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Large-scale survey, animal models and computational modeling identify histological neurodegenerative biomarkers for traumatic optic neuropathy

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Abstract

Background: Traumatic optic neuropathy (TON) is a leading cause of blindness following closed traumatic brain injury, with no effective treatments available. Previous interventional clinical trials were complicated by its low prevalence, variability in neurodegenerative severity, and unavailability of reliable biomarkers. Methods: We analyzed data from 1226 patients enrolled in the prospective National Multi-Center Collaborative Clinical Research Program of China (2017-2024) to establish a clinical profile and identify non-invasive biomarkers for neurodegenerative severity. Subgroup analysis of monocular TON patients revealed potential biomarkers including visual functional parameters, inner retinal thickness, and time post-injury. Results: The ganglion cell complex (GCC) thickness showed a strong correlation with retinal ganglion cell somata (R² = 0.87, p < .0001) and axon density (R² = 0.89, p < .0001) in a clinically relevant large animal model. Computational analysis demonstrated that using GCC thickness as a biomarker could substantially enhance the statistical power of clinical trials (by up to 4.5-fold), as confirmed by real-world data. Conclusion: This study presented the largest epidemiological analysis of TON to date and established GCC thickness as a crucial biomarker for stratifying disease severity and improving the efficiency of clinical trials. Trial registration: Chinese Clinical Trial Registry (ChiCTR-OOC-17013437). Funding: National Key R&D Program of China (Grant No.2022YFA1105500); Key Science and Technology Program of Wenzhou (Grant No.ZY2022021); National Natural Science Foundation of China (Grant No.82471080).

Authors

YiKui Zhang, BoYue Xu, ShiWei Huang, ZhaoHui Shi, Wei Xiong, Ruijun Wang, GuiQin Liu, Linlin Chen, ZhenHua Ge, YongJie Zhang, HongLei Liu, BaoYun Jia, Chunxia Wang, HaiHong Shi, Jun Kang, NingYu An, Shuirui Huang, De-Fu Chen, Shenghai Huang, YuTing Luo, MingYue Liu, ZhuoWei Wang, Zhonghao Yu, Jingwei Zheng, Wentao Yan, Gen Li, Hao Chen, XingGuang Deng, Shihui Wei, YunHai Tu, EnDe Wu, Kang Zhang, Wencan Wu

The MUC5B promoter variant results in proteomic changes in the non-fibrotic lung

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Abstract

The gain-of-function MUC5B promoter variant is the dominant risk factor for the development of idiopathic pulmonary fibrosis (IPF). However, its impact on protein expression in both non-fibrotic control and IPF lung specimens have not been well characterized. Utilizing laser capture microdissection coupled to mass spectrometry (LCM-MS), we investigated the proteomic profiles of airway and alveolar epithelium in non-fibrotic controls (n = 12) and IPF specimens (n = 12), stratified by the MUC5B promoter variant. Through qualitative and quantitative analyses, as well as pathway analysis and immunohistological validation, we have identified a distinct MUC5B-associated protein profile. Notably, the non-fibrotic control alveoli exhibited substantial MUC5B-associated protein changes, with an increase of IL-3 signaling. Additionally, we found that epithelial cells overlying IPF fibroblastic foci cluster closely to alveolar epithelia and express proteins associated with cellular stress pathways. In conclusion, our findings suggest that the MUC5B promoter variant leads to protein changes in alveolar and airway epithelium that appears to be associated with initiation and progression of lung fibrosis.

Authors

Jeremy A. Herrera, Mark Maslanka, Rachel Z. Blumhagen, Rachel Blomberg, Nyan Ye Lwin, Janna Brancato, Carlyne D. Cool, Jonathan P. Huber, Jonathan S. Kurche, Chelsea M. Magin, Kirk C. Hansen, Ivana V. Yang, David A. Schwartz

Fine mapping of heterozygous IL6ST nonsense variants underlying autosomal dominant hyper-IgE syndrome

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Abstract

Loss-of-function (LOF) variants in IL6ST, encoding GP130, can cause hyper-IgE syndrome (HIES). Monoallelic LOF variants in IL6ST lead to HIES when located in the intracellular domain downstream of box 1/2 and upstream of the STAT3 phosphorylation sites and the recycling motif, due to their dominant negative (DN) activity. In this region, two previously unreported IL6ST variants, p.K702Sfs7* and p.Y759Wfs26*, were identified in two families with autosomal dominant (AD) HIES. Both variants were LOF, exhibited DN effects, leading to the accumulation of mutant GP130 on the cell surface. The p.K702Sfs7* mutation was the most upstream N-terminal mutation linked to HIES caused by heterozygous IL6ST variants. Comprehensive screening of IL6ST mutants revealed that most premature terminations downstream of amino acid F641, at the end of the transmembrane domain, resulted in LOF and DN effects via GP130 accumulation on the cell surface. The absence of the recycling motif (positions 782-787) in surface-expressed LOF GP130 led to its accumulation, contributing to the DN effect. The importance of intracellular truncating IL6ST variants can possibly be predicted based on the location of the premature stop codon. GP130 accumulation on the cell surface is a characteristic and potentially diagnostic finding in HIES patients with heterozygous IL6ST variants.

Authors

Kosuke Ashihara, Takaki Asano, Kanako Takeuchi, Kosuke Noma, Miyuki Tsumura, Wenjie Wang, Wei-Te Lei, Hisao Higo, Toshio Kubo, Yoko Mizoguchi, Shuhei Karakawa, Aurélie Cobat, Clément Conil, Etsushi Toyofuku, Akimasa Sekine, Kohsuke Imai, Dusan Bogunovic, Jean-Laurent Casanova, Cheng-Lung Ku, Vivien Béziat, Satoshi Okada

Identification of hypoxia-inducible factor HIF1A as a therapeutic target during SARS-CoV-2-associated lung injury

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Abstract

Hypoxia-inducible factors (HIFs) promote lung protection and pathogen eradication during acute lung injury. We therefore tested the theory that pharmacologic stabilization of HIFs dampens lung injury during SARS-CoV-2 pneumonia. Initial studies in murine SARS-CoV-2 models showed improved outcomes after treatment with the FDA-approved HIF-stabilizer vadadustat. Subsequent studies in genetic models implicated alveolar-expressed Hif1a in mediating lung protection. Therefore, we performed a randomized, double-blinded, multicenter phase 2 trial in patients admitted for SARS-CoV-2 infection and concomitant hypoxia (SpO2 ≤ 94%). Patients (n=448) were randomized to oral vadadustat (900 mg/day) or placebo for up to 14 days. Safety events were similar between the two groups. Vadadustat treatment induced surrogate HIF-target genes. The primary outcome of severe lung injury requiring high oxygen support on day 14 occurred in 43 patients in the vadadustat group and 53 patients in the placebo group (estimated probability, 13.3% vs. 16.9%). Among patients with baseline FiO2 ≥ 80% (n=106), the estimated probability of the primary outcome was 12.1% (vadadustat) vs. 79.1% (placebo), indicating an even greater benefit in patients with more severe baseline hypoxia. HIF1A is a likely therapeutic target during SARS-CoV-2-associated lung injury. Robust clinical trials of HIF stabilizers during pathogen-associated lung injury are warranted.

Authors

Bentley Bobrow, Samuel D. Luber, Paul Potnuru, Katherine Figarella, Jieun Kim, Yanyu Wang, In Hyuk Bang, David Robinson, Paulina B. Sergot, Steven K. Burke, Tingting Mills, Constanza de Dios, Robert Suchting, George W. Williams, Adit A. Ginde, Yafen Liang, Hongfang Liu, Charles Green, Marie-Francoise Doursout, Alparslan Turan, Daniel I. Sessler, Xiaoyi Yuan, Holger K. Eltzschig

Estrogens determine the efficacy of cancer immunotherapy in obese males with melanoma

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Abstract

Although obesity is a major risk factor for cancer, it may also improve the response to cancer therapy. Here we investigated the impact of obesity on the efficacy of immune checkpoint inhibitors (ICI). In male mice, obesity promoted tumor growth but enhanced the response to ICI. This was associated with higher expression of immune-related genes within the tumor and enhanced infiltration of tumor-specific CD8+ T cells. Further, obesity in mice was associated with higher estrogen levels and enrichment of estrogen response genes in the tumor, and anti-PD-1 efficacy was reduced upon administration of the aromatase inhibitor letrozole, which blocks the production of estrogens. Mechanistically, adipocyte-derived estrogens increased antigen presentation by dendritic cells and tumor-specific CD8+ T-cell cytotoxicity. Lastly, overweight and obese male melanoma patients responded better to ICI, high estrogen levels being associated with improved response and survival. Our results suggest that estrogens may serve as a predictive factor of response to ICI in men with melanoma.

Authors

Eloïse Dupuychaffray, Hélène Poinot, Aurélie Vuilleumier, Maxime Borgeaud, Montserrat Alvarez, Betül Taskoparan, Olivier Preynat-Seauve, Clarissa D. Voegel, Eliana Marinari, Denis Migliorini, Valérie Dutoit, Carole Bourquin, Aurélien Pommier

Role of progesterone action in inguinal hernia formation via skeletal muscle fibrosis and atrophy

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Abstract

More than one in four men will undergo surgery for inguinal hernia, which is commonly associated with fibrotic degeneration of the lower abdominal muscle (LAM) in the groin region. Utilizing a male mouse model expressing the human aromatase gene (Aromhum), previous studies showed that locally produced estradiol acting via estrogen receptor alpha in LAM fibroblasts leads to fibrosis, myofiber atrophy, and hernia development. Here, we found that upregulation of progesterone receptor (PGR) in a LAM fibroblast population mediates this estrogenic effect. A PGR-selective progesterone antagonist in Aromhum mice decreased LAM fibrosis and atrophy, preventing hernia formation and stopping progression of existing hernias. Addition of progesterone to estradiol treatment was essential for early-onset development of LAM fibrosis and large hernias in wild type mice, which was averted by a progesterone antagonist. Single-nuclei multiomics sequencing of herniated LAM revealed a unique population of Pgr-expressing fibroblasts that promotes fibrosis and myofiber atrophy through transforming growth factor beta-2 signaling. Multiomics findings were validated in vivo in herniated LAM tissues of both mice and adult men. Our findings suggest an important and rare pathologic role of progesterone signaling in males and provide evidence for progesterone antagonists as a non-surgical alternative for inguinal hernia management.

Authors

Tianming You, Mehrdad Zandigohar, Tanvi Potluri, Natalie Piehl, John S. Coon V, Elizabeth Baker, Maya Kafali, Yang Dai, Jonah J. Stulberg, David J. Escobar, Richard L. Lieber, Hong Zhao, Serdar E. Bulun

Device-free isolation of photoreceptor cells from patient iPSC-derived retinal organoids

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Abstract

Autologous photoreceptor cell replacement is one of the most promising strategies currently being developed for the treatment of patients with inherited retinal degenerative blindness. Induced pluripotent stem cell (iPSC) derived retinal organoids, which faithfully recapitulate the structure of the neural retina, are an ideal source of transplantable photoreceptors required for these therapies. However, retinal organoids contain other retinal cell types, including bipolar, horizontal and amacrine cells, which are unneeded and may reduce the potency of the final therapeutic product. Therefore, approaches for isolating fate committed photoreceptor cells from dissociated retinal organoids are desirable. In this work, we present partial dissociation, a technique which leverages the high level of organization found in retinal organoids to enable selective enrichment of photoreceptor cells without the use of specialized equipment or reagents such as antibody labels. We demonstrate up to 90% photoreceptor cell purity by simply selecting cell fractions liberated from retinal organoids during enzymatic digestion in the absence of mechanical dissociation. As the presented approach relies on the use of standard plasticware and commercially available cGMP compliant reagents, we believe that it is ideal for use in the preparation of clinical photoreceptor cell replacement therapies.

Authors

Nicholas E. Stone, Laura R. Bohrer, Nathaniel K. Mullin, Alexander Berthold, Allison T. Wright, Ian C. Han, Edwin M. Stone, Robert F. Mullins, Budd A. Tucker

Csk-mediated Src family kinase regulation dampens neutrophil infiltration during pulmonary infection

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Abstract

Neutrophil recruitment is crucial for pathogen elimination. However, precise control of the inflammatory response prevents overshooting reactions. Neutrophil activation initiates signaling resulting in integrin beta 2 (Itgb2) activation and neutrophil arrest. Src family kinases are involved in multiple cellular processes and are negatively regulated by the C-terminal Src kinase (Csk). During this study, we investigated the mechanism by which Csk regulates integrin activation and neutrophil recruitment. Here, we demonstrated that Csk deficiency in murine neutrophils resulted in increased neutrophil adhesion to the endothelium along with decreased neutrophil transmigration into inflamed tissues compared to their littermate controls. In bacterial pneumonia, infected Csk-deficient mice showed higher bacterial burdens and decreased neutrophil recruitment, while other immune cell counts and cytokine levels were not significantly different compared to control. Analyses of Csk-deficient neutrophils revealed an increased Itgb2 affinity, leading to reduced migration and intravascular crawling. Mechanistically, elevated cAMP-levels increased protein kinase A activity, which subsequently enhanced Csk activation. Csk, in turn, suppressed Src family kinases activation through phosphorylation (Y529). Hence, Csk-mediated regulation of neutrophil infiltration contributes to maintain a balanced immune response during bacterial pneumonia.

Authors

Wida Amini, Lena Schemmelmann, Jan-Niklas Heming, Marina Oguama, Katharina Thomas, Helena Block, Pia Lindental, Bernadette Bardel, Andreas Margraf, Oliver Soehnlein, Anika Cappenberg, Alexander Zarbock

Cord blood proteomics identifies biomarkers of early-onset neonatal sepsis

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Abstract

BACKGROUND. Symptoms of early-onset sepsis (EOS) in preterm infants are nonspecific, overlapping with normal postnatal physiological adaptations and noninfectious pathologies. This clinical uncertainty and the lack of reliable EOS diagnostics results in liberal use of antibiotics in the first days to weeks of life, leading to increased risk of antibiotic-related morbidities in infants who do not have an invasive infection. METHODS. To identify potential biomarkers for EOS in newborn infants, we used unlabelled tandem mass spectrometry proteomics to identify differentially abundant proteins in the umbilical cord blood of infants with and without culture-confirmed EOS. Proteins were then confirmed using immunoassay, and logistic regression and random forest models were built including both biomarker concentration and clinical variables to predict EOS. RESULTS. These data identified five proteins that were significantly upregulated in infants with EOS, three of which (serum amyloid A, C-reactive protein, and lipopolysaccharide-binding protein) were confirmed using a quantitative immunoassay. The random forest classifier for EOS was applied to a cohort of infants with culture-negative presumed sepsis (PS). Most PS infants were classified as resembling control infants, having low EOS biomarker concentrations. CONCLUSION. These results suggest that cord blood biomarker screening may be useful for early stratification of EOS risk among neonates, improving targeted, evidence-based use of antibiotics early in life. FUNDING. National Institutes of Health, Gerber Foundation, Friends of Prentice, Thrasher Research Fund, Ann & Robert H. Lurie Children’s Hospital, Stanley Manne Children’s Research Institute of Lurie Children’s.

Authors

Leena B. Mithal, Mark E. Becker, Ted Ling-Hu, Young Ah Goo, Sebastian Otero, Aspen Kremer, Surya Pandey, Nicola Lancki, Yawei Li, Yuan Luo, William Grobman, Denise Scholtens, Karen K. Mestan, Patrick C. Seed, Judd F. Hultquist

Pentose phosphate pathway inhibition metabolically reprograms CD8⁺ T cells and disrupts CNS autoimmunity

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Abstract

Multiple sclerosis is characterized by CNS infiltration of auto-reactive immune cells that drive both acute inflammatory demyelination and chronic progressive axonal and neuronal injury. Expanding evidence implicates CD8+ anti-neural T cells in the irreversible neurodegeneration that underlies progression in multiple sclerosis, yet therapies specifically targeting this cell population are limited. CD8+ T cells from patients with MS exhibit increased engagement of the pentose phosphate pathway. Pharmacologic inhibition of the pentose phosphate pathway reduced glycolysis, glucose uptake, NADPH production, ATP production, proliferation, and proinflammatory cytokine secretion in CD8+ T cells activated by ligation of CD3 and CD28. Pentose phosphate pathway inhibition also prevented CD8+ T cell-mediated antigen-specific neuronal injury in vitro and in both an adoptive transfer-based cuprizone model of demyelination and in mice with experimental autoimmune encephalomyelitis. Notably, transcriptional profiling of CNS-infiltrating CD8+ T cells in patients with MS indicated increased pentose phosphate pathway engagement, suggesting that this pathway is involved in CD8+ T cell-mediated injury of axons and neurons in the demyelinated CNS. Inhibiting the pentose phosphate pathway disrupts CD8+ T cell metabolic reprogramming and effector functions, suggesting that such inhibition may serve as a therapeutic strategy to prevent neurodegeneration in patients with progressive MS.

Authors

Ethan M. Grund, Benjamin D.S. Clarkson, Susanna Pucci, Maria S. Westphal, Carolina Muniz Partida, Sara A. Muhammad, Charles L. Howe

MicroRNA-6084 orchestrates angiogenesis and liver metastasis in colorectal cancer via extracellular vesicles

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Abstract

The prognosis for colorectal cancer (CRC) patients with liver metastasis remains poor, and the molecular mechanisms driving CRC liver metastasis are still not fully understood. Hypoxia-induced extracellular vesicles (H-EVs) derived from tumors have emerged as key players in inducing angiogenesis by transferring non-coding RNAs. However, the specific role of CRC-derived hypoxic EVs (H-EVs) in regulating the formation of the pre-metastatic microenvironment (PMN) by inducing angiogenesis remains unclear. Our study demonstrates that H-EVs induce both angiogenesis and liver metastasis. Through microRNA microarray analysis, we identified a reduction in miR-6084 levels within H-EVs. We found that miR-6084 inhibits angiogenesis by being transferred to endothelial cells via EVs. In endothelial cells, miR-6084 directly targets ANGPTL4 mRNA, thereby suppressing angiogenesis through ANGPTL4-mediated JAK2/STAT3 pathway. Furthermore, we uncovered that SP1 acts as a transcription factor regulating miR-6084 transcription, while HIF1A decreases miR-6084 expression by promoting SP1 protein dephosphorylation and facilitating ubiquitin‒proteasome degradation in SW620 cells. In clinical samples, we observed low expression of miR-6084 in plasma-derived EVs from CRC patients with liver metastasis. In summary, our findings suggest that CRC-derived hypoxic EVs promote angiogenesis and liver metastasis through the HIF1A/SP1/miR-6084/ANGPTL4 axis. Additionally, miR-6084 holds promise as a potential diagnostic and prognostic biomarker for CRC liver metastasis.

Authors

Yang Zhang, Xuyang Yang, Su Zhang, Qing Huang, Sicheng Liu, Lei Qiu, Mingtian Wei, Xiangbing Deng, Wenjian Meng, Hai-Ning Chen, Yaguang Zhang, Junhong Han, Ziqiang Wang

Emerin is an effector of oncogenic KRAS-driven nuclear dynamics in pancreatic cancer

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Abstract

For over a century, scientists reported the disruption of normal nuclear shape and size in cancer. These changes have long been used as tools for diagnosis and staging of malignancies. However, to date, the mechanisms underlying these aberrant nuclear phenotypes and their biological significance remain poorly understood. Using a model of pancreatic ductal adenocarcinoma (PDAC), the major histological subtypes of pancreatic cancer, we found oncogenic mutant KRAS reduces nuclear size. Transcriptomic and protein expression analysis of mutant KRAS-expressing PDAC cells revealed differential levels of several nuclear envelope-associated genes. Further analysis demonstrated the nuclear lamina protein, Emerin (EMD), acted downstream of KRAS to mediate nuclear size reduction in PDAC. Analysis of human PDAC samples showed that increased EMD expression associates with reduced nuclear size. Finally, in vivo genetic depletion of EMD in a mutant KRAS-driven PDAC model resulted in an increased nuclear size and a reduced incidence of poorly differentiated PDAC. Thus, our data provides evidence of a novel mechanism underlying nuclear size regulation and its impact in PDAC carcinogenesis.

Authors

Luis F. Flores, David L. Marks, Renzo E. Vera, Ashley N. Sigafoos, Ezequiel J. Tolosa, Luciana L. Almada, David R. Pease, Merih D. Toruner, Brian Chang, Brooke R. Tader, Kayla C. LaRue-Nolan, Ryan M. Carr, Rondell P. Graham, Catherine E. Hagen, Matthew R. Brown, Aleksey V. Matveyenko, Katherine L. Wilson, David W. Dawson, Christopher L. Pin, Kyle J. Roux, Martin E. Fernandez-Zapico

Donor genetics and storage conditions influence mitochondrial DNA and extracellular vesicle levels in RBC units

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Abstract

Mitochondrial DNA (mtDNA) shares characteristics with bacterial DNA and activates immune cells via Toll like receptor (TLR)-9. Extracellular vesicles (EVs) and mtDNA have been found in blood products and can activate immune cells; we sought to characterize their evolution in stored blood products. From a previous study of hemolysis in 13,403 blood donors, a second blood unit was drawn from 651 donors and sampled at days 10, 21, and 42. EV counts and RBC-EVs increased with storage time, and EV levels were higher in males and in RBC units processed in AS-1 compared to AS-3. mtDNA levels were higher in females and RBC units processed in AS-3. EV populations and mtDNA levels were highly correlated within donors for 98 donations obtained 2-12 months apart. Quantitative trait locus analysis revealed several genetic associations, most notably linking mtDNA levels with polymorphisms in ANKLE1, which encodes an erythroid-specific protein that preferentially cleaves mtDNA. These data suggest that donor-intrinsic factors may influence mtDNA and EV levels found in RBC units. This finding lends impetus to determining if genetic or environmental factors control levels of these immune mediators in blood donors.

Authors

Xutao Deng, Clara Di Germanio, Erika G. Marques de Menezes, Pamela Milani, Mars Stone, Heather Tanner, Sonia Coco Bakkour, Daniel M. Chafets, Sarah E. Reese, Nareg H. Roubinian, Steven Kleinman, Tamir Kanias, Michael P. Busch, Eric J. Earley, Grier P. Page, Travis Nemkov, Angelo D’Alessandro, Philip J. Norris

Cytoskeleton-associated protein 4 (CKAP4) affects podocyte cytoskeleton dynamics in diabetic kidney disease

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Abstract

Podocytes are kidney glomerular cells that depend on rigorously regulated cytoskeleton components and integrins to form and maintain the so-called foot processes, apparatuses that attach podocytes to the glomerular basement membrane and connect them to neighboring podocytes. In diabetic kidney disease (DKD) these foot processes are effaced as a result of cytoskeleton dysregulation, a phenomenon that gradually reduces glomerular filtration. Cytoskeleton-associated protein 4 (CKAP4) is a known linker between the endoplasmic reticulum, integrins and microtubular cytoskeleton. Since CKAP4 gene expression is downregulated in glomeruli from DKD patients but not in other chronic kidney diseases, we hypothesized a role for CKAP4 in the mechanisms leading to foot process effacement (FPE) in DKD. CKAP4 mRNA reduction in podocytes in DKD was demonstrated in human kidney biopsies. Knockdown of CKAP4 in vivo in zebrafish resulted in edema, proteinuria and foot process effacement, all typical features of DKD. Knockdown of CKAP4 in vitro led to disruption of the actin cytoskeleton and of the microtubular orientation. Moreover, it caused a downregulation of several integrins. These findings indicate that CKAP4 is crucial for foot process dynamics of podocytes. Its reduction, unique to DKD, is mechanistically connected to the pathophysiological processes leading to podocyte FPE.

Authors

Roberto Boi, Emelie Lassén, Alva Johansson, Peidi Liu, Aditi Chaudhari, Ramesh Tati, Janina Müller-Deile, Mario Schiffer, Kerstin Ebefors, Jenny Nyström

Impact of the IL-15 superagonist N-803 on lymphatic reservoirs of HIV

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Abstract

Background. NK cell function is impaired in people with HIV (PWH), hindering their potential to reduce the lymphoid tissue (LT) reservoir. The IL-15 superagonist N-803 has been shown to enhance NK and T cell function, and thus may reduce viral reservoirs. Methods. To determine the impact of N-803 on LTs, we conducted a clinical trial where 10 PWH on effective antiretroviral therapy (ART) were given three 6 mcg/kg doses of N-803 subcutaneously. We obtained PBMCs and lymph node (LN) and gut biopsies at baseline and after the last N-803 dose. Results. We found a non-statistically significant ~0.50 median log reduction in the frequency of viral(v)RNA+ and vDNA+ cells/g in the 6 participants with baseline and post-treatment LNs. In the ileum, we observed reductions of vRNA+ cells in 8/10 participants and vDNA+ cells in all participants. We also found significant inverse correlations between NK cell proliferation and the frequency of vRNA+ cells, and between NKG2A expression on NK cells and the frequency of vRNA+ cells. Conclusions. Our findings suggest N-803 may reduce the HIV reservoir in LTs of PWH on ART, an effect likely mediated by enhanced NK cell function. Controlled studies assessing the impact of NK cell therapy on HIV LTs are needed.

Authors

Joshua Rhein, Jeffrey G. Chipman, Gregory J. Beilman, Ross Cromarty, Kevin Escandón, Jodi Anderson, Garritt Wieking, Jarrett Reichel, Rodolfo Batres, Alexander Khoruts, Christopher M. Basting, Peter Hinderlie, Zachary B. Davis, Anne Eaton, Byron P. Vaughn, Elnaz Eilkhani, Jeffrey T. Safrit, Patrick Soon-Shiong, Jason V. Baker, Nichole R. Klatt, Steven G. Deeks, Jeffrey S. Miller, Timothy W. Schacker

Distinct cell types along thick ascending limb express pathways for monovalent and divalent cation transport

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Abstract

Kidney thick ascending limb cells reabsorb sodium, potassium, calcium, magnesium and contribute to urinary concentration. These cells are typically viewed as a single type that recycles potassium across the apical membrane and generates a lumen-positive transepithelial voltage driving calcium and magnesium reabsorption, although variability in potassium channel expression has been reported. Additionally, recent transcriptomic analyses suggest that different cell types exist along this segment, but classifications have varied and have not led to a new consensus model. We used immunolocalization, electrophysiology and enriched single nucleus RNA-Seq to identify thick ascending limb cell types in rat, mouse and human. We identified three major TAL cell types defined by expression of potassium channels and claudins. One has apical potassium channels, low basolateral potassium conductance, and is bordered by a monovalent cation-permeable claudin. A second lacks apical potassium channels, has high basolateral potassium conductance and is bordered by calcium- and magnesium-permeable claudins. A third type also lacks apical potassium channels and has high basolateral potassium conductance, but these cells are ringed by monovalent cation-permeable claudins. The recognition of diverse cell types may resolve longstanding questions about how solute transport can be modulated selectively and how disruption of these cells leads to human disease.

Authors

Hasan Demirci, Jessica P. Bahena-Lopez, Alina Smorodchenko, Xiao-Tong Su, Jonathan W. Nelson, Chao-Ling Yang, Joshua N. Curry, Xin-Peng Duan, Wen-Hui Wang, Yuliya Sharkovska, Ruisheng Liu, Duygu Elif Yilmaz, Catarina Quintanova, Katie Emberley, Ben Emery, Nina Himmerkus, Markus Bleich, David H. Ellison, Sebastian Bachmann

Pan-H7 influenza human antibody virus neutralization depends on avidity and steric hindrance

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Abstract

H7N9 avian influenza virus is a zoonotic influenza virus of public health concern, with a 39% mortality rate in humans. H7N9-specific prevention or treatments for humans have not been approved. We previously isolated a human monoclonal antibody (mAb) designated H7-235 that broadly reacts to diverse H7 viruses and neutralizes H7N9 viruses in vitro. Here, we report the crystal structure of H7 HA1 bound to the fragment antigen-binding region (Fab) of recombinant H7-235 (rH7-235). The crystal structure revealed that rH7-235 recognizes residues near but outside of the receptor binding site (RBS). Nevertheless, the rH7-235 IgG potently inhibits hemagglutination mediated by H7N9 viruses due to avidity effect and Fc steric hindrance. This mAb prophylactically protects mice against weight loss and death caused by challenge with lethal H7N9 viruses in vivo. rH7-235 mAb neutralizing activity alone is sufficient for protection when used at high dosed in a prophylactic setting. This study provides insights into mechanisms of viral neutralization by protective broadly reactive anti-H7 antibodies informing the rational design of therapeutics and vaccines against H7N9 influenza virus.

Authors

Iuliia M. Gilchuk, Jinhui Dong, Ryan P. Irving, Cameron D. Buchman, Erica Armstrong, Hannah L. Turner, Sheng Li, Andrew B. Ward, Robert H. Carnahan, James E. Crowe Jr.

Stearoyl-CoA-Desaturase Inhibition normalizes brain lipid saturation, alpha-synuclein homeostasis, and motor function in mutant Gba1-Parkinson mice

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Abstract

Loss-of-function mutations in the GBA1 gene are a prevalent risk factor for Parkinson’s disease (PD). Defining features are Lewy bodies that can be rich in α-synuclein (αS), vesicle- and other lipid membranes coupled with striatal dopamine loss and progressive motor dysfunction. Of these, lipid abnormalities are the least understood. An altered lipid metabolism in PD patient-derived neurons, carrying mutations in either GBA1, encoding for glucocerebrosidase, or αS can shift the physiological αS tetramer-monomer (T:M) equilibrium, resulting in PD phenotypes. We previously reported inhibition of stearoyl-CoA desaturase (SCD), the rate-limiting enzyme for fatty acid desaturation, stabilized αS tetramers and improved motor deficits in αS mice. Here we show that mutant GBA-PD cultured neurons have increased SCD products (monounsaturated fatty acids, MUFAS) and reduced αS T:M ratios that were improved by inhibiting SCD. Oral treatment of symptomatic L444P- and E326K Gba1 mutant mice with 5b also improved the αS T:M homeostasis and dopaminergic striatal integrity. Moreover, SCD inhibition normalized GCase maturation and dampened lysosomal and lipid-rich clustering, key features of neuropathology in GBA-PD. In conclusion, this study supports brain MUFA metabolism links GBA1 genotype and wildtype αS homeostasis to downstream neuronal and behavioral impairments, identifying SCD as a therapeutic target for GBA-PD.

Authors

Silke Nuber, Harrison Hsiang, Esra'a Keewan, Tim E. Moors, Sydney J. Reitz, Anupama Tiwari, Gary P. H. Ho, Elena Su, Wolf Hahn, Marie-Alexandre Adom, Riddhima Pathak, Matthew Blizzard, Sangjune Kim, Han Seok Ko, Xiaoqun Zhang, Per Svenningsson, Dennis J. Selkoe, Saranna Fanning

mTOR signaling regulates the development of airway mucous cell metaplasia associated with severe asthma

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Abstract

In asthma, airway epithelial remodeling is characterized by aberrant goblet cell metaplastic differentiation accompanied by epithelial cell hyperplasia and hypertrophy. These pathologic features in severe asthma indicate a loss of control of proliferation, cell size, differentiation, and migration. mTOR is a highly conserved pathway that regulates protein synthesis, cell size, and proliferation. We hypothesized that the balance between mTOR and autophagy regulates mucous cell metaplasia. Airways from individuals with severe asthma showed increased mTOR signaling by RPS6 phosphorylation, which was reproduced using an IL-13-activated model of primary human airway epithelial cells (hAECs). mTOR inhibition by rapamycin led to a decrease of IL-13-mediated cell hypertrophy, hyperplasia, and MUC5AC mucous metaplasia. BrdU labeling during IL-13-induced mucous metaplasia confirmed that mTOR was associated with increased basal-to-apical hAEC migration. mTOR activation by genetic deletion of Tsc2 in cultured mouse AECs increased IL-13-mediated hyperplasia, hypertrophy, and mucous metaplasia. Transcriptomic analysis of IL-13-stimulated hAEC identified mTOR-dependent expression of genes associated with epithelial migration and cytoskeletal organization. In summary, these findings point to IL-13-dependent and independent roles of mTOR signaling in the development of pathogenic epithelial changes contributing to airway obstruction in severe asthma.

Authors

Katrina M. Kudrna, Luis F. Vilches, Evan M. Eilers, Shailendra K. Maurya, Steven L. Brody, Amjad Horani, Kristina L. Bailey, Todd A. Wyatt, John D. Dickinson

NEXN regulates vascular smooth muscle cells phenotypic switching and neointimal hyperplasia

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Abstract

Vascular smooth muscle cells (VSMCs) exhibit significant heterogeneity and plasticity, enabling them to switch between contractile and synthetic states, which is crucial for vascular remodeling. NEXN has been identified as a high confidence gene associated with dilated cardiomyopathy (DCM). Existing evidence indicate NEXN is involved in phenotypic switching of VSMCs. However, a comprehensive understanding of the cell-specific roles and precise mechanisms of NEXN in vascular remodeling remains elusive. Using integrative transcriptomics analysis and smooth muscle specific lineage tracing mice, we demonstrate NEXN is highly expressed in VSMCs, and the expression of NEXN is significantly reduced during the phenotypic transformation of VSMCs and intimal hyperplasia induced by vascular injury. VSMC-specific NEXN deficiency promoted the phenotypic transition of VSMCs and exacerbated neointimal hyperplasia in mice following vascular injury. Mechanistically, we found NEXN primarily mediated VSMCs proliferation and phenotypic transition through endoplasmic reticulum (ER) stress and KLF4 signaling. Inhibiting ER stress ameliorated VSMCs phenotypic transition by reducing cell cycle activity and proliferation caused by NEXN deficiency. These findings indicate targeting NEXN could be explored as a promising therapeutic approach for proliferative arterial diseases.

Authors

Zexuan Lin, Chaojie Wang, Zhuohua Wen, Zhaohui Cai, Wenjie Guo, Xin Feng, Zengyan Huang, Rongjun Zou, Xiaoping Fan, Canzhao Liu, Hanyan Yang