Abstract
Systemic inflammation is now recognized as a key contributor to epilepsy pathophysiology, yet the role of innate immune cells, particularly neutrophils, remains poorly defined in epilepsy. Preclinical studies in rodent models have implicated neutrophils in seizure activity, but their phenotype in human epilepsy has not been thoroughly investigated. In this study, we aimed to characterize systemic inflammatory profiles and neutrophil-associated immune signatures in the blood of patients with drug-resistant epilepsy compared with healthy controls. We identified a systemic low-grade inflammatory profile in patients characterized by elevated neutrophil-to-lymphocyte ratio, C-reactive protein, proinflammatory cytokines (IL-6, CXCL8/IL-8, TNF-α), and activated neutrophils (CXCR4+CD62Llo). Neutrophil phenotyping revealed two distinct immune profiles. Patients with longer disease duration exhibited a more immature systemic signature characterized by immature neutrophils (CD15+CD10–), resting neutrophils (CXCR4+CD62L+), and elevated IL-6 levels. In contrast, patients with higher seizure frequency displayed a more inflammatory profile, marked by increased IL-12 and activated (CXCR4+CD62Llo) and hyperactivated (CXCR4hiCD62Llo) neutrophil subsets. Moreover, elevated presurgical levels of inflammatory profile TNF-α, IL-6, and hyperactivated CXCR4hiCD62Llo neutrophils were associated with seizure recurrence 1 year after surgery. This pioneering study highlights the heterogeneity of peripheral immune responses in drug-resistant epilepsy and identifies neutrophil-related signatures as promising prognostic biomarkers in this context.
Authors
Coraly Simoës Da Gama, Aurélie Hanin, Gwen Goudard, Véronique Masson, Aurore Besnard, Karim Dorgham, Guy Gorochov, Guillaume Dorothée, Valerio Frazzini, Vincent Navarro, Mélanie Morin-Brureau
Abstract
HIV infection rapidly impairs the gastrointestinal barrier, contributing to persistent mucosal immune dysfunction, microbial translocation, and systemic inflammation despite antiretroviral therapy (ART). Using SIV-infected rhesus macaques on long-term ART, we investigated mechanisms underlying impairment in gut barrier–protective IL-17/IL-22 responses and the potential modulation of this pathway by dietary indoles. Longitudinal profiling of colonic epithelial and lamina propria cells revealed a selective loss of IL-17/IL-22–producing γδ T cells and type 3 innate lymphoid cells (ILC3s). This loss correlated with reduced expression of the transcription factors AHR and RORγt and was associated with elevated plasma markers of intestinal epithelial barrier disruption (IEBD), including intestinal fatty acid–binding protein (iFABP), zonulin, and LPS-binding protein (LBP). Targeting this transcriptional deficiency, dietary indole supplementation for 1 month restored colonic AHR+ IL-22–producing γδ T cells, RORγt+ ILC3s, and Vδ1 T cells, and was associated with reduced iFABP and zonulin levels. Immunohistochemical analyses further demonstrated enrichment of AHR/RORγt-coexpressing cells in the colon of indole-supplemented animals during chronic SIV infection on ART. Collectively, these findings indicate that disruption of the AHR-RORγt axis is a key pathogenic mechanism underlying persistent IEBD in chronic SIV/HIV infection. Modulation of AHR and RORγt signaling pathways in the gut may therefore represent a promising therapeutic strategy to reinforce mucosal barrier function and mitigate chronic inflammation in people living with HIV.
Authors
Siva Thirugnanam, Alison R. Van Zandt, Alexandra B. McNally, Victoria A. Hart, Isabelle Berthelot, Cecily C. Midkiff, Lara A. Doyle-Meyers, David A. Welsh, Robert V. Blair, Andrew G. MacLean, Namita Rout
Abstract
Subendothelial retention of cholesterol-rich apolipoprotein-B–containing lipoproteins drives atherosclerotic arterial disease. In peripheral interstitial fluid from patients with type 2 diabetes (T2D), levels of such particles have been shown to be paradoxically reduced relative to those in serum, presumably reflecting their increased retention within the arterial wall. To identify possible mechanisms involved in lipoprotein retention in T2D, we obtained serum and skin blister fluid from such patients and matched controls, together with skin biopsies in a subset of individuals. In T2D, smaller LDL and VLDL remnant particles were more prominent in serum but not in interstitial fluid, reflecting their enhanced vascular entrapment. The interstitial-fluid-to-serum ratio of apolipoprotein-B was 58% lower in T2D than in controls (0.14 versus 0.33), concomitant with increased susceptibility for LDL binding to proteoglycans. The most marked differences were seen in patients with clinically evident cardiovascular disease. The degree of transvascular retention was positively related to the propensity of isolated serum LDL to bind aortic proteoglycans, both in T2D and in controls. Skin unesterified cholesterol levels were higher in patients with T2D relative to healthy controls. With aging, both proteoglycan binding and apparent vascular retention of LDL increased in controls but not in T2D, indicating that these mechanisms may also be relevant for atherogenesis in nondiabetic individuals.
Authors
Pär Björklund, Jennifer Härdfeldt, Lauri Äikäs, Sara Straniero, Minna Holopainen, Katariina Öörni, Mats Rudling, Bo Angelin
Abstract
Spinocerebellar ataxia type 14 (SCA14) is an autosomal dominant neurodegenerative disease caused by mutations in the gene encoding protein kinase C γ (PKCγ), a Ca2+- and diacylglycerol-dependent Ser/Thr kinase dominantly expressed in cerebellar Purkinje cells. These mutations impair autoinhibitory constraints to increase the basal activity of the kinase, resulting in deficits in the cerebellum that are not observed upon simple deletion of the gene, and severe ataxia. To better understand the impact of aberrant PKCγ signaling in disease pathology, we developed a knockin murine model of the SCA14 mutation ΔF48 in PKCγ. This fully penetrant mutation is severe in humans and is mechanistically informative, as it has high basal activity but is unresponsive to agonist stimulation. Genetic, behavioral, and molecular testing revealed that ΔF48 PKCγ mice have ataxia-related phenotypes and an altered cerebellar phosphoproteome driven primarily by enhanced Ca2+/calmodulin-dependent kinase 2 signaling, effects that were more severe in male mice. Analysis of existing human data revealed that SCA14 has a significantly earlier age of onset for males compared with females. Data from this clinically relevant mutation suggested that enhanced basal activity of PKCγ is sufficient to cause ataxia and that treatment strategies to modulate aberrant PKCγ may be particularly beneficial in males.
Authors
Sarah A. Wolfe, Yuliang Ma, Tomer M. Yaron-Barir, Carly Chang, Caila A. Pilo, Majid Ghassemian, Amanda J. Roberts, Sang Ryeul Lee, Benjamin A. Henson, Kristen Jepsen, Jared L. Johnson, Lewis C. Cantley, Susan S. Taylor, George Gorrie, Alexandra C. Newton
Abstract
Spinal muscular atrophy (SMA) is a devastating neuromuscular disorder caused by mutations in the survival motor neuron 1 (SMN1) gene leading to decreased SMN protein levels and motor neuron dysfunction. SMN-restoring therapies offer clinical benefit, but the downstream molecular consequences of SMN reduction remain incompletely understood. SMN deficiency resulted in downregulation of kinesin heavy chain isoform 5A (KIF5A) in human neurons and in a mouse model of SMA. SMN associated with KIF5A mRNA and contributed to its stability. Reduced SMN levels impaired axon regeneration, which was rescued by KIF5A overexpression. Because KIF5A has also been connected to ALS, these findings provide evidence of a molecular link between SMA and ALS pathophysiology, highlighting KIF5A as an SMN-regulated factor. Our findings suggest that SMN-independent interventions targeting KIF5A could represent a complementary therapeutic approach for SMA and other motor neuron diseases.
Authors
Tetsuya Akiyama, Yi Zeng, Caiwei Guo, Olivia Gautier, Lauren Koepke, Heankel Lyons, Elana Molotsky, Juliane S. Bombosch, Odilia Sianto, Jay P. Ross, Phuong Hoang, Luke Zhao, Cole Spencer, Charlotte J. Sumner, Michelle Monje, John W. Day, Aaron D. Gitler
Abstract
Dedifferentiated liposarcoma (DDLS), myxofibrosarcoma (MFS), and undifferentiated pleomorphic sarcoma (UPS) are the most common types of genetically complex sarcoma. There is an urgent need to develop effective targeted therapy for these deadly sarcoma types. Despite their genetic complexity, these sarcomas share genomic alterations causing PI3K/Akt/mTOR and MAPK pathway activation, and both pathways control translation mediated by the RNA helicase eIF4A. We therefore investigated eIF4A inhibition as a therapeutic strategy. The eIF4A inhibitor CR-1-31B effectively suppressed tumor growth and induced apoptosis in DDLS, MFS, and UPS patient–derived cell lines and mouse xenografts. Transcriptome-scale ribosome footprinting identified eIF4A-dependent mRNAs such as the Hippo pathway transcriptional coactivators YAP1 (YAP) and WWTR1 (TAZ). Combined knockdown of YAP and TAZ induced apoptosis in DDLS, MFS, and UPS cell lines, and their ectopic expression partially rescued cells from apoptosis induced by CR-1-31B. Genomic analysis of patient tumors revealed that YAP and WWTR1 were frequently amplified or gained in DDLS, MFS, and UPS and were associated with worse clinical outcomes. Together, our findings identify a strategy for targeting the Hippo pathway in incurable forms of sarcoma based on inhibition of eIF4A-dependent translation of the key oncogenic transcription factors YAP and TAZ.
Authors
Young-Mi Kim, Prathibha Mohan, Urmila Sehrawat, Evan Seffar, Rafaela Muniz De Queiroz, Kalyani Chadalavada, Nikita Persaud, Tomoyo Okada, Anirudh Kulkarni, Jianan Lin, Nathalie Lailler, Shaleigh Smith, Bhumika Jadeja, Nicholas D. Socci, Zhengqing Ouyang, Hans-Guido Wendel, Samuel Singer
Abstract
We investigated whether destroying malignant cells and the associated tumor microenvironment (TME) by focal gene therapy would broaden immune checkpoint inhibitor (ICI) effectiveness. We show that ICI antitumor activity against syngeneic (murine) triple-negative breast cancer (TNBC) was augmented when a therapeutic transgene (purine nucleoside phosphorylase, referred to here as E. coli PNP) was used to cleave fludarabine (2-fluoro-arabinofuranosyl adenine) to the anticancer purine base, 2-fluoroadenine (F-Ade). We also established strong repression of anatomically distant, non-PNP-expressing tumors being treated by the same strategy. TNBC cytoreduction was associated with decreased intratumoral PD1+ Tregs, increased granzyme B+ NK cells, elevated MKI67+ T8 cells, and rapid immune clearance. Because F-Ade works by a mechanism that destroys quiescent neoplastic and supporting cells in the microenvironment, and since resistance to ICIs depends upon an intact TME, tumor killing by this approach offers a means to sensitize refractory malignancies to immune ablation and points to broad applicability against numerous cancer subtypes.
Authors
Regina Rab, Jeong S. Hong, Brendan L.C. Kinney, Nicole C. Schmitt, William B. Parker, Adrianna Westbrook, Kelsey B. Bennion, Mandy L. Ford, Douglas H. Weitzel, Paula L. Miliani de Marval, Eric J. Sorscher, Annette Ehrhardt
Abstract
Hyperglycemia is a principal driver of β cell failure and multiple-organ complications in diabetes. Chronic exposure to hyperglycemia overstimulates mTORC1, disrupting glucose metabolism and promoting ER stress, oxidative stress, and inflammation; however, the upstream metabolic signal(s) linking glucose to mTORC1 activation remains unclear. Here, we identified glucosamine as a key metabolite connecting elevated glucose to mTORC1 signaling in pancreatic islets and kidney, both major targets of hyperglycemic damage. Using 13C6-glucose metabolic labeling in diabetic rodents treated with or without the SGLT2 inhibitor dapagliflozin or insulin, combined with targeted metabolomics and metabolic flux analysis, we found that tissue glucose concentrations strongly correlated with glucosamine. A similar correlation with plasma glucose was conserved in humans with or without type 2 diabetes, and inversely associated with β cell function. In vitro, low-dose glucosamine stimulated mTORC1 in islets and kidney proximal tubule cells in an O-GlcNAcylation–dependent manner. Broad phosphoproteomics and transcriptomics analyses in β cells showed that glucosamine activated mTORC1-regulating pathways, induced oxidative stress, ER stress, and dedifferentiation. Genetic inhibition of β cell mTORC1 via heterozygous Raptor knockout, as well as pharmacologic inhibition of the glucosamine/mTORC1 axis through SGLT2 inhibition, alleviated β cell stress, improved glycemic control, and restored β cell function. These findings identified the glucosamine/mTORC1 pathway as an important mediator of β cell and kidney dysfunction in diabetes.
Authors
Yael Riahi, Aviram Kogot-Levin, Ziv Teselpapa, Elisheva Zemelman, Fatema Gamal, Tamar Cohen, Abed Nasereddin, Idit Shiff, Ifat Abramovich, Bella Agranovich, Dana Avrahami, Liad Hinden, Erol Cerasi, Daljeet Kaur, Lihi Grinberg, Ron Piran, Joseph Tam, Ernesto Bernal-Mizrachi, Erez Dror, Gil Leibowitz
Abstract
Small-conductance Ca2+-activated K+ (SK) channels regulate neuronal excitability and act as a feedback mechanism to limit firing during sustained stimulation. In the present study, we demonstrated that SK2 plays an important role in the control of bladder function and visceral pain processing. SK2 channels are expressed in bladder-innervating afferent neurons, and ablation of this subunit results in elevated afferent firing rates in response to physiological levels of bladder distension, supporting a role for SK2 in modulating mechanosensory excitability. Mice overexpressing SK2 exhibit increased bladder capacity and reduced voiding frequency. Furthermore, overexpression of SK2 prevents the onset of pelvic mechanical allodynia and attenuates the exaggerated visceromotor response to bladder distension seen in wild-type mice with chemical cystitis. Thus, SK2 may be a promising target for treating overactive bladder and pain originating from the urinary bladder and other pelvic organs.
Authors
Guadalupe Manrique-Maldonado, Xuejiao Sun, Allison L. Marciszyn, Nicolas Montalbetti, Marcelo D. Carattino
Abstract
Brain metastases (BrMs) occur in approximately 30% of cancer patients, causing nearly one-fifth of cancer deaths. While immune checkpoint inhibitors (ICIs) benefit some BrM patients, responses remain highly variable. This variability partly reflects distinct histopathological growth patterns that include minimally invasive (MI) and highly invasive (HI) brain BrMs. Here we show that MI BrMs exhibit robust immune infiltration, whereas HI lesions are immunosuppressed. However, histological differentiation between MI and HI can be challenging because of subjective margin assessment. Here, using highly multiplexed spatial proteomics on 119 tumor sections from 46 patients with BrMs, we identify CHI3L1 as a key mediator of the immunosuppressive microenvironment in HI BrMs. In preclinical models, genetic deletion of CHI3L1 converts immune-cold metastases into lymphocyte-rich, ICI-responsive lesions infiltrated by granzyme B+ CD8+ T cells. In BrM patients treated with ICI, immunohistochemical quantification of CHI3L1 expression was a stronger predictor of ICI response than traditional MI/HI classification. Thus, CHI3L1 represents a promising biomarker and therapeutic target for BrMs.
Authors
Sarah M. Maritan, Elham Karimi, Matthew Dankner, Aldo Hernandez-Corchado, Miranda W. Yu, Matthew G. Annis, Yashar Aghazadeh Habashi, Morteza Rezanejad, Bridget Liu, Nebras Koudieh, Emilie Pichette, Parvaneh Fallah, Benoit Fiset, Yuhong Wei, Ali Nehme, Chun Geun Lee, Jack A. Elias, Morag Park, Yasser Riazalhosseini, Hamed Najafabadi, Kevin Petrecca, Marie-Christine Guiot, Daniela F. Quail, Logan A. Walsh, Peter M. Siegel
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