Functional monocytic myeloid-derived suppressor cells increase in blood but not airways and predict COVID-19 severity

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Abstract

The immunopathology of COVID-19 remains enigmatic, exhibiting immunodysregulation and T cell lymphopenia. Monocytic myeloid-derived suppressor cells (M-MDSC) are T cell suppressors that expand in inflammatory conditions, but their role in acute respiratory infections remains unclear. We studied blood and airways of COVID-19 patients across disease severity at multiple timepoints. M-MDSC frequencies were elevated in blood but not in nasopharyngeal or endotracheal aspirates of COVID-19 patients compared to controls. M-MDSCs isolated from COVID-19 patients suppressed T cell proliferation and IFNg production partly via an arginase-1 (Arg-1) dependent mechanism. Furthermore, patients showed increased Arg-1 and IL-6 plasma levels. COVID-19 patients had fewer T cells, and displayed downregulated expression of the CD3ζ chain. Ordinal regression showed that early M-MDSC frequency predicted subsequent disease severity. In conclusion, M-MDSCs expand in blood of COVID-19 patients, suppress T cells and strongly associate with disease severity, suggesting a role for M-MDSCs in the dysregulated COVID-19 immune response.

Authors

Sara Falck-Jones, Sindhu Vangeti, Meng Yu, Ryan Falck-Jones, Alberto Cagigi, Isabella Badolati, Björn Österberg, Maximilian Julius Lautenbach, Eric Ahlberg, Ang Lin, Rico Lepzien, Inga Szurgot, Klara Lenart, Fredrika Hellgren, Holden T. Maecker, Jörgen Sälde, Jan Albert, Niclas Johansson, Max Bell, Karin Lore, Anna Färnert, Anna Smed-Sörensen

Lipid-induced endothelial vascular cell adhesion molecule 1 promotes nonalcoholic steatohepatitis pathogenesis

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Abstract

Monocyte homing to the liver and adhesion to the liver sinusoidal endothelial cells (LSEC) are key elements in nonalcoholic steatohepatitis (NASH) pathogenesis. We reported previously that vascular cell adhesion molecule 1 (VCAM-1) mediates monocyte adhesion to LSEC. However, the pathogenic role of VCAM-1 in NASH is unclear. Herein, we report that VCAM-1 was a top upregulated adhesion molecule in the NASH mouse liver transcriptome. Open chromatin landscape profiling combined with genome-wide transcriptome analysis showed robust transcriptional upregulation of LSEC-VCAM-1 in murine NASH. Moreover, LSEC-VCAM-1 expression was significantly increased in human NASH. LSEC-VCAM-1 expression was upregulated by palmitate treatment in vitro, and reduced with inhibition of the mitogen-activated protein 3 kinase, mixed lineage kinase 3 (MLK3). Likewise, LSEC-VCAM-1 expression was reduced in the Mlk3-/- mice with diet-induced NASH. Furthermore, VCAM-1 neutralizing antibody or pharmacological inhibition attenuated diet-induced NASH in mice, mainly via reducing the proinflammatory monocyte hepatic population as examined by mass cytometry by time of flight (CyTOF). Moreover, endothelium-specific Vcam1 knockout mice were also protected against NASH. In summary, lipotoxic stress enhances the expression of LSEC-VCAM-1, in part, through MLK3 signaling. Inhibition of VCAM-1 was salutary in murine NASH, and might serve as a potential therapeutic strategy for human NASH.

Authors

Kunimaro Furuta, Qianqian Guo, Kevin D. Pavelko, Jeong-Heon Lee, Keith D. Robertson, Yasuhiko Nakao, Jan Melek, Vijay H. Shah, Petra Hirsova, Samar H. Ibrahim

Radiotherapy-exposed CD8⁺ and CD4⁺ neoantigens enhance tumor control

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Abstract

Neoantigens generated by somatic non-synonymous mutations are key targets of tumor-specific T cells, but only a small number of mutations predicted to be immunogenic are presented by MHC molecules on cancer cells. Vaccination studies in mice and patients have shown that the majority of neoepitopes that elicit T cell responses fail to induce significant anti-tumor activity, for incompletely understood reasons. We report that radiotherapy upregulates the expression of genes containing immunogenic mutations in a poorly immunogenic mouse model of triple negative breast cancer. Vaccination with neoepitopes encoded in these genes elicited CD8+ and CD4+ T cells that, whereas ineffective in preventing tumor growth, improved the therapeutic efficacy of radiotherapy. Mechanistically, neoantigen-specific CD8+ T cells preferentially killed irradiated tumor cells. Neoantigen-specific CD4+ T cells were required for the therapeutic efficacy of vaccination and acted by producing Th1 cytokines, killing irradiated tumor cells and promoting epitope spread. Such a cytotoxic activity relied on the ability of radiation to upregulate class II MHC molecules as well as the death receptors FAS/CD95 and DR5 on the surface of tumor cells. These results provide proof-of-principle evidence that radiotherapy works in concert with neoantigen vaccination to improve tumor control.

Authors

Claire Lhuillier, Nils-Petter Rudqvist, Takahiro Yamazaki, Tuo Zhang, Maud Charpentier, Lorenzo Galluzzi, Noah Dephoure, Cristina C. Clement, Laura Santambrogio, Xi K. Zhou, Silvia C. Formenti, Sandra Demaria

Smoothened-activating lipids drive resistance to CDK4/6 inhibition in Hedgehog-associated medulloblastoma cells and preclinical models

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Abstract

Medulloblastoma is an aggressive pediatric brain tumor that can be driven by misactivation of the Hedgehog (HH) pathway. CDK6 is a critical effector of oncogenic Hedgehog signaling, but attempts to target the Hedgehog pathway in medulloblastoma have been encumbered by resistance to single-agent molecular therapy. We identified resistance mechanisms to CDK6 inhibition in HH-associated medulloblastoma by performing orthogonal CRISPR and CRISPR interference screens in medulloblastoma cells treated with a CDK4/6 inhibitor, and RNA-sequencing of a mouse model of HH-associated medulloblastoma with genetic deletion of Cdk6. Our concordant in vitro and in vivo data revealed decreased ribosomal protein expression underlies resistance to CDK6 inhibition in HH-associated medulloblastoma, leading to endoplasmic reticular (ER) stress and activation of the unfolded protein response (UPR). These pathways increased the activity of enzymes producing Smoothened-activating sterol lipids that sustained oncogenic HH signaling in medulloblastoma despite cell cycle attenuation. Consistently, we demonstrated concurrent genetic deletion or pharmacological inhibition of CDK6 and HSD11ß2, an enzyme producing Smoothened-activating lipids, additively blocked cancer growth in multiple mouse genetic models of HH-associated medulloblastoma. Our data reveal a resistance pathway to CDK4/6 inhibition and a combination therapy to treat the most common malignant brain tumor in children that we believe are novel.

Authors

Vikas Daggubati, Jordan Hochstetler, Anirudh Bommireddy, Abrar Choudhury, Alexis Leigh Krup, Pervinder K. Choksi, Pakteema Tong, Amy Li, Libin Xu, Jeremy F. Reiter, David R. Raleigh

CC17 Group B Streptococcus exploits integrins for neonatal meningitis development

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Abstract

Group B Streptococcus (GBS) is the major cause of human neonatal infections. A single clone, designated CC17-GBS, accounts for more than 80% of meningitis cases, the most severe form of the infection. However, the events allowing blood-borne GBS to penetrate the brain remain largely elusive. In this study, we identified the host transmembrane receptors α5β1 and αvβ3 integrins as the ligands of Srr2, a major CC17-GBS specific adhesin. Two motifs located in the binding region of Srr2 were responsible for the interaction between CC17-GBS and these integrins. We demonstrated, in a blood-brain barrier cellular model, that both integrins contributed to the adhesion and internalization of CC17-GBS. Strikingly, both integrins were overexpressed during the post-natal period in the brain vessels of the blood-brain and blood-cerebrospinal fluid barriers and contributed to the juvenile susceptibility to CC17-meningitis. Finally, blocking these integrins decreased CC17-GBS crossing into the juvenile mice central nervous system in an in vivo model of meningitis.Our study demonstrates that CC17-GBS exploits integrins for crossing the brain vessels leading to meningitis. Importantly, it provides host molecular insights into neonate’s susceptibility to CC17-GBS meningitis, thereby opening new perspectives for therapeutic and prevention strategies of GBS-elicited meningitis.

Authors

Romain Deshayes de Cambronne, Agnès Fouet, Amandine Picart, Anne-Sophie Bourrel, Cyril Anjou, Guillaume Bouvier, Cristina Candeias, Abdelouhab Bouaboud, Lionel Costa, Anne-Cécile Boulay, Martine Cohen-Salmon, Isabelle Plu, Caroline Rambaud, Eva Faurobert, Corinne Albiges-Rizo, Asmaa Tazi, Claire Poyart, Julie Guignot

Antibody-mediated depletion of viral reservoirs is limited in SIV-infected macaques treated early with antiretroviral therapy

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Abstract

The effectiveness of virus-specific strategies, including administered HIV-specific mAbs, to target cells that persistently harbor latent, rebound competent HIV genomes during combination antiretroviral therapy (cART) has been limited by inefficient induction of viral protein expression. To examine antibody-mediated viral reservoir targeting without a need for viral induction, we used an anti-CD4 mAb to deplete both infected and uninfected CD4+ T cells. Ten rhesus macaques infected with barcoded SIVmac239M received cART for 93 weeks starting 4 days post-infection. During cART, five animals received 5-6 anti-CD4 antibody administrations and CD4+ T cell populations were then allowed one year on cART to recover. Despite profound CD4+ T cell depletion in blood and lymph nodes, time to viral rebound following cART cessation was not significantly delayed in anti-CD4 treated animals compared with controls. Viral reactivation rates, determined based on rebounding SIVmac239M clonotype proportions, also were not significantly different in CD4 depleted animals. Notably, antibody-mediated depletion was limited in rectal tissue and negligible in lymphoid follicles. These results suggest that even if robust viral reactivation can be achieved, antibody-mediated viral reservoir depletion may be limited in key tissue sites.

Authors

Adrienne E. Swanstrom, Taina T. Immonen, Kelli Oswald, Cathi Pyle, James A. Thomas, William J. Bosche, Lorna Silipino, Michael Hull, Laura Newman, Vicky Coalter, Adam Wiles, Rodney Wiles, Jacob Kiser, David R. Morcock, Rebecca Shoemaker, Randy Fast, Matthew W. Breed, Joshua Kramer, Duncan Donohue, Tyler Malys, Christine M. Fennessey, Charles M. Trubey, Claire Deleage, Jacob D. Estes, Jeffrey D. Lifson, Brandon F. Keele, Gregory Q. Del Prete

Endoplasmic reticulum protein TXNDC5 promotes renal fibrosis by enforcing TGFβ signaling in kidney fibroblasts

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Abstract

Renal fibrosis, a common pathological manifestation of virtually all types of chronic kidney diseases (CKD), often results in diffuse kidney scarring and predisposes to end-stage renal disease. Currently, there is no effective therapy against renal fibrosis. Recently, our laboratory identified an ER-resident protein, thioredoxin domain containing 5 (TXNDC5), as a critical mediator of cardiac fibrosis. Transcriptome analyses of renal biopsy specimens from CKD patients revealed marked TXNDC5 upregulation in fibrotic kidneys, suggesting a potential role of TXNDC5 in renal fibrosis. Employing multiple fluorescent reporter mouse lines, we showed that TXNDC5 was specifically upregulated in collagen-secreting fibroblasts in fibrotic mouse kidneys. In addition, we showed that TXNDC5 was required for TGFβ1-induced fibrogenic responses in human kidney fibroblasts (HKF), whereas TXNDC5 over-expression was sufficient to promote HKF activation, proliferation and collagen production. Mechanistically, we showed that TXNDC5, transcriptionally controlled by ATF6-dependent ER stress pathway, mediates its pro-fibrogenic effects by enforcing TGFβ signaling activity through post-translational stabilization and upregulation of type I TGFβ receptor in kidney fibroblasts. Using a tamoxifen-inducible, fibroblast-specific Txndc5 knockout mouse line, we demonstrated that deletion of Txndc5 in kidney fibroblasts mitigated the progression of established kidney fibrosis, suggesting the therapeutic potential of TXNDC5 targeting for renal fibrosis and CKD.

Authors

Yen-Ting Chen, Pei-Yu Jhao, Chen-Ting Hung, Yueh-Feng Wu, Sung-Jan Lin, Wen-Chih Chiang, Shuei-Liong Lin, Kai-Chien Yang

Renal tubule Cpt1a overexpression protects from kidney fibrosis by restoring mitochondrial homeostasis

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Abstract

Chronic kidney disease (CKD) remains a major epidemiological, clinical and biomedical challenge. During CKD, renal tubular epithelial cells (TECs) suffer a persistent inflammatory and profibrotic response. Fatty acid oxidation (FAO), the main source of energy for TECs, is reduced in kidney fibrosis and contributes to its pathogenesis. To determine if FAO gain-of-function (FAO-GOF) could protect from fibrosis, we generated a conditional transgenic mouse model with overexpression of the fatty acid shuttling enzyme carnitine palmitoyl-transferase 1 A (CPT1A) in TECs. Cpt1a knock-in mice subjected to three different models of renal fibrosis (unilateral ureteral obstruction, folic acid nephropathy-FAN and adenine induced nephrotoxicity) exhibited decreased expression of fibrotic markers, a blunted pro-inflammatory response and reduced epithelial cell damage and macrophage influx. Protection from fibrosis was also observed when Cpt1a overexpression was induced after FAN. FAO-GOF restituted oxidative metabolism and mitochondrial number and enhanced bioenergetics increasing palmitate oxidation and ATP levels, changes also recapitulated in TECs exposed to profibrotic stimuli. Studies in patients evidenced decreased CPT1 levels and increased accumulation of short and middle chain acyl-carnitines, reflecting impaired FAO in human CKD. We propose that strategies based on FAO-GOF may constitute powerful alternatives to combat fibrosis inherent to CKD.

Authors

Verónica Miguel, Jessica Tituaña, J. Ignacio Herrero, Laura Herrero, Dolors Serra, Paula Cuevas-Delgado, Coral Barbas, Diego Rodriguez-Puyol, Laura Marquez-Exposito, Marta Ruiz-Ortega, Carolina Castillo, Xin Sheng, Katalin Susztak, Miguel Ruiz-Canela, Jordi Salas-Salvado, Miguel A. Martinez-Gonzalez, Sagrario Ortega, Ricardo Ramos-Ruiz, Santiago Lamas

Impaired complex I repair causes recessive Leber’s hereditary optic neuropathy

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Abstract

Leber’s hereditary optic neuropathy (LHON) is the most frequent mitochondrial disease and was the first to be genetically defined by a point mutation in the mitochondrial DNA (mtDNA). A molecular diagnosis is reached in up to 95%, the vast majority of which are accounted for by three mutations within mitochondrial complex I (CI) subunit encoding genes in the mtDNA (mtLHON). Here, we resolve the enigma of LHON in the absence of pathogenic mtDNA mutations. We describe biallelic mutations in a nuclear encoded gene, DNAJC30, in 33 unsolved patients from 29 families and establish an autosomal recessive mode of inheritance for LHON (arLHON), which to date has been a prime example of a maternally inherited disorder. Remarkably, all hallmarks of mtLHON are recapitulated, including incomplete penetrance, male predominance, and significant idebenone responsivity. Moreover, by tracking protein turnover in patient-derived cell lines and a DNAJC30-knock-out cellular model, we measure reduced turnover of specific CI N-module subunits and a resultant impairment of CI function. This demonstrates DNAJC30 is to be a chaperone protein needed for the efficient exchange of CI subunits exposed to reactive oxygen species and integral to a mitochondrial CI repair mechanism, thereby providing the first example of a disease resulting from impaired exchange of assembled respiratory chain subunits.

Authors

Sarah L. Stenton, Natalia L. Sheremet, Claudia B. Catarino, Natalia Andreeva, Zahra Assouline, Piero Barboni, Ortal Barel, Riccardo Berutti, Igor O. Bychkov, Leonardo Caporali, Mariantonietta Capristo, Michele Carbonelli, Maria Lucia Cascavilla, Peter Charbel Issa, Peter Freisinger, Sylvie Gerber, Daniele Ghezzi, Elisabeth Graf, Juliana Heidler, Maja Hempel, Elise Heon, Yulia S. Itkis, Elisheva Javasky, Josseline Kaplan, Robert Kopajtich, Cornelia Kornblum, Reka Kovacs-Nagy, Tatiana Krylova, Wolfram S. Kunz, Chiara La Morgia, Costanza Lamperti, Christina Ludwig, Pedro F. Malacarne, Alessandra Maresca, Johannes A. Mayr, Jana Meisterknecht, Tatiana Nevinitsyna, Flavia Palombo, Ben Pode-Shakked, Maria S. Shmelkova, Tim M. Strom, Francesca Tagliavini, Michal Tzadok, Amelie T. van der Ven, Catherine Vignal-Clermont, Matias Wagner, Ekaterina Zakharova, Nino Zhorzholadze, Jen-Michel Rozet, Valerio Carelli, Polina Tsygankova, Thomas Klopstock, Ilka Wittig, Holger Prokisch

Disrupting the DREAM transcriptional repressor complex induces apolipoprotein overexpression and systemic amyloidosis in mice

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Abstract

DREAM is a transcriptional repressor complex that regulates cell proliferation and its loss causes neonatal lethality in mice. To investigate DREAM function in adult mice, we utilized an assembly defective p107 protein and conditional deletion of its redundant family member p130. In the absence of DREAM assembly, mice displayed shortened survival characterized by systemic amyloidosis, but no evidence of excessive cellular proliferation. Amyloid deposits were found in the heart, liver, spleen, and kidneys, but not the brain or bone marrow. Using laser capture microdissection followed by mass spectrometry, we identified apolipoproteins as the most abundant components of amyloids. Intriguingly, apoA-IV was the most detected amyloidogenic protein in amyloid deposits, suggesting AApoAIV amyloidosis. AApoAIV is a recently described form whereby wildtype apoA-IV has been shown to predominate in amyloid plaques. We determined that DREAM directly regulates Apoa4 by chromatin immunoprecipitation and that the histone variant H2AZ is reduced from the Apoa4 gene body in DREAM’s absence, leading to overexpression. Collectively, we describe a mechanism by which epigenetic misregulation causes apolipoprotein overexpression and amyloidosis, potentially explaining the origins of non-genetic amyloid subtypes.

Authors

Pirunthan Perampalam, Haider M. Hassan, Grace E. Lilly, Daniel T. Passos, Joseph Torchia, Patti K. Kiser, Andrea Bozovic, Vathany Kulasingam, Frederick A. Dick