Research Article
Abstract
Bardet-Biedl syndrome (BBS), a ciliopathy characterized by obesity, hyperphagia, and learning deficits, arises from mutations in Bbs genes. Exacerbated symptoms occur with mutations in genes encoding the BBSome, a complex regulating primary cilia function. We investigated the mechanisms underlying BBS-induced obesity using a Bbs5-knockout (Bbs5–/–) mouse model. Bbs5–/– mice were characterized by hyperphagia, learning deficits, glucose/insulin intolerance, and disrupted metabolic hormones, phenocopying human BBS. White adipose tissue in these mice had a unique immunophenotype, with increased proinflammatory macrophages and dysfunctional Tregs, suggesting a mechanism for adiposity distinct from those of typical obesity models. Additionally, Bbs5–/– mice exhibited pancreatic islet hyperplasia but failed to normalize blood glucose, suggesting defective insulin action. Hypothalamic transcriptomics revealed dysregulation of endocrine signaling pathways, with functional analyses confirming defects in insulin, leptin, and cholecystokinin (CCK) signaling, while glucagon-like peptide-1 receptor (GLP-1R) responsiveness was preserved. Notably, treatment with a GLP-1RA effectively alleviated hyperphagia and body weight gain, improved glucose tolerance, and regulated circulating metabolic hormones in Bbs5–/– mice. This study suggests that Bbs5–/– mice represent a valuable translational model of BBS for understanding pathogenesis and developing better treatments. Our findings highlight the therapeutic potential of GLP-1RAs for managing BBS-associated metabolic dysregulation, indicating that further investigation for clinical application is warranted.
Authors
Arashdeep Singh, Naila Haq, Mingxin Yang, Shelby Luckey, Samira Mansouri, Martha Campbell-Thompson, Lei Jin, Sofia Christou-Savina, Guillaume de Lartigue
Abstract
In the kidney, cells of thick ascending limb of the loop of Henle (TAL) are resistant to ischemic injury, despite high energy demands. This adaptive metabolic response is not fully understood even though the integrity of TAL cells is essential for recovery from acute kidney injury (AKI). TAL cells uniquely express uromodulin, the most abundant protein secreted in healthy urine. Here, we demonstrate that alternative splicing generates a conserved intracellular isoform of uromodulin, which contributes to metabolic adaptation of TAL cells. This splice variant was induced by oxidative stress and was upregulated by AKI that is associated with recovery, but not by severe AKI and chronic kidney disease (CKD). This intracellular variant was targeted to the mitochondria, increased NAD+ and ATP levels, and protected TAL cells from hypoxic injury. Augmentation of this variant using antisense oligonucleotides after severe AKI improved the course of injury. These findings underscore an important role of condition-specific alternative splicing in adaptive energy metabolism to hypoxic stress. Enhancing this protective splice variant in TAL cells could become a therapeutic intervention for AKI.
Authors
Azuma Nanamatsu, George J. Rhodes, Kaice A. LaFavers, Radmila Micanovic, Virginie Lazar, Shehnaz Khan, Daria Barwinska, Shinichi Makino, Amy Zollman, Ying-Hua Cheng, Emma H. Doud, Amber L. Mosley, Matthew J. Repass, Malgorzata M. Kamocka, Aravind Baride, Carrie L. Phillips, Katherine J. Kelly, Michael T. Eadon, Jonathan Himmelfarb, Matthias Kretzler, Robert L. Bacallao, Pierre C. Dagher, Takashi Hato, Tarek M. El-Achkar
Abstract
Patients with systemic lupus erythematosus (SLE) are photosensitive, developing skin inflammation with even ambient ultraviolet radiation (UVR), and this cutaneous photosensitivity can be associated with UVR-induced flares of systemic disease, which can involve increased autoantibodies and further end-organ injury. Mechanistic insight into the link between the skin responses and autoimmunity is limited. Signals from skin are transmitted directly to the immune system via lymphatic vessels, and here we show evidence for potentiation of UVR-induced lymphatic flow dysfunction in SLE patients and murine models. Improving lymphatic flow by manual lymphatic drainage (MLD) or with a transgenic model with increased lymphatic vessels reduces both cutaneous inflammation and lymph node B and T cell responses, and long-term MLD reduces splenomegaly and titers of a number of autoantibodies. Mechanistically, improved flow restrains B cell responses in part by stimulating a lymph node fibroblastic reticular cell-monocyte axis. Our results point to lymphatic modulation of lymph node stromal function as a link between photosensitive skin responses and autoimmunity and as a therapeutic target in lupus, provide insight into mechanisms by which the skin state regulates draining lymph node function, and suggest the possibility of MLD as an accessible and cost-effective adjunct to add to ongoing medical therapies for lupus and related diseases.
Authors
Mir J. Howlader, William G. Ambler, Madhavi Latha S. Chalasani, Aahna Rathod, Ethan S. Seltzer, Ji Hyun Sim, Jinyeon Shin, Noa Schwartz, William D. Shipman III, Dragos C. Dasoveanu, Camila B. Carballo, Ecem Sevim, Salma Siddique, Yurii Chinenov, Scott A. Rodeo, Doruk Erkan, Raghu P. Kataru, Babak J. Mehrara, Theresa T. Lu
Abstract
Diffuse midline gliomas (DMGs) are lethal brain tumors characterized by p53-inactivating mutations and oncohistone H3.3K27M mutations that rewire the cellular response to genotoxic stress. We used RCAS/tv-a retroviruses and Cre recombinase to inactivate p53 and induce native H3.3K27M mutations in a lineage- and spatially directed manner. We generated primary mouse tumors that recapitulated human DMG. Disrupting ataxia-telangiectasia mutated (ATM) kinase enhanced the efficacy of radiation therapy (RT) in murine and patient-derived DMG models and increased survival. Microscopy-based in situ sequencing was used to spatially resolve transcriptional profiles in more than 750,000 single cells with or without ATM disruption and RT, revealing altered immune-neoplastic and endothelial cell interactions after treatment. An allelic series of primary murine DMG models with different p53 mutations confirmed that transactivation-independent p53 activity was a key mediator of radiosensitivity after ATM disruption. We generated primary DMG mouse models and performed deep profiling that revealed mechanisms of response to ATM disruption and RT that can be utilized as a therapeutic strategy.
Authors
Avani Mangoli, Vennesa Valentine, Spencer M. Maingi, Sophie R. Wu, Harrison Q. Liu, Michael Aksu, Vaibhav Jain, Bronwen E. Foreman, Joshua A. Regal, Loren B. Weidenhammer, Connor E. Stewart, Maria E. Guerra Garcia, Emily Hocke, Karen Abramson, Tal Falick Michaeli, Nerissa T. Williams, Lixia Luo, Megan Romero, Katherine Deland, Samantha Gadd, Eita Uchida, Laura Attardi, Kouki Abe, Rintaro Hashizume, David M. Ashley, Oren J. Becher, David G. Kirsch, Simon G. Gregory, Zachary J. Reitman
Abstract
The presence of B cells is essential for the formation of CD8+ T cell memory after infection and vaccination. In this study, we investigated whether B cells influence the programming of naive CD8+ T cells prior to their involvement in an immune response. RNA sequencing indicated that B cells are necessary for sustaining the FOXO1-controlled transcriptional program, which is critical for homeostasis of these T cells. Without an appropriate B cell repertoire, mouse naive CD8+ T cells exhibit a terminal, effector-skewed phenotype, which significantly impacts their response to vaccination. A similar effector-skewed phenotype with reduced FOXO1 expression was observed in naive CD8+ T cells from human patients undergoing B cell–depleting therapies. Furthermore, we show that patients without B cells have a defect in generating long-lived CD8+ T cell memory following COVID vaccination. In summary, we demonstrate that B cells promote the quiescence of naive CD8+ T cells, poising them to become memory cells upon vaccination.
Authors
Cameron Manes, Miguel Guerrero Moreno, Jennifer Cimons, Marc A. D’Antonio, Tonya M. Brunetti, Michael G. Harbell, Sean Selva, Christopher Mizenko, Tyler L. Borko, Erika L. Lasda, Jay R. Hesselberth, Elena W.Y. Hsieh, Michael R. Verneris, Amanda L. Piquet, Laurent Gapin, Ross M. Kedl, Jared Klarquist
Abstract
Autosomal dominant optic atrophy (ADOA), the most prevalent hereditary optic neuropathy, leads to retinal ganglion cell (RGC) degeneration and vision loss. ADOA is primarily caused by mutations in the optic atrophy type 1 (OPA1) gene, which encodes a conserved GTPase important for mitochondrial inner membrane dynamics. To date, the disease mechanism remains unclear, and no therapies are available. We generated a mouse model carrying the pathogenic Opa1R290Q/+ allele that recapitulated key features of human ADOA, including mitochondrial defects, age-related RGC loss, optic nerve degeneration, and reduced RGC functions. We identified sterile alpha and TIR motif containing 1 (SARM1), a neurodegeneration switch, as a key driver of RGC degeneration in these mice. Sarm1 KO nearly completely suppressed all the degeneration phenotypes without reversing mitochondrial fragmentation. Additionally, we show that a portion of SARM1 localized within the mitochondrial intermembrane space. These findings indicated that SARM1 was activated downstream of mitochondrial dysfunction in ADOA, highlighting it as a promising therapeutic target.
Authors
Chen Ding, Papa S. Ndiaye, Sydney R. Campbell, Michelle Y. Fry, Jincheng Gong, Sophia R. Wienbar, Whitney Gibbs, Philippe Morquette, Luke H. Chao, Michael Tri H. Do, Thomas L. Schwarz
Abstract
Hormone receptor–positive and human epidermal growth factor receptor 2–negative breast cancer (HR+/HER2− BC) is the most common subtype, with a high risk of long-term recurrence and metastasis. Endocrine therapy (ET) combined with cyclin-dependent kinase 4/6 (CDK4/6) inhibitors is a standard treatment for advanced/metastatic HR+/HER2– BC, but resistance remains a major clinical challenge. We report that kinesin family member C2 (KIFC2) was amplified in approximately 50% of patients with HR+/HER2– BC, and its high expression was associated with poor disease outcome, increased tumor protein p53 (TP53) somatic mutation, and active pyrimidine metabolism. Functional assays revealed that depletion of KIFC2 suppressed growth and enhanced sensitivity of HR+/HER2– BC cells to tamoxifen and CDK4/6 inhibitors. Mechanistically, KIFC2 stabilized CDK4 by enhancing its interaction with ubiquitin-specific peptidase 9 X-linked (USP9X). Importantly, reexpression of CDK4 in KIFC2-depleted cells partially rescued the decreased growth and increased sensitivity to tamoxifen and CDK4/6 inhibitors caused by KIFC2 depletion. Clinically, high KIFC2 mRNA expression was negatively associated with the survival rate of patients with HR+/HER2– BC who received adjuvant ET alone or in combination with CDK4/6 inhibitors. Collectively, these findings identify an important role for KIFC2 in HR+/HER2– BC growth and therapeutic resistance, and support its potential as a therapeutic target and predictive biomarker.
Authors
Shao-Ying Yang, Ming-Liang Jin, Lisa Andriani, Qian Zhao, Yun-Xiao Ling, Cai-Jin Lin, Min-Ying Huang, Jia-Yang Cai, Yin-Ling Zhang, Xin Hu, Zhi-Ming Shao, Fang-Lin Zhang, Xi Jin, A Yong Cao, Da-Qiang Li
Abstract
As antimicrobial resistance rises, new antibacterial candidates are urgently needed. Using sequence space information from over 14,743 functional antimicrobial peptides (AMPs), we improved the antimicrobial properties of citropin 1.1, an AMP with weak antimethicillin resistant Staphylococcus aureus (MRSA) activity, producing a short and potent antistaphylococcal peptide, CIT-8 (13 residues). At 40 μg/mL, CIT-8 eradicated 1 × 108 drug-resistant MRSA and vancomycin resistant S. aureus (VRSA) persister cells within 30 minutes of exposure and reduced the number of viable biofilm cells of MRSA and VRSA by 3 log10 and 4 log10 in established biofilms, respectively. CIT-8 (at 32 μg/mL) depolarized and permeated the S. aureus MW2 membrane. In a mouse model of MRSA skin infection, CIT-8 (2% w/w in petroleum jelly) significantly reduced the bacterial burden by 2.3 log10 (P < 0.0001). Our methodology accelerated AMP design by combining traditional peptide design strategies, such as truncation, substitution, and structure-guided alteration, with machine learning–backed sequence optimization.
Authors
Biswajit Mishra, Anindya Basu, Fadi Shehadeh, LewisOscar Felix, Sai Sundeep Kollala, Yashpal Singh Chhonker, Mandar T. Naik, Charilaos Dellis, Liyang Zhang, Narchonai Ganesan, Daryl J. Murry, Jianhua Gu, Michael B. Sherman, Frederick M. Ausubel, Paul P. Sotiriadis, Eleftherios Mylonakis
Abstract
Weight loss medications are emerging candidates for pharmacotherapy of sleep-disordered breathing (SDB). A melanocortin 4 receptor (MC4R) agonist, setmelanotide (Set), is used to treat obesity caused by abnormal melanocortin and leptin signaling. We hypothesized that Set can treat SDB in mice with diet-induced obesity. We performed a proof-of-concept randomized crossover trial of a single dose of Set versus vehicle and a 2-week daily Set versus vehicle trial, examined colocalization of Mc4r mRNAs with the markers of CO2-sensing neurons Phox2b and neuromedin B in the brainstem, and expressed Cre-dependent designer receptors exclusively activated by designer drugs (DREADDs) or caspase in obese Mc4r-Cre mice. Set increased minute ventilation across sleep/wake states, enhanced the hypercapnic ventilatory response (HCVR), and abolished apneas during sleep. Phox2b+ neurons in the nucleus of the solitary tract (NTS) and the parafacial region expressed Mc4r. Chemogenetic stimulation of the MC4R+ neurons in the parafacial region, but not in the NTS, augmented HCVR without any changes in metabolism. Caspase elimination of the parafacial MC4R+ neurons abolished effects of Set on HCVR. Parafacial MC4R+ neurons projected to the respiratory premotor neurons retrogradely labeled from C3–C4. In conclusion, MC4R agonists enhance the HCVR and treat SDB by acting on the parafacial MC4R+ neurons.
Authors
Mateus R. Amorim, Noah R. Williams, O. Aung, Melanie Alexis Ruiz, Frederick Anokye-Danso, Junia L. de Deus, Jiali Xiong, Olga Dergacheva, Shannon Bevans-Fonti, Sean M. Lee, Jeffrey S. Berger, Mark N. Wu, Rexford S. Ahima, David Mendelowitz, Vsevolod Y. Polotsky
Abstract
The efficacy of T cell–activating therapies against glioma is limited by an immunosuppressive tumor microenvironment and tumor-induced T cell sequestration. We investigated whether peripherally infused nonantigen specific autologous lymphocytes could accumulate in intracranial tumors. We observed that nonspecific autologous CD8+ ALT cells can indeed accumulate in this context, despite endogenous T cell sequestration in bone marrow. Rates of intratumoral accumulation were markedly increased when expanding lymphocytes with IL-7 compared with IL-2. Pretreatment with IL-7 ALT also enhanced the efficacy of multiple tumor-specific and nontumor-specific T cell–dependent immunotherapies against orthotopic murine and human xenograft gliomas. Mechanistically, we detected increased VLA-4 on mouse and human CD8+ T cells following IL-7 expansion, with increased transcription of genes associated with migratory integrin expression (CD9). We also observed that IL-7 increases S1PR1 transcription in human CD8+ T cells, which we have shown to be protective against tumor-induced T cell sequestration. These observations demonstrate that expansion with IL-7 enhances the capacity of ALT to accumulate within intracranial tumors and that pretreatment with IL-7 ALT can boost the efficacy of subsequent T cell–activating therapies against glioma. Our findings will inform the development of future clinical trials where ALT pretreatment can be combined with T cell–activating therapies.
Authors
Kirit Singh, Kelly M. Hotchkiss, Sarah L. Cook, Pamy Noldner, Ying Zhou, Eliese M. Moelker, Chelsea O. Railton, Emily E. Blandford, Bhairavy J. Puviindran, Shannon E. Wallace, Pamela K. Norberg, Gary E. Archer, Beth H. Shaz, Katayoun Ayasoufi, John H. Sampson, Mustafa Khasraw, Peter E. Fecci
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