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Abstract
Resistance to chemotherapy of pancreatic ductal adenocarcinoma (PDAC) is largely driven by intratumoral heterogeneity (ITH) due to tumor cell plasticity and clonal diversity. In order to develop novel strategies to overcome this defined mechanism of resistance, tools to monitor and quantify ITH in a rapid and scalable fashion are needed urgently. Here, we employed label-free digital holographic microscopy (DHM) to characterize ITH in PDAC. We established a robust experimental and machine learning analysis pipeline to perform single cell phenotyping based on DHM-derived phase images of PDAC cells in suspension. Importantly, we are able to detect dynamic changes in tumor cell differentiation and heterogeneity of distinct PDAC subtypes upon induction of epithelial-to-mesenchymal transition and under treatment-imposed pressure in murine and patient-derived model systems. This platform allows us to assess phenotypic ITH in PDAC on a single cell level in real-time. Implementing this technology into the clinical workflow has the potential to fundamentally increase our understanding of tumor heterogeneity during evolution and treatment response.
Authors
Katja Wittenzellner, Manuel Lengl, Stefan Röhrl, Carlo Maurer, Christian Klenk, Aristeidis Papargyriou, Laura Schmidleitner, Nicole Kabella, Akul Shastri, David E. Fresacher, Farid Harb, Nawal Hafez, Stefanie Bärthel, Daniele Lucarelli, Carmen Escorial-Iriarte, Felix Orben, Rupert Öllinger, Ellen Emken, Lisa Fricke, Joanna Madej, Patrick Wustrow, I. Ekin Demir, Helmut Friess, Tobias Lahmer, Roland M. Schmid, Roland Rad, Günter Schneider, Bernhard Kuster, Dieter Saur, Oliver Hayden, Klaus Diepold, Maximilian Reichert
Abstract
Single dose radiotherapy (SDRT) is a highly-curative modality that may transform radiotherapy practice. Unfortunately, only ~50% of oligometastatic lesions are SDRT treatable due to adjacent radiosensitive normal organs at risk. Here we address extent to which an anti-angiogenic drug, VEGFR2-antagonist DC101, radiosensitizes SDRT using murine MCA/129 fibrosarcomas and Lewis Lung Carcinomas, which display a dose range for SDRT lesional eradication virtually identical to that employed clinically (10-30Gy). SDRT induces unique tumor cure, stimulating rapid endothelial acid sphingomyelinase (ASMase)/ceramide signaling that yields marked vasoconstriction and perfusion defects in tumor xenografts and human oligometastases. Ensuing tumor parenchymal oxidative damage initiates a SUMO Stress Response (SSR), which inactivates multiple homologous recombination repair enzymes, radiosensitizing all tumor types. While VEGF inhibits neo-angiogenic ASMase, optimal radiosensitization occurs only upon anti-angiogenic drug delivery at ~1h preceding SDRT. Obeying these principles, we find DC101 radiosensitizes SSR, DNA double strand break unrepair and tumor cure by 4-8Gy at all clinically-relevant doses. Critically, DC101 fails to sensitize small intestinal endothelial injury or lethality from the gastrointestinal-acute radiation syndrome.
Authors
Jin Cheng, Liyang Zhao, Sahra Bodo, Prashanth K. B. Nagesh, Rajvir Singh, Adam O. Michel, Regina Feldman, Zhigang Zhang, Simon N. Powell, Zvi Fuks, Richard Kolesnick
Abstract
There is an emerging role for Stimulator of interferon genes (STING) signaling in pulmonary hypertension (PH) development. Related, prior resesarch has demonstrated the relevance of the immune checkpoint protein Programmed death ligand 1 (PD-L1) expression by immunoregulatory myeloid cells in PH. However, there remains a need to elucidate the cell-specific role of STING expression, and the STING/PD-L1 signaling axis in PH, before readily available disease-modifying therapies can be applied to patients with disease. Here, through generation of bone marrow chimeric mice, we show that STING-/- mice receiving wild-type (WT) bone marrow are protected against PH secondary to chronic hypoxia. We further demonstrate a cellular dichotomous role for STING in PH development with STING expression by smooth muscle cells contributing to PH, and its activation on myeloid cells being pivotal in severe disease prevention. Finally, we provide evidence that a STING-PD-L1 axis modulates disease severity, suggesting future potential therapeutic applications. Overall, these data provide concrete evidence of STING involvement in PH in a cell-specific manner, establishing biologic plausibility for cell-targeted STING-related therapies in PH treatment.
Authors
Ann T. Pham, Shiza Virk, Aline C. Oliveira, Matthew D. Alves, Chunhua Fu, Yutao Zhang, Jimena Alvarez-Castanon, Brian B. Lee, Keira L. Lee, Radwan Mashina, Katherine E. Ray, Patrick Donabedian, Elnaz Ebrahimi, Harsh Patel, Reeha Patel, Duncan Lewis, Zhiguang Huo, Harry Karmouty-Quintana, Li Chen, Lei Jin, Andrew J. Bryant
Abstract
CD16 is an activating Fc receptor on natural killer cells that mediates antibody-dependent cellular cytotoxicity (ADCC), a key mechanism in antiviral immunity. However, the role of NK cell-mediated ADCC in SARS-CoV-2 infection remains unclear, particularly whether it limits viral spread and disease severity or contributes to the immunopathogenesis of COVID-19. We hypothesized that the high-affinity CD16AV176 polymorphism influences these outcomes. Using a novel in vitro reporter system, we demonstrated that CD16AV176 is a more potent and sensitive activator than the common CD16AF176 allele. To assess its clinical relevance, we analyzed 1,027 hospitalized COVID-19 patients from the Immunophenotyping Assessment in a COVID-19 Cohort (IMPACC), a comprehensive longitudinal dataset with extensive transcriptomic, proteomic, and clinical data. The high-affinity CD16AV176 allele was associated with a significantly reduced risk of ICU admission, mechanical ventilation, and severe disease trajectories. Lower anti-SARS-CoV-2 IgG titers were correlated to CD16AV176; however, there was no difference in viral load across CD16 genotypes. Proteomic analysis revealed that participants homozygous for CD16AV176 had lower levels of inflammatory mediators. These findings suggest that CD16AV176 enhances early NK cell-mediated immune responses, limiting severe respiratory complications in COVID-19. This study identifies a protective genetic factor against severe COVID-19, informing future host-directed therapeutic strategies.
Authors
Anita E. Qualls, Tasha Tsao, Irene Lui, Shion A. Lim, Yapeng Su, Ernie Chen, Dylan Duchen, Holden T. Maecker, Seunghee Kim-Schulze, Ruth R. Montgomery, Florian Krammer, Charles R. Langelier, Ofer Levy, Lindsey R. Baden, Esther Melamed, Lauren I.R. Ehrlich, Grace A. McComsey, Rafick P. Sekaly, Charles B. Cairns, Elias K. Haddad, Albert C. Shaw, David A. Hafler, David B. Corry, Farrah Kheradmand, Mark A. Atkinson, Scott C. Brakenridge, Nelson I. Agudelo Higuita, Jordan P. Metcalf, Catherine L. Hough, William B. Messer, Bali Pulendran, Kari C. Nadeau, Mark M. Davis, Ana Fernandez-Sesma, Viviana Simon, Monica Kraft, Christian Bime, Carolyn S. Calfee, David J. Erle, Joanna Schaenmann, Al Ozonoff, Bjoern Peters, Steven H. Kleinstein, Alison D. Augustine, Joann Diray-Arce, Patrice M. Becker, Nadine Rouphael, IMPACC Network, Jason D. Goldman, Daniel R. Calabrese, James R. Heath, James A. Wells, Elaine F. Reed, Lewis L. Lanier, Harry Pickering, Oscar A. Aguilar
Abstract
Diabetic kidney disease (DKD) is the leading cause of end stage kidney disease. Kidney tubular cells have a high energy demand, dependent on fatty acid oxidation (FAO). Although carnitine is indispensable for FAO, the pathological role of carnitine deficiency in DKD is not fully understood. We showed here that ectopic lipid accumulation due to impaired FAO increased in patients with DKD and inversely correlated with renal function. OCTN2 deficient mice exhibited systemic carnitine deficiency with increased renal lipid accumulation. Cell death and inflammation were induced in OCTN2-deficient, but not wild-type tubular cells exposed to high salt and high glucose. Compared with SDT fatty rats, uninephrectomized SDT fatty rats fed with 0.3% NaCl had higher lipid accumulation and exhibited increased urinary albumin excretion with renal dysfunction and tubulointerstitial injury, all of which were ameliorated by L-carnitine supplementation via stimulating FAO and mitochondrial biogenesis. In our single-center randomized control trial with patients undergoing peritoneal dialysis, L-carnitine supplementation preserved residual renal function and increased urine volume, the latter of which was correlated with improvement of tubular injury. The present study demonstrates the pathological role of impairment of carnitine-induced FAO in DKD, suggesting that L-carnitine supplementation is a potent therapeutic strategy for this devastating disorder.
Authors
Sakuya Ito, Kensei Taguchi, Goh Kodama, Saori Kubo, Tomofumi Moriyama, Yuya Yamashita, Yunosuke Yokota, Yosuke Nakayama, Yusuke Kaida, Masami Shinohara, Kyoko Tashiro, Keisuke Ohta, Sho-ichi Yamagishi, Kei Fukami
Abstract
Community-acquired infectious meningoencephalitis is associated with high rates of mortality and morbidity, compounded by limited access to diagnostic resources. The current study assessed acute central nervous system (CNS) infections in patients with meningoencephalitis enrolled in a hospital-based diagnostic surveillance study in São Paulo, Brazil. Cerebrospinal fluid (CSF) was collected from 600 subjects between March 2018 and November 2019 and initially screened for a broad range of pathogens according to a local diagnostic algorithm. Standard microbiological and molecular diagnostic methods were applied. Metagenomic sequencing was used as a complementary approach to investigating etiology in cases where no pathogen was initially identified. Standard testing identified infectious etiologies in 292 cases (48.6%), with 227 (77.7%) confirmed as viral infections, predominantly caused by enteroviruses (n=144) and herpesviruses (n=40). Non-viral agents were identified in 65 cases (22.3%). Metagenomic sequencing (mNGS) of 279 out of 308 undiagnosed cases revealed several additional potential etiologies, including Parvovirus B19, Toxoplasma gondii, Picobirnavirus, other enterovirus species and Vesivirus, the latter being associated with CNS infection for the first time. These findings underscore the complexity of CNS infections and highlight the potential of metagenomics to improve diagnostic accuracy, inform treatment strategies, and support efforts to address future pandemics.
Authors
Noely Evangelista Ferreira, Michael G. Berg, Antonio C. da Costa, Mary A. Rodgers, Esper G. Kallas, Cassia G. Terrasani Silveira, Mateus Vailant Thomazella, Ana Carolina Soares de Oliveira, Layla Honorato, Heuder G.O. Paião, Renan Barros Domingues, Carlos Senne, Marina F. Côrtes, Tania R. Tozetto-Mendoza, Hélio R. Gomes, Maria Laura Mariano Matos, Geovani de Oliveria Ribeiro, Steven S. Witkin, Gavin A. Cloherty, Maria Cassia Mendes-Correa
Abstract
Benign prostatic hyperplasia (BPH) is the most common urologic condition in elderly men, characterized by the reactivation of developmental programs such as prostatic budding and branching. However, the molecular mechanisms underlying this reactivation in BPH remain unclear. In this study, we identified TIAM1 (T-lymphoma invasion and metastasis-inducing protein-1) as a critical regulator of prostatic budding and branching. By generating an unbiased BPH transcriptomic signature from patient datasets, we discovered an upregulation of TIAM1, which was subsequently validated at the protein level. Functional assays using organoid cultures derived from human prostatic cell lines revealed that TIAM1 is essential for prostatic budding and branching. Additionally, the BPH transcriptomic signature identified NSC23766, a small molecule inhibitor of TIAM1-RAC1 signaling, as a therapeutic proof-of-concept agent for BPH. Genetic knockdown of TIAM1 in human prostatic cell lines markedly reduced organoid branching, an effect mirrored by administration of NSC23766. The translational relevance of these findings is underscored by the growth inhibition observed in patient-derived BPH organoids treated with NSC23766. In conclusion, our findings identify TIAM1 as a key driver of prostatic branching and growth, and suggest that targeting TIAM1-RAC1 signaling could be a promising therapeutic strategy for BPH.
Authors
Hamed Khedmatgozar, Sayanika Dutta, Michael Dominguez, Murugananthkumar Raju, Girijesh Kumar Patel, Daniel Latour, Melanie Johnson, Mohamed Fokar, Irfan Warraich, Allan Haynes, Barry J. Maurer, Werner de Riese, Luis Brandi, Robert J. Matusik, Srinivas Nandana, Manisha Tripathi
Abstract
DNA repair is essential for preserving genome integrity. Podocytes, post-mitotic epithelial cells of the kidney filtration unit, bear limited regenerative capacity, yet their survival is indispensable for kidney health. Podocyte loss is a hallmark of the aging process and of many diseases, but the underlying factors remain unclear. We investigated the consequences of DNA damage in a podocyte-specific knockout mouse model for Ercc1 and in cultured podocytes under genomic stress. Furthermore, we characterized DNA damage-related alterations in mouse and human renal tissue of different ages and patients suffering from minimal change disease and focal segmental glomerulosclerosis. Ercc1 knockout resulted in accumulation of DNA damage, ensuing albuminuria and kidney disease. Podocytes reacted to genomic stress by activating mTORC1 signaling in vitro and in vivo. This was abrogated by inhibiting DNA damage signaling through DNA-PK and ATM kinases and inhibition of mTORC1 modulated the development of glomerulosclerosis. Perturbed DNA repair gene expression and genomic stress in podocytes was also detected in focal segmental glomerulosclerosis. Beyond that, DNA damage signaling occurred in podocytes of healthy aging mice and humans. We provide evidence that genome maintenance in podocytes is linked to the mTORC1 pathway, involved in the aging process and the development of glomerulosclerosis.
Authors
Fabian Braun, Amrei M. Mandel, Linda Blomberg, Milagros N. Wong, Georgia Chatzinikolaou, David H. Meyer, Anna Reinelt, Viji Nair, Roman Akbar-Haase, Phillip J. McCown, Fabian Haas, He Chen, Mahdieh Rahmatollahi, Damian Fermin, Robin Ebbestad, Gisela G. Slaats, Tillmann Bork, Christoph Schell, Sybille Koehler, Paul T. Brinkkoetter, Maja T. Lindenmeyer, Clemens D. Cohen, Martin Kann, David Unnersjö-Jess, Wilhelm Bloch, Matthew G. Sampson, Martijn E.T. Dollé, Victor G. Puelles, Matthias Kretzler, George A. Garinis, Tobias B. Huber, Bernhard Schermer, Thomas Benzing, Björn Schumacher, Christine E. Kurschat
Abstract
Dysregulation of T follicular helper (Tfh) and T follicular regulatory (Tfr) cell homeostasis in germinal centers (GCs) can lead to antibody-mediated autoimmunity. While interleukin-1β (IL-1β) modulates the GC response via IL-1R1 and IL-1R2 receptors on follicular T cells in animal models, its role in humans remains unclear. We analyzed Tfh and Tfr phenotypes in human secondary lymphoid organs (tonsils, spleen, and mesenteric lymph nodes) using flow cytometry, single-cell transcriptomics, and in vitro culture, comparing findings with samples from autoimmune patients. We observed organ-specific Tfh/Tfr phenotypes according to activation status and IL-1 receptor expression. An excess of IL-1R1 over IL-1R2 expression promoted a unique activated Tfr subset with Treg and GC-Tfh features. IL-1β signaling via IL-1R1 enhanced follicular T-cell activation and Tfh-to-Tfr differentiation, while IL-1β inhibition upregulated IL-1R1, indicating a tightly regulated process. In autoimmune patients, high IL-1β and circulating Tfr levels correlated with increased autoantibody production, linking inflammation, IL-1β signaling, and Tfr/Tfh balance. Our findings highlight the critical role of IL-1β in follicular T-cell activation and suggest that targeting IL-1β signaling in Tfh and Tfr cells could be a promising strategy for treating antibody-mediated autoimmune diseases.
Authors
Romain Vaineau, Raphaël Jeger-Madiot, Samir Ali-Moussa, Laura Prudhomme, Hippolyte Debarnot, Nicolas Coatnoan, Johanna Dubois, Marie Binvignat, Hélène Vantomme, Bruno Gouritin, Gwladys Fourcade, Paul Engeroff, Aude Belbézier, Romain Luscan, Françoise Denoyelle, Roberta Lorenzon, Claire Ribet, Michelle Rosenzwajg, Bertrand Bellier, David Klatzmann, Nicolas Tchitchek, Stéphanie Graff-Dubois
Abstract
JAK inhibitors (JAKi) are widely used anti-inflammatory drugs. Recent data suggest JAKi have superior effects on pain reduction in rheumatoid arthritis (RA). However, the underlying mechanisms for this observation are not fully understood. We investigated whether JAKi can act directly on human sensory neurons. We analysed RNA sequencing datasets of sensory neurons and found they expressed JAK1 and STAT3. Addition of cell-free RA synovial fluid to human induced pluripotent stem cell (iPSC)-derived sensory neurons led to phosphorylation of STAT3 (pSTAT3), which was completely blocked by the JAKi tofacitinib. Compared to paired serum, RA synovial fluid was enriched for the STAT3 signalling cytokines IL-6, IL-11, LIF, IFN-alpha and IFN-beta, with their requisite receptors present in peripheral nerves post-mortem. Accordingly, these recombinant cytokines induced pSTAT3 in iPSC-derived sensory neurons. Furthermore, IL-6+sIL-6R and LIF upregulated expression of pain-relevant genes with STAT3-binding sites, an effect which was blocked by tofacitinib. LIF also induced neuronal sensitisation, highlighting this molecule as a putative pain mediator. Finally, over time, tofacitinib reduced the firing rate of sensory neurons stimulated with RA synovial fluid. Together, these data indicate that JAKi can act directly on human sensory neurons, providing a potential mechanistic explanation for their suggested superior analgesic properties.
Authors
Yuening Li, Elizabeth H. Gray, Rosie Ross, Irene Zebochin, Amy Lock, Laura Fedele, Louisa Janice Kamajaya, Rebecca J. Marrow, Sarah Ryan, Pascal Röderer, Oliver Brüstle, Susan John, Franziska Denk, Leonie S. Taams
Abstract
Epstein-Barr virus (EBV) infection precedes multiple sclerosis (MS) onset and plays a poorly understood etiologic role. To investigate possible viral pathogenesis, we analyzed single-cell expression in peripheral B cells from people with early MS collected longitudinally during the Immune Tolerance Network (ITN) STAyCIS Trial. Expression profiles were compared to scRNA-seq from in vitro EBV models, autoimmune disorders, chronic infectious diseases, and healthy controls. Analyses focused on CD19+/CD20+/CD21lo/CD11c+/T-bet+ atypical B cells (ABCs). ABCs were significantly enriched in early MS PBMCs versus healthy controls by scRNA-seq and flow cytometry, establishing ABC expansion as a clinical feature. EBV-associated ABC expression including CXCR3, PD-L1, and PD-L2 was enriched in early MS; however, direct EBV infection of ABCs was not detected. Early MS ABCs exhibited significantly upregulated inflammatory cytokine mRNAs (CXCL8, IL18, VEGFA). Further de novo EBV-infected B cells secreted IL-8 and VEGF. MS activity stratification revealed rare distinctive inflammatory ABCs significantly underrepresented in individuals with no evidence of activity long-term (LTNA) versus people with additional RRMS activity at the primary endpoint. Moreover, CXCR3+ ABCs increased after baseline diagnosis and were significantly enriched in people with disease exacerbation during the study. Thus, ABC expansion and inflammatory responses correlate to early MS activity, possibly as a bystander response to EBV.
Authors
Elliott D. SoRelle, Ellora Haukenfrers, Gillian Q. Horn, Vaibhav Jain, James Giarraputo, Karen Abramson, Emily Hocke, Laura A. Cooney, Kristina M. Harris, Scott S. Zamvil, Simon G. Gregory, Micah A. Luftig
Abstract
Many chemotherapeutic agents impair cancer growth by inducing DNA damage. The impact of these agents on mutagenesis in normal cells, including sperm, is largely unknown. Here, we applied high-fidelity duplex sequencing to 94 samples from 36 individuals exposed to diverse chemotherapies and 32 controls. We found that many of the sperm samples from men exposed to chemotherapy, the mutation burden was elevated as compared to controls and the expected burden based on trio studies, with one subject having >10-fold increase over expected for age. Saliva from this same individual also had a markedly higher mutation burden. We then validated this finding using other tissues, also finding an increased mutation burden in the blood and liver of many subjects exposed to chemotherapy as compared to unexposed controls. Similarly, mice treated with three cycles of cisplatin had an increased mutation burden in sperm but also in the liver, and hematopoietic progenitor cells. These results suggest an association between cancer therapies and mutation burden, with implications for counseling cancer patients considering banking sperm prior to therapy and for cancer survivors considering the tradeoffs of using banked sperm as compared to conceiving naturally.
Authors
Shany Picciotto, Camilo Arenas-Gallo, Amos Toren, Ruty Mehrian-Shai, Bryan Daly, Stephen Rhodes, Megan Prunty, Ruolin Liu, Anyull Bohorquez, Marta Grońska-Pęski, Shana Melanaphy, Pamela Callum, Emilie Lassen, Anne-Bine Skytte, Rebecca C. Obeng, Christopher Barbieri, Molly Gallogly, Brenda Cooper, Katherine Daunov, Lydia Beard, Koen Van-Besien, Joshua Halpern, Quintin Pan, Gilad D. Evrony, Viktor A. Adalsteinsson, Jonathan E. Shoag
Abstract
High rates of physician-scientist attrition from the investigative workforce remains a significant problem despite the development of dedicated programs and initiatives designed to address the unique challenges faced by physician-scientists. However, many of these efforts are restricted to single career stages of physician-scientist training or to a single medical specialty, which may limit opportunities for beneficial vertical and horizontal mentorship regarding overcoming common career obstacles. Here, we outline the development of a physician-scientist symposium to break down silos and enable productive interactions between physician-scientists across career/training stages, academic and scientific disciplines, and medical specialties. Participants were mixed in (1) small-group problem-based discussions, (2) a cross-specialty keynote panel on overcoming barriers in a physician-scientist career, and (3) skill-building workshops. Attendees indicated that they fostered new connections, developed new skills to overcome career challenges, and increased their commitment to persevering in a career as a physician-scientist. Positive evaluations were not dependent on attendee career/training stage or gender. We suggest these elements of the symposium curriculum may be easily adapted for inclusion in a wide variety of physician-scientist training formats.
Authors
Kevin F. Dowling, Shohini K. Ghosh-Choudhary, Neil Carleton, Kathleen Prigg, Richard A. Steinman
Abstract
Background: Traffic-related air pollution (TRAP) is a risk factor for Alzheimer disease (AD), where unresolved brain inflammation has been linked to deficits in the levels of free lipid mediators that enable the resolution of inflammation. It is unknown whether these deficits are due to reductions in esterified lipid pools, the main source of free bioactive pro-resolving lipids in the brain, and whether they are related AD pathophysiology. Methods: This unknown was tested by measuring brain esterified lipid mediators and pathogenic markers of AD in TgF344-AD and wildtype (WT) male and female rats exposed to filtered air or TRAP for 14 months, and in human postmortem pre-frontal cortex of individuals with or without AD. Results: Significant reductions in pro-resolving lipid mediators esterified to neutral lipids and/or phospholipids were seen in AD and TRAP-exposed female rats, where levels were associated with inflammation, synaptic loss and impaired glucose metabolism. Lower esterified pro-resolving lipid mediator concentrations were associated with older age in pre-frontal cortex of humans with AD. Conclusion: Impaired resolution in AD is due to depletion of esterified pro-resolving lipid pools that supply the brain with free bioactive mediators involved in inflammation resolution. TRAP exposure alters the same esterified resolution pathways, reflecting convergent mechanisms underlying AD.
Authors
Ameer Y. Taha, Qing Shen, Yurika Otoki, Nuanyi Liang, Kelley T. Patten, Anthony E. Valenzuela, Christopher D. Wallis, Douglas J. Rowland, Abhijit J. Chaudhari, Keith J. Bein, Anthony S. Wexler, Lee-Way Jin, Brittany N. Dugger, Danielle J. Harvey, Pamela J. Lein
Abstract
Airway smooth muscle (ASM) hyperplasia is a hallmark of airway remodeling in asthma, which still lacks an effective treatment. Low-density lipoprotein receptor-related protein 1 (LRP1) is involved in regulating the proliferation of various cell types, and the intracellular domain of LRP1 (LRP1-ICD) also exhibits unique biological functions. However, the role of LRP1 in asthma airway remodeling remains unclear. In the present study, LRP1 was increased in ASM cells of mice with OVA-induced chronic asthma, with the elevation in LRP1-ICD protein levels being significantly greater than that of the LRP1 β-chain. In vivo experiments demonstrated that inhibiting LRP1 reduced ASM proliferation in these mice. Mechanistically, LRP1 knockdown inhibited the FGF2/ERK signaling pathway, thereby arresting cell cycle progression and suppressing ASM cell proliferation. Additionally, in vitro experiments revealed that the inhibitory effect of LRP1-ICD overexpression on ASM cell proliferation was lost after adjusting the levels of the LRP1. LRP1-ICD overexpression inhibited full-length LRP1 protein levels by promoting its protein degradation rather than by suppressing its transcription, thus preventing further exacerbation of asthma. In conclusion, this study clarifies the molecular biological mechanism by which LRP1 regulates ASM proliferation, suggesting targeting full-length LRP1 as a novel strategy for therapeutic intervention in asthma airway remodeling.
Authors
Ya Deng, Jiaying Zhao, Chen Gong, Wenqian Ding, Lulu Fang, Huaqing Liu, Ming Li, Bing Shen, Shenggang Ding
Abstract
PCSK9 induces the hepatic degradation of the low-density lipoprotein receptor (LDLR), thereby increasing the concentration of LDL-cholesterol in the blood. Beyond its effects on LDL, recent studies have reported pleiotropic effects of PCSK9, notably in septic shock, vascular inflammation, viral infection, and cancer. While the functional and structural integrity of peripheral nerves are critically influenced by circulating lipids, the impact of PCSK9 on the peripheral nervous system remains unknown. In this study, we investigated the consequences of PCSK9 deficiency on peripheral nerves. We found that PCSK9 deletion in mice leads to peripheral neuropathy, characterized by reduced thermal and mechanical pain sensations. PCSK9 deficient mice also presented with skin structural changes, including a reduction in the number of nociceptive Schwann cells, Remak fiber axonal swelling, as well as hypomyelination of small nerve fibers. Interestingly, the peripheral nerves of PCSK9-deficient mice showed an upregulation of CD36, a fatty acid transporter, which correlated with increased nerve lipid content, structural mitochondrial abnormalities, and acylcarnitine accumulation. Our findings demonstrate that PCSK9 plays a critical role in peripheral nerves by regulating lipid homeostasis and its deficiency results in symptoms related to peripheral neuropathy.
Authors
Ali K. Jaafar, Aurélie Paulo-Ramos, Guillaume Rastoldo, Bryan Veeren, Cynthia Planesse, Matthieu Bringart, Philippe Rondeau, Kévin Chemello, Olivier Meilhac, Gilles C. Lambert, Steeve Bourane
Abstract
Spinal cord injury (SCI) evokes profound dysfunction in hollow organs such as the urinary bladder and gut. Current treatments are limited by a lack of molecular data to inform novel therapeutic avenues. Previously, we showed systemic treatment with the neuroprotective agent inosine improved bladder function following SCI in rats. Here, we applied integrated multi-omics analysis to explore molecular alterations in the bladder over time and their sensitivity to inosine following SCI. Canonical signaling pathways regulated by SCI included those associated with protein synthesis, neuroplasticity, wound healing, and neurotransmitter degradation. Upstream regulator and causal network analysis predicted multiple effectors of DNA damage response signaling following injury, including PARP1. Markers of DNA damage (gammaH2AX, ATM/ATR substrates) and PARP activity (poly-ADP-ribose) were increased in bladder tissue following SCI and attenuated with inosine treatment. Inosine treatment also attenuated oxidative DNA damage in rat bladder cells in vitro. Proteomics analysis suggested that SCI induced changes in protein synthesis-, neuroplasticity-, and oxidative stress-associated pathways, a subset of which were shown in transcriptomics data to be inosine-sensitive. These findings provide insights into the molecular landscape of the bladder following SCI and identify key inosine-sensitive pathways associated with injury.
Authors
Ali Hashemi Gheinani, Bryan S. Sack, Alexander Bigger-Allen, Hatim Thaker, Hussein Atta, George Lambrinos, Kyle Costa, Claire Doyle, Mehrnaz Gharaee-Kermani, Susan Patalano, Mary Piper, Justin F. Cotellessa, Dijana Vitko, Haiying Li, Manubhai Kadayil Prabhakaran, Vivian Cristofaro, John Froehlich, Richard S. Lee, Wei Yang, Maryrose P. Sullivan, Jill A. Macoska, Rosalyn M. Adam
Abstract
INTRODUCTION: Maladaptive hypertrophy, podocyte stress, and depletion contribute to kidney function decline. Although IGF-1 plays a key role in early hypertrophic responses in the single kidney state, its impact on KTx outcomes remains uncertain. This report tests the hypothesis that early IGF-1 exposure reduces KTx survival. METHODS. Population datasets compared incident Death Censored Graft Failure (DCGF) rates by age at KTx (n=366,404) with IGF-1 levels by age (n=15,014). A clinical study of 216 KTx recipients evaluated the association of IGF-1 exposure with DCGF and secondary outcomes of proteinuria and Biopsy-Proven Acute Rejection. IGF-1 exposure was modeled using pre-KTx IGF-1 levels and donor kidney dose estimated from the donor:recipient body surface area ratio reflecting allograft hyperfiltration. The association of DCGF with an IGF1 SNP linked to high IGF-1 levels was assessed in 724 genotyped allograft recipients. Single-cell transcriptomic data from first-year post-KTx patients and binephric donors were compared to assess intrarenal cellular expression of IGF1, IGF1R, and GHR transcripts. RESULTS. DCGF risk by age at KTx paralleled IGF-1 levels by age. Higher IGF-1 exposure was associated with significantly increased risks of DCGF, proteinuria, and T-cell-mediated rejection. Genotypic analysis showed a 50% increase in DCGF risk per risk allele at IGF1 eQTL rs35767. First-year biopsy results revealed no increase in intrarenal IGF1 transcript, while GHR and IGF-1R transcripts were suppressed, consistent with circulating IGF-1 (vs. graft-derived IGF-1) being the primary source of IGF-1 exposure. CONCLUSION. We identify a role for the GH-IGF-1 axis in reducing KTx survival.
Authors
Matthew Cusick, Viji Nair, Damian Fermin, John Hartman, Jeffrey A. Beamish, Zeguo Sun, Zhongyang Zhang, Edgar Otto, Rajasree Menon, Sudha Nadimidla, Nicholas Demchuk, Kelly Shaffer, Peter Heeger, Weija Zhang, Madhav C. Menon, Matthias Kretzler, Roger C. Wiggins, Abhijit S. Naik
Abstract
Inherited retinal degenerations (IRDs) are important causes of progressive, irreversible blindness. Hereditary macular diseases in particular are significant in their effect on the specialized, central cone photoreceptor-rich macula responsible for high resolution vision. Autosomal dominant Best vitelliform macular dystrophy (BVMD), caused by variants in the BEST1 gene, is one of the most common inherited macular dystrophies. Gene therapies have emerged as promising treatments for IRDs, but a lack of suitable animal models has hindered progress both in treatments and in understanding the mechanisms underlying macular diseases. Here, we report a Macaca fascicularis carrying a heterozygous potential pathogenic BEST1p.Q327E variant that disrupts the BEST1 ion channel by destabilizing the A195 helix, mirroring the structural perturbations seen in certain human pathological mutants. Longitudinal imaging over two years revealed progressive macular changes, including subfoveal cleft enlargement, lipid-rich deposit accumulation, retinal pigment epithelium (RPE) disruption, and central-to-peripheral photoreceptor degeneration, recapitulating early human BVMD pathology. Histopathology demonstrated diminished BEST1 expression, attenuation of the RPE-photoreceptor interface, and two distinct types of lipid deposits, including heretofore unappreciated cone mitochondrial-enriched lesions, highlighting selective cone mitochondria vulnerability. This first non-human primate model of inherited macular dystrophy links BEST1 mutations, mitochondrial dysfunction, and progressive macular degeneration, offering new insights into BVMD pathophysiology and highlighting its utility for studying disease progression and potential therapeutic interventions.
Authors
Wei Yi, Mingming Xu, Ying Xue, Yingxue Cao, Ziqi Yang, Lingli Zhou, Yang Zhou, Le Shi, Xiaomei Mai, Zehui Sun, Wenjie Qing, Yuying Li, Aolun Qing, Kaiwen Zhang, Lechun Ou, Shoudeng Chen, Elia J. Duh, Xialin Liu
Abstract
Toll-like receptors (TLRs) are being explored to enhance immunity in HIV cure strategies. The TLR7 agonist GS-9620 promotes immune activation, reactivates latent HIV, and delays viral rebound in some people with HIV. Previous work has shown that biological sex influences TLR7 signaling. This study examined the interplay between biological sex, age, and the sex hormones 17β-estradiol, progesterone, and testosterone on GS-9620’s ability to promote cytokine secretion and activate CD4, CD8, and NK cells ex vivo. Interestingly, sex hormones had no effect on GS-9620-mediated immune activation or cytokine induction. However, we found that GS-9620 activity was influenced by age only in female donors. Further, we found that GS-9620-mediated CD4 T cell activation was positively correlated with the induction of IFN-γ and IL-12, while CD4 T cell activation and IL-12 production were negatively correlated with age. Additionally, CD8 T cell activation was positively correlated with IFN-γ production. Mechanistically, IFN-γ was sufficient to promote higher immune activation of both CD4 and CD8 T cells in female versus male donors. In conclusion, biological sex and age, but not sex hormones, influence GS-9620-mediated immune activation. Understanding these factors will help design and evaluate future clinical trials using GS-9620 for an HIV cure.
Authors
Carissa S. Holmberg, Callie Levinger, Adam R. Ward, Alberto Bosque
