL-7 modulated the differentiation of Th1- and Th17-like PLZF+CD4+ T cells and thus likely contributes to the anatomic compartmentalization of human prenatal CD4+ T cell effector function.
Veronica Locher, Sara Park, Daniel G. Bunis, Stephanie Makredes, Margareta Mayer, Trevor D. Burt, Gabriela K. Fragiadakis, Joanna Halkias
MAD2L1BP-encoded p31comet mediates Trip13-dependent disassembly of Mad2- and Rev7-containing complexes and, through this antagonism, promotes timely spindle assembly checkpoint (SAC) silencing, faithful chromosome segregation, insulin signaling, and homology-directed repair (HDR) of DNA double-strand breaks. We identified a homozygous MAD2L1BP nonsense variant, R253*, in 2 siblings with microcephaly, epileptic encephalopathy, and juvenile granulosa cell tumors of ovary and testis. Patient-derived cells exhibited high-grade mosaic variegated aneuploidy, slowed-down proliferation, and instability of truncated p31comet mRNA and protein. Corresponding recombinant p31comet was defective in Trip13, Mad2, and Rev7 binding and unable to support SAC silencing or HDR. Furthermore, C-terminal truncation abrogated an identified interaction of p31comet with tp53. Another homozygous truncation, R227*, detected in an early-deceased patient with low-level aneuploidy, severe epileptic encephalopathy, and frequent blood glucose elevations, likely corresponds to complete loss of function, as in Mad2l1bp–/– mice. Thus, human mutations of p31comet are linked to aneuploidy and tumor predisposition.
Ghada M. H. Abdel-Salam, Susanne Hellmuth, Elise Gradhand, Stephan Käseberg, Jennifer Winter, Ann-Sophie Pabst, Maha M. Eid, Holger Thiele, Peter Nürnberg, Birgit S. Budde, Mohammad Reza Toliat, Ines B. Brecht, Christopher Schroeder, Axel Gschwind, Stephan Ossowski, Friederike Häuser, Heidi Rossmann, Mohamed S. Abdel-Hamid, Ibrahim Hegazy, Ahmed G. Mohamed, Dominik T. Schneider, Aida Bertoli-Avella, Peter Bauer, Jillian N. Pearring, Rolph Pfundt, Alexander Hoischen, Christian Gilissen, Dennis Strand, Ulrich Zechner, Soha A. Tashkandi, Eissa A. Faqeih, Olaf Stemmann, Susanne Strand, Hanno J. Bolz
Postictal apnea is thought to be a major cause of sudden unexpected death in epilepsy (SUDEP). However, the mechanisms underlying postictal apnea are unknown. To understand causes of postictal apnea, we used a multimodal approach to study brain mechanisms of breathing control in 20 patients (ranging from pediatric to adult) undergoing intracranial electroencephalography for intractable epilepsy. Our results indicate that amygdala seizures can cause postictal apnea. Moreover, we identified a distinct region within the amygdala where electrical stimulation was sufficient to reproduce prolonged breathing loss persisting well beyond the end of stimulation. The persistent apnea was resistant to rising CO2 levels, and air hunger failed to occur, suggesting impaired CO2 chemosensitivity. Using es-fMRI, a potentially novel approach combining electrical stimulation with functional MRI, we found that amygdala stimulation altered blood oxygen level–dependent (BOLD) activity in the pons/medulla and ventral insula. Together, these findings suggest that seizure activity in a focal subregion of the amygdala is sufficient to suppress breathing and air hunger for prolonged periods of time in the postictal period, likely via brainstem and insula sites involved in chemosensation and interoception. They further provide insights into SUDEP, may help identify those at greatest risk, and may lead to treatments to prevent SUDEP.
Gail I.S. Harmata, Ariane E. Rhone, Christopher K. Kovach, Sukhbinder Kumar, Md Rakibul Mowla, Rup K. Sainju, Yasunori Nagahama, Hiroyuki Oya, Brian K. Gehlbach, Michael A. Ciliberto, Rashmi N. Mueller, Hiroto Kawasaki, Kyle T.S. Pattinson, Kristina Simonyan, Paul W. Davenport, Matthew A. Howard III, Mitchell Steinschneider, Aubrey C. Chan, George B. Richerson, John A. Wemmie, Brian J. Dlouhy
Ventricular arrhythmias (VAs) in heart failure are enhanced by sympathoexcitation. However, radiotracer studies of catecholamine uptake in failing human hearts demonstrate a proclivity for VAs in patients with reduced cardiac sympathetic innervation. We hypothesized that this counterintuitive finding is explained by heterogeneous loss of sympathetic nerves in the failing heart. In a murine model of dilated cardiomyopathy (DCM), delayed PET imaging of sympathetic nerve density using the catecholamine analog [11C]meta-Hydroxyephedrine demonstrated global hypoinnervation in ventricular myocardium. Although reduced, sympathetic innervation in 2 distinct DCM models invariably exhibited transmural (epicardial to endocardial) gradients, with the endocardium being devoid of sympathetic nerve fibers versus controls. Further, the severity of transmural innervation gradients was correlated with VAs. Transmural innervation gradients were also identified in human left ventricular free wall samples from DCM versus controls. We investigated mechanisms underlying this relationship by in silico studies in 1D, 2D, and 3D models of failing and normal human hearts, finding that arrhythmogenesis increased as heterogeneity in sympathetic innervation worsened. Specifically, both DCM-induced myocyte electrical remodeling and spatially inhomogeneous innervation gradients synergistically worsened arrhythmogenesis. Thus, heterogeneous innervation gradients in DCM promoted arrhythmogenesis. Restoration of homogeneous sympathetic innervation in the failing heart may reduce VAs.
Al-Hassan J. Dajani, Michael B. Liu, Michael A. Olaopa, Lucian Cao, Carla Valenzuela-Ripoll, Timothy J. Davis, Megan D. Poston, Elizabeth H. Smith, Jaime Contreras, Marissa Pennino, Christopher M. Waldmann, Donald B. Hoover, Jason T. Lee, Patrick Y. Jay, Ali Javaheri, Roger Slavik, Zhilin Qu, Olujimi A. Ajijola
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