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Dual dysregulation of TNF/interferon signaling and classical monocytes are implicated in Reactive Infectious Mucocutaneous Eruptions

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Authors

Zhehao Tan, Gio Wu, Daniela Salgado Figueroa, Paramita Dutta, Zachary Jaeger, Marissa Mazurie, David Schairer, Dawn Eichenfield, Wynnis L. Tom, Lauren Galli, Lawrence Eichenfield, Bob Geng, Brian Hinds, Hal M. Hoffman, Lori Broderick, Ben Croker, Ferhat Ay, Reid Oldenburg

PhIP-Seq uncovers marked heterogeneity in Acute Rheumatic Fever Autoantibodies

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Acute rheumatic fever (ARF) and associated rheumatic heart disease are serious sequelae of a Group A Streptococcus (GAS/Strep A) infection. Autoantibodies are thought to contribute to pathogenesis, with deeper exploration of the autoantibody repertoire needed to improve mechanistic understanding and identify new biomarkers. Phage immunoprecipitation and Sequencing (PhIP-Seq) with the HuScan library (>250,000 overlapping 90-mer peptides spanning the human proteome) was utilised to analyse autoreactivity in sera from children with ARF, uncomplicated Strep A pharyngitis and matched healthy controls. A global proteome-wide increase in autoantigen reactivity was observed in ARF, as was marked heterogeneity between patients. Public epitopes, common between individuals with ARF were rare, and comprised < 1% of all enriched peptides. Differential analysis identified both novel and previously identified ARF autoantigens, including PPP1R12B, a myosin phosphatase complex regulatory subunit expressed in cardiac muscle, and members of the collagen-protein family, respectively. Pathway analysis found antigens from the disease-relevant processes encompassing sarcomere and heart-morphogenesis were targeted. In sum, PhIP-Seq has substantially expanded the spectrum of autoantigens in ARF, and reveals the rarity of public epitopes in the disease. It provides further support for the role of epitope spreading in pathogenesis and has identified PPP1R12B as a novel, enriched autoantigen.

Authors

Reuben McGregor, Lauren H. Carlton, Timothy J. O'Donnell, Elliot Merritt, Campbell R. Sheen, Florina Chan Mow, William John Martin, Michael G. Baker, Nigel Wilson, Uri Laserson, Nicole J. Moreland

Inhibition of the angiotensin-converting enzyme N-terminal catalytic domain prevents endogenous opioid degradation in brain tissue

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Authors

Filip Hanak, Jessica L. Swanson, Krzysztof Felczak, Prakashkumar Dobariya, Ursula C.H. Girdwood, Kenneth E. Bernstein, Swati S. More, Patrick E. Rothwell

Cannabidiol exerts anti-inflammatory effects but maintains T effector memory cell differentiation in humans

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BACKGROUND. Cannabidiol (CBD) is increasingly used for pain management, including in transplant recipients with limited analgesic options. Its immunomodulatory effects in humans are not well defined at a single cell level at CBD steady state with concomitant tacrolimus treatment. METHODS. In a Phase 1 ex vivo study, peripheral blood mononuclear cells from 23 participants who received oral CBD (Epidiolex®) up to 5 mg/kg twice daily for 11 days were collected before CBD (pre-CBD) and at steady state (post-CBD). Lymphocytes were isolated and stimulated with anti-CD3/CD28 antibodies, with or without tacrolimus (5 ng/mL). Pharmacodynamic responses were assessed using CellTiter-Glo® proliferation, single-cell/nucleus RNA sequencing, cytokine assays, and flow cytometry. Steady-state plasma concentrations of CBD were quantified via tandem mass spectrometry. RESULTS. We identified an increased proportion of T effector memory (TEM) cells post-cannabidiol (22% increase), which correlated with CBD plasma concentrations (R = 0.77, P-value = 0.01). Cannabidiol reduced proliferation of T (37% decrease) and CD70hi B (17% decrease) lymphocytes with additive immunosuppressive effects to tacrolimus. Single-cell RNA sequencing revealed reduced IL2 and TNF signaling and altered receptor–ligand networks in TEM cells. Post-cannabidiol cytokine assays revealed elevated proinflammatory IL-6 protein levels and anti-inflammatory IL-10 levels, with reduced TNF-α, LTA, and IL-2. In flow cytometry, the proportion of TEM and TEMRA increased post-cannabidiol with tacrolimus. CONCLUSION. Cannabidiol exhibits mixed immunomodulatory effects with pro- and anti-inflammatory signals. Understanding the clinical safety of cannabidiol use is important given the paucity of pain control options available for immunocompromised transplant populations.

Authors

Debora L. Gisch, Sachiko Koyama, Jumar Etkins, Gerald C. So, Daniel J. Fehrenbach, Jessica Bo Li Lu, Ying-Hua Cheng, Ricardo Melo Ferreira, Evan Rajadhyaksha, Kelsey McClara, Mahla Asghari, Asif A. Sharfuddin, Pierre C. Dagher, Laura M. Snell, Meena S. Madhur, Rafael B. Polidoro, Zeruesenay Desta, Michael T. Eadon

FMT promotes type 2 mucosal immune responses with colonic epithelium proliferation in recurrent CDI patients

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BACKGROUND. Fecal Microbiota Transplantation (FMT) is the most effective therapy for recurrent Clostridioides difficile infection (rCDI), yet its mechanism of action remains poorly understood. METHODS. We report the results of a clinical trial of subjects undergoing FMT therapy for rCDI (n=16), analyzing colon biopsies, plasma, peripheral blood mononuclear cells, and stool at the time of FMT and two-month follow-up. Plasma and colon biopsy samples were also collected from healthy controls for comparison with rCDI patients. Microbiome composition, colonic gene expression, and immune changes were evaluated through high-throughput sequencing and immunoprofiling via flow cytometry. RESULTS. No subjects experienced recurrence at follow-up. FMT significantly altered the intestinal microbiome but had no significant impact on the systemic immune system. In contrast, FMT promoted broad changes in colonic transcriptional profiles compared to both pre-FMT and healthy control biopsies, inhibiting genes associated with pro-inflammatory signaling and upregulating type 2 immunity and proliferative pathways (Myc and mTORC1). FMT increased expression of IL-33 and the type 2 immune EGFR family ligand amphiregulin, potentially explaining upregulation of Myc and mTORC1 pathways. Spatial transcriptomics demonstrated that these changes were localized to the colonic epithelium. Comparison of transcriptional profiles with available single cell gene sets determined that post-FMT biopsies were enriched in signatures associated with proliferative cell types while repressing signatures of differentiated colonocytes. CONCLUSIONS. We conclude that FMT promotes proliferation of the colonic epithelium in rCDI patients, which may drive regeneration and protect against subsequent CDI. REGISTRATION. Clinicaltrials.gov NCT02797288. FUNDING. NIH grants R01 AI152477, R01 AI124214, and K23 AI163368.

Authors

G. Brett Moreau, Jiayi Tian, Nick R. Natale, Farha Naz, Mary K. Young, Uma Nayak, Mehmet Tanyüksel, Isaura Rigo, Gregory R. Madden, Mayuresh M. Abhyankar, Nicholas Hagspiel, Savannah Brovero, Mark Worthington, Brian Behm, Chelsea Marie, William A. Petri Jr., Girija Ramakrishnan

Genital herpes shedding episodes associate with altered spatial organization and activation of mucosal immune cells

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Herpes Simplex Virus 2 (HSV-2) infection results in variable rates of local viral shedding in anogenital skin. The impact of episodic viral exposures on immune cells in adjacent mucosal tissues, including the genital tract, is unknown. However, any immune responses at this site could impact protective mucosal immunity, tissue homeostasis, and adverse health outcomes. To investigate the impact of HSV-2 on cervicovaginal tract immunity, we applied flow cytometry, immunofluorescent imaging, analysis of soluble immune factors, and spatial transcriptomics to cervicovaginal tissue and blood samples provided by a total of 232 HSV-2-seropositive and seronegative participants, with genital HSV-2 shedding evaluated at the time of biopsy. This unique dataset was used to define and spatially map immune cell subsets and localized gene expression via spatial transcriptomics. HSV-2-seropositivity alone was associated with minimal differences in cervicovaginal and circulating T cell phenotypes. However, the vaginal mucosa during active HSV-2 shedding was associated with alterations in T cell, macrophage, and dendritic cell localization and gene expression consistent with increased immune surveillance, with immune activating and suppressing signals potentially reinforcing mucosal tissue homeostasis.

Authors

Finn MacLean, Rachael M. Zemek, Adino Tesfahun Tsegaye, Jessica B. Graham, Jessica L. Swarts, Sarah C. Vick, Nicole B. Potchen, Irene Cruz Talavera, Lakshmi Warrier, Julien Dubrulle, Lena K. Schroeder, Anna Elz, David Sowerby, Ayumi Saito, Katherine K. Thomas, Matthias Mack, Joshua T. Schiffer, R. Scott McClelland, Keith R. Jerome, Bhavna H. Chohan, Kenneth Ngure, Nelly Rwamba Mugo, Evan W. Newell, Jairam R. Lingappa, Jennifer M. Lund

Aebp1 loss in osteoprogenitors leads to skeletal defects resembling Ehlers-Danlos Syndrome by diminishing Wnt/β-catenin signaling

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Ehlers-Danlos Syndrome, Classic-Like, 2 (clEDS2) is a rare genetic disorder caused by biallelic mutations in the AEBP1 gene, which encodes Aortic carboxypeptidase-like protein (ACLP). Patients with clEDS2 exhibit hallmark features such as loose connective tissues, osteoporosis, and scoliosis. Despite its clinical significance, the molecular mechanisms underlying AEBP1 mutations in skeletal development remain poorly understood, and effective therapeutic strategies are currently unavailable. Here, using OsxCre conditional knockout mice, we show that Aebp1 deletion in osteoprogenitors reduces body size and bone mass, recapitulating key skeletal features reported in clEDS2. In primary osteoblasts, both genetic deletion and siRNA-mediated knockdown of Aebp1 impair osteoblast differentiation. Mechanistically, Aebp1 loss attenuates Wnt/β-catenin signaling in bone. Restoration of Wnt/β-catenin signaling by injecting BIO, a small molecule inhibitor of GSK3, substantially rescued bone mass reduction in Aebp1 knockout mice. These findings support a model in which Aebp1 sustains baseline Wnt/β-catenin tone in osteoblast-lineage cells and suggest that Wnt-targeted approaches may help mitigate clEDS2-related skeletal defects.

Authors

Shuhao Feng, Zihang Feng, Zhonghao Deng, Yiran Wei, Ru Lian, Yangchen Jin, Shiqi Zhao, Yu Jin, Zhongmin Zhang, Liang Zhao

LAIR1 prevents excess inflammatory tissue damage in Staphylococcus aureus skin infection and Cutaneous T-cell Lymphoma

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Patients with cutaneous T cell lymphoma (CTCL) experience high morbidity and mortality due to S. aureus skin infections and sepsis, but the underlying mechanisms remain unclear. We have previously identified high levels of LAIR2, a decoy protein for the inhibitory receptor LAIR1, in advanced CTCL. Mice lack a LAIR2 homolog, so we used Lair1 knock-out (KO) mice to model LAIR2 overexpression. In a model of S. aureus skin infection, Lair1 KO mice had significantly larger abscesses and areas of dermonecrosis compared to WT despite similar bacterial burdens. Lair1 KO exhibited a pattern of increased inflammatory responses in infection and sterile immune stimulation, with increased production of proinflammatory cytokines and myeloid chemokines, neutrophil ROS, and collagen/ECM pathway proteins, including collagens and complement factors. These findings support the notion that loss of LAIR1 signaling causes an excessive inflammatory response that exacerbates tissue damage and does not improve infection control. Underscoring the clinical relevance of our findings, CTCL skin lesions exhibited similarly increased expression in cytokine and collagen/ECM remodeling pathways, suggesting that high levels of LAIR2 promote excessive inflammatory tissue damage and compromise host defense against S. aureus infection. LAIR signaling represents a promising target for therapeutic development in CTCL and other inflammatory diseases.

Authors

Hannah K. Dorando, Evan C. Mutic, Kelly L. Tomaszewski, Yulia Korshunova, Ling Tian, Mellisa K. Stefanov, Chaz C. Quinn, Deborah J. Veis, Juliane Bubeck Wardenburg, Amy C. Musiek, Neha Mehta-Shah, Jacqueline E. Payton

Bronchial Epithelial Transcriptome Reveals Dysregulated Interferon and Inflammatory Responses to Rhinovirus in Exacerbation-Prone Pediatric Asthma

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Host factors influencing susceptibility to rhinovirus-induced asthma exacerbations remain poorly characterized. Using organotypic bronchial epithelial cultures from well-characterized children with asthma and healthy children, this study investigated viral load kinetics and resultant host responses by bulk and single-cell transcriptomics and targeted protein analyses. Bronchial epithelium from exacerbation-prone children exhibited greater rhinovirus replication and a cascade of exaggerated downstream interferon (IFN), inflammatory, epithelial stress, and remodeling responses. These transcriptional patterns were confirmed and further refined using single-cell transcriptomics, revealing cell type-specific contributions—particularly from non- ciliated cell populations including secretory immune response, tuft, and basal cells. We observed that these post-infection differences were associated with lower pre-infection IFN- stimulated gene (ISG) expression and protein levels of the ISG CXCL10. Prophylactic IFN-β treatment reduced viral replication and normalized downstream responses, supporting low baseline (pre-infection) IFN tone as a modifiable causal determinant of host susceptibility to adverse rhinovirus-induced responses in exacerbation-prone children with asthma.

Authors

Naresh Doni Jayavelu, Basilin Benson, Patricia C. dela Cruz, Weston T. Powell, Lucille M. Rich, Elizabeth R. Vanderwall, Camile R. Gates, Andrew J. Nagel, Maria P. White, Nyssa B. Samanas, Kourtnie Whitfield, Teal S. Hallstrand, Steven F. Ziegler, Matthew C. Altman, Jason S. Debley

Mutation in IR or IGF1R produces features of long-lived mice while maintaining metabolic health

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Insulin/insulin growth factor signaling is a conserved pathway that regulates lifespan. Yet, long-lived loss-of-function mutants often produce insulin-resistance, slow growth, and impair reproduction. Recently, a gain-of-function mutation in the kinase insert domain (KID) of the Drosophila insulin/IGF receptor was seen to dominantly extend lifespan without impairing insulin-sensitivity, growth and reproduction. This substitution occurs within residues conserved in mammalian insulin receptor (IR) and insulin growth factor-1 receptor (IGF-1R). We produced two knock-in mouse strains that carry the homologous KID Arg/Cys substitution in murine IR or IGF-1R, and we replicated these genotypes in human cells. Cells with heterodimer receptors of IR or IGF-1R induce receptor phosphorylation and phospho-Akt when stimulated with insulin or IGF. Heterodimer receptors of IR fully induce pERK but ERK was less phosphorylated in cells with IGF-1R heterodimers. Adults with a single KID allele (producing heterodimer receptors) have normal growth and glucose regulation. At four months, these mice variably display hormonal markers that associate with successful aging counteraction, including elevated adiponectin, FGF21, and reduced leptin and IGF-1. Livers of IGF-1R females show decreased transcriptome-based biological age, which may point toward delayed aging and warrants an actual lifespan experiment. These data suggest that KID mutants may slow mammalian aging while they avoid the complications of insulin resistance.

Authors

Ulalume Hernández-Arciga, Jun Kyoung Kim, Jacob L. Fisher, Alexander Tyshkovskiy, Alibek Moldakozhayev, Catherine Hall, Souvik Ghosh, Yashvandhini Govindaraj, Ian J. Sipula, Jake Kastroll, Diana Cooke, Jinping Luo, Jonathan K. Alder, Stacey J. Sukoff Rizzo, Gene P. Ables, Eunhee Choi, Vadim N. Gladyshev, Michael J. Jurczak, Marc Tatar, Andrey A. Parkhitko

Preclinical assessment of oral TLR7 agonist SA-5 in a non-human primate model

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Toll-like receptor 7 (TLR7) agonists are promising immunostimulatory agents for the treatment of chronic infections and cancer. However, their systemic toxicity remains a challenge. In this study, SA-5, a novel liver-targeted, orally available TLR7 agonist, was evaluated for pharmacokinetics, safety, and efficacy in young and aged macaques across 1–10 mg/kg repeated doses. Safety was evaluated through hematologic, biochemical, and flow cytometric profiling, while efficacy was assessed via IFN-α production, gene expression of interferon-stimulated genes, and plasmacytoid dendritic cell activation. A principal component analysis (PCA)-based composite scoring system was used to integrate multimodal parameters. SA-5 induced dose-dependent type I IFN with limited systemic inflammation, with 3 mg/kg showing optimal balance. SA-5 had comparable immunostimulatory activity to GS-9620 but with reduced adverse biomarker shifts. In aged macaques, efficacy was maintained with modestly increased safety responses. These findings support SA-5 as a safer next-generation TLR7 agonist effective across age groups, highlighting integrated biomarker profiling in preclinical immunomodulatory drug development.

Authors

Shokichi Takahama, Takahiro Tomiyama, Sachiyo Yoshio, Yuta Nagatsuka, Hirotomo Murakami, Takuto Nogimori, Mami Kochi, Shoko Ochiai, Hidenori Kimura, Akihisa Fukushima, Tatsuya Kanto, Takuya Yamamoto

GDF15 induces excessive activation of osteoclasts within the vertebral endplates leading to early endplate degeneration

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Modic type 1 and 2 changes (MC-1 and MC-2) are highly prevalent in individuals with chronic low back pain, yet the cellular and molecular mechanisms underlying vertebral endplate degeneration remain poorly defined. Here, we report that osteoclastogenesis is markedly elevated in MC-1 and MC-2 lesions compared to MC-3, suggesting an active role for osteoclasts in the early stages of degeneration. Using a lumbar spine instability (LSI) mouse model, we demonstrate enhanced osteoclast activity in degenerating endplates. RNA sequencing of mononuclear cells isolated from the endplate and adjacent subchondral bone identifies Gdf15 as a potential upstream regulator of this process. Conditional knockout of Gdf15 in monocytes reduces osteoclast formation, aberrant CD31hiEmcnhi angiogenesis, and pain-associated neurogenesis, ultimately mitigating endplate degeneration and mechanical allodynia. Mechanistically, GDF15 promotes the fusion of preosteoclasts by modulating the expression of Rho-family small GTPases. In a humanized GDF15 knock-in mouse model, therapeutic neutralization of GDF15 leads to a reduction in osteoclast burden, improved endplate structure, and attenuated pain behavior. Together, these findings uncover a previously unrecognized role for GDF15 in driving osteoclast-mediated endplate degeneration and highlight its potential as a therapeutic target for the treatment of endplate-related chronic low back pain.

Authors

Xiaoqun Li, Jinhui Wu, Qingjie Kong, Miao Hu, Yuhong Li, Ziheng Wei, Heng Jiang, Xuhui Zhou, Jun Ma

Cross-species blood transcriptional correlates of BCG-mediated protection against tuberculosis include innate and adaptive immune processes

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The immune mechanisms induced by the Bacillus Calmette-Guérin (BCG) vaccine, and the subset of which mediate protection against tuberculosis (TB), remain poorly understood. This is further complicated by difficulties to verify vaccine-induced protection in humans. Although research in animal models, namely mice and non-human primates (NHPs), has begun to close this knowledge gap, discrepancies in the relative importance of biological pathways across species limit the utility of animal model-derived biological insights in humans. To address these challenges, we applied a systems modeling framework, Translatable Components Regression (TransCompR), to identify human blood transcriptional variability which could predict Mtb challenge outcomes in BCG-vaccinated NHPs. These protection-associated pathways included both innate and adaptive immune activation mechanisms, along with signaling via type I interferons and anti-mycobacterial T helper cytokines. We further partially validated the associations between these mechanisms and protection in humans using publicly available microarray data collected from BCG-vaccinated infants who either developed TB or remained healthy during two years of follow-up. Overall, our work demonstrates how species translation modeling can leverage animal studies to generate hypotheses about the mechanisms that underlie human infectious disease and vaccination outcomes, which may be difficult or impossible to ascertain using human data alone.

Authors

Kate Bridges, Denis Awany, Anele Gela, Temwa-Dango Mwambene, Sherry L. Kurtz, Richard E. Baker, Karen L. Elkins, Christopher M. Sassetti, Thomas J. Scriba, Douglas A. Lauffenburger

Palmitate impairs autophagic degradation via oxidative stress-perilysosomal Ca²⁺ overload-mTORC1 activation in pancreatic β-cells

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Saturated fatty acids impose lipotoxic stress on pancreatic β-cells, leading to β-cell failure and diabetes. In this study, we investigate the critical role of organellar Ca2+ disturbance on defective autophagy and β-cell lipotoxicity. Palmitate, a saturated fatty acid, induced perilysosomal Ca2+ elevation, sustained mTORC1 activation on the lysosomal membrane, suppression of the lysosomal transient receptor potential mucolipin 1 (TRPML1) channel, and accumulation of undigested autophagosomes in β-cells. These Ca2+ aberrations with autophagy defects by palmitate were prevented by an mTORC1 inhibitor or a mitochondrial superoxide scavenger. To alleviate perilysosomal Ca2+ overload, strategies such as lowering extracellular Ca2+, employing voltage-gated Ca2+ channel blocker or ATP-sensitive K+ channel opener effectively abrogated mTORC1 activation and preserved autophagy. Furthermore, redirecting perilysosomal Ca2+ into the endoplasmic reticulum (ER) with an ER Ca2+ ATPase activator, restores TRPML1 activity, promotes autophagic flux, and improves survival of β-cells exposed to palmitate-induced lipotoxicity. Our findings suggest oxidative stress-Ca2+ overload-mTORC1 pathway involvement in TRPML1 suppression and defective autophagy during β-cell lipotoxicity. Restoring perilysosomal Ca2+ homeostasis emerges as a promising therapeutic strategy for metabolic diseases.

Authors

Ha Thu Nguyen, Luong Dai Ly, Thuy Thi Thanh Ngo, Soo Kyung Lee, Carlos Noriega Polo, Subo Lee, Taesic Lee, Seung-Kuy Cha, Xaviera Riani Yasasilka, Kae Won Cho, Myung-Shik Lee, Andreas Wiederkehr, Claes B. Wollheim, Kyu-Sang Park

Central SELENOT deficiency impairs gonadotrope axis function, sexual behavior and fertility in male and female mice

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Reproductive disorders can result from a defective action of the neuropeptide gonadotropin-releasing hormone (GnRH), the master regulator of reproduction. We have previously shown that SELENOT, a newly-described thioredoxin-like selenoprotein highly expressed in endocrine and neuroendocrine cells, plays a role in hormone secretion and neuroprotection. However, whether SELENOT is involved in neuro-endocrine regulations in vivo is totally unknown. We found that SELENOT deficiency in the brain impaired sexual behavior, leading to a decline in fertility in both male and female mice. Biochemical and histological analyses of the gonadotrope axis of these mice revealed a higher expression of GnRH, which is associated with circulating luteinizing hormone (LH) excess, and elevated steroid hormones in males and a polycystic ovary syndrome (PCOS)-like phenotype in females. In addition, SELENOT deficiency impaired LH pulse secretion in both male and female mice. These alterations are reverted after administration of a GnRH antagonist. Together, our data demonstrate for the first time the role of a selenoprotein in the central control of sexual behavior and reproduction, and identify a new redox effector of GnRH neuron activity impacting both male and female reproductive function.

Authors

Ben Yamine Mallouki, Loubna Boukhzar, Ludovic Dumont, Azénor Abgrall, Marjorie Gras, Agathe Prieur, David Alexandre, David Godefroy, Yves Tillet, Luca Grumolato, Nathalie Rives, Fatiha Chigr, Youssef Anouar

A rebrand for proteasome inhibition in solid tumors via continuous hepatic artery infusion

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Authors

Carolina M. Larrain, Jack H. Victory, Priyanka P. Desai, Lindsay R. Friedman, Hannah Stepp, Rachel Ashe, Kirsten Remmert, Surajit Sinha, Emily C. Smith, Nicole Russell, Tracey Pu, Alyssa V. Eade, Justine F. Burke, Jason Ho, Michael B. Yaffe, David E. Kleiner, Keith Schmidt, William D. Figg, Jonathan M. Hernandez

H3K18 lactylation potentiates microglial polarization via the TLR4 pathway in diabetes-induced cognitive impairment

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The present study aims to explore the role and possible underlying mechanisms of histone lactylation modifications in diabetes-associated cognitive impairment (DACD). In this study, behavioral tests, Hematoxylin & Eosin (HE) staining, and immunohistochemistry were used to evaluate cognitive function and the extent of cerebral tissue injury. We quantified the levels of lactic acid and Pan-lysine lactylation (Pan Kla) in the brains of type 2 diabetes mellitus (T2DM) mice and in high glucose–treated microglia. We also identified all Kla sites in isolated microglia. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were subsequently conducted to identify the functions and pathways that were enriched at the differentially expressed modification sites. cleavage under targets and tagmentation (CUT&Tag) technology was used to identify candidate genes that are regulated by H3K18la. Small interfering RNA (siRNA) and H3K18R mutant sequences were used to knock down crucial components in key signaling pathways to assess the effects of histone lactylation on microglial polarization. We found that lactic acid levels were significantly greater in the brains of T2DM mice and high glucose-treated microglia than in those of their corresponding controls, which increased the level of Pan-Kla. We discovered that lactate can directly stimulate an increase in H3K18la. The global landscape of the lactylome reveals information about modification sites, indicating a correlation between the upregulation of H3K18la and protein lactylation and Toll-like receptor signaling. CUT&Tag demonstrated that enhanced H3K18la directly stimulates the nuclear factor kappa-B (NF-κB) signaling pathway by increasing binding to the promoter of Toll Like Receptor 4 (TLR4), thereby promoting M1 microglial polarization. The present study demonstrated that enhanced H3K18la directly stimulates TLR4 signaling to promote M1 microglial polarization, thereby facilitating DACD phenotypes. Targeting such loop may be a potential therapeutic approach for the treatment of DACD.

Authors

Ying Yang, Fei Chen, Lulu Song, Liping Yu, Jinping Zhang, Bo Zhang

Myosin inhibition partially rescues the myofibre proteome in X-linked myotubular myopathy

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X-linked myotubular myopathy (XLMTM) due to MTM1 mutations is a rare and often lethal congenital myopathy. Its downstream molecular and cellular mechanisms are currently incompletely understood. The most abundant protein in muscle, myosin, has been implicated in the pathophysiology of other congenital myopathies. Hence, in the present study, we aimed to define whether myosin is also dysfunctional in XLMTM and whether it thus may constitute a potential drug target. To this end, we used skeletal muscle tissue from human patients and canine/mouse models; we performed Mant-ATP chase experiments coupled with X-ray diffraction analyses and LC/MS-based proteomics studies. In XLMTM humans, we found that myosin molecules are structurally disordered and preferably adopt their ATP-consuming biochemical state. This phosphorylation-related (mal)adaptation was mirrored by a striking remodelling of the myofibre energetic proteome in XLMTM dogs. In line with these, we confirmed an accrued myosin ATP consumption in mice lacking MTM1. Hence, we treated these, with a myosin ATPase inhibitor, mavacamten. After a four-week treatment period, we observed a partial restoration of the myofibre proteome, especially proteins involved in cytoskeletal, sarcomeric and energetic pathways. Altogether, our study highlights myosin inhibition as a new potential drug mechanism for the complex XLMTM muscle phenotype.

Authors

Elise Gerlach Melhedegaard, Fanny Rostedt, Charlotte Gineste, Robert A.E. Seaborne, Hannah F. Dugdale, Vladimir Belhac, Edmar Zanoteli, Michael W. Lawlor, David L. Mack, Carina Wallgren-Pettersson, Anthony L. Hessel, Heinz Jungbluth, Jocelyn Laporte, Yoshihiko Saito, Ichizo Nishino, Julien Ochala, Jenni Laitila

ETS1 suppresses Hepatic Stellate Cell activation and liver fibrosis

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Chronic liver injury results in activation of quiescent Hepatic Stellate Cells (qHSCs) into Collagen Type I-producing activated HSCs that make liver fibrotic. We identified ETS1/2 (E26 transformation-specific transcription factors 1/2) as lineage-specific transcription factors regulating HSC phenotypes. Here we investigated the role of ETS1/2 in HSCs in liver fibrosis using toxic liver injury models and 3D human liver spheroids. Liver fibrosis was induced in wild-type and HSC-specific Ets1 (Ets1ΔHSC) and Ets2 (Ets2ΔHSC) knockout mice by administration of carbon tetrachloride for 6 weeks, following cessation of liver injury for 2 weeks. Liver fibrosis was more severe in Ets1ΔHSC, and to lesser extent in Ets2ΔHSC, compared to wild-type mice. Regression of liver fibrosis was suppressed only in Ets1ΔHSC, indicating Ets1 as the predominant isoform maintaining quiescent-like phenotype in HSCs. Similar results were obtained in a MASH model using 3D human liver spheroids. Knockdown of ETS1 in human HSCs caused upregulation of fibrogenic genes in MASH human liver spheroids and prevented fibrosis regression. ETS1 regulated the qHSC phenotype via CRTC2/PGC1α/PPARγ pathway. Knockdown of CRTC2 (cAMP response element-binding protein (CREB)-regulated transcription co-activator 2) abrogated PPARγ responses and facilitated HSC activation. These findings suggest that ETS1 may represent a therapeutic target for anti-fibrotic therapy.

Authors

Wonseok Lee, Xiao Liu, Sara Brin Rosenthal, Charlene Miciano, Sadatsugu Sakane, Kanani Hokutan, Debanjan Dhar, Hyun Young Kim, David A. Brenner, Tatiana Kisseleva

Chromosomal instability in circulating tumor cells and cabazitaxel resistance in metastatic castration-resistant prostate cancer

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BACKGROUND. Predictive biomarkers to guide chemotherapy decisions for metastatic castration resistant prostate cancer (mCRPC) are lacking. Preclinical studies indicate that circulating tumor cell (CTC) studies of chromosomal instability (CTC-CIN) can predict taxane resistance. METHODS. The CARD trial randomized subjects with mCRPC progressing within a year of treatment with an androgen receptor pathway inhibitor (ARPI; enzalutamide or abiraterone acetate plus prednisolone/prednisone) to cabazitaxel or the alternative ARPI. As a pre-planned biomarker analysis, CTCs were isolated from blood samples obtained at baseline; cycle two, and end of treatment. Associations between baseline CTC and CTC-CIN counts with imaging-based progression free survival (ibPFS), overall survival (OS), time to prostate-specific antigen (PSA) progression, RECIST 1.1 objective response rate (ORR), and PSA50 response rate (PRR) were assessed. RESULTS. High baseline CTC-CIN counts significantly associated with worse OS after adjustment for confounding variables (median OS, 15.3 vs 8.9 months; univariate HR, 2.16; 95% CI, 1.52 – 3.06; p < 0.001; multivariate HR, 1.56; 95% CI, 1.01 – 2.43; p = 0.047). Detectable CTC-CIN counts at baseline may predict a lack of ibPFS and OS benefit when comparing cabazitaxel to ARPI. CONCLUSION. This preplanned biomarker analysis of CARD confirms that CTC-CIN counts are a clinically useful prognostic and predictive biomarker of taxane resistance in mCRPC. Detectable CTC-CIN at baseline defines a patient subpopulation with unmet clinical needs in which alternative therapeutics should be tested. TRIAL REGISTRATION. CARD ClinicalTrials.gov number, NCT02485691. FUNDING. Funded by Sanofi and Epic Sciences.

Authors

Ossian Longoria, Jan Rekowski, Santosh Gupta, Nick Beije, Klaus Pantel, Eleni Efstathiou, Cora Sternberg, Daniel Castellano, Karim Fizazi, Bertrand Tombal, Adam Sharp, Oliver Sartor, Sandrine Macé, Christine Geffriaud-Ricouard, Richard Wenstrup, Ronald de Wit, Johann de Bono