Severe lung damage in COVID-19 involves complex interactions between diverse populations of immune and stromal cells. In this study, we used a spatial transcriptomics approach to delineate the cells, pathways and genes present across the spectrum of histopathological damage in COVID-19 lung tissue. We applied correlation network-based approaches to deconvolve gene expression data from 46 areas of interest covering >62,000 cells within well preserved lung samples from three patients. Despite substantial inter-patient heterogeneity, we discovered evidence for a common immune cell signaling circuit in areas of severe tissue that involves crosstalk between cytotoxic lymphocytes and pro-inflammatory macrophages. Expression of IFNG by cytotoxic lymphocytes was associated with induction of chemokines including CXCL9, CXCL10 and CXCL11 which are known to promote the recruitment of CXCR3+ immune cells. The tumour necrosis factor (TNF) superfamily members BAFF (TNFSF13B) and TRAIL (TNFSF10) were found to be consistently upregulated in the areas with severe tissue damage. We used published spatial and single cell SARS-CoV-2 datasets to confirm our findings in the lung tissue from additional cohorts of COVID-19 patients. The resulting model of severe COVID-19 immune-mediated tissue pathology may inform future therapeutic strategies.
Amy R. Cross, Carlos E. de Andrea, María Villalba-Esparza, Manuel F. Landecho, Lucia Cerundolo, Praveen Weeratunga, Rachel E. Etherington, Laura Denney, Graham Ogg, Ling-Pei Ho, Ian S.D. Roberts, Joanna Hester, Paul Klenerman, Ignacio Melero, Stephen N. Sansom, Fadi Issa
Vascular smooth muscle cells (vSMC) exert a critical role in sensing and maintaining vascular integrity. These cells abundantly express the low-density lipoprotein receptor-related protein 1 (LRP1), a large endocytic and signaling receptor that recognizes numerous ligands including ApoE-rich lipoproteins, proteases, and protease-inhibitor complexes. We observed the spontaneous formation of aneurysms in the superior mesenteric artery (SMA) of both male and female mice in which LRP1 was genetically deleted in v SMC (smLRP1-/- mice). Quantitative proteomics revealed elevated abundance of several proteins in smLRP1-/- mice that are known to be induced by angiotensin II (AngII)-mediated signaling, suggesting that this pathway is dysregulated. Administration of losartan, an AngII type I receptor antagonist, or an angiotensinogen antisense oligonucleotide to reduce plasma angiotensinogen concentrations restored the normal SMA phenotype in smLRP1-/- mice and prevented aneurysm formation. Additionally, employing a vascular injury model, we noted excessive vascular remodeling and neointima formation in smLRP1-/- mice that was restored by losartan administration. Together, these findings reveal that LRP1 regulates vascular integrity and remodeling of the SMA by attenuating excessive AngII-mediated signaling.
Jackie M. Zhang, Dianaly T. Au, Hisashi Sawada, Michael K. Franklin, Jessica J. Moorleghen, Deborah A. Howatt, Pengjun Wang, Brittany O. Aicher, Brian Hampton, Mary Migliorini, Fenge Ni, Adam E. Mullick, Mashhood M. Wani, Areck A. Ucuzian, Hong S. Lu, Selen C. Muratoglu, Alan Daugherty, Dudley K. Strickland
Hepatocellular carcinoma (HCC) is the most common lethal form of liver cancer. Apart from surgical removal and transplantation, other treatments have not yet been well established for patients with HCC. Herein, we found that carboxylesterase 1 (CES1) was expressed at various levels in HCC. We further revealed that blockage of CES1 by pharmacological and genetical approaches leads to altered lipid profiles that are directly linked to impaired mitochondrial function. Mechanistically, LC-MS/MS and lipidomic analyses revealed that lipid signaling molecules, including polyunsaturated fatty acids (PUFAs), which activate PPARα/γ, were dramatically reduced upon CES1 inhibition. As a result, SCD, a PPARα/γ target gene involved in tumor progression and chemoresistance, was significantly downregulated. Consistently, clinical analysis demonstrated a strong correlation between the protein levels of CES1 and SCD in HCC. Interference with lipid signaling by targeting the “CES1-PPARα/γ-SCD” axis sensitized HCC cells to cisplatin treatment. As a demonstration, the growth of HCC xenograft tumors in NU/J mice was potently limited by co-administration of cisplatin and CES1 inhibition. Our results suggest that CES1 is a promising therapeutic target for HCC treatment.
Gang Li, Xin Li, Iqbal Mahmud, Jazmin Ysaguirre, Baharan Fekry, Shuyue Wang, Bo Wei, Kristin L. Eckel-Mahan, Philip L. Lorenzi, Richard Lehner, Kai Sun
Podocyte injury and loss are key drivers of primary and secondary glomerular diseases, such as focal segmental glomerulosclerosis (FSGS) and diabetic kidney disease (DKD). We previously demonstrated the renoprotective role of protein S (PS) and its cognate tyrosine-protein kinase receptor, TYRO3, in models of FSGS and DKD and that their signaling exerts anti-apoptotic and anti-inflammatory effects to confer protection against podocyte loss. Among the three TAM receptors (TYRO3, AXL, and MER), only TYRO3 expression is largely restricted to podocytes, and glomerular TYRO3 mRNA expression negatively correlates with human glomerular disease progression. We, therefore, posited that the agonism PS-TYRO3 signaling could serve as a potential therapeutic approach to attenuate glomerular disease progression. As PS function is not limited to TYRO3-mediated signal transduction but includes its anticoagulant activity, we focused on the development of TYRO3 agonist as an optimal therapeutic approach to glomerular disease. Among the small molecule TYRO3 agonist compounds screened, compound-10 (C-10) showed a select activation of TYRO3 without any effects on AXL or MER. We also confirmed that C-10 directly binds to TYRO3, but not the other receptors. In vivo, C-10 attenuated proteinuria, glomerular injury, and podocyte loss in mouse models of adriamycin-induced nephropathy and db/db model of type 2 diabetes. Moreover, these renoprotective effects of C-10 are lost in Tyro3 knockout mice, indicating that C-10 is a select agonist of TYRO3 activity that mitigates podocyte injury and glomerular disease. Therefore, C-10, a novel TYRO3 agonist, could be potentially developed as a new therapy for glomerular disease.
Fang Zhong, Hong Cai, Jia Fu, Zeguo Sun, Zhengzhe Li, David Bauman, Lois Wang, Bhaskar Das, Kyung Lee, John He
Dysfunction of alveolar epithelial type 2 cells (AEC2s), the facultative progenitors of lung alveoli, is implicated in pulmonary disease pathogenesis, highlighting the importance of human in vitro models. However, AEC2-like cells in culture have yet to be directly compared to their in vivo counterparts at single cell resolution. Here, we perform head-to-head comparisons between the transcriptomes of fresh primary (1o) adult human AEC2s, their cultured progeny, and human induced pluripotent stem cell-derived AEC2s (iAEC2s). We find each population occupies a distinct transcriptomic space with cultured AEC2s (1o and iAEC2s) exhibiting similarities to and differences from freshly purified 1o cells. Across each cell type, we find an inverse relationship between proliferative and maturation states, with pre-culture 1o AEC2s being most quiescent/mature and iAEC2s being most proliferative/least mature. Cultures of either type of human AEC2 do not generate detectable alveolar type 1 cells in these defined conditions; however, a subset of iAEC2s co-cultured with fibroblasts acquires a “transitional cell state” described in mice and humans to arise during fibrosis or following injury. Hence, we provide direct comparisons of the transcriptomic programs of 1o and engineered AEC2s, two in vitro models that can be harnessed to study human lung health and disease.
Konstantinos-Dionysios Alysandratos, Carolina Garcia-de-Alba, Changfu Yao, Patrizia Pessina, Jessie Huang, Carlos Villacorta-Martin, Olivia T. Hix, Kasey Minakin, Claire L. Burgess, Pushpinder Bawa, Aditi Murthy, Bindu Konda, Michael F. Beers, Barry R. Stripp, Carla F. Kim, Darrell N. Kotton
Understanding persistence and evolution of B cell clones after COVID-19 infection and vaccination is crucial for predicting responses against emerging viral variants and optimizing vaccines. Here, we collected longitudinal samples from severe COVID-19 patients every third to seventh day during hospitalization and every third month after recovery. We profiled the antigen-specific immune cell dynamics by combining single cell RNA-Seq, Cellular Indexing of Transcriptomes and Epitopes by Sequencing (CITE)-Seq, B cell receptor (BCR)-Seq with oligo-tagged antigen baits. While the proportion of Spike Receptor Binding Domain-specific memory B cells (MBC) increased from 3 months after infection, the other Spike- and Nucleocapsid-specific B cells remained constant. All patients showed ongoing class switching and sustained affinity maturation of antigen specific cells, which was not significantly increased early after vaccine. B cell analysis revealed a polyclonal response with limited clonal expansion; nevertheless, some clones detected during hospitalization, as plasmablasts, persisted for up to one year, as MBC. Monoclonal antibodies derived from persistent B cell families increased their binding and neutralization breadth and started recognizing viral variants by 3 months after infection. Overall, our findings provide important insights into the clonal evolution and dynamics of antigen specific B cell responses in longitudinally sampled COVID-19 infected patients.
Lydia Scharf, Hannes Axelsson, Aikaterini Emmanouilidi, Nimitha R. Mathew, Daniel J. Sheward, Susannah Leach, Pauline Isakson, Ilya V. Smirnov, Emelie Marklund, Nicolae Miron, Lars-Magnus Andersson, Magnus Gisslén, Ben Murrell, Anna Lundgren, Mats Bemark, Davide Angeletti
FOXD1+ derived stromal cells give rise to pericytes and fibroblasts that support the kidney vasculature and interstitium but are also major precursors of myofibroblasts. ZEB2 is a SMAD-interacting transcription factor that is expressed in developing kidney stromal progenitors. Here we show that Zeb2 is essential for normal FOXD1+ stromal progenitor development. Specific deletion of mouse Zeb2 in FOXD1+ stromal progenitors (Zeb2 cKO) leads to abnormal interstitial stromal cell development, differentiation, and kidney fibrosis. Immunofluorescent staining analyses revealed abnormal expression of interstitial stromal cell markers MEIS1/2/3, CDKN1C, and CSPG4 (NG2) in newborn and 3-week-old Zeb2 cKO mouse kidneys. Zeb2 deficient FOXD1+ stromal progenitors also took on a myofibroblast fate that led to kidney fibrosis and kidney failure. Cell marker studies further confirmed that these myofibroblasts expressed pericyte and resident fibroblast markers including PDGFRβ, CSPG4, Desmin, GLI1, and NT5E. Notably, increased interstitial collagen deposition associated with loss of Zeb2 in FOXD1+ stromal progenitors was accompanied by increased expression of activated SMAD1/5/8, SMAD2/3, SMAD4, and AXIN2. Thus, our study identifies a key role of ZEB2 in maintaining the cell fate of FOXD1+ stromal progenitors during kidney development whereas loss of ZEB2 leads to differentiation of FOXD1+ stromal progenitors into myofibroblasts and kidney fibrosis.
Sudhir Kumar, Xueping Fan, Hila Milo Rasouly, Richa Sharma, David J. Salant, Weining Lu
Autosomal dominant polycystic kidney disease (ADPKD), the most common monogenic nephropathy, is characterized by phenotypic variability exceeding genic effects. Dysregulated metabolism and immune cell function are key disease modifiers. The tryptophan metabolites, kynurenines, produced through IDO1, are known immunomodulators. Here, we study the role of tryptophan metabolism in PKD using an orthologous disease model (C57Bl/6J Pkd1RC/RC). We found elevated kynurenine and IDO1 levels in Pkd1RC/RC kidneys versus wildtype. Further, IDO1 levels were increased in ADPKD cell lines. Genetic Ido1 loss in Pkd1RC/RC animals resulted in reduced PKD severity as measured by %kidney weight/body weight and cystic index. Consistent with an immunomodulatory role of kynurenines, Pkd1RC/RC;Ido1-/- mice presented with significant changes in the cystic immune microenvironment (CME) versus controls. Kidney macrophage numbers decreased and CD8+ T cell numbers increased, both known PKD modulators. Also, pharmacological IDO1 inhibition in Pkd1RC/RC mice and kidney specific Pkd2 knockout mice with rapidly progressive PKD resulted in less severe PKD versus controls with similar changes in the CME as in the genetic model. Our data suggest that tryptophan metabolism is dysregulated in ADPKD and that its inhibition results in changes to the CME and slows disease progression, making IDO1 a novel therapeutic target for ADPKD.
Dustin T. Nguyen, Emily K. Kleczko, Nidhi Dwivedi, Marie-Louise T. Monaghan, Berenice Y. Gitomer, Michel B. Chonchol, Eric T. Clambey, Raphael A. Nemenoff, Jelena Klawitter, Katharina Hopp
Mitochondria are dynamic organelles responsible for energy production and many processes central to cellular function. Alterations in mitochondrial function is associated with human fibrotic lung diseases, including idiopathic pulmonary fibrosis (IPF). Pulmonary fibrosis is characterized by stiffening of the extracellular matrix (ECM). Fibroblasts migrate in the direction of greater stiffness, a phenomenon termed durotaxis. The mechanically guided fibroblast migration could be a crucial step in the progression of lung fibrosis. In this study, we identified mitochondria as an important mechanotransducer at the intersection between extracellular mechanical signals and durotactic lung fibroblast migration. Primary human lung fibroblasts sense increasing matrix stiffness with a change of mitochondrial dynamics in favor of mitochondrial fission and increased production of ATP. Mitochondria polarize in the direction of a physiologically relevant stiffness gradient, with conspicuous localization to the leading edge, primarily lamellipodia and filopodia, of migrating lung fibroblasts. Matrix stiffness-regulated mitochondrial fission and durotactic lung fibroblast migration are mediated by a DRP1/MFF-dependent pathway. Importantly, we found that the DRP1/MFF pathway is activated in fibrotic lung myofibroblasts in both human IPF and bleomycin-induced mouse lung fibrosis. Our findings suggest that energy-producing mitochondria need to be sectioned via fission and repositioned in durotactic lung fibroblasts to meet the higher energy demand. This represents a new mechanism through which mitochondria may contribute to the progression of fibrotic lung diseases. Inhibition of durotactic migration of lung fibroblasts may play an important role in preventing the progression of IPF.
Ting Guo, Chun-sun Jiang, Shan-Zhong Yang, Yi Zhu, Chao He, A. Brent Carter, Veena B. Antony, Hong Peng, Yong Zhou
Although glycogen synthase kinase β (Gsk3β) has been shown to regulate tissue inflammation, whether and how it regulates inflammation resolution vs. inflammation activation is unclear. In a murine liver partial warm ischemia/reperfusion injury (IRI) model, we found that Gsk3β inhibitory phosphorylation increased at both the early activation and late resolution stages of the disease. Myeloid Gsk3β deficiency not only alleviated liver injuries, but also facilitated the restoration of liver homeostasis. Depletion of Kupffer cells (KCs) prior to the onset of liver ischemia diminished the differences between the WT and Gsk3β KO mice in the activation of liver IRI. However, the resolution of liver IRI remained accelerated in the Gsk3β KO mice. In CD11b-DTR mice, Gsk3β deficient bone marrow-derived macrophages (BMMs) facilitated the resolution of liver IRI as compared with WT cells. Furthermore, Gsk3β deficiency promoted the reparative phenotype differentiation in vivo in liver infiltrating macrophages and in vitro in BMMs. Gsk3 pharmacological inhibition promoted the resolution of liver IRI in WT, but not myeloid MerTK deficient, mice. Thus, Gsk3β regulates liver IRI at both activation and resolution stages of the disease. Gsk3 inactivation enhances the pro-resolving function of liver infiltrating macrophages in MerTK–dependent manner.
Hanwen Zhang, Ming Ni, Han Wang, Jing Zhang, Dan Jin, Ronald W. Busuttil, Jerzy W. Kupiec-Weglinski, Wei Li, Xuehao Wang, Yuan Zhai
No posts were found with this tag.