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Proteomic analyses of human islets reveal potential markers of β-cell dysfunction during prediabetes
Abstract
The mechanisms driving progressive beta-cell dysfunction in type 2 diabetes (T2D) remain incompletely understood. This study aimed to identify pancreatic islet proteome changes that could predict diabetes onset. We isolated islets from non-diabetic subjects undergoing partial pancreatectomy, previously characterized for glucose tolerance, insulin sensitivity, and insulin secretion, using laser capture microdissection (LCM) and analyzed them via high-performance liquid chromatography-mass spectrometry (HPLC-MS). Proteomic analysis revealed that subjects with impaired glucose tolerance (IGT) had reductions in proteins regulating glycolysis (PGK1, G3P), lipid metabolism (ACBP, ARF1), glucose transport (14-3-3B), and insulin secretion (STARD10, CAPDS) compared to normal glucose tolerant (NGT) subjects. Additionally, IGT islets showed impaired expression of proteins involved in glucose- and incretin-stimulated insulin response (CREB1, IQGA1). Stratification by beta-cell glucose sensitivity (βGS) indicated that subjects with lower βGS exhibited reduced levels of insulin maturation (ERO1B) and anti-apoptotic proteins (CASP8, PAK2, SKP1), along with increased SEL1L, a factor promoting endocrine precursor differentiation. These findings suggest that early defects in glucose metabolism and insulin secretion characterize IGT, while reduced βGS may trigger compensatory mechanisms, through enhanced beta-cell survival or neogenesis, to delay T2D progression. Overall, proteomic alterations in prediabetic islets provide potential early predictive markers and targets for interventions aimed at preserving beta-cell function.
Authors
Chiara Maria Assunta Cefalo, Teresa Mezza, Giuseppe Quero, Sergio Alfieri, Donatella Lucchetti, Filomena Colella, Alessandro Sgambato, Wei-Jun Qian, Andrea Mari, Alfredo Pontecorvi, Andrea Giaccari, Rohit N. Kulkarni
Abstract
Cytotoxic T lymphocytes form a critical component of SARS-CoV-2 immunity by recognizing viral peptides bound to HLA class I molecules. Here, we identified the Spike-derived peptide NYNYLYRLF448-456 (NF9) as the immunodominant HLA-A*24:02-restricted epitope in both convalescent and vaccinated donors. Across cohorts, A24/NF9-specific responses were dominated by public TCR motifs featuring TRAV12-1 (or TRAV6-1) paired with TRBJ2-7 and a conserved CDR3β sequence (CASSXXXGYEQYF). Using a panel of thirteen TCRs, we mapped recognition of single amino acid substitutions within NF9 and identified residue 5 (L452) as the principal determinant of escape. The L452R substitution, characteristic of the Delta variant, abolished recognition across all tested TCRs despite preserved HLA binding. Crystallography of a representative public TCR (P1-15) revealed that mutation at position 5 reoriented the peptide within HLA-A*24:02, flipping the adjacent Y453 side chain into the peptide-binding groove and eliminating the dominant TCR contact. This position-5-driven conformational switch provided a structural mechanism for universal loss of NF9 recognition by HLA-A*24:02-restricted T-cells. Consistent with this, Delta-infected convalescents failed to mount de novo NF9-5R-specific responses while retaining responses to the conserved A24/QI9 Spike epitope. Together, these findings defined the basis of A24/NF9 recognition and showed how one mutation remodelled peptide presentation to abrogate TCR responses.
Authors
Takeshi Nakama, Aaron Wall, Garry Dolton, Li-Rong Tan, Hannah Thomas, Hiroshi Hamana, Yoshiki Aritsu, Toong Seng Tan, Mako Toyoda, Yoshihiko Goto, Huanyu Li, Mizuki Kitamatsu, Keiko Udaka, Yusuke Miyashita, Hiroyuki Oshiumi, Kimitoshi Nakamura, Yoji Nagasaki, Rumi Minami, Hirotomo Nakata, Pierre J. Rizkallah, Hiroyuki Kishi, Takamasa Ueno, Andrew K. Sewell, Chihiro Motozono
Abstract
Sleep disturbance is a prevalent yet poorly understood comorbidity in autism spectrum disorders (ASD). Here, we uncover a bidirectional regulatory axis connecting the ASD risk gene POGZ to core circadian mechanisms. We demonstrate that Pogz is widely expressed in the suprachiasmatic nucleus (SCN), the central pacemaker of the circadian rhythms and exhibits circadian oscillations in both the hypothalamus and liver with its transcription directly regulated by the circadian molecule DBP through a D-box element in its proximal enhancer. Pogz-deficient mice exhibited prolonged circadian periodicity, impaired light-induced phase shift, delayed adaption to an 8-hour advance jet-lag, and reduced SCN c-Fos activation in response to light pulses. Mechanistically, POGZ interacts with and enhances the transcription activity of CREB, a key regulator of light-induced phase resetting. Notably, Pogz deletion leads to ASD-related deficits in social novelty and cognition, with cognitive impairments influenced by both photoperiod and behavioral paradigm. Our findings thus reveal a critical, previously unrecognized intersection between an ASD risk gene and circadian clock, offering new insights into the pathogenesis of core ASD symptoms and comorbid sleep disturbances.
Authors
Ting Wu, Jiao He, Chu-Jun Xu, Chi-Yu Li, Pingchuan Zhang, Yanfeng Wang, Shanshan Zhu, Lusi Zhang, Jingtan Zhu, Jing Zhang, Jia-Da Li, Huadie Liu
Abstract
The lung alveoli are continually exposed to inhaled pathogens and environmental hazards and rely on coordinated communication between alveolar macrophages and type 2 alveolar epithelial cells (AT2s) to maintain homeostasis. Disruption of these interactions can impair immunity and repair, contributing to acute and chronic respiratory diseases. To better define these mechanisms and support therapeutic discovery, we established a human iPSC-derived air-liquid interface platform that captures key features of AT2-macrophage crosstalk. Using this system, we show that coculture enhances AT2-specific transcriptional programs including lipid synthesis, while macrophages actively phagocytose AT2-derived surfactant. iPSC-derived macrophages adopt an alveolar macrophage-like phenotype and respond to AT2-derived M-CSF. During respiratory infection, macrophages played a crucial role in modulating epithelial inflammatory responses, augmenting antiviral immunity, and limiting viral replication. We further identify a previously unrecognized role for macrophages in epithelial repair, where VEGF-mediated signaling to macrophages increases epithelial permeability during viral infection. Together, these findings reveal dimensions of AT2-macrophage cooperation in homeostasis, infection, and repair, and demonstrate how this iPSC-derived platform can be used to dissect mechanisms that may initiate or drive the progression of respiratory diseases.
Authors
Declan L. Turner, Hannah Baric, Katelyn Patatsos, Sahel Amoozadeh, Michael See, Kathleen A. Strumila, Jack T. Murphy, Jeremy J. Wiyana, Liam Gubbels, Elizabeth S. Ng, Andrew G. Elefanty, Melanie R. Neeland, Shivanthan Shanthikumar, Sarah L. Londrigan, Mirana Ramialison, Fernando J. Rossello, Edouard G. Stanley, Rhiannon B. Werder
Abstract
Maternal opioid use disorder (OUD) poses substantial risks to maternal and fetal health. These adverse outcomes are believed to be mediated, in part, by changes in placenta structure and function; however, few studies have addressed this question. Here, we utilized flow cytometry, histology, spatial and single-cell transcriptomics to uncover the impact of OUD on placental tissues. Given that half of subjects with chronic OUD contract hepatitis C (HCV), we further stratified our findings by maternal HCV status. Our results indicate that OUD leads to higher incidence of vascular malperfusion accompanied by increased levels of inflammatory markers and dysregulated secretion of placental development factors. Spatial transcriptomics revealed that OUD disrupts the communication between trophoblasts and immune cells important for placental vascular development. Additionally, CellChat analysis revealed aberrant vascular remodeling, neuropeptide, and chemotactic signaling across trophoblast, endothelial, and myeloid cells. Processes associated with tissue homeostasis and repair were also upregulated across trophoblast and leukocytes. In addition, placental leukocytes were rewired towards regulatory/tissue surveillant phenotypes. Finally, frequencies and responses to ex-vivo stimulation of decidual macrophages and cytolytic NKcells, critical for tissue remodeling and fetal tolerance, were decreased. Altogether, these results highlight substantial disruptions to placental health by maternal OUD.
Authors
Heather E. True, Brianna M. Doratt, Sheridan B. Wagner, Delphine C. Malherbe, Nathan R. Shelman, Mahdi Eskandarian Boroujeni, Cynthia Cockerham, John M. O'Brien, Ilhem Messaoudi
Abstract
Resident cardiac fibroblast (RCF)-derived cardiac myofibroblasts (CMF) contribute to myocardial repair but also drive adverse ventricular remodeling and contractile dysfunction after myocardial infarction (MI). The sodium-activated potassium channel Slick (Slo2.1) has been described in cardiomyocyte (CM) mitochondria; however, transcriptomic analyses indicate higher Slick expression in RCFs/CMFs. Here, we investigated the role of Slick in cardiac fibroblast function and post-MI remodeling. Using live-cell imaging and whole-cell patch-clamp recordings, we found that plasma membrane Slick channels in RCFs and CMFs regulated potassium (K+) efflux and modulated store-operated calcium entry (SOCE), particularly in CMFs. Global Slick KO and conditional CMF-specific KO hearts exhibited reduced fibrosis and preserved left ventricular function following ischemia/reperfusion injury. This cardioprotection was associated with diminished CMF activation and proliferation, reduced inflammation, and improved CM survival post-MI. Collectively, these findings identify fibroblast Slick channels as regulators of SOCE-dependent fibrogenesis and demonstrate that their deletion mitigates maladaptive remodeling and functional decline after MI.
Authors
Jiaqi Yang, Lin Zhu, David Spähn, Melanie Cruz Santos, Sophia Schanz, Selina Maier, Lena Birkenfeld, Helmut Bischof, Anna Roslan, Nina Wettschureck, Oliver Borst, Lucas Matt, Robert Lukowski
Abstract
Glucagon-like peptide-1 (GLP-1) and glucose-induced insulinotropic polypeptide (GIP) receptor agonists have revolutionized obesity therapy but causes for obesity-associated dysregulation of endogenous incretin production remain incompletely understood. Here we show that intestinal transmembrane serine protease 2 (TMPRSS2) plays a pivotal role in deregulating anti-diabetic GLP-1 production in obesity. TMPRSS2 is widely co-expressed in intestinal epithelial cells (IEC) along with its signaling target protease activated receptor 2 (PAR2). In addition to its role in regulating coagulation protease-mediated adipose tissue inflammation, PAR2 signaling in the gut controls postprandial GIP secretion. TMPRSS2, but not the epithelial-expressed proteases FXa or matriptase, activates PAR2 and thereby promotes postprandial GIP release. Accordingly, a PAR2 mutant mouse resistant to TMPRSS2 cleavage is protected from GIP upregulation and diet induced obesity. In the context of obesity, TMPRSS2 also attenuates bioavailability of ghrelin pathway and thereby suppresses GLP-1-mediated control of glucose homeostasis. Pharmacological inhibition or genetic deletion of TMPRSS2 restores ghrelin signaling dependent GLP-1 secretion and GLP-1’s anti-diabetic effects on nutritional glucose homeostasis. Thus, epithelial cell-expressed TMPRSS2, which critically contributes to the lung pathology in SARS-CoV-2 infection, emerges as an intestinal incretin regulator and a potential link between infection and chronic cardiometabolic diseases.
Authors
Dilraj Kaur, Sagarika Chakrabarty, Claudius Witzler, Hongjie Wang, Mengwen Wang, Romina Wolz, Petra Wilgenbus, Jens J.N. Posma, P. Sivaramakrishna Rachakonda, Federico Marini, Valeriya V. Zinina, Sabine Reyda, Rajinikanth Gogiraju, Claudine Graf, Fahumiya Samad, Katrin Schäfer, Christoph Reinhardt, Natalia Soshnikova, Wolfram Ruf, Thati Madhusudhan
Abstract
Dengue virus (DENV) vaccines should be designed to induce balanced protective immunity against all four dengue serotype to mitigate the risk of vaccine-mediated enhanced dengue disease. The first tetravalent vaccine (Dengvaxia) tested in humans was efficacious in children who were partially immune to DENV at baseline. In DENV-naive children, the vaccine was not efficacious and placed some naïve children at risk of experiencing more severe wild-type DENV breakthrough infections. To define dengue vaccine responses at the individual subject level and their relationship to mild and severe dengue infections, we prospectively studied a cohort of DENV-naive children who received one dose of Dengvaxia. The vaccine stimulated variable responses that neutralized 0, 1 (monotypic), or 2+ (multitypic) serotypes in individual children. We used a logistic regression model to evaluate whether vaccine status and serotype-specific NAb status at the end of study period 1 influenced the probability of experiencing a virologically confirmed dengue disease (VCD) case thereafter (months 20 - 60). Vaccinated children with NAb response to only one serotype were at greater risk of being a case compared to the DENV-naïve control group (Odds Ratio 5.07). This risk was not observed in vaccinated children with no NAb or NAb to 2 or more serotypes. We propose that individuals with durable NAb to one serotype have an abundance of serotype cross-reactive, non-neutralizing Abs implicated in the enhanced replication of heterologous serotypes. We discuss the implications of our findings for flagging vaccine candidates that are likely to pose a special risk to seronegative subjects.
Authors
Laura J. White, Lindsay D. Hein, Maria Abad Fernandez, Cameron Adams, Elizabeth Adams, Emily Freeman, Ruby Shah, Lakshmanane Premkumar, Kristal An Agrupis, Maria Vinna Crisostomo, Jedas Veronica Daag, Michelle Ylade, Jacqueline Deen, Ana Lena Lopez, Leah Katzelnick, Aravinda M. de Silva
Abstract
Allogeneic islet transplantation is an effective treatment for type 1 diabetes, but its clinical use is limited by rejection involving innate and adaptive immune responses, requiring life-long immunosuppression. We herein engineered islets by transiently display two immunomodulators chimeric with streptavidin (SA), thrombomodulin (SA-TM) and CD47 (SA-CD47) for localized modulation of both innate and adaptive immune responses. The engineering process did not impact islet viability, glucose responsiveness, and metabolic activity. Intraportal transplantation into allogeneic recipients achieved sustained survival, with 8/11 grafts surviving 120-330 days without immunosuppression. In contrast, non-engineered islets were acutely rejected [median survival time (MST) = 12 days], while islets engineered with SA-TM showed delayed rejection [Median survival time (MST) = 13.5 days], and those with SA-CD47 exhibited prolonged survival (MST = 24 days). Double-engineered islets generated a localized tolerogenic immune environment characterized by low frequencies of inflammatory innate immune cells and increased frequencies of M2 macrophages, myeloid-derived suppressor cells, and CD4+FoxP3+ T regulatory cells. The transcriptomic analysis showed downregulation of proinflammatory and upregulation of immune regulatory pathways. Our results demonstrate that transient co-display of immunomodulatory molecules on the islet surface is a versatile platform with significant translational potential for islet transplantation.
Authors
Shadab Kazmi, Mohammad Tarique, Darshan Badal, Vahap Ulker, Ali Turan, Kathleen M. Yee-Flores, Abdalmonam Jadou Nejma, Esma S. Yolcu, Haval Shirwan
Abstract
BACKGROUND In chronic alcohol consumers, immune cells may drive the progression from mild liver injury to more severe alcohol-associated liver diseases (ALD), including alcohol-associated hepatitis (AAH) and cancer. Liver macrophages, both resident and infiltrating, express Allograft Inflammatory Factor 1 (AIF1), which is upregulated during inflammation and enhances immune activation. METHODS Using serum and urine samples from 868 individuals classified as having alcohol use disorder or not based on DSM-IV/V criteria, along with serum and liver biopsy tissue from a second cohort of 27 patients diagnosed with AAH, we evaluated the impact of the AIF1 promoter single nucleotide polymorphism (SNP) (rs3132451; C/C, C/G, G/G) on liver function markers and immune cell profiles. RESULTS AIF1 transcript levels were genotype-dependent: C/C homozygotes expressed 5.2% of the levels observed in G/G individuals, while C/G heterozygotes expressed 46%. Unlike most SNPs associated with harmful effects, the G/G genotype is highly prevalent, present in ~70% of patients. Among chronic alcohol users, G/G individuals exhibited elevated markers of liver injury and a more than threefold increase in hepatic immune cells, including infiltrating AIF1⁺ macrophages and neutrophils. Despite similar durations of alcohol misuse, G/G individuals had higher Model for End-Stage Liver Disease scores compared to C/G individuals, indicating a significantly greater 90-day mortality risk. Notably, some immune abnormalities, such as elevated neutrophils, persisted in G/G males even after alcohol abstinence. CONCLUSION These findings suggest that functional genetic variation in AIF1 may contribute to the severity and persistence of ALD. TRIAL REGISTRATION ClinicalTrials.gov NCT02231840. FUNDING Research support was provided from the National Institute on Alcohol Abuse and Alcoholism (NIAAA) of the National Institutes of Health (NIH) under grant 1ZIAAA000440-02 and R24AA025017.
Authors
Priscila C. Antonello, Colin A. Hodgkinson, Dechun Feng, Cheryl Marietta, Baskar Mohana Krishnan, Maria A. Parra, Zhaoli Sun, Bin Gao, David Goldman, Michelle W. Antoine
Abstract
Introduction: BK polyomavirus (BKV) infection is associated with injury and subsequent graft loss due to the extent of injury or rejection. However, the molecular mechanisms driving injury and subsequent adverse outcomes remain poorly understood. Methods: In a cross-sectional study, single-cell RNA sequencing from kidney allograft biopsies was used to assess cell type-specific responses between uninfected controls and two distinct phases of BKV infection: peaking (increasing viral blood titers) and resolving (decreasing viral titers following immunosuppression reduction). Results: Genes upregulated in BK viral nephropathy (BKVN) were enriched for polyomavirus infection hallmarks, including ribosome biogenesis, translation, and energy restructuring. Additionally enriched pathways included wound healing, cellular stress, antigen presentation and immune signaling. Even without BKVN (peaking BK viremia alone), epithelial cells expressed signatures for wound healing, cellular stress, and extracellular matrix remodeling. In vivo tubular cell responses at single-cell resolution were validated against single cell transcriptomic data of BKV infected cells in a cell culture model. Despite similarities, in vivo tubular cells underwent metabolic adaptation favoring fatty acid oxidation and proinflammatory responses not observed in culture models likely due to an absent innate and adaptive immune system. Despite lymphopenia and immunosuppressive therapies, the proportion of recipient derived intrarenal adaptive immune cells was increased in biopsies associated with peaking viremia alongside activation of innate immune responses. Adaptive immune cells exhibited persistent inflammatory signaling and remodeling of energy metabolism during the resolving phase of infection. Conclusion: These not previously reported insights into BKV-associated injury may have implications for clinical management and improved allograft outcomes.
Authors
Tess Marvin, Rachel Sealfon, Phillip J. McCown, Fadhl AlAkwaa, Evan A. Farkash, Edgar A. Otto, Felix Eichinger, Ping An, Rajasree Menon, Celine C. Berthier, Tavis J. Reed, Paula Arrowsmith, Lalita Subramanian, Kelly J. Shaffer, Silas P. Norman, Ramnika Gumber, Michael J. Imperiale, James M. Pipas, Olga G. Troyanskaya, Matthias Kretzler, Chandra L. Theesfeld, Abhijit S. Naik
Abstract
Adipocytes exist along a functional spectrum: white adipocytes are energy storing while brown adipocytes have thermogenic capacity such that activation may counteract obesity-related disease. In between are UCP1-expressing beige adipocytes, which can transition between these two energetic states. We previously showed that bone morphogenetic protein 7 (BMP7), a member of the transforming growth factor-β (TGFβ) superfamily, enables differentiation of brown preadipocytes to mature thermogenic cells. To see if immortalized, clonal human white and brown preadipocytes (hWA and hBA, respectively) would become more thermogenic in response to BMP exposure, we treated them with BMP7 or BMP4 for the first 7d of a 30d differentiation protocol. In hBA, absence of either BMP7 or BMP4 led to lower expression of brown-specific markers and oxygen consumption relative to 7d with either BMP. hWA treated for 7d with either BMP did not increase expression of thermogenic protein UCP1 nor induce a brown-like transcription profile. However, BMP-treated hWA produced adipocytes that had higher basal and drug-induced maximal oxygen consumption, which was UCP1-independent and due substantially to the futile creatine cycle (FCC). Our results demonstrate that energetically quiescent human white preadipocytes can be pushed into an energy expending phenotype without transdifferentiation into beige adipocytes, providing a new approach to treat obesity-related metabolic disease.
Authors
Kelly T. Long, Cheryl Cero, Sahara L. Ali, Nhuquynh Nguyen, Adrienne R. Guarnieri, Ju Hee Kim, Young Jae Bahn, Jurgen Heymann, Jonathan M. Dreyfuss, Sushil G. Rane, Yu-Hua Tseng, Aaron M. Cypess
Abstract
BACKGROUND. Transplanting kidneys from donors with HIV to recipients with HIV has become standard clinical practice. However, donors with HIV may have higher prevalence of viral and bacterial infections and autoimmunity that could increase allograft rejection in recipients. METHODS. We included deceased kidney donors (60 with HIV and 41 without HIV) who participated in a multicenter prospective study of HIV kidney transplantation between April 2018-September 2021. Using Phage ImmunoPrecipitation Sequencing, we compared the human antibody repertoire (allergens, autoantibodies, viruses and bacterial toxins) between donors with and without HIV, and evaluated their association with recipient allograft rejection. Moderated t-tests were used to assess reactivity and a multivariate logistic regression model adjusted for donor sex and KDPI assessed the association between donor adenovirus reactivity and recipient allograft rejection. RESULTS. Compared to donors without HIV, donors with HIV had lower BMI and were more likely to be African American. The median number of positive autoantibodies was marginally higher among donors with HIV (499 [IQR = 357, 579]) compared to donors without HIV (395 [IQR = 256, 538] (P = 0.058). Donors with HIV additionally had significantly higher antibody reactivity to Epstein-Barr virus and cytomegalovirus (q < 0.05). Among all donors with and without HIV, antibodies to adenovirus were significantly associated with increased rejection among recipients, including after adjusting for false discovery (q < 0.05) and also adjusting for demographic factors using multivariable logistic regression (odds ratio = 4.97, 95% CI = 1.89–13.61). CONCLUSION. The presence of antibodies to adenovirus infection in kidney donors with HIV may be associated with allograft rejection. TRIAL REGISTRATION. ClinicalTrials.gov NCT03500315. FUNDING. US National Institute of Health
Authors
Xianming Zhu, William R. Morgenlander, Diane M. Brown, Yolanda Eby, Megan Morsheimer, Jonah Odim, Serena M. Bagnasco, Meenakshi M. Rana, Sander S. Florman, Rachel J. Friedman-Moraco, Peter G. Stock, Alexander J. Gilbert, Shikha Mehta, Valentina Stosor, Sapna A. Mehta, Marcus R. Pereira, Catherine B. Small, Michele I. Morris, Jonathan Hand, Saima Aslam, Ghady Haidar, Maricar Malinis, Carlos A.Q. Santos, Joanna Schaenman, David Wojciechowski, Karthik M. Ranganna, Emily Blumberg, Nahel Elias, Josa A. Castillo-Lugo, Emmanouil Giorgakis, Senu Apewokin, M. Kate Grabowski, Dorry L. Segev, Andrew D. Redd, Christine M. Durand, H. Benjamin Larman, Aaron A.R. Tobian
Abstract
The molecular mechanisms responsible for the “atopic march” of allergic skin disease to allergic airway disease are incompletely understood. Secreted phospholipase A2 group X (sPLA2-X) is implicated in human asthma and modulates airway hyperresponsiveness (AHR) and inflammation in murine models of allergic asthma. We developed a complete proteolytic allergen model of dermal sensitization followed by airway challenge to mimic the “atopic march” and examined the role of sPLA2-X in regulating peripheral allergen sensitization, AHR, and airway inflammation. Pla2g10-/- mice receiving both house dust mite (HDM) peripheral sensitization and airway challenge had attenuated AHR relative to WT mice and lower airway eosinophils. Transgenic C57BL/6 PLA2G10 mice (only expressing the human sPLA2-X gene) receiving treatment with a small molecule inhibitor of sPLA2-X (ROC0929) during the dermal sensitization phase demonstrated attenuated AHR and a reduction in lung tissue dust mite-specific tissue resident memory CD4+ T cells. Thus, sPLA2-X acts as an endogenous adjuvant to facilitate allergic sensitization in the periphery, which leads to AHR and airway inflammation following inhalation of the allergen. These results provide proof of concept that inhibition of sensitization in the periphery with a sPLA2-X inhibitor modulates subsequent allergen-induced airway dysfunction.
Authors
Ryan C. Murphy, Ying Lai, Yu-Hua Chow, Matt Liu, Brian D. Hondowicz, Dowon An, Marion Pepper, William A. Altemeier, Teal S. Hallstrand
Abstract
Background: The molecular landscape of lung adenocarcinoma (LUAD) is often illustrated as a driver-oncogene “pie chart,” but identical mutations exhibit heterogeneous signaling shaped by co-mutations, transcriptional programs, and lineage context. We propose a lineage-integrated signaling framework using an EGFR mutation signature (mSig). Methods: We defined EGFR mSig using differentially expressed genes in EGFR-mutant (mt) LUADs. Semi-supervised clustering and machine learning models were used to test reproducibility in different combinations of datasets. We analyzed molecular subtypes, lineage markers, co-occurring mutations and EGFR copy number alterations in EGFR mSig-defined subtypes of LUAD. Results: EGFR mSig showed robust classification performance (AUROC = 0.83-0.95; mean NPV = 96.3%). Validated gene expression subtypes and lung lineage markers were closely aligned with EGFR mSig status. Most EGFR mSig(+) tumors, including many without EGFR mutations belonged to Bronchioid subtype. A subset of canonical RAS mutations were mSig(+) and mirrored the EGFR mutation pattern. EGFR wild-type (WT)/mSig(-) tumors were enriched for non-Bronchioid subtypes and had co-mutations in TP53 or RAS/RAF/RTKs. We highlighted a parsimonious collection of coordinated mutations identified including RAS, KEAP1, STK11, TP53, and CDKN2A, supportive of prior reports. Conclusions: A novel EGFR mSig that captures the transcriptional footprint of EGFR activation revealed a subset of EGFR WT LUADs with “mt-like” features. mSig refines LUAD taxonomy beyond mutation-only pie-chart models by incorporating lineage and co-mutation context. Lineage-directed stratification with co-alteration identifies clinically relevant groups across EGFR and RAS states and highlights new treatment opportunities for patients currently considered “oncogene-negative.” Funding: NCI U01CA272541, R01CA262296, U24CA264021, UG1CA233333, R01CA211939.
Authors
Minjeong Kim, Wisut Lamlertthon, Heejoon Jo, Yan Cui, Miyeon Yeon, Hyo Young Choi, Katherine A. Hoadley, Matthew P. Smeltzer, Michele C. Hayward, Matthew D. Wilkerson, Liza Makowski, D. Neil Hayes
Abstract
Macrophages (MΦ), endowed with remarkable phenotypic plasticity are essential for orchestrating injury responses and regulating iron homeostasis. Given the central role of ferritin heavy chain (FtH) as a molecular rheostat linking iron sequestration to redox-dependent signaling, we examined how myeloid FtH governs renal iron trafficking and ensuing oxidative-stress pathways during acute kidney injury (AKI). Transcriptome analysis revealed coupling of FtH deficiency in monocytes and MΦ with ferroptosis activation, a regulated cell death associated with iron accumulation. Moreover, myeloid FtH deletion worsened AKI, increasing leukocyte infiltration and iron deposition, together with ferroptosis‐associated gene induction, oxidative stress, and lipid peroxidation. Notably, ⍺-synuclein (SNCA), an iron-binding protein and the main pathological driver of Parkinson’s disease, was robustly induced by both FtH deficiency and following AKI. Mechanistic studies showed that monomeric SNCA exhibits ferrireductase activity, amplifying redox cycling and promoting ferroptotic cell death. Furthermore, SNCA expression was elevated in kidney pathologies characterized by leukocyte expansion in both mouse models and human cohorts, suggesting that inflammatory microenvironments promote SNCA accumulation and redox imbalance. These findings define a MΦ FtH-SNCA regulatory axis as a key driver of ferroptosis in AKI, implicating SNCA as a pathological nexus between iron dyshomeostasis and inflammatory kidney injury.
Authors
Tanima Chatterjee, Sarah Machado, Kellen Cowen, Mary E. Miller, Bronte Johnson, Yanfeng Zhang, Laura A. Volpicelli-Daley, Lauren A. Fielding, Rudradip Pattanayak, Frida Rosenblum, László Potor, György Balla, Jozsef Balla, Christian Faul, Abolfazl Zarjou
Abstract
Moderate hyperoxia (30–60% O₂) in premature infants promotes bronchial airway hyperresponsiveness (AHR) via airway smooth muscle (ASM), a key regulator of bronchoconstriction, bronchodilation, and remodeling. Understanding how O2 exposure drives long-term bronchial changes in prematurity is critical for developing therapies for airway disease across the lifespan. Premature lungs have immature antioxidant defenses, potentially due to disrupted mitochondrial dynamics, increasing susceptibility to O2-induced oxidative stress. Thus, mitochondrial homeostasis is highly relevant to ASM dysfunction and airway disease. We propose that hyperoxia in prematurity promotes mitochondrial dysfunction, and that the gasotransmitter hydrogen sulfide (H₂S) mitigates O2-induced mitochondrial damage in developing ASM. Human fetal ASM (fASM) were exposed to moderate hyperoxia to investigate the effects of exogenous H₂S donors (GYY4137, AP39) and stabilization of cystathionine β-synthase (CBS), an H₂S biosynthetic enzyme, on mitochondrial structure and function. Hyperoxia impaired fASM mitochondrial integrity, while H₂S donors in particular, or CBS stabilization attenuated adverse O2 effects on mitochondrial morphology, reactive oxygen species, respiration, calcium regulation, and contractility. These findings highlight the therapeutic potential of H₂S in the premature lung exposed to moderate hyperoxia.
Authors
Colleen M. Bartman, Michael Thompson, Samantha K. Hamrick, Niyati A. Borkar, Daniel Pfeffer-Kleemann, Preetham Ravi, Marta Schiliro, Yak Nak, Christian Vivar Ramon, Li Drake, Y. S. Prakash, Christina Pabelick
Abstract
Heterogeneity in disease severity and treatment response in inflammatory bowel disease (IBD) likely evolve from individual differences in host-microbiota-immune interactions. Histological evaluation of intestinal biopsies is central to diagnosis, but histological parameters that define underlying immune mechanisms are limited. We investigated histological features that distinguish individual patient immune profiles in therapy-naive paediatric IBD patients (age 6‒18-years-old) using biopsy immunohistochemistry and transcriptomics and plasma proteomics across two cohorts. High colonic epithelial expression of Secretory Leukocyte Protease Inhibitor (SLPI), a microbiota-induced regulator of epithelial function, occurred in IBD patients with high clinical disease activity and more severe endoscopic and microscopic disease activity. SLPI expression related to increased neutrophil infiltration, transcriptomic signatures of activation and genes known to associate with therapeutic resistance. High SLPI co-localized with high densities of IL-17-secreting cells and was associated with high plasma concentrations of Th17-related immune proteins. Additionally, patients with high intestinal SLPI had an intrinsically different immunotype, in which circulating neutrophils exhibited altered transcription of genes involved in neutrophil granule formation, phagocytosis, oxidative phosphorylation, and interferon signalling. Thus, high colonic SLPI expression at diagnosis associates with severe IBD, increased IL-17A-neutrophil pathway responses and altered transcriptomic wiring of circulating neutrophils.
Authors
Sandrine Nugteren, Beatriz Calado, Ytje Simons-Oosterhuis, Daniëlle H. Hulleman-van Haaften, Willem K. Smits, Renz C.W. Klomberg, Bastiaan Tuk, Mohammed Charrout, Dicky J. Lindenbergh-Kortleve, Michail Doukas, Mathijs A. Sanders, Gregory van Beek, Johanna C. Escher, Lissy de Ridder, Maria Fernanda Pascutti, Janneke N. Samsom
Abstract
Viral lower respiratory tract infections are common early in life and are associated with long-term development of asthma, a chronic condition defined by reversible airflow obstruction secondary to inflammation. Understanding the immunologic mechanism connecting these two pathologies observed early in life becomes imperative to guide therapeutic measures. To investigate this connection, neonatal (day of life 4-6) or adult mice were infected with human metapneumovirus (HMPV) followed by a secondary HMPV infection 6 weeks later. Mice initially infected as neonates demonstrate increased mucus production, eosinophil recruitment, airway hyperresponsiveness, and Th2 T-cell differentiation following re-challenge compared to adult mice rechallenged with HMPV. Neonatal HMPV infection led to formation of Th2 clonally expanded tissue resident memory (TRM) T cells that were absent after adult HMPV. FTY720-mediated disruption of lymphocyte circulation demonstrated TRMs contribute to pathology. Local depletion of lung CD4+ T cells and JAK2-inhibition mitigated pathology. These findings suggest TRMs uniquely generated after early life viral infection can contribute to Th2-driven asthma pathology.
Authors
Emma E. Brown, Jie Lan, Olivia B. Parks, Li Fan, Dequan Lou, Alysia McCray, Lisa Mathews, Alexander J. Wardropper, Anna Shull, Michelle L. Manni, Hēth R. Turnquist, Kong Chen, Taylor Eddens
Abstract
Metabolic dysfunction-associated steatotic liver disease (MASLD) is a global health concern with limited interventions. While the role of gut bacteria in MASLD has been extensively studied, the contribution of gut fungi remains largely unexplored. This study investigates the impact of fungal dysbiosis and the role of CARD9, a key adaptor protein in fungal sensing on gut-liver axis dysfunction in MASLD. Patients with advanced liver fibrosis exhibited distinct mycobiota profiles. Using a CARD9-deficient mouse model subjected to high-fat high-glucose/fructose feeding, we observed exacerbated liver injury and fibrosis accompanied by fungal dysbiosis, paralleling our findings in human patients. Beyond its established expression in myeloid cells, CARD9 was also detected in intestinal enterocytes where its expression was diminished under metabolic stress. Intestinal organoids with CARD9 inhibition had reduced expression of antimicrobial Reg3g, the tight junction protein ZO-1, and the antifungal enteroendocrine hormone PYY. These findings suggest that CARD9 maintains gut barrier integrity, preventing microbial translocation and subsequent liver injury and fibrosis. Our results provide new insights into the interplay between fungal dysbiosis, gut barrier dysfunction, and MASLD, and identify CARD9 as a key protein within this axis.
Authors
Vijay Pandyarajan, So Yeon Kim, Takashi Tsuchiya, Selena Liu, Sadam H. Bhat, Jieun Kim, David M. Underhill, Mazen Noureddin, Shelly C. Lu, Ekihiro Seki
