Research Article
Abstract
Inflammation promotes adverse ventricular remodeling, a common antecedent of heart failure. Here, we set out to determine how inflammatory cells affect cardiomyocytes in the remodeling heart. Pathogenic cardiac macrophages induced an IFN response in cardiomyocytes, characterized by upregulation of the ubiquitin-like protein IFN-stimulated gene 15 (ISG15), which posttranslationally modifies its targets through a process termed ISGylation. Cardiac ISG15 is controlled by type I IFN signaling, and ISG15 or ISGylation is upregulated in mice with transverse aortic constriction or infused with angiotensin II; rats with uninephrectomy and DOCA-salt, or pulmonary artery banding; cardiomyocytes exposed to IFNs or CD4+ T cell–conditioned medium; and ventricular tissue of humans with nonischemic cardiomyopathy. By nanoscale liquid chromatography–tandem mass spectrometry, we identified the myofibrillar protein filamin-C as an ISGylation target. ISG15 deficiency preserved cardiac function in mice with transverse aortic constriction and led to improved recovery of mouse hearts ex vivo. Metabolomics revealed that ISG15 regulates cardiac amino acid metabolism, whereas ISG15 deficiency prevented misfolded filamin-C accumulation and induced cardiomyocyte autophagy. In sum, ISG15 upregulation is a feature of pathological ventricular remodeling, and protein ISGylation is an inflammation-induced posttranslational modification that may contribute to heart failure development by altering cardiomyocyte protein turnover.
Authors
Veera Ganesh Yerra, Sri Nagarjun Batchu, Harmandeep Kaur, MD Golam Kabir, Youan Liu, Suzanne L. Advani, Duc Tin Tran, Shadi Sadeghian, Phelopater Sedrak, Filio Billia, Uros Kuzmanov, Anthony O. Gramolini, Deema O. Qasrawi, Evgeniy V. Petrotchenko, Christoph H. Borchers, Kim A. Connelly, Andrew Advani
Abstract
Adaptation of the islet β cell insulin-secretory response to changing insulin demand is critical for blood glucose homeostasis, yet the mechanisms underlying this adaptation are unknown. Here, we have shown that nutrient-stimulated histone acetylation plays a key role in adapting insulin secretion through regulation of genes involved in β cell nutrient sensing and metabolism. Nutrient regulation of the epigenome occurred at sites occupied by the chromatin-modifying enzyme lysine-specific demethylase 1 (Lsd1) in islets. β Cell–specific deletion of Lsd1 led to insulin hypersecretion, aberrant expression of nutrient-response genes, and histone hyperacetylation. Islets from mice adapted to chronically increased insulin demand exhibited shared epigenetic and transcriptional changes. Moreover, we found that genetic variants associated with type 2 diabetes were enriched at LSD1-bound sites in human islets, suggesting that interpretation of nutrient signals is genetically determined and clinically relevant. Overall, these studies revealed that adaptive insulin secretion involves Lsd1-mediated coupling of nutrient state to regulation of the islet epigenome.
Authors
Matthew Wortham, Fenfen Liu, Austin R. Harrington, Johanna Y. Fleischman, Martina Wallace, Francesca Mulas, Medhavi Mallick, Nicholas K. Vinckier, Benjamin R. Cross, Joshua Chiou, Nisha A. Patel, Yinghui Sui, Carolyn McGrail, Yesl Jun, Gaowei Wang, Ulupi S. Jhala, Roland Schüle, Orian S. Shirihai, Mark O. Huising, Kyle J. Gaulton, Christian M. Metallo, Maike Sander
Abstract
Clearance of senescent cells (SnCs) can prevent several age-related pathologies, including bone loss. However, the local versus systemic roles of SnCs in mediating tissue dysfunction remain unclear. Thus, we developed a mouse model (p16-LOX-ATTAC) that allowed for inducible SnC elimination (senolysis) in a cell-specific manner and compared the effects of local versus systemic senolysis during aging using bone as a prototype tissue. Specific removal of Sn osteocytes prevented age-related bone loss at the spine, but not the femur, by improving bone formation without affecting osteoclasts or marrow adipocytes. By contrast, systemic senolysis prevented bone loss at the spine and femur and not only improved bone formation, but also reduced osteoclast and marrow adipocyte numbers. Transplantation of SnCs into the peritoneal cavity of young mice caused bone loss and also induced senescence in distant host osteocytes. Collectively, our findings provide proof-of-concept evidence that local senolysis has health benefits in the context of aging, but, importantly, that local senolysis only partially replicates the benefits of systemic senolysis. Furthermore, we establish that SnCs, through their senescence-associated secretory phenotype (SASP), lead to senescence in distant cells. Therefore, our study indicates that optimizing senolytic drugs may require systemic instead of local SnC targeting to extend healthy aging.
Authors
Joshua N. Farr, Dominik Saul, Madison L. Doolittle, Japneet Kaur, Jennifer L. Rowsey, Stephanie J. Vos, Mitchell N. Froemming, Anthony B. Lagnado, Yi Zhu, Megan Weivoda, Yuji Ikeno, Robert J. Pignolo, Laura J. Niedernhofer, Paul D. Robbins, Diana Jurk, João F. Passos, Nathan K. LeBrasseur, Tamara Tchkonia, James L. Kirkland, David G. Monroe, Sundeep Khosla
Abstract
Pancreatic ductal adenocarcinoma (PDAC) frequently presents with metastasis, but the molecular programs in human PDAC cells that drive invasion are not well understood. Using an experimental pipeline enabling PDAC organoid isolation and collection based on invasive phenotype, we assessed the transcriptomic programs associated with invasion in our organoid model. We identified differentially expressed genes in invasive organoids compared with matched noninvasive organoids from the same patients, and we confirmed that the encoded proteins were enhanced in organoid invasive protrusions. We identified 3 distinct transcriptomic groups in invasive organoids, 2 of which correlated directly with the morphological invasion patterns and were characterized by distinct upregulated pathways. Leveraging publicly available single-cell RNA-sequencing data, we mapped our transcriptomic groups onto human PDAC tissue samples, highlighting differences in the tumor microenvironment between transcriptomic groups and suggesting that non-neoplastic cells in the tumor microenvironment can modulate tumor cell invasion. To further address this possibility, we performed computational ligand-receptor analysis and validated the impact of multiple ligands (TGF-β1, IL-6, CXCL12, MMP9) on invasion and gene expression in an independent cohort of fresh human PDAC organoids. Our results identify molecular programs driving morphologically defined invasion patterns and highlight the tumor microenvironment as a potential modulator of these programs.
Authors
Yea Ji Jeong, Hildur Knutsdottir, Fatemeh Shojaeian, Michael G. Lerner, Maria F. Wissler, Elodie Henriet, Tammy Ng, Shalini Datta, Bernat Navarro-Serer, Peter Chianchiano, Benedict Kinny-Köster, Jacquelyn W. Zimmerman, Genevieve Stein-O’Brien, Matthias M. Gaida, James R. Eshleman, Ming-Tseh Lin, Elana J. Fertig, Andrew J. Ewald, Joel S. Bader, Laura D. Wood
Abstract
Lysosomal inhibition elicited by palmitoyl-protein thioesterase 1 (PPT1) inhibitors such as DC661 can produce cell death, but the mechanism for this is not completely understood. Programmed cell death pathways (autophagy, apoptosis, necroptosis, ferroptosis, and pyroptosis) were not required to achieve the cytotoxic effect of DC661. Inhibition of cathepsins, or iron or calcium chelation, did not rescue DC661-induced cytotoxicity. PPT1 inhibition induced lysosomal lipid peroxidation (LLP), which led to lysosomal membrane permeabilization and cell death that could be reversed by the antioxidant N-acetylcysteine (NAC) but not by other lipid peroxidation antioxidants. The lysosomal cysteine transporter MFSD12 was required for intralysosomal transport of NAC and rescue of LLP. PPT1 inhibition produced cell-intrinsic immunogenicity with surface expression of calreticulin that could only be reversed with NAC. DC661-treated cells primed naive T cells and enhanced T cell–mediated toxicity. Mice vaccinated with DC661-treated cells engendered adaptive immunity and tumor rejection in “immune hot” tumors but not in “immune cold” tumors. These findings demonstrate that LLP drives lysosomal cell death, a unique immunogenic form of cell death, pointing the way to rational combinations of immunotherapy and lysosomal inhibition that can be tested in clinical trials.
Authors
Monika Bhardwaj, Jennifer J. Lee, Amanda M. Versace, Sandra L. Harper, Aaron R. Goldman, Mary Ann S. Crissey, Vaibhav Jain, Mahendra Pal Singh, Megane Vernon, Andrew E. Aplin, Seokwoo Lee, Masao Morita, Jeffrey D. Winkler, Qin Liu, David W. Speicher, Ravi K. Amaravadi
Abstract
Peripheral neuropathy is a frequent complication of type 2 diabetes mellitus (T2DM). We investigated whether human islet amyloid polypeptide (hIAPP), which forms pathogenic aggregates that damage pancreatic islet β cells in T2DM, is involved in T2DM-associated peripheral neuropathy. In vitro, hIAPP incubation with sensory neurons reduced neurite outgrowth and increased levels of mitochondrial reactive oxygen species. hIAPP-transgenic mice, which have elevated plasma hIAPP levels without hyperglycemia, developed peripheral neuropathy as evidenced by pain-associated behavior and reduced intraepidermal nerve fiber (IENF) density. Similarly, hIAPP Ob/Ob mice, which have hyperglycemia in combination with elevated plasma hIAPP levels, had signs of neuropathy, although more aggravated. In wild-type mice, intraplantar and intravenous hIAPP injections induced long-lasting allodynia and decreased IENF density. Non-aggregating murine IAPP, mutated hIAPP (pramlintide), or hIAPP with pharmacologically inhibited aggregation did not induce these effects. T2DM patients had reduced IENF density and more hIAPP oligomers in the skin compared with non-T2DM controls. Thus, we provide evidence that hIAPP aggregation is neurotoxic and mediates peripheral neuropathy in mice. The increased abundance of hIAPP aggregates in the skin of T2DM patients supports the notion that hIAPP is a potential contributor to T2DM neuropathy in humans.
Authors
Mohammed M.H. Albariqi, Sabine Versteeg, Elisabeth M. Brakkee, J. Henk Coert, Barend O.W. Elenbaas, Judith Prado, C. Erik Hack, Jo W.M. Höppener, Niels Eijkelkamp
Abstract
Circadian rhythmicity in renal function suggests rhythmic adaptations in renal metabolism. To decipher the role of the circadian clock in renal metabolism, we studied diurnal changes in renal metabolic pathways using integrated transcriptomic, proteomic, and metabolomic analysis performed on control mice and mice with an inducible deletion of the circadian clock regulator Bmal1 in the renal tubule (cKOt). With this unique resource, we demonstrated that approximately 30% of RNAs, approximately 20% of proteins, and approximately 20% of metabolites are rhythmic in the kidneys of control mice. Several key metabolic pathways, including NAD+ biosynthesis, fatty acid transport, carnitine shuttle, and β-oxidation, displayed impairments in kidneys of cKOt mice, resulting in perturbed mitochondrial activity. Carnitine reabsorption from primary urine was one of the most affected processes with an approximately 50% reduction in plasma carnitine levels and a parallel systemic decrease in tissue carnitine content. This suggests that the circadian clock in the renal tubule controls both kidney and systemic physiology.
Authors
Yohan Bignon, Leonore Wigger, Camille Ansermet, Benjamin D. Weger, Sylviane Lagarrigue, Gabriel Centeno, Fanny Durussel, Lou Götz, Mark Ibberson, Sylvain Pradervand, Manfredo Quadroni, Meltem Weger, Francesca Amati, Frédéric Gachon, Dmitri Firsov
Abstract
The rapid evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variants has emphasized the need to identify antibodies with broad neutralizing capabilities to inform future monoclonal therapies and vaccination strategies. Herein, we identified S728-1157, a broadly neutralizing antibody (bnAb) targeting the receptor-binding site (RBS) that was derived from an individual previously infected with WT SARS-CoV-2 prior to the spread of variants of concern (VOCs). S728-1157 demonstrated broad cross-neutralization of all dominant variants, including D614G, Beta, Delta, Kappa, Mu, and Omicron (BA.1/BA.2/BA.2.75/BA.4/BA.5/BL.1/XBB). Furthermore, S728-1157 protected hamsters against in vivo challenges with WT, Delta, and BA.1 viruses. Structural analysis showed that this antibody targets a class 1/RBS-A epitope in the receptor binding domain via multiple hydrophobic and polar interactions with its heavy chain complementarity determining region 3 (CDR-H3), in addition to common motifs in CDR-H1/CDR-H2 of class 1/RBS-A antibodies. Importantly, this epitope was more readily accessible in the open and prefusion state, or in the hexaproline (6P)-stabilized spike constructs, as compared with diproline (2P) constructs. Overall, S728-1157 demonstrates broad therapeutic potential and may inform target-driven vaccine designs against future SARS-CoV-2 variants.
Authors
Siriruk Changrob, Peter J. Halfmann, Hejun Liu, Jonathan L. Torres, Joshua J.C. McGrath, Gabriel Ozorowski, Lei Li, G. Dewey Wilbanks, Makoto Kuroda, Tadashi Maemura, Min Huang, Nai-Ying Zheng, Hannah L. Turner, Steven A. Erickson, Yanbin Fu, Atsuhiro Yasuhara, Gagandeep Singh, Brian Monahan, Jacob Mauldin, Komal Srivastava, Viviana Simon, Florian Krammer, D. Noah Sather, Andrew B. Ward, Ian A. Wilson, Yoshihiro Kawaoka, Patrick C. Wilson
Abstract
How phosphate levels are detected in mammals is unknown. The bone-derived hormone fibroblast growth factor 23 (FGF23) lowers blood phosphate levels by reducing kidney phosphate reabsorption and 1,25(OH)2D production, but phosphate does not directly stimulate bone FGF23 expression. Using PET scanning and LC-MS, we found that phosphate increases kidney-specific glycolysis and synthesis of glycerol-3-phosphate (G-3-P), which then circulates to bone to trigger FGF23 production. Further, we found that G-3-P dehydrogenase 1 (Gpd1), a cytosolic enzyme that synthesizes G-3-P and oxidizes NADH to NAD+, is required for phosphate-stimulated G-3-P and FGF23 production and prevention of hyperphosphatemia. In proximal tubule cells, we found that phosphate availability is substrate-limiting for glycolysis and G-3-P production and that increased glycolysis and Gpd1 activity are coupled through cytosolic NAD+ recycling. Finally, we show that the type II sodium-dependent phosphate cotransporter Npt2a, which is primarily expressed in the proximal tubule, conferred kidney specificity to phosphate-stimulated G-3-P production. Importantly, exogenous G-3-P stimulated FGF23 production when Npt2a or Gpd1 were absent, confirming that it was the key circulating factor downstream of glycolytic phosphate sensing in the kidney. Together, these findings place glycolysis at the nexus of mineral and energy metabolism and identify a kidney-bone feedback loop that controls phosphate homeostasis.
Authors
Wen Zhou, Petra Simic, Iris Y. Zhou, Peter Caravan, Xavier Vela Parada, Donghai Wen, Onica L. Washington, Maria Shvedova, Kerry A. Pierce, Clary B. Clish, Michael Mannstadt, Tatsuya Kobayashi, Marc N. Wein, Harald Jüppner, Eugene P. Rhee
Abstract
The origin of breast cancer, whether primary or recurrent, is unknown. Here, we show that invasive breast cancer cells exposed to hypoxia release small extracellular vesicles (sEVs) that disrupt the differentiation of normal mammary epithelia, expand stem and luminal progenitor cells, and induce atypical ductal hyperplasia and intraepithelial neoplasia. This was accompanied by systemic immunosuppression with increased myeloid cell release of the alarmin S100A9 and oncogenic traits of epithelial-mesenchymal transition, angiogenesis, and local and disseminated luminal cell invasion in vivo. In the presence of a mammary gland driver oncogene (MMTV-PyMT), hypoxic sEVs accelerated bilateral breast cancer onset and progression. Mechanistically, genetic or pharmacologic targeting of hypoxia-inducible factor-1α (HIF1α) packaged in hypoxic sEVs or homozygous deletion of S100A9 normalized mammary gland differentiation, restored T cell function, and prevented atypical hyperplasia. The transcriptome of sEV-induced mammary gland lesions resembled luminal breast cancer, and detection of HIF1α in plasma circulating sEVs from luminal breast cancer patients correlated with disease recurrence. Therefore, sEV-HIF1α signaling drives both local and systemic mechanisms of mammary gland transformation at high risk for evolution to multifocal breast cancer. This pathway may provide a readily accessible biomarker of luminal breast cancer progression.
Authors
Irene Bertolini, Michela Perego, Yulia Nefedova, Cindy Lin, Andrew Milcarek, Peter Vogel, Jagadish C. Ghosh, Andrew V. Kossenkov, Dario C. Altieri
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