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Abstract
Adeno-associated viruses (AAVs) have been used in gene therapy, especially for inherited retinal diseases. Despite their effectiveness in gene transduction, immune responses to the AAV capsid and transgene products have been reported, which can compromise both the efficacy and safety of AAV-mediated therapies. The eye is regarded as an immune-privileged organ where immune activity is constitutively suppressed. Here, we highlight that immunomonitoring in an ocular gene transfer reveals variable immune responses, whatever the species (human clinical trial, non-human primates, mice), the site of injection, the cassette, and the dose. We further explored factors contributing to this variability, investigating the potential correlation among immune parameters in a controlled experimental setting. In a syngeneic murine model after a subretinal injection of AAV, our results highlight an inter-individual variability of immune parameters, emphasizing the importance of considering inherent variability among individuals while designing personalized therapies.
Authors
Duohao REN, Gaelle A. CHAUVEAU, Julie VENDOMELE, Emilie CABON, Audrey PINEIRO, Catherine VIGNAL-CLERMONT, Hanadi SALIBA, Giuseppe RONZITTI, Anne GALY, Deniz DALKARA, Juliette PULMAN, Divya AIL, Sylvain FISSON
Abstract
Oxidative signaling is a central mechanism in alcohol-induced injury and has a strong implication in blood-brain barrier (BBB) dysregulation and neuroinflammation. Here, by targeting oxidative signaling, we hypothesized an innovative approach to develop a clinically relevant therapeutic strategy for alleviating alcohol-mediated neurovascular damage. To accomplish this, we enhanced the endogenous activity of Nrf2 (nuclear factor E2-related factor 2) by treating with an Nrf2 activator III TAT peptide (Nrf2 peptide, NP) and investigated the neuroprotective role of Nrf2 in promoting antioxidant defense properties and reducing BBB damage and transmigration of leukocytes to the brain following alcohol ingestion. We administered the NP subcutaneously to alcohol-ingested mice and evaluated its therapeutic potential in alleviating alcohol-associated neurovascular impairments. We compared the results with control peptide (random sequence with TAT)-treated animals. The studies showed that the NP treatment preserved the oxidant-antioxidant balance, downregulated ICAM-1 and its receptors, and mitigated BBB damage and leukocyte infiltration into the brain. We validated the effect of the NP in Nrf2 knock-out (KO) mice (Nrf2−/−). Thus, this study demonstrates NP’s neurovascular protective effects by regulating the oxidant-antioxidant balance, reducing oxidative stress-induced BBB disruption, and limiting transmigration of immune cells to the brain in a mouse model of alcohol ingestion.
Authors
Bibhuti B. Saikia, Saleena Alikunju, Yemin A. Poovanthodi, Zayan Kassim, P. M. Abdul Muneer
Abstract
Systemic sclerosis (SSc) is a complex and heterogeneous condition characterized by progressive fibrosis in multiple organs. Recent studies implicate plasminogen activator inhibitor 1 (PAI-1) in the pathogenesis of SSc, and PAI-1 is considered as a potential target for therapy. Here, using single-cell and spatial RNA-seq analysis of skin biopsies from 18 healthy individuals and 22 SSc patients, we found elevated PAI-1 co-localizing to myofibroblasts with enriched extracellular matrix-associated biological processes. Treatment of SSc dermal fibroblasts with the small molecule PAI-1 inhibitor MDI-2517 reduced the expression of the profibrotic markers COL1A1 and ACTA2. To investigate the therapeutic potential of MDI-2517, we evaluated its efficacy in reducing fibrosis in a preclinical model of SSc. Treatment of mice with MDI-2517 significantly reduced both skin and lung fibrosis and was superior to treatment with either pirfenidone or mycophenolate mofetil. Additionally, MDI-2517 attenuated weight loss and significantly reduced the expression of key profibrotic markers. Compared to tiplaxtinin, another PAI-1 inhibitor previously shown to be effective in a model of SSc, MDI-2517 was found to have superior efficacy at a 10-fold lower dose. These findings highlight the role of PAI-1 in the pathogenesis of SSc, and the potential of MDI-2517 for the treatment of SSc.
Authors
Enming J. Su, Pei-Suen Tsou, Mark Warnock, Natalya Subbotina, Kris Mann, Sirapa Vichaikul, Alyssa Rosek, Lisa Leung, Xianying Xing, Enze Xing, Olesya Plazyo, Rachael Bogle, Lam C. Tsoi, Cory D. Emal, Dinesh Khanna, John Varga, Thomas H. Sisson, Johann E. Gudjonsson, Daniel A. Lawrence
Abstract
Proper development of the umbilical cord and placental vasculature is essential for embryonic development. While the allantois is known give rise to endothelial cells (ECs) within the placenta, whether the allantois gives rise to ECs in the umbilical cord is debated. Furthermore, a lack of genetic tools to study placental vascular development independent of the embryo proper has hindered robust investigation into the primary cause of vascular defects from early studies utilizing global knockouts. In this study, we delineate the contribution of the allantois to the umbilical vessels and utilize a mouse genetic tool previously developed by our lab to revisit the role of Notch signaling during placental development. We show that the allantois has mosaic contribution to the umbilical endothelium with higher contributions closer to the placenta. Allantoic deletion of Dll4 disrupts umbilical cord and placental vascular formation with secondary defects in the heart. Lastly, we identify Unc5b downstream of Notch signaling that restricts EC migration while promoting chemokine signaling for smooth muscle cell (SMC) recruitment to arteries. These findings identify a genetic tool for investigating placental vascular development and give new insights into the ontogeny and mechanisms of placental vascular and umbilical cord development.
Authors
Derek C. Sung, Hana A. Ahanger, Sweta Narayan, Jesse A. Pace, Mei Chen, Jisheng Yang, Siqi Gao, T.C.S. Keller IV, Jenna Bockman, Xiaowen Chen, Erica Nguyen, Alan T. Tang, Patricia Mericko-Ishizuka, Ivan Maillard, Mark L. Kahn
Abstract
The contribution of 9p deletion to B-cell acute lymphoblastic leukemia (B-ALL) has remained elusive since its discovery more than 40 years ago. Here we show that loss of CD72 is recurrent in B-ALL cases containing PAX5 deletions, and that Cd72 haploinsufficiency drives B-ALL development in Pax5+/− mice. Mechanistically, Cd72+/-;Pax5+/- precursor B cells exhibit an inflammatory transcriptional profile characterized by a decrease in Myd88 expression, a finding that aligns with our previous studies of B-ALL development in Pax5+/- mice following exposure to immune stressors. These combined genomic analyses and functional models provide compelling evidence that co-deletion of two contiguous genes, Pax5 and Cd72, drives B-cell leukemogenesis.
Authors
Belén Ruiz-Corzo, Ana Casado-García, Ninad Oak, Paula Somoza-Cotillas, Andrea López-Álvarez de Neyra, Jorge Martínez-Cano, Alba Pérez-Pons, Elena G. Sánchez, Oscar Blanco, Diego Alonso-López, Javier De Las Rivas, Susana Riesco, Pablo Prieto-Matos, Francisco Javier Garcia-Criado, Maria Begoña Garcia-Cenador, Alberto Orfao, Manuel Ramírez-Orellana, César Cobaleda, Carolina Vicente-Dueñas, Kim E. Nichols, Isidro Sánchez-García
Abstract
Degradation of cellular waste from phagocytosis, endocytosis and autophagy occurs through hydrolases that become activated during acidification of late endosomes and lysosomes (LELs). In a cross-sectional study we show diminished LEL acidification and the accumulation of surface-bound nucleosome on monocytes, dendritic cells, and B cells from SLE patients. Diminished acidification and exocytosis of undegraded IgG-ICs is evident in active, but not inactive disease. This is supported by our murine study where LEL acidification is diminished, promoting exocytosis and the accumulation of cell surface IgG-immune complexes. Mechanistically, LEL dysfunction is induced by chronic PI3k activation in lupus-prone MRL/lpr mice. We also show that on a non-autoimmune C57BL/6 background, deficiency in SHP-1 and inhibition of SHIP-1 activity is sufficient to recapitulate LEL dysfunction found in MRL/lpr mice. Non-acidic LELs are evident in 67% of patients, and associate with SLEDAI arthritis, rash, and nephritis. The high frequency of LEL dysfunction in SLE suggests it could serve as a biomarker identifying a specific disease endotype.
Authors
SunAh Kang, Andrew J. Monteith, Liubov Arbeeva, Karissa Grier, Shruti Saxena Beem, Anthony C. Trujillo, Xinyun Bi, Kai Sun, Rebecca E. Sadun, Mithu Maheswaranathan, Megan E.B. Clowse, Saira Z. Sheikh, Jennifer L. Rogers, Barbara J. Vilen
Abstract
Sarcomas are a heterogeneous group of cancers with few shared therapeutic targets. We show that PI3K signaling is frequently activated in sarcomas due to PTEN loss (in 30-60%), representing a common therapeutic target. The PI3K pathway has lacked a downstream oncogenic transcription factor. We show TAZ and YAP are transcriptional co-activators regulated by PI3K and drive a transcriptome necessary for tumor growth in a PI3K-driven sarcoma mouse model. This PI3K-TAZ/YAP axis exists in parallel to the known PI3K-Akt-mTORC1 axis providing a rationale for combination therapy targeting the TAZ/YAP-TEAD interaction and mTORC1. Combination therapy using IK-930 (TEAD inhibitor) and everolimus (mTORC1 inhibitor) synergistically diminished proliferation and anchorage dependent growth of PI3K-activated sarcoma cell lines at low, physiologically achievable doses. Furthermore, this combination therapy showed a synergistic effect in vivo, suggesting that an integrated view of PI3K and Hippo signaling can be leveraged therapeutically in PI3K activated sarcomas.
Authors
Keith C. Garcia, Ali A. Khan, Krishnendu Ghosh, Souradip Sinha, Nicholas Scalora, Gillian DeWane, Colleen Fullenkamp, Nicole Merritt, Yuliia Drebot, Samuel Y. Yu, Mariah Leidinger, Michael D. Henry, Patrick J. Breheny, Michael S. Chimenti, Munir R. Tanas
Abstract
Hirschsprung disease (HSCR) is a congenital intestinal disorder characterized by the absence of ganglia in the distal intestine. Despite surgical resection of the aganglionic intestine and pull-through surgery, HSCR patients still experience bowel dysfunction, indicating that latent abnormalities may also exist in the proximal ganglionic intestine. To elucidate possible causes of postoperative bowel dysfunction in HSCR, we investigated differences in the proximal ganglionic intestine using an animal model of HSCR (Ednrb-null mice) and validated our findings in tissue from human HSCR patients. We found that the proximal ganglionic colon of HSCR mice exhibited greater stiffness and fibrosis than their wild-type littermates. Similarly, submucosal fibrosis was significantly greater in the proximal ganglionic intestine of HSCR patients than in intestinal tissue from age and site-matched controls. Furthermore, we observed dysregulated expression of extracellular matrix (ECM)-related genes in the proximal ganglionic intestine of HSCR mice compared to controls. We conclude that increased fibrosis, stiffness, and alterations in ECM composition may contribute to persistent dysfunction of the ganglionic intestine in HSCR. These findings add to the growing body of literature that describe abnormalities in the proximal ganglionic intestine of HSCR and suggest that HSCR is not limited to the aganglionic intestine alone.
Authors
Prisca C. Obidike, Britney A. Hsu, Chioma Moneme, Oluyinka O. Olutoye II, Walker D. Short, Mary Hui Li, Swathi Balaji, Yuwen Zhang, Sundeep G. Keswani, Lily S. Cheng
Abstract
Dysfunctional white adipose tissue contributes to the development of obesity-related morbidities, including insulin resistance, dyslipidemia, and other metabolic disorders. Adipose tissue macrophages (ATMs) accumulate in obesity and play both beneficial and harmful roles in the maintenance of adipose tissue homeostasis and function. Despite their importance, the molecules and mechanisms that regulate these diverse functions are not well understood. Lipid-associated macrophages (LAMs), the dominant subset of obesity-associated ATMs, accumulate in crown-like structures and are characterized by a metabolically activated and proinflammatory phenotype. We previously identified CD9 as a surface marker of LAMs. However, the contribution of CD9 to the activation and function of LAMs during obesity is unknown. Using a myeloid-specific CD9 knockout model, we show that CD9 supports ATM-adipocyte adhesion and crown-like structure formation. Furthermore, CD9 promotes the expression of pro-fibrotic and extracellular matrix remodeling genes. Loss of myeloid CD9 reduces adipose tissue fibrosis, increases visceral adipose tissue accumulation, and improves global metabolic outcomes during diet-induced obesity. These results identify CD9 as a causal regulator of pathogenic LAM functions, highlighting CD9 as a potential therapeutic target for treating obesity-associated metabolic disease.
Authors
Julia Chini, Nicole DeMarco, Dana V. Mitchell, Sam J. McCright, Kaitlyn M. Shen, Divyansi Pandey, Rachel L. Clement, Jessica Miller, Rajan Jain, Deanne M. Taylor, Mitchell A. Lazar, David A. Hill
Abstract
Latently infected cells persist in people living with HIV (PWH) despite suppressive antiretroviral therapy (ART) and evade immune clearance. Shock and Kill cure strategies are hampered by insufficient enhancement of targeted immune responses following latency reversal. We previously demonstrated autologous Vδ2 T cells from PWH retain anti-HIV activity and can reduce CD4+ T cell reservoirs, although their use in cure approaches is limited due to their dual role as a viral reservoir. However, promising clinical data in oncology shows their unique MHC- unrestricted antigen recognition affords potent on-target cytotoxicity in the absence of graft-versus-host disease when used as an allogeneic adoptive cell therapy modality. Here, we found expanded allogeneic Vδ2 T cells specifically eliminated HIV-infected CD4+ T cells and monocyte-derived macrophages (MDM), overcoming inherent resistance to killing by other cell types such as NK and CD8+ T cells. Notably, we demonstrated allogeneic Vδ2 T cells recognized and eliminated the HIV-latent CD4+ T cell reservoir following latency reversal. Our study provides evidence for developing an allogeneic γδ T cell therapy for HIV cure and warrants pre-clinical investigation in combination approaches.
Authors
Brendan T. Mann, Marta Sanz, Alisha Chitrakar, Kayley Langlands, Marc Siegel, Natalia Soriano-Sarabia
Abstract
Infectious diseases remain a global health challenge, driven by increasing antimicrobial-resistance and the threat of emerging epidemics. Mycobacterium tuberculosis and Staphylococcus aureus are leading causes of mortality worldwide. Trained immunity—a form of innate immune memory—offers a promising approach to enhance pathogen clearance. Here, we demonstrate that IFN-γ induces trained immunity in human monocytes through a mechanism involving mTORC1 activation, glutaminolysis, and epigenetic remodeling. Macrophages derived from IFN-γ–trained monocytes exhibited increased glycolytic activity with enhanced cytokine and chemokine responses upon stimulation or infection. Crucially, trained macrophages had increased production of reactive oxygen species which mediated enhanced bactericidal activity against methicillin-resistant S. aureus. Furthermore, ATAC-sequencing analysis of IFN-γ trained macrophages revealed increased chromatin accessibility in regions associated with host defence. Lastly, IFN-γ training restored impaired innate responses in macrophages from individuals homozygous for the TIRAP 180L polymorphism, a genetic variant associated with increased susceptibility to infection. These findings establish IFN-γ as a potent inducer of trained immunity in human monocytes and support its potential as a host-directed strategy to strengthen antimicrobial defenses, particularly in genetically susceptible individuals and high-risk clinical contexts.
Authors
Dearbhla M. Murphy, Isabella Batten, Aoife O'Farrell, Simon R. Carlile, Sinead A. O'Rourke, Chloe Court, Brenda Morris, Gina Leisching, Gráinne Jameson, Sarah A. Connolly, Adam H. Dyer, John P. McGrath, Emma McNally, Olivia Sandby-Thomas, Anjali Yennemadi, Conor M. Finlay, Clíona Ni Cheallaigh, Jean Dunne, Cilian Ó Maoldomhnaigh, Laura E. Gleeson, Aisling Dunne, Nollaig Bourke, Reinout van Crevel, Donal J. Cox, Niall Conlon, Arjun Raj, Rachel M. McLoughlin, Joseph Keane, Sharee A. Basdeo
Abstract
Nicotinamide adenine dinucleotide (NAD⁺) is essential for cellular metabolism, DNA repair, and stress responses. NAD+ is synthesized from nicotinamide, nicotinic acid (collectively termed niacin), and tryptophan. In humans, deficiencies in these nutrients result in pellagra, marked by dermatitis, diarrhea, and dementia. The dermatitis associated with pellagra typically manifests as photodermatosis in sun-exposed areas. This study examined the effects of NAD+ deficiency on skin homeostasis using epidermis-specific Nampt conditional knockout (cKO) mice. These mice displayed substantial NAD⁺ depletion, reduced poly(ADP-ribose) polymerase (PARP) activity, and increased DNA damage. Consequently, Nampt cKO mice developed spontaneous skin inflammation and epidermal hyperplasia. RNA sequencing and immunohistochemical analyses demonstrated increased interleukin-36 (IL-36) cytokine expression, suggesting that DNA repair-related genomic stress triggers keratinocyte-driven IL-36 production, which promotes inflammation. Furthermore, reduced collagen17A1 expression and elevated thymic stromal lymphopoietin (TSLP) levels were observed. NAD+ repletion by transdermal supplementation of nicotinamide mononucleotide (NMN) suppressed the rise of IL-36 levels and skin inflammation. These findings underscore the importance of Nampt-mediated NAD⁺ metabolism for epidermal stability and indicate that NAD⁺ depletion may contribute to IL-36-mediated skin inflammation, offering insights for therapeutic strategies in inflammatory skin disorders.
Authors
Taiki Seki, Jun-Dal Kim, Yasuhito Yahara, Hitoshi Uchida, Keisuke Yaku, Mariam Karim, Teruhiko Makino, Tadamichi Shimizu, Takashi Nakagawa
Abstract
Adaptive remodeling of retrodiscal tissue following anterior disc displacement (ADD) of the temporomandibular joint (TMJ) has been recognized for decades, yet the underlying cellular dynamics and molecular mechanisms remain unclear. Using a porcine ADD model, this study investigated the cellular and molecular basis driving retrodiscal tissue adaptation. Histological staining revealed adaptive remodeling of retrodiscal tissue after ADD induction, with dense connective tissue and cartilaginous masses replacing loose connective tissue. Furthermore, single-cell RNA sequencing (scRNA-seq) captured pronounced fibroblast expansion during tissue remodeling, notably the FB2 subcluster with high developmental potential, and the emergence of a mural cell subcluster MC4 associated with extracellular matrix (ECM) remodeling. CellChat analysis highlighted MC4-FB2 crosstalk via FGF2 and BMP5 signaling. The combination of pathway-aware multi-layered hierarchical network (P-NET) and Seurat with drug database screening identified five promising compounds. Among them, Zaprinast demonstrated the most robust effects by enhancing the remodeling capability of fibroblasts in vitro, and also alleviated TMJ deformation in vivo. Collectively, fibroblast activation is pivotal for early retrodiscal tissue adaptation following ADD, which is driven by MC4-derived FGF2/BMP5 signaling. Zaprinast treatment potentiates this remodeling process. These findings provide new insights into cellular basis of TMJ adaptation and identify potential therapeutic targets for ADD management.
Authors
Wenlin Yuan, Yilin Chen, Ruojin Yan, Wei Liu, Chenyu Wang, Ying Wang, Qiaoli Dai, Wen Li, Mengqi Zhu, Xiao Chen, Jiejun Shi
Abstract
The chronic inflammation of Crohn’s disease frequently leads to fibrosis and muscular hypertrophy of the intestinal wall. This often culminates in strictures, a serious condition lacking directed therapy. Severe pathological changes occur in the submucosa and muscularis propria intestinal wall layers of strictures, yet stricture-associated proteome changes in these layers is unexplored. We perform unbiased proteomics on submucosa and muscularis propria microdissected from transmural sections of strictured and non-strictured ileum. Proteome changes in stricture submucosa reflect a transition from homeostasis to tissue remodeling, inflammation and smooth muscle alterations. Top submucosa features include reduced vascular components and lipid metabolism proteins accompanied by increased proteins with immune-, matrix- or stress functions including CTHRC1, TNC, IL16, MZB1 and TXNDC5. In parallel, predominant changes in stricture muscularis propria include increased matrix (POSTN) and immune (mast cell CPA3) proteins alongside decreased proteins with lipid metabolic, mitochondrial or key muscle functions. Finally, trends of differentially expressed proteins along non-stricture submucosa suggest progressive profibrotic tissue remodeling and muscle expansion as proximity to stricture increases. The comprehensive proteome map presented here offers unique layer-resolved insight into the stricture microenvironment and potential drivers of fibrotic disease, providing a valuable resource to fuel biomarker and therapeutic target research. Keywords: Crohn’s disease strictures, proteomics, intestinal fibrosis, fibrostenosis
Authors
Johannes Alfredsson, Carina Sihlbom Wallem, Maja Östling, Hanna de la Croix, Elinor Bexe-Lindskog, Mary Jo Wick
Abstract
Cardiovascular disease (CVD), both atherosclerosis-related and heart failure with preserved ejection fraction (HFpEF) linked to cardiac fibrosis, contributes to morbidity and mortality in people with HIV (PWH) receiving antiretroviral therapy (ART). In the REPRIEVE trial, pitavastatin reduces atherosclerotic CVD risk to a magnitude inconsistent with pitavastatin’s impact solely on LDL-cholesterol and inflammation. We hypothesized that HFpEF in PWH is related to HIV-induced fibrosis mediated by platelet TGFβ1, is accelerated by certain contemporary ART, and may also be inhibited by statins. ART drugs used in REPRIEVE, including a nucleoside/nucleotide, integrase inhibitor-based regimen (tenofovir (TDF), emtricitabine (FTC), and dolutegravir (DTG)), and the protease inhibitors ritonavir (RTV) and darunavir (DRV), and the impact of atorvastatin, were examined in two HIV mouse models: transgenic HIV-Tg26 mice and HIV-PDX mice engrafted with T cells isolated from PWH. HIV-Tg26 and HIV-PDX mice had higher cardiac fibrosis than littermate controls without HIV (p<0.05). Administration of TDF-FTC-DTG or RTV, but not DRV, resulted in a further ~2-fold increase in fibrosis (p<0.01). Cardiac fibrosis and intracardiac fat accumulation correlated with reduced diastolic function. Mice depleted of platelet TGFβ1 (TGFβ1Platelet-ΔTg26), or treated with atorvastatin, were partially protected from HIV- and ART-induced cardiac fibrosis, steatosis, and diastolic dysfunction. Atorvastatin effects were independent of changes in inflammatory cytokines. These effects correlated with reduced platelet activation and TGFβ signaling in cardiac endothelial cells, fibroblasts, and macrophages undergoing mesenchymal transition. Our results indicate that certain ART regimens accelerate HIV-associated CVD characterized by HFpEF via platelet TGFβ1-dependent processes, which were mitigated by atorvastatin. We postulate that our findings provide a potential mechanism for the pleiotropic effects of statins in HIV/ART-linked CVD which could be targeted by antiplatelet agents or inhibition of TGFβ signaling.
Authors
Kumar Subramani, Denys Babii, Brienne Cole, Tayyab A. Afzal, Thamizhiniyan Venkatesan, Trevor Word, Sandra Gostynska, Sixia Chen, Kar-Ming Fung, Ali Danesh, Itzayana G. Miller, Paul Klotman, Brad R. Jones, Jeffrey Laurence, Jasimuddin Ahamed
Abstract
Mutation of KRAS in endothelial cells (KRAS-EC) leads to intracerebral hemorrhage (ICH) in brain arteriovenous malformations (bAVM), resulting in severe disabilities or even death. However, it is unclear what causes this hemorrhagic conversion of bAVM. Here, using a locally established, clinically-relevant sporadic bAVM mouse model, created by overexpressing mutant KRAS (KRASG12V) in the brain EC, we demonstrate that KRAS-EC act as trigger for microglia (MG) activation and infiltration of macrophages (Mϕ). Using three-dimensional immunostaining approach with cleared human and mouse bAVM tissues, we demonstrate an abundance of MG/Mϕ around the bAVM nidus. The presence of MG/Mϕ are correlated to the blood-brain barrier leakage in bAVM area. Time-lapsed intravital imaging in Cx3cr1-gfp;Ccr2-rfp reporter mice demonstrate the dynamic activation of MG and infiltration of Mϕ toward mutant KRAS-modified dysplastic vessels. Importantly, a time course analysis showed that these activated/infiltrated MG/Mϕ are present around the bAVMs prior to hemorrhagic conversion, and controlled depletion of MG/Mϕ reduced ICH incidence in bAVM. Inhibition of MG/Mϕ with long-term minocycline treatment attenuated the incidence of ICHs around bAVMs. Our study indicates that MG/Mϕ are involved in destabilization of KRAS-induced bAVM, leading to hemorrhagic conversion/ICH. Thus, modulation of MG/Mϕ may reduce ICH risk in bAVM patients.
Authors
Hyejin Park, Jung-Eun Park, Bridger H. Freeman, Bosco Seong Kyu Yang, Shun-Ming Ting, Alexander K. Suh, Jude P.J. Savarraj, Shuning Huang, Jakob Körbelin, Huimahn Alex Choi, Sean P. Marrelli, Jaroslaw Aronowski, Peng Roc Chen, Eunhee Kim, Eun S. Park
Abstract
Angelman syndrome (AS) is a neurodevelopmental disorder caused by loss of the maternal UBE3A allele, the sole source of UBE3A in mature neurons due to epigenetic silencing of the paternal allele. Although emerging therapies are being developed to restore UBE3A expression by activating the dormant paternal UBE3A allele, existing mouse models for such preclinical studies have limited throughput and utility, creating bottlenecks for both in vitro therapeutic screening and in vivo characterization. To address this, we developed the Ube3a-INSG dual-reporter knock-in mouse, in which an IRES-Nanoluciferase-T2A-Sun1-sfGFP (INSG) cassette was inserted downstream of the endogenous Ube3a stop codon. The INSG model preserves UBE3A protein levels and function while enabling two complementary allele-specific readouts: Sun1-sfGFP and Nanoluciferase. We show that Sun1-sfGFP, a nuclear envelope-localized reporter, enables single-cell fluorescence analysis, whole-brain light-sheet imaging, and nuclear quantification by flow cytometry. Further, Nanoluciferase supports high-throughput luminescence assays for sensitive pharmacological profiling in cultured neurons and non-invasive in vivo bioluminescence imaging for pharmacodynamic assessment. By combining scalable screening, cellular analysis, and real-time in vivo monitoring in a single model, the Ube3a-INSG dual-reporter mouse provides a powerful platform to accelerate therapeutic development centered on UBE3A.
Authors
Hanna Vihma, Lucas M. James, Hannah C. Nourie, Audrey L. Smith, Siyuan Liang, Carlee A. Friar, Tasmai Vulli, Lei Xing, Dale O. Cowley, Alain C. Burette, Benjamin D. Philpot
Abstract
Immunosuppression and metastasis are critical hallmarks of breast cancer, often linked to poor patient outcomes. The secreted cytokine chitinase-3 like 1 (CHI3L11) is frequently overexpressed in breast cancer samples and promotes an immunosuppressed tumor microenvironment. Notably, CHI3L1 expression is elevated in metastatic patient samples when compared to the matched primary breast tumor. To investigate its role in breast cancer metastasis, we generated an inducible Genetically Engineered Mouse Model (GEMM) that overexpresses CHI3L1 in the mammary epithelium. Ectopic expression of CHI3L1 in the Polyomavirus Middle T (PyMT) mouse model of breast cancer suppressed anti-tumor immune responses, accelerated mammary tumor onset and enhanced lung metastasis. Mechanistically, elevated CHI3L1 expression in the mammary epithelium enhanced neutrophil recruitment, which subsequently degraded the extracellular matrix and increased the number of circulating tumor cells. These findings reveal a key mechanism driving metastatic dissemination and argue that therapeutically targeting Chi3l1 could enhance anti-tumor immunity and suppress metastasis.
Authors
Tarek Taifour, Adéline Massé, Yu Gu, Virginie Sanguin-Gendreau, Dongmei Zuo, Bin Xiao, Emilie Solymoss, Yunyun Shen, Hailey Proud, Sherif Samer Attalla, Vasilios Papavasiliou, Nancy U. Lin, Melissa E. Hughes, Kalie Smith, Chun Geun Lee, Suchitra Kamle, Josie Ursini-Siegel, Jack A. Elias, Peter M. Siegel, Rinath Jeselsohn, William J. Muller
Abstract
Authors
Rimas V. Lukas, Ruochen Du, Harrshavasan Congivaram, Kathleen McCortney, Karan Dixit, Craig Horbinski, Margaret Schwartz, Raymond Lezon, Lauren Singer, Ditte Primdahl, Jigisha Thakkar, Amy B. Heimberger, Roger Stupp, Priya Kumthekar
Potentiation of fentanyl-induced respiratory depression by alcohol is not fully reversed by naloxone
Abstract
The high frequency of opioid overdose deaths often involves co-use of alcohol, which is reported in approximately 30% of fentanyl fatalities. Both substances depress respiratory function, and their combined effects can be lethal. The present study investigated physiological parameters of respiratory-depressant effects of fentanyl when co-administered with alcohol and its sensitivity to naloxone reversal using whole-body plethysmography in male and female Long-Evans rats. Administration of a high, sedative-like dose of alcohol alone or fentanyl alone resulted in no mortality, but fentanyl+alcohol led to mortality rates of 42% and 33% in females and males, respectively. The fentanyl+alcohol combination reduced minute ventilation and increased apneic pauses compared with either drug alone. Lower, binge-like alcohol doses, when combined with fentanyl, also amplified respiratory depression. Pretreatment with naloxone did not fully restore normal respiration. Naloxone administered after fentanyl+alcohol transiently reversed the decrease in minute ventilation but did not reverse apneic pauses. Fentanyl-dependent rats were partially tolerant to fentanyl- and fentanyl+alcohol-induced respiratory depression, but alcohol-dependent rats exhibited sensitization to alcohol- and fentanyl+alcohol-induced apnea. These findings highlight physiological parameters of severe respiratory risks with fentanyl+alcohol co-use, which are inadequately reversed by naloxone, underscoring the need for targeted strategies to manage opioid+alcohol overdoses.
Authors
Emma V. Frye, Lyndsay E. Hastings, Aniah N. Matthews, Adriana Gregory-Flores, Janaina C.M. Vendruscolo, Lindsay A. Kryszak, Shelley N Jackson, Aidan J. Hampson, Nora D. Volkow, Leandro F. Vendruscolo, Renata C.N. Marchette, George F. Koob
