Concise Communication

Abstract

Cutaneous leishmaniasis (CL) is caused by Leishmania donovani in Sri Lanka. Pentavalent antimonials (e.g. sodium stibogluconate; SSG) remain first line drugs for CL with no new effective treatments emerging. We studied whole blood and lesion transcriptomes from Sri Lankan CL patients at presentation and during SSG treatment. From lesions but not whole blood, we identified differential expression of immune-related genes, including immune checkpoint molecules, after onset of treatment. Using spatial profiling and RNA-FISH, we confirmed reduced expression of PD-L1 and IDO1 proteins on treatment in lesions of a second validation cohort and further demonstrated significantly higher expression of these checkpoint molecules on parasite-infected compared to non-infected lesional CD68+ monocytes / macrophages. Crucially, early reduction in PD-L1 but not IDO1 expression was predictive of rate of clinical cure (HR = 4.88) and occurred in parallel with reduction in parasite load. Our data support a model whereby the initial anti-leishmanial activity of antimonial drugs alleviates checkpoint inhibition on T cells, facilitating immune-drug synergism and clinical cure. Our findings demonstrate that PD-L1 expression can be used as predictor of rapidity of clinical response to SSG treatment in Sri Lanka and support further evaluation of PD-L1 as a host directed therapy target in leishmaniasis.

Authors

Nidhi S. Dey, Sujai Senaratne, Vijani Somaratne, Nayani P. Madarasinghe, Bimalka Seneviratne, Sarah Forrester, Marcela Montes de Oca, Luiza Campos Reis, Srija Moulik, Pegine B. Walrad, Mitali Chatterjee, Hiro Goto, Renu Wickremasinghe, Dimitris Lagos, Paul M. Kaye, Shalindra Ranasinghe

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Abstract

The cardiac conduction system (CCS) ensures regular contractile function, and injury to any of its components can cause cardiac dysrhythmia. Although all cardiomyocytes (CMs) originate from common progenitors, the CCS is composed of biologically distinct cell types with unique functional and developmental characteristics. In contrast to ventricular cardiomyocytes, which continue to proliferate after birth, most CCS cells terminally exit the cell cycle during fetal development. Although the CCS should thus provide a poor substrate for postnatal injury repair, its regenerative capacity remains untested. Here, we describe a genetic system for ablating CMs that reside within the atrioventricular conduction system (AVCS). Adult mouse AVCS ablation resulted in regenerative failure characterized by persistent atrioventricular conduction defects and contractile dysfunction. In contrast, AVCS injury in neonatal mice led to recovery in a subset of these mice, thus providing evidence for CCS plasticity. Furthermore, CM proliferation did not appear to completely account for the observed functional recovery, suggesting that mechanisms regulating recovery from dysrhythmia are likely to be distinct from cardiac regeneration associated with ventricular injury. Taken together, we anticipate that our results will motivate further mechanistic studies of CCS plasticity and enable the exploration of rhythm restoration as an alternative therapeutic strategy.

Authors

Lin Wang, Minoti Bhakta, Antonio Fernandez-Perez, Nikhil V. Munshi

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Abstract

In this study, we demonstrate that Forkhead Box F1 (FOXF1), a mesenchymal transcriptional factor essential for lung development, is retained in a topographically distinct mesenchymal stromal cell population along the bronchovascular space in an adult lung and identify this distinct subset of collagen-expressing cells as a key player in lung allograft remodeling and fibrosis. Utilizing Foxf1_tdTomato BAC (Foxf1_tdTomato) and Foxf1_tdTomato;Col1a1_GFP mice, we show that Lin-Foxf1+ cells encompass the Sca1+CD34+ subset of collagen I-expressing mesenchymal cells (MCs) with capacity to generate colony forming units and lung epithelial organoids. Histologically, Foxf1-expressing MCs formed a three-dimensional network along the conducting airways; FOXF1 was noted to be conspicuously absent in MCs in the alveolar compartment. Bulk and single-cell RNA sequencing confirmed distinct transcriptional signatures of Foxf1pos/neg MCs, with Foxf1-expressing cells delineated by their high Gli1 and low Integrin α8 expression, from other collagen-expressing MCs. Foxf1+Gli1+ MCs demonstrated proximity to Sonic hedgehog (Shh) expressing bronchial epithelium, and mesenchymal Foxf1/Gli1 expression was found to be dependent on the paracrine Shh signaling in epithelial organoids. Utilizing a murine lung transplant model, we show dysregulation of the epithelial mesenchymal Shh/Gli1/Foxf1 crosstalk and expansion of this specific peri-bronchial MC population in chronically rejecting fibrotic lung allografts.

Authors

Russell R. Braeuer, Natalie M. Walker, Keizo Misumi, Serina Mazzoni-Putman, Yoshiro Aoki, Ruohan Liao, Ragini Vittal, Gabriel G. Kleer, David S. Wheeler, Jonathan Z. Sexton, Carol F. Farver, Joshua D. Welch, Vibha N. Lama

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Abstract

The heterogeneity of human hematopoietic stem (HSCs) and progenitor cells (HPCs) under stress conditions such as ex vivo expansion is poorly understood. Here we report that the frequencies of SCID repopulating cells (SRCs) were greatly decreased in cord blood(CB)CD34+ HSCs and HPCs upon ex vivo culture. Transcriptome analysis and metabolic profiling demonstrated that mitochondrial oxidative stress of human CB HSCs and HPCs notably increased along with loss of stemness. Limiting dilution analysis (LDA) revealed that functional human HSCs were enriched in cell populations with low levels of mitochondrial reactive oxygen species (mitoROS) during ex vivo culture. Using single cell RNA sequencing (scRNA-seq) analysis of mitoROS low cell population, we demonstrated that functional HSCs were substantially enriched in the Adhesion G protein-coupled receptor G1 positive (ADGRG1+) population of CD34+CD133+ CB cells upon ex vivo expansion stress. GSEA analysis revealed that HSC signature genes including MSI2 and MLLT3 are enriched in CD34+CD133+ ADGRG1+ CB HSCs. Our study reveals that ADGRG1 enriches for functional human HSCs under oxidative stress during ex vivo culture, which can be a reliable target for drug screening of agonists of HSC expansion.

Authors

Yandan Chen, Shuyi Fang, Qingwei Ding, Rongzhen Jiang, Jiefeng He, Qin Wang, Yuting Jin, Xinxin Huang, Sheng Liu, Maegan Capitano, Thao Trinh, Yincheng Teng, Qingyou Meng, Jun Wan, Hal Broxmeyer, Bin Guo

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Abstract

Dementia resulting from small vessel diseases of the brain (SVDs) is an emerging epidemic for which there is no treatment. Hypertension is the major risk factor for SVDs, but how hypertension damages the brain microcirculation is unclear. Here, we show that chronic hypertension in a mouse model progressively disrupts on-demand delivery of blood to metabolically active areas of the brain (functional hyperemia) through diminished activity of the capillary endothelial cell inward-rectifier potassium channel, Kir2.1. Despite similar efficacy in reducing blood pressure, amlodipine, a voltage-dependent calcium-channel blocker, prevented hypertension-related damage to functional hyperemia whereas losartan, an angiotensin II type-1 receptor blocker, did not. We attribute this drug class effect to losartan-induced ‘aldosterone breakthrough’, a phenomenon triggered by pharmacological interruption of the renin-angiotensin pathway leading to elevated plasma aldosterone levels. This hypothesis is supported by the finding that combining losartan with the aldosterone receptor antagonist eplerenone prevented the hypertension-related decline in functional hyperemia. Collectively, these data suggest Kir2.1 as a possible therapeutic target in vascular dementia and indicate that concurrent mineralocorticoid aldosterone receptor blockade may aid in protecting against late-life cognitive decline in hypertensive patients treated with angiotensin II type-1 receptor blockers.

Authors

Masayo Koide, Osama F. Harraz, Fabrice Dabertrand, Thomas A. Longden, Hannah R. Ferris, George C. Wellman, David C. Hill-Eubanks, Adam S. Greenstein, Mark Nelson

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Abstract

Natural killer (NK) cell suppression of T cells is a key determinant of viral pathogenesis and vaccine efficacy. This process involves perforin-dependent elimination of activated CD4 T cells during the first three days of infection. Although this mechanism requires cell-cell contact, NK cells and T cells typically reside in different compartments of lymphoid tissues at steady state. Here, we showed that NK-cell suppression of T cells is associated with transient accumulation of NK cells within T cell-rich sites of the spleen during lymphocytic choriomeningitis virus infection. The chemokine receptor CXCR3 was required for this relocation and suppression of antiviral T cells. Accordingly, NK-cell migration was mediated by type I interferon (IFN)-dependent promotion of CXCR3 ligand expression. In contrast, adenoviral vectors that weakly induced type I IFN and did not stimulate NK-cell inhibition of T cells also did not promote measurable redistribution of NK cells to T-cell zones. Exogenous IFN rescued NK-cell migration during adenoviral vector immunization. Thus, type I IFN and CXCR3 were critical for properly positioning NK cells to constrain antiviral T-cell responses. Development of strategies to curtail migration of NK cells between lymphoid compartments may enhance vaccine-elicited immune responses.

Authors

Ayad Ali, Laura M. Canaday, H. Alex Feldman, Hilal Cevik, Michael T. Moran, Sanjeeth Rajaram, Nora Lakes, Jasmine A. Tuazon, Harsha Seelamneni, Durga Krishnamurthy, Eryn Blass, Dan H. Barouch, Stephen N. Waggoner

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Abstract

Intratumoral T cells that might otherwise control tumors are often identified in an ‘exhausted’ state, defined by specific epigenetic modifications and upregulation of genes such as CD38, CTLA-4 and PD-1. While the term might imply inactivity, there has been little study of this state at the phenotypic level in tumors to understand the extent of their incapacitation. Starting with the observation that T cells move more quickly through mouse tumors as residence time increases and they progress towards exhaustion, we elaborated a non-stimulatory live-biopsy method for real-time study of T cell behaviors within individual patient tumors. Using two-photon microscopy, we studied native CD8 T cells interacting with APCs and cancer cells in different micro-niches of human tumors, finding that T cell speed was variable by region and by patient and was inversely correlated with local tumor density. Across a range of tumor types, we found a strong relationship between CD8 T cell motility and exhausted T cell state that corresponds to observations made in mouse models where exhausted T cells move faster. Our study demonstrates T cell dynamic states in individual human tumors and supports the existence of an active program in ‘exhausted’ T cells that extends beyond incapacitating them.

Authors

Ran You, Jordan Artichoker, Adam Fries, Austin W. Edwards, Alexis J. Combes, Gabriella C. Reeder, Bushra Samad, Matthew F. Krummel

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Abstract

A recent report found that rare predicted loss-of-function (pLOF) variants across 13 candidate genes in TLR3- and IRF7-dependent type I IFN pathways explain up to 3.5% of severe COVID-19 cases. We performed whole-exome or whole-genome sequencing of 1,934 COVID-19 cases (713 with severe and 1,221 with mild disease) and 15,251 ancestry-matched population controls across four independent COVID-19 biobanks. We then tested if rare pLOF variants in these 13 genes were associated with severe COVID-19. We identified only one rare pLOF mutation across these genes amongst 713 cases with severe COVID-19 and observed no enrichment of pLOFs in severe cases compared to population controls or mild COVID-19 cases. We find no evidence of association of rare loss-of-function variants in the proposed 13 candidate genes with severe COVID-19 outcomes.

Authors

Gundula Povysil, Guillaume Butler-Laporte, Ning Shang, Chen Wang, Atlas Khan, Manal Alaamery, Tomoko Nakanishi, Sirui Zhou, Vincenzo Forgetta, Robert J. M. Eveleigh, Mathieu Bourgey, Naveed Aziz, Steven J.M. Jones, Bartha Knoppers, Stephen W. Scherer, Lisa J. Strug, Pierre Lepage, Jiannis Ragoussis, Guillaume Bourque, Jahad Alghamdi, Nora Aljawini, Nour Albesher, Hani M. Al-Afghani, Bader Alghamdi, Mansour S. Almutair, Ebrahim Sabri Mahmoud, Leen Abu-Safieh, Hadeel El Bardisy, Fawz S. Al Harthi, Abdulraheem Alshareef, Bandar Ali Suliman, Saleh A. Alqahtani, Abdulaziz Almalik, May M. Alrashed, Salam Massadeh, Vincent Mooser, Mark Lathrop, Mohamed Fawzy, Yaseen M. Arabi, Hamdi Mbarek, Chadi Saad, Wadha Al-Muftah, Junghyun Jung, Serghei Mangul, Radja Badji, Asma Al Thani, Said I. Ismail, Ali G. Gharavi, Malak S. Abedalthagafi, J Brent Richards, David B. Goldstein, Krzysztof Kiryluk

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Abstract

Autoantibodies to interferon(IFN)-α and IFN-ω (type I IFNs) were recently reported as causative for severe COVID-19 in the general population. Autoantibodies against IFN-α and IFN-ω are present in almost all patients with Autoimmune-Polyendocrine-Syndrome Type 1 (APS-1) caused by biallelic deleterious or heterozygous dominant mutations in AIRE. We therefore hypothesized that autoantibodies against type I IFNs also predispose patients with APS-1 to severe COVID-19. We prospectively studied six patients with APS-1 between April 1st, 2020 and April 1st, 2021. Biobanked pre-COVID-19 sera of APS-1 subjects were tested for neutralizing autoantibodies to IFN-αand IFN-ω. The patients ́ sera ability to block recombinant human IFN-α and IFN-ω was assessed by assays quantifying phosphorylation of signal transducer and activator of transcription 1 (STAT1) as well as infection-based IFN-neutralization assays. We describe four patients with APS-1 and pre-existing high titers of neutralizing autoantibodies to IFN-α and IFN-ω who contracted SARS-CoV-2, yet developed only mild symptoms of COVID-19. None of the patients developed dyspnoea, oxygen requirement or high temperature. All infected patients with APS-1 shared female sex and age younger than 26 years. Clinical penetrance of neutralizing autoantibodies against type I IFNs for severe COVID-19 is not complete.

Authors

Christian Meisel, Bengisu Akbil, Tim Meyer, Erwin Lankes, Victor M. Corman, Olga Staudacher, Nadine Unterwalder, Uwe Kölsch, Christian Drosten, Marcus A. Mall, Tilmann Kallinich, Dirk Schnabel, Christine Goffinet, Horst von Bernuth

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Abstract

The four serotypes of dengue virus (DENV1-4) are mosquito-borne flaviviruses that infect humans. Live attenuated tetravalent DENV vaccines are at different phases of clinical testing. DENV vaccine developers have relied on neutralizing antibodies (NAbs) as a correlate of protection. A leading tetravalent vaccine (Dengvaxia) stimulated NAbs to the 4 DENV serotypes, yet overall vaccine efficacy was low in children who were DENV seronegative at baseline before vaccination. We compared the properties of 1) NAbs induced by wild type DENV1 or 3 infections, which are strongly correlated with protection from repeat infections, and 2) NAbs induced by Dengvaxia in individuals who subsequently experienced DENV1 or DENV3 breakthrough infections. Wild type infections induced NAbs that recognized epitopes unique (type-specific) to each serotype, whereas the vaccine stimulated qualitatively different NAbs that recognized epitopes conserved (cross-reactive) between serotypes. Our results indicate that among children who were DENV seronegative at baseline, unbalanced replication of the DENV type 4 vaccine component in the tetravalent vaccine stimulates Abs capable of cross neutralizing DENV1 and 3 in vitro but not protect in vivo. In DENV seronegative individuals who are vaccinated, we propose that type specific NAbs are a better correlate of protection than total levels of NAbs.

Authors

Sandra Henein, Cameron Adams, Matthew Bonaparte, Janice M. Moser, Alina Munteanu, Ralph Baric, Aravinda M. Desilva

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