47 results on '"Billiar TR"'
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2. NETWORK ANALYSIS OF SINGLE-NUCLEOTIDE POLYMORPHISMS ASSOCIATED WITH ABERRANT INFLAMMATION IN TRAUMA PATIENTS SUGGESTS A ROLE FOR VESICLE-ASSOCIATED INFLAMMATORY PROGRAMS INVOLVING CD55.
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El-Dehaibi F, Zamora R, Yin J, Namas RA, Billiar TR, and Vodovotz Y
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Background: Critical illness stemming from severe traumatic injury is a leading cause of morbidity and mortality worldwide, and involves the dysfunction of multiple organ systems, driven, at least in part, by dysregulated inflammation. We and others have shown a key role for genetic predisposition to dysregulated inflammation and downstream adverse critical illness outcomes. Recently, we demonstrated an association among genotypes at the single-nucleotide polymorphism (SNP) rs10404939 in LYPD4, dysregulated systemic inflammation, and adverse clinical outcomes in a broad sample of ~1000 critically ill patients., Methods: We sought to gain mechanistic insights into the role of LYPD4 in critical illness by bioinformatically analyzing potential interactions among rs10404939 and other SNPs. We analyzed a dataset of common (i.e., not rare) SNPs previously defined to be associated with genotype-specific, significantly dysregulated systemic inflammation trajectories in trauma patients, in comparison to a control dataset of common SNPs determined to exhibit an absence of genotype-specific inflammatory responses., Results: In the control dataset, this analysis implicated SNPs associated with phosphatidylinositol and various membrane transport proteins, but not LYPD4. In the patient subset with genotypically dysregulated inflammation, our analysis suggested the co-localization to lipid rafts of LYPD4 and the complement receptor CD55, as well as the neurally related CNTNAP2 and RIMS4. Segregation of trauma patients based on genotype of the CD55 SNP rs11117564 showed distinct trajectories of organ dysfunction and systemic inflammation despite similar demographics and injury characteristics., Conclusion: These analyses define novel interactions among SNPs that could enhance our understanding of the response to traumatic injury and critical illness., (Copyright © 2024 by the Shock Society.)
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- 2024
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3. Cardiolipin oxidized by ROS from complex II acts as a target of gasdermin D to drive mitochondrial pore and heart dysfunction in endotoxemia.
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Tang Y, Wu J, Sun X, Tan S, Li W, Yin S, Liu L, Chen Y, Liu Y, Tan Q, Jiang Y, Yang W, Huang W, Weng C, Wu Q, Lu Y, Yuan H, Xiao Q, Chen AF, Xu Q, Billiar TR, and Cai J
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- Animals, Mice, Humans, Mice, Inbred C57BL, Male, Apoptosis, NLR Family, Pyrin Domain-Containing 3 Protein metabolism, Mitochondria metabolism, Gasdermins, Cardiolipins metabolism, Reactive Oxygen Species metabolism, Endotoxemia metabolism, Endotoxemia pathology, Phosphate-Binding Proteins metabolism, Intracellular Signaling Peptides and Proteins metabolism, Oxidation-Reduction, Myocytes, Cardiac metabolism, Myocytes, Cardiac pathology
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Cardiac dysfunction, an early complication of endotoxemia, is the major cause of death in intensive care units. No specific therapy is available at present for this cardiac dysfunction. Here, we show that the N-terminal gasdermin D (GSDMD-N) initiates mitochondrial apoptotic pore and cardiac dysfunction by directly interacting with cardiolipin oxidized by complex II-generated reactive oxygen species (ROS) during endotoxemia. Caspase-4/11 initiates GSDMD-N pores that are subsequently amplified by the upregulation and activation of NLRP3 inflammation through further generation of ROS. GSDMD-N pores form prior to BAX and VDAC1 apoptotic pores and further incorporate into BAX and VDAC1 oligomers within mitochondria membranes to exacerbate the apoptotic process. Our findings identify oxidized cardiolipin as the definitive target of GSDMD-N in mitochondria of cardiomyocytes during endotoxin-induced myocardial dysfunction (EIMD), and modulation of cardiolipin oxidation could be a therapeutic target early in the disease process to prevent EIMD., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)
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- 2024
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4. Activation of TLR9 signaling suppresses the immunomodulating functions of CD55 lo fibroblastic reticular cells during bacterial peritonitis.
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Jiang T, Li Y, Huang X, Jayakumar P, Billiar TR, and Deng M
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- Animals, Humans, Male, Mice, Disease Models, Animal, Immunomodulation, Mice, Inbred C57BL, Mice, Knockout, Fibroblasts metabolism, Fibroblasts immunology, Peritonitis immunology, Peritonitis metabolism, Signal Transduction, Toll-Like Receptor 9 metabolism, Toll-Like Receptor 9 genetics
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Fibroblastic reticular cells (FRCs) are a subpopulation of stromal cells modulating the immune environments in health and disease. We have previously shown that activation of TLR9 signaling in FRC in fat-associated lymphoid clusters (FALC) regulate peritoneal immunity via suppressing immune cell recruitment and peritoneal resident macrophage (PRM) retention. However, FRCs are heterogeneous across tissues and organs. The functions of each FRC subset and the regulation of TLR9 in distinct FRC subsets are unknown. Here, we confirmed that specific deletion of TLR9 in FRC improved bacterial clearance and survival during peritoneal infection. Furthermore, using single-cell RNA sequencing, we found two subsets of FRCs (CD55
hi and CD55lo ) in the mesenteric FALC. The CD55hi FRCs were enriched in gene expression related to extracellular matrix formation. The CD55lo FRCs were enriched in gene expression related to immune response. Interestingly, we found that TLR9 is dominantly expressed in the CD55lo subset. Activation of TLR9 signaling suppressed proliferation, cytokine production, and retinoid metabolism in the CD55lo FRC, but not CD55hi FRC. Notably, we found that adoptive transfer of Tlr9-/- CD55lo FRC from mesenteric FALC more effectively improved the survival during peritonitis compared with WT-FRC or Tlr9-/- CD55hi FRC. Furthermore, we identified CD55hi and CD55lo subsets in human adipose tissue-derived FRC and confirmed the suppressive effect of TLR9 on the proliferation and cytokine production in the CD55lo subset. Therefore, inhibition of TLR9 in the CD55lo FRCs from adipose tissue could be a useful strategy to improve the therapeutic efficacy of FRC-based therapy for peritonitis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Jiang, Li, Huang, Jayakumar, Billiar and Deng.)- Published
- 2024
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5. IRG1/ACOD1 Promotes Neutrophil Reverse Migration and Alleviates Local Inflammation.
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Ji J, Zhong H, Li Y, Billiar TR, Wilson MA, Scott MJ, and Fan J
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Polymorphonuclear neutrophil (PMN) infiltration at inflammatory site plays a critical role in inflammation. PMN reverse migration (rM) describes the phenomenon that PMNs migrate away from inflammatory site back into the vasculature, and its role within inflammatory scenarios remains to be fully determined. This study aimed to investigate the mechanism underlying PMN rM and its role in inflammation. First, we demonstrated PMN rM in a mouse model of LPS-induced acute lung inflammation. By single-cell RNA sequencing (scRNA-seq), we demonstrated that reverse migrated (rM-ed) PMNs in blood expressed high level of immuneresponsive gene 1 (Irg1), the encoding gene of cis-aconitate decarboxylase (ACOD1). Using a mouse air pouch model, which enables us to directly track rM-ed PMNs in vivo, we detected higher expression of ACOD1 in the rM-ed PMNs in circulation. Furthermore, mice with Irg1 knockout exhibited decreased PMN rM and higher levels of inflammatory cytokine in inflammatory site. Mechanistically, we found that itaconate, the product of ACOD1 catalyzation, decreased PMN ICAM-1 expression at the inflammation site. Furthermore, inflammatory site showed a high level of shed CD11a, the ligand of ICAM-1. Neutralization of either ICAM-1 or CD11a leading to increased PMN rM. These findings suggest that the binding of ICAM-1 and shed CD11a serves as a retaining force to hold PMNs in the site of inflammation, and ACOD1-decreased PMN surface expression of ICAM-1 weakens the retaining force, so promoting PMNs to leave the inflammatory site. These results indicate a regulatory role of IRG1 in PMN rM and subsequent contributions to inflammation resolution., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)
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- 2024
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6. Transcriptomic and Proteomic Characterization of the Immune Response to Elective Spinal Reconstructive Surgery: Impact of Aging and Comparison with Traumatic Injury Response.
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Bonaroti JW, Ozel M, Chen T, Darby JL, Sun X, Moheimani H, Reitz KM, Kar UK, Zuckerbraun BS, Das J, Okonkwo DO, and Billiar TR
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- Humans, Aged, Adolescent, Young Adult, Adult, Transcriptome, Leukocytes, Mononuclear, Proteomics, Aging, Gene Expression Profiling, Immunity, Inflammation, Surgery, Plastic, Multiple Trauma surgery
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Background: Major surgery triggers trauma-like stress responses linked to age, surgery duration, and blood loss, resembling polytrauma. This similarity suggests elective surgery as a surrogate model for studying polytrauma immune responses. We investigated stress responses across age groups and compared them with those of polytrauma patients., Study Design: Patients undergoing major spinal reconstruction surgery were divided into older (age >65 years, n = 5) and young (age 18 to 39 years, n = 6) groups. A comparison group consisted of matched trauma patients (n = 8). Blood samples were collected before, during, and after surgery. Bone marrow mononuclear cells and peripheral blood mononuclear cells were analyzed using cellular indexing of transcriptomes and epitopes sequencing or single-cell RNA sequencing. Plasma was subjected to dual-platform proteomic analysis (SomaLogic and O-link)., Results: Response to polytrauma was highest within 4 hours. By comparison, the response to surgery was highest at 24 hours. Both insults triggered significant changes in cluster of differentiation 14 monocytes, with increased inflammation and lower major histocompatibility complex-class 2 expression. Older patient's cluster of differentiation 14 monocytes displayed higher inflammation and less major histocompatibility complex-class 2 suppression; a trend was also seen in bone marrow mononuclear cells. Although natural killer cells were markedly activated after polytrauma, they were suppressed after surgery, especially in older patients. In plasma, innate immunity proteins dominated at 24 hours, shifting to adaptive immunity proteins by 6 weeks with heightened inflammation in older patients. Senescence-associated secretory phenotype proteins were higher in older patients at baseline and further elevated during and after surgery., Conclusions: Although both major surgery and polytrauma initiate immune and stress responses, substantial differences exist in timing and cellular profiles, suggesting major elective surgery is not a suitable surrogate for the polytrauma response. Nonetheless, distinct responses in young vs older patients highlight the utility of elective spinal in studying patient-specific factors affecting outcomes after major elective surgery., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American College of Surgeons.)
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- 2024
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7. Fibroblastic reticular cell-derived exosomes are a promising therapeutic approach for septic acute kidney injury.
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Li Y, Hu C, Zhai P, Zhang J, Jiang J, Suo J, Hu B, Wang J, Weng X, Zhou X, Billiar TR, Kellum JA, Deng M, and Peng Z
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- Mice, Animals, Inflammasomes metabolism, NLR Family, Pyrin Domain-Containing 3 Protein metabolism, Lipopolysaccharides, Mice, Inbred C57BL, Exosomes metabolism, Acute Kidney Injury metabolism, Sepsis complications, Sepsis metabolism
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Sepsis-induced acute kidney injury (S-AKI) is highly lethal, and effective drugs for treatment are scarce. Previously, we reported the robust therapeutic efficacy of fibroblastic reticular cells (FRCs) in sepsis. Here, we demonstrate the ability of FRC-derived exosomes (FRC-Exos) to improve C57BL/6 mouse kidney function following cecal ligation and puncture-induced sepsis. In vivo imaging confirmed that FRC-Exos homed to injured kidneys. RNA-Seq analysis of FRC-Exo-treated primary kidney tubular cells (PKTCs) revealed that FRC-Exos influenced PKTC fate in the presence of lipopolysaccharide (LPS). FRC-Exos promoted kinase PINK1-dependent mitophagy and inhibited NLRP3 inflammasome activation in LPS-stimulated PKTCs. To dissect the mechanism underlying the protective role of Exos in S-AKI, we examined the proteins within Exos by mass spectrometry and found that CD5L was the most upregulated protein in FRC-Exos compared to macrophage-derived Exos. Recombinant CD5L treatment in vitro attenuated kidney cell swelling and surface bubble formation after LPS stimulation. FRCs were infected with a CD5L lentivirus to increase CD5L levels in FRC-Exos, which were then modified in vitro with the kidney tubular cell targeting peptide LTH, a peptide that binds to the biomarker protein kidney injury molecule-1 expressed on injured tubule cells, to enhance binding specificity. Compared with an equivalent dose of recombinant CD5L, the modified CD5L-enriched FRC-Exos selectively bound PKTCs, promoted kinase PINK-ubiquitin ligase Parkin-mediated mitophagy, inhibiting pyroptosis and improved kidney function by hindering NLRP3 inflammasome activation, thereby improving the sepsis survival rate. Thus, strategies to modify FRC-Exos could be a new avenue in developing therapeutics against kidney injury., (Copyright © 2023 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.)
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- 2024
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8. Mechanism matters: mortality and endothelial cell damage marker differences between blunt and penetrating traumatic injuries across three prehospital clinical trials.
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Donohue JK, Gruen DS, Iyanna N, Lorence JM, Brown JB, Guyette FX, Daley BJ, Eastridge BJ, Miller RS, Nirula R, Harbrecht BG, Claridge JA, Phelan HA, Vercruysse GA, O'Keeffe T, Joseph B, Neal MD, Billiar TR, and Sperry JL
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- Humans, Proportional Hazards Models, Endothelial Cells, Retrospective Studies, Wounds, Penetrating complications, Wounds, Nonpenetrating complications, Emergency Medical Services
- Abstract
Injury mechanism is an important consideration when conducting clinical trials in trauma. Mechanisms of injury may be associated with differences in mortality risk and immune response to injury, impacting the potential success of the trial. We sought to characterize clinical and endothelial cell damage marker differences across blunt and penetrating injured patients enrolled in three large, prehospital randomized trials which focused on hemorrhagic shock. In this secondary analysis, patients with systolic blood pressure < 70 or systolic blood pressure < 90 and heart rate > 108 were included. In addition, patients with both blunt and penetrating injuries were excluded. The primary outcome was 30-day mortality. Mortality was characterized using Kaplan-Meier and Cox proportional-hazards models. Generalized linear models were used to compare biomarkers. Chi squared tests and Wilcoxon rank-sum were used to compare secondary outcomes. We characterized data of 696 enrolled patients that met all secondary analysis inclusion criteria. Blunt injured patients had significantly greater 24-h (18.6% vs. 10.7%, log rank p = 0.048) and 30-day mortality rates (29.7% vs. 14.0%, log rank p = 0.001) relative to penetrating injured patients with a different time course. After adjusting for confounders, blunt mechanism of injury was independently predictive of mortality at 30-days (HR 1.84, 95% CI 1.06-3.20, p = 0.029), but not 24-h (HR 1.65, 95% CI 0.86-3.18, p = 0.133). Elevated admission levels of endothelial cell damage markers, VEGF, syndecan-1, TM, S100A10, suPAR and HcDNA were associated with blunt mechanism of injury. Although there was no difference in multiple organ failure (MOF) rates across injury mechanism (48.4% vs. 42.98%, p = 0.275), blunt injured patients had higher Denver MOF score (p < 0.01). The significant increase in 30-day mortality and endothelial cell damage markers in blunt injury relative to penetrating injured patients highlights the importance of considering mechanism of injury within the inclusion and exclusion criteria of future clinical trials., (© 2024. The Author(s).)
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- 2024
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9. Transforming the Future of Surgeon-Scientists.
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Ladner DP, Goldstein AM, Billiar TR, Cameron AM, Carpizo DR, Chu DI, Coopersmith CM, DeMatteo RP, Feng S, Gallagher KA, Gillanders WE, Lal BK, Lipshutz GS, Liu A, Maier RV, Mittendorf EA, Morris AM, Sicklick JK, Velazquez OC, Whitson BA, Wilke LG, Yoon SS, Zeiger MA, Farmer DL, and Hwang ES
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- Humans, United States, Mentors, Faculty, Academic Medical Centers, Career Mobility, National Institutes of Health (U.S.), Surgeons, Biomedical Research
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Objective: To create a blueprint for surgical department leaders, academic institutions, and funding agencies to optimally support surgeon-scientists., Background: Scientific contributions by surgeons have been transformative across many medical disciplines. Surgeon-scientists provide a distinct approach and mindset toward key scientific questions. However, lack of institutional support, pressure for increased clinical productivity, and growing administrative burden are major challenges for the surgeon-scientist, as is the time-consuming nature of surgical training and practice., Methods: An American Surgical Association Research Sustainability Task Force was created to outline a blueprint for sustainable science in surgery. Leaders from top NIH-sponsored departments of surgery engaged in video and in-person meetings between January and April 2023. A strength, weakness, opportunities, threats analysis was performed, and workgroups focused on the roles of surgeons, the department and institutions, and funding agencies., Results: Taskforce recommendations: (1) SURGEONS: Growth mindset : identifying research focus, long-term planning, patience/tenacity, team science, collaborations with disparate experts; Skill set : align skills and research, fill critical skill gaps, develop team leadership skills; DEPARTMENT OF SURGERY (DOS): (2) MENTORSHIP: Chair : mentor-mentee matching/regular meetings/accountability, review of junior faculty progress, mentorship training requirement, recognition of mentorship (eg, relative value unit equivalent, awards; Mentor: dedicated time, relevant scientific expertise, extramural funding, experience and/or trained as mentor, trusted advisor; Mentee : enthusiastic/eager, proactive, open to feedback, clear about goals; (3) FINANCIAL SUSTAINABILITY: diversification of research portfolio, identification of matching funding sources, departmental resource awards (eg, T-/P-grants), leveraging of institutional resources, negotiation of formalized/formulaic funds flow investment from academic medical center toward science, philanthropy; (4) STRUCTURAL/STRATEGIC SUPPORT: Structural: grants administrative support, biostats/bioinformatics support, clinical trial and research support, regulatory support, shared departmental laboratory space/equipment; Strategic: hiring diverse surgeon-scientist/scientists faculty across DOS, strategic faculty retention/ recruitment, philanthropy, career development support, progress tracking, grant writing support, DOS-wide research meetings, regular DOS strategic research planning; (5) COMMUNITY AND CULTURE: Community: right mix of faculty, connection surgeon with broad scientific community; Culture: building research infrastructure, financial support for research, projecting importance of research (awards, grand rounds, shoutouts); (6) THE ROLE OF INSTITUTIONS: Foundation: research space co-location, flexible start-up packages, courses/mock study section, awards, diverse institutional mentorship teams; Nurture: institutional infrastructure, funding (eg, endowed chairs), promotion friendly toward surgeon-scientists, surgeon-scientists in institutional leadership positions; Expectations: RVU target relief, salary gap funding, competitive starting salaries, longitudinal salary strategy; (7) THE ROLE OF FUNDING AGENCIES: change surgeon research training paradigm, offer alternate awards to K-awards, increasing salary cap to reflect market reality, time extension for surgeon early-stage investigator status, surgeon representation on study section, focused award strategies for professional societies/foundations., Conclusions: Authentic recommitment from surgeon leaders with intentional and ambitious actions from institutions, corporations, funders, and society is essential in order to reap the essential benefits of surgeon-scientists toward advancements of science., Competing Interests: S.H.F. reports past compensation to her institution from Novartis, Randox, and Transplant Genomics. S.H.F. also serves as an Editor-in-Chief for American Journal of Transplantation and a member of Editorial Board for New England Journal of Medicine . W.E.G. reports ongoing compensation to his institution from National Cancer Institute, Leidos Biomedical, and 1440 Foundation. E.A.M. reports compensated service on scientific advisory boards for Astra Zeneca, BioNTech and Merck; uncompensated service on steering committees for Bristol Myers Squibb and Roche/Genentech; and institutional research support from Roche/Genentech (via SU2C grant) and Gilead. E.A.M. also reports research funding from Susan Komen for the Cure for which she serves as a Scientific Advisor, and uncompensated participation as a member of the American Society of Clinical Oncology Board of Directors. J.K.S. receives consultant fees from Deciphera, Aadi and Grand Rounds, serves as a consultant for CureMatch, received speaker’s fees from Deciphera, La-Hoffman Roche, Foundation Medicine, Merck, QED, and Daiichi Sankyo; and owns stock in Personalis and CureMatch. L.G.W. reports ongoing compensation to her institution from Perimeter Medical and compensated services from Elucent Medical. D.L.F. reports compensated services from Briopryme and Artemis Cures; uncompensated services as a consultant for the National Institutes of Health; and research funding from California Institute for Regenerative Medicine (CIRM). The other authors report no conflicts of interest., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)
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- 2024
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10. NOS2 and COX2 Provide Key Spatial Targets that Determine Outcome in ER- Breast Cancer.
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Ridnour LA, Heinz WF, Cheng RY, Wink AL, Kedei N, Pore M, Imtiaz F, Femino EL, Gonzalez AL, Coutinho L, Butcher D, Edmondson EF, Rangel MC, Kinders RJ, Lipkowitz S, Wong ST, Anderson SK, McVicar DW, Li X, Glynn SA, Billiar TR, Chang JC, Hewitt SM, Ambs S, Lockett SJ, and Wink DA
- Abstract
Estrogen receptor-negative (ER-) breast cancer is an aggressive breast cancer subtype with limited therapeutic options. Upregulated expression of both inducible nitric oxide synthase (NOS2) and cyclo-oxygenase (COX2) in breast tumors predicts poor clinical outcomes. Signaling molecules released by these enzymes activate oncogenic pathways, driving cancer stemness, metastasis, and immune suppression. The influence of tumor NOS2/COX2 expression on the landscape of immune markers using multiplex fluorescence imaging of 21 ER- breast tumors were stratified for survival. A powerful relationship between tumor NOS2/COX2 expression and distinct CD8+ T cell phenotypes was observed at 5 years post-diagnosis. These results were confirmed in a validation cohort using gene expression data showing that ratios of NOS2 to CD8 and COX2 to CD8 are strongly associated with poor outcomes in high NOS2/COX2-expressing tumors. Importantly, multiplex imaging identified distinct CD8+ T cell phenotypes relative to tumor NOS2/COX2 expression in Deceased vs Alive patient tumors at 5-year survival. CD8+NOS2-COX2- phenotypes defined fully inflamed tumors with significantly elevated CD8+ T cell infiltration in Alive tumors expressing low NOS2/COX2. In contrast, two distinct phenotypes including inflamed CD8+NOS2+COX2+ regions with stroma-restricted CD8+ T cells and CD8-NOS2-COX2+ immune desert regions with abated CD8+ T cell penetration, were significantly elevated in Deceased tumors with high NOS2/COX2 expression. These results were supported by applying an unsupervised nonlinear dimensionality-reduction technique, UMAP, correlating specific spatial CD8/NOS2/COX2 expression patterns with patient survival. Moreover, spatial analysis of the CD44v6 and EpCAM cancer stem cell (CSC) markers within the CD8/NOS2/COX2 expression landscape revealed positive correlations between EpCAM and inflamed stroma-restricted CD8+NOS2+COX2+ phenotypes at the tumor/stroma interface in deceased patients. Also, positive correlations between CD44v6 and COX2 were identified in immune desert regions in deceased patients. Furthermore, migrating tumor cells were shown to occur only in the CD8-NOS2+COX2+ regions, identifying a metastatic hot spot. Taken together, this study shows the strength of spatial localization analyses of the CD8/NOS2/COX2 landscape, how it shapes the tumor immune microenvironment and the selection of aggressive tumor phenotypes in distinct regions that lead to poor clinical outcomes. This technique could be beneficial for describing tumor niches with increased aggressiveness that may respond to clinically available NOS2/COX2 inhibitors or immune-modulatory agents.
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- 2023
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11. Spatial analysis of NOS2 and COX2 interaction with T-effector cells reveals immunosuppressive landscapes associated with poor outcome in ER- breast cancer patients.
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Ridnour LA, Cheng RYS, Heinz WF, Pore M, Gonzalez AL, Femino EL, Moffat R, Wink AL, Imtiaz F, Coutinho L, Butcher D, Edmondson EF, Rangel MC, Wong STC, Lipkowitz S, Glynn S, Vitek MP, McVicar DW, Li X, Anderson SK, Paolocci N, Hewitt SM, Ambs S, Billiar TR, Chang JC, Lockett SJ, and Wink DA
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Multiple immunosuppressive mechanisms exist in the tumor microenvironment that drive poor outcomes and decrease treatment efficacy. The co-expression of NOS2 and COX2 is a strong predictor of poor prognosis in ER- breast cancer and other malignancies. Together, they generate pro-oncogenic signals that drive metastasis, drug resistance, cancer stemness, and immune suppression. Using an ER- breast cancer patient cohort, we found that the spatial expression patterns of NOS2 and COX2 with CD3+CD8+PD1- T effector (Teff) cells formed a tumor immune landscape that correlated with poor outcome. NOS2 was primarily associated with the tumor-immune interface, whereas COX2 was associated with immune desert regions of the tumor lacking Teff cells. A higher ratio of NOS2 or COX2 to Teff was highly correlated with poor outcomes. Spatial analysis revealed that regional clustering of NOS2 and COX2 was associated with stromal-restricted Teff, while only COX2 was predominant in immune deserts. Examination of other immunosuppressive elements, such as PDL1/PD1, Treg, B7H4, and IDO1, revealed that PDL1/PD1, Treg, and IDO1 were primarily associated with restricted Teff, whereas B7H4 and COX2 were found in tumor immune deserts. Regardless of the survival outcome, other leukocytes, such as CD4 T cells and macrophages, were primarily in stromal lymphoid aggregates. Finally, in a 4T1 model, COX2 inhibition led to a massive cell infiltration, thus validating the hypothesis that COX2 is an essential component of the Teff exclusion process and, thus, tumor evasion. Our study indicates that NOS2/COX2 expression plays a central role in tumor immunosuppression. Our findings indicate that new strategies combining clinically available NOS2/COX2 inhibitors with various forms of immune therapy may open a new avenue for the treatment of aggressive ER-breast cancers.
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- 2023
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12. Nuclear localization of STING1 competes with canonical signaling to activate AHR for commensal and intestinal homeostasis.
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Zhang R, Yu C, Zeh HJ, Wang H, Kroemer G, Klionsky DJ, Billiar TR, Kang R, and Tang D
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- Animals, Mice, DNA, Homeostasis, Intestines, Proteomics, Receptors, Aryl Hydrocarbon genetics, Receptors, Aryl Hydrocarbon metabolism
- Abstract
Extensive studies demonstrate the importance of the STING1 (also known as STING) protein as a signaling hub that coordinates immune and autophagic responses to ectopic DNA in the cytoplasm. Here, we report a nuclear function of STING1 in driving the activation of the transcription factor aryl hydrocarbon receptor (AHR) to control gut microbiota composition and homeostasis. This function was independent of DNA sensing and autophagy and showed competitive inhibition with cytoplasmic cyclic guanosine monophosphate (GMP)-AMP synthase (CGAS)-STING1 signaling. Structurally, the cyclic dinucleotide binding domain of STING1 interacted with the AHR N-terminal domain. Proteomic analyses revealed that STING1-mediated transcriptional activation of AHR required additional nuclear partners, including positive and negative regulatory proteins. Although AHR ligands could rescue colitis pathology and dysbiosis in wild-type mice, this protection was abrogated by mutational inactivation of STING1. These findings establish a key framework for understanding the nuclear molecular crosstalk between the microbiota and the immune system., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 Elsevier Inc. All rights reserved.)
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- 2023
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13. Nitric oxide and thioredoxin modulate the activity of caspase 9 in HepG2 cells.
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Chakraborty S, Choudhuri A, Mishra A, Bhattacharyya C, Billiar TR, Stoyanovsky DA, and Sengupta R
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- Humans, Caspase 9 metabolism, Hep G2 Cells, NADP metabolism, Thioredoxins metabolism, Thioredoxin-Disulfide Reductase, Nitric Oxide metabolism, Thioctic Acid
- Abstract
The interdependent and finely tuned balance between the well-established redox-based modification, S-nitrosylation, and its counteractive mechanism of S-nitrosothiol degradation, i.e., S-denitrosylation of biological protein or non-protein thiols defines the cellular fate in the context of redox homeostasis. S-nitrosylation of cysteine residues by S-nitrosoglutathione, S-nitroso-L-cysteine-like physiological and S-nitroso-L-cysteine ethyl ester-like synthetic NO donors inactivate Caspase-3, 8, and 9, thereby hindering their apoptotic activity. However, spontaneous restoration of their activity upon S-denitrosylation of S-nitrosocaspases into their reduced, free thiol active states, aided by the members of the ubiquitous cellular redoxin (thioredoxin/ thioredoxin reductase/ NADPH) and low molecular weight dithiol (lipoic acid/ lipoamide dehydrogenase/ dihydrolipoic acid/ NADPH) systems imply a direct relevance to their proteolytic activities and further downstream signaling cascades. Additionally, our previous and current findings offer crucial insight into the concept of redundancy between thioredoxin and lipoic acid systems, and the redox-modulated control of the apoptotic and proteolytic activity of caspases, triggering their cyto- and neurotoxic effects in response to nitro-oxidative stress. Thus, this might lay the foundation for the exogenous introduction of precise and efficient NO or related donor drug delivery systems that can directly participate in catering to the S-(de)-nitrosylation-mediated functional outcomes of the cysteinyl proteases in pathophysiological settings., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)
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- 2023
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14. A common single nucleotide polymorphism is associated with inflammation and critical illness outcomes.
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El-Dehaibi F, Zamora R, Radder J, Yin J, Shah AM, Namas RA, Situ M, Zhao Y, Bain W, Morris A, McVerry BJ, Barclay DA, Billiar TR, Zhang Y, Kitsios GD, and Vodovotz Y
- Abstract
Acute inflammation is heterogeneous in critical illness and predictive of outcome. We hypothesized that genetic variability in novel, yet common, gene variants contributes to this heterogeneity and could stratify patient outcomes. We searched algorithmically for significant differences in systemic inflammatory mediators associated with any of 551,839 SNPs in one derivation (n = 380 patients with blunt trauma) and two validation (n = 75 trauma and n = 537 non-trauma patients) cohorts. This analysis identified rs10404939 in the LYPD4 gene. Trauma patients homozygous for the A allele (rs10404939AA; 27%) had different trajectories of systemic inflammation along with persistently elevated multiple organ dysfunction (MOD) indices vs. patients homozygous for the G allele (rs10404939GG; 26%). rs10404939AA homozygotes in the trauma validation cohort had elevated MOD indices, and non-trauma patients displayed more complex inflammatory networks and worse 90-day survival compared to rs10404939GG homozygotes. Thus, rs10404939 emerged as a common, broadly prognostic SNP in critical illness., Competing Interests: The authors declare no competing interests associated with this article., (© 2023 The Authors.)
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- 2023
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15. A Prospective Cohort Protocol for the Remnant Investigation in Sepsis Study.
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Seymour CW, Urbanek KL, Nakayama A, Kennedy JN, Powell R, Robinson RAS, Kapp KL, Billiar TR, Vodovotz Y, Gelhaus SL, Cooper VS, Tang L, Mayr F, Reitz KM, Horvat C, Meyer NJ, Dickson RP, Angus D, and Palmer OP
- Abstract
Background: Sepsis is a common and deadly syndrome, accounting for more than 11 million deaths annually. To mature a deeper understanding of the host and pathogen mechanisms contributing to poor outcomes in sepsis, and thereby possibly inform new therapeutic targets, sophisticated, and expensive biorepositories are typically required. We propose that remnant biospecimens are an alternative for mechanistic sepsis research, although the viability and scientific value of such remnants are unknown., Methods and Results: The Remnant Biospecimen Investigation in Sepsis study is a prospective cohort study of 225 adults (age ≥ 18 yr) presenting to the emergency department with community sepsis, defined as sepsis-3 criteria within 6 hours of arrival. The primary objective was to determine the scientific value of a remnant biospecimen repository in sepsis linked to clinical phenotyping in the electronic health record. We will study candidate multiomic readouts of sepsis biology, governed by a conceptual model, and determine the precision, accuracy, integrity, and comparability of proteins, small molecules, lipids, and pathogen sequencing in remnant biospecimens compared with paired biospecimens obtained according to research protocols. Paired biospecimens will include plasma from sodium-heparin, EDTA, sodium fluoride, and citrate tubes., Conclusions: The study has received approval from the University of Pittsburgh Human Research Protection Office (Study 21120013). Recruitment began on October 25, 2022, with planned release of primary results anticipated in 2024. Results will be made available to the public, the funders, critical care societies, laboratory medicine scientists, and other researchers., Competing Interests: Dr. Seymour reports personal fees for consulting from Beckman Coulter Inc. and Inotrem, Inc., unrelated to the study. Dr. Meyer reports personal fees for consulting from Endpoint Health Inc and AstraZeneca BioPharmaceuticals Medical and payments to her institution by Quantum Leap Healthcare Collaborative for work unrelated to the study., (Copyright © 2023 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the Society of Critical Care Medicine.)
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- 2023
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16. Direct Trauma Center Access by Helicopter Emergency Medical Services is Associated With Improved Survival After Severe Injury.
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Deeb AP, Teng CY, Peitzman AB, Billiar TR, Sperry JL, Lu L, Beiriger J, and Brown JB
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- Humans, Retrospective Studies, Aircraft, Triage, Trauma Centers, Injury Severity Score, Air Ambulances, Emergency Medical Services, Wounds and Injuries therapy
- Abstract
Objective: Evaluate the association of survival with helicopter transport directly to a trauma center compared with ground transport to a non-trauma center (NTC) and subsequent transfer., Summary Background Data: Helicopter transport improves survival after injury. One potential mechanism is direct transport to a trauma center when the patient would otherwise be transported to an NTC for subsequent transfer., Methods: Scene patients 16 years and above with positive physiological or anatomic triage criteria within PTOS 2000-2017 were included. Patients transported directly to level I/II trauma centers by helicopter were compared with patients initially transported to an NTC by ground with a subsequent helicopter transfer to a level I/II trauma center. Propensity score matching was used to evaluate the association between direct helicopter transport and survival. Individual triage criteria were evaluated to identify patients most likely to benefit from direct helicopter transport., Results: In all, 36,830 patients were included. Direct helicopter transport was associated with a nearly 2-fold increase in odds of survival compared with NTC ground transport and subsequent transfer by helicopter (aOR 2.78; 95% CI 2.24-3.44, P <0.01). Triage criteria identifying patients with a survival benefit from direct helicopter transport included GCS≤13 (1.71; 1.22-2.41, P <0.01), hypotension (2.56; 1.39-4.71, P <0.01), abnormal respiratory rate (2.30; 1.36-3.89, P <0.01), paralysis (8.01; 2.03-31.69, P <0.01), hemothorax/pneumothorax (2.34; 1.36-4.05, P <0.01), and multisystem trauma (2.29; 1.08-4.84, P =0.03)., Conclusions: Direct trauma center access is a mechanism driving the survival benefit of helicopter transport. First responders should consider helicopter transport for patients meeting these criteria who would otherwise be transported to an NTC., Competing Interests: The authors report no conflicts of interest., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)
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- 2023
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17. Targeting inflammation in traumatic injury: entering a new era.
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Billiar TR, Hunt BJ, and Bailly S
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- Humans, Inflammation, Brain Injuries, Traumatic, Brain Injuries
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- 2023
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18. High-dimensional proteomics identifies organ injury patterns associated with outcomes in human trauma.
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Li SR, Moheimani H, Herzig B, Kail M, Krishnamoorthi N, Wu J, Abdelhamid S, Scioscia J, Sung E, Rosengart A, Bonaroti J, Johansson PI, Stensballe J, Neal MD, Das J, Kar U, Sperry J, and Billiar TR
- Subjects
- Humans, Biomarkers, Critical Care, Injury Severity Score, Proteomics, Inflammation
- Abstract
Introduction: Severe traumatic injury with shock can lead to direct and indirect organ injury; however, tissue-specific biomarkers are limited in clinical panels. We used proteomic and metabolomic databases to identify organ injury patterns after severe injury in humans., Methods: Plasma samples (times 0, 24, and 72 hours after arrival to trauma center) from injured patients enrolled in two randomized prehospital trials were subjected to multiplexed proteomics (SomaLogic Inc., Boulder, CO). Patients were categorized by outcome: nonresolvers (died >72 hours or required ≥7 days of critical care), resolvers (survived to 30 days and required <7 days of critical care), and low Injury Severity Score (ISS) controls. Established tissue-specific biomarkers were identified through a literature review and cross-referenced with tissue specificity from the Human Protein Atlas. Untargeted plasma metabolomics (Metabolon Inc., Durham, NC), inflammatory mediators, and endothelial damage markers were correlated with injury biomarkers. Kruskal-Wallis/Mann-Whitney U tests with false discovery rate correction assessed differences in biomarker expression across outcome groups (significance; p < 0.1)., Results: Of 142 patients, 78 were nonresolvers (median ISS, 30), 34 were resolvers (median ISS, 22), and 30 were low ISS controls (median ISS, 1). A broad release of tissue-specific damage markers was observed at admission; this was greater in nonresolvers. By 72 hours, nine cardiac, three liver, eight neurologic, and three pulmonary proteins remained significantly elevated in nonresolvers compared with resolvers. Cardiac damage biomarkers showed the greatest elevations at 72 hours in nonresolvers and had significant positive correlations with proinflammatory mediators and endothelial damage markers. Nonresolvers had lower concentrations of fatty acid metabolites compared with resolvers, particularly acyl carnitines and cholines., Conclusion: We identified an immediate release of tissue-specific biomarkers with sustained elevation in the liver, pulmonary, neurologic, and especially cardiac injury biomarkers in patients with complex clinical courses after severe injury. The persistent myocardial injury in nonresolvers may be due to a combination of factors including metabolic stress, inflammation, and endotheliopathy., (Copyright © 2023 Wolters Kluwer Health, Inc. All rights reserved.)
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- 2023
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19. Targeting Nitric Oxide: Say NO to Metastasis.
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Reddy TP, Glynn SA, Billiar TR, Wink DA, and Chang JC
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- Humans, Nitric Oxide Synthase metabolism, Nitric Oxide Synthase Type II metabolism, Tumor Microenvironment, Nitric Oxide metabolism, Neoplasms drug therapy
- Abstract
Utilizing targeted therapies capable of reducing cancer metastasis, targeting chemoresistant and self-renewing cancer stem cells, and augmenting the efficacy of systemic chemo/radiotherapies is vital to minimize cancer-associated mortality. Targeting nitric oxide synthase (NOS), a protein within the tumor microenvironment, has gained interest as a promising therapeutic strategy to reduce metastatic capacity and augment the efficacy of chemo/radiotherapies in various solid malignancies. Our review highlights the influence of nitric oxide (NO) in tumor progression and cancer metastasis, as well as promising preclinical studies that evaluated NOS inhibitors as anticancer therapies. Lastly, we highlight the prospects and outstanding challenges of using NOS inhibitors in the clinical setting., (©2022 The Authors; Published by the American Association for Cancer Research.)
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- 2023
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20. Interferon-gamma is quintessential for NOS2 and COX2 expression in ER - breast tumors that lead to poor outcome.
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Cheng RYS, Ridnour LA, Wink AL, Gonzalez AL, Femino EL, Rittscher H, Somasundaram V, Heinz WF, Coutinho L, Rangel MC, Edmondson EF, Butcher D, Kinders RJ, Li X, Wong STC, McVicar DW, Anderson SK, Pore M, Hewitt SM, Billiar TR, Glynn SA, Chang JC, Lockett SJ, Ambs S, and Wink DA
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- Female, Humans, CD8-Positive T-Lymphocytes, Cell Line, Tumor, Cyclooxygenase 2 genetics, Cyclooxygenase 2 metabolism, Nitric Oxide Synthase Type II genetics, Nitric Oxide Synthase Type II metabolism, Tumor Necrosis Factor-alpha metabolism, Interferon-gamma genetics, Interferon-gamma metabolism, Triple Negative Breast Neoplasms genetics, Triple Negative Breast Neoplasms metabolism, Triple Negative Breast Neoplasms pathology, Tumor Microenvironment genetics, Tumor Microenvironment immunology
- Abstract
A strong correlation between NOS2 and COX2 tumor expression and poor clinical outcomes in ER breast cancer has been established. However, the mechanisms of tumor induction of these enzymes are unclear. Analysis of The Cancer Genome Atlas (TCGA) revealed correlations between NOS2 and COX2 expression and Th1 cytokines. Herein, single-cell RNAseq analysis of TNBC cells shows potent NOS2 and COX2 induction by IFNγ combined with IL1β or TNFα. Given that IFNγ is secreted by cytolytic lymphocytes, which improve clinical outcomes, this role of IFNγ presents a dichotomy. To explore this conundrum, tumor NOS2, COX2, and CD8
+ T cells were spatially analyzed in aggressive ER-, TNBC, and HER2 + breast tumors. High expression and clustering of NOS2-expressing tumor cells occurred at the tumor/stroma interface in the presence of stroma-restricted CD8+ T cells. High expression and clustering of COX2-expressing tumor cells extended into immune desert regions in the tumor core where CD8+ T cell penetration was limited or absent. Moreover, high NOS2-expressing tumor cells were proximal to areas with increased satellitosis, suggestive of cell clusters with a higher metastatic potential. Further in vitro experiments revealed that IFNγ + IL1β/TNFα increased the elongation and migration of treated tumor cells. This spatial analysis of the tumor microenvironment provides important insight into distinct neighborhoods where stroma-restricted CD8+ T cells exist proximal to NOS2-expressing tumor niches that could have increased metastatic potential., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)- Published
- 2023
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21. Central role for neurally dysregulated IL-17A in dynamic networks of systemic and local inflammation in combat casualties.
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Zamora R, Forsberg JA, Shah AM, Unselt D, Grey S, Lisboa FA, Billiar TR, Schobel SA, Potter BK, Elster EA, and Vodovotz Y
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- Humans, Tumor Necrosis Factor-alpha pharmacology, Interferon-gamma pharmacology, Biomarkers, Th17 Cells, Interleukin-17 pharmacology, Inflammation
- Abstract
Dynamic Network Analysis (DyNA) and Dynamic Hypergraphs (DyHyp) were used to define protein-level inflammatory networks at the local (wound effluent) and systemic circulation (serum) levels from 140 active-duty, injured service members (59 with TBI and 81 non-TBI). Interleukin (IL)-17A was the only biomarker elevated significantly in both serum and effluent in TBI vs. non-TBI casualties, and the mediator with the most DyNA connections in TBI wounds. DyNA combining serum and effluent data to define cross-compartment correlations suggested that IL-17A bridges local and systemic circulation at late time points. DyHyp suggested that systemic IL-17A upregulation in TBI patients was associated with tumor necrosis factor-α, while IL-17A downregulation in non-TBI patients was associated with interferon-γ. Correlation analysis suggested differential upregulation of pathogenic Th17 cells, non-pathogenic Th17 cells, and memory/effector T cells. This was associated with reduced procalcitonin in both effluent and serum of TBI patients, in support of an antibacterial effect of Th17 cells in TBI patients. Dysregulation of Th17 responses following TBI may drive cross-compartment inflammation following combat injury, counteracting wound infection at the cost of elevated systemic inflammation., (© 2023. The Author(s).)
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- 2023
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22. Interferon-gamma is Quintessential for NOS2 and COX2 Expression in ER - Breast Tumors that Lead to Poor Outcome.
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Cheng RY, Ridnour LA, Wink AL, Gonzalez AL, Femino EL, Rittscher H, Somasundarum V, Heinz WF, Coutinho L, Cristina Rangel M, Edmondson EF, Butcher D, Kinders RJ, Li X, Wong STC, McVicar DW, Anderson SK, Pore M, Hewitt SM, Billiar TR, Glynn S, Chang JC, Lockett SJ, Ambs S, and Wink DA
- Abstract
A strong correlation between NOS2 and COX2 tumor expression and poor clinical outcomes in ER-breast cancer has been established. However, mechanisms of tumor induction of these enzymes are unclear. Analysis of The Cancer Genome Atlas (TCGA) revealed correlations between NOS2 and COX2 expression and Th1 cytokines. Herein, single cell RNAseq analysis of TNBC cells shows potent NOS2 and COX2 induction by IFNγ combined with IL1β or TNFα. Given that IFNγ is secreted by cytolytic lymphocytes, which improve clinical outcomes, this role of IFNγpresents a dichotomy. To explore this conundrum, tumor NOS2, COX2, and CD8
+ T cells were spatially analyzed in aggressive ER-, TNBC, and HER2+ breast tumors. High expression and clustering of NOS2-expressing tumor cells occurred at the tumor/stroma interface in the presence of stroma-restricted CD8+ T cells. High expression and clustering of COX2-expressing tumor cells extended into immune desert regions in the tumor core where CD8+ T cell penetration was limited or absent. Moreover, high NOS2-expressing tumor cells were proximal to areas with increased satellitosis suggestive of cell clusters with a higher metastatic potential. Further in vitro experiments revealed that IFNγ+IL1β/TNFα increased elongation and migration of treated tumor cells. This spatial analysis of the tumor microenvironment provides important insight of distinct neighborhoods where stroma-restricted CD8+ T cells exist proximal to NOS2-expressing tumor niches that could have increased metastatic potential.- Published
- 2023
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23. Low Titer Group O Whole Blood In Injured Children Requiring Massive Transfusion.
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Gaines BA, Yazer MH, Triulzi DJ, Sperry JL, Neal MD, Billiar TR, and Leeper CM
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- Humans, Child, Infant, Child, Preschool, Adolescent, Plasma, Blood Transfusion, Resuscitation, Prospective Studies, ABO Blood-Group System, Blood Component Transfusion, Wounds and Injuries therapy
- Abstract
Objective: The aim of this study was to assess the survival impact of low-titer group O whole blood (LTOWB) in injured pediatric patients who require massive transfusion., Summary Background Data: Limited data are available regarding the effectiveness of LTOWB in pediatric trauma., Methods: A prospective observational study of children requiring massive transfusion after injury at UPMC Children's Hospital of Pittsburgh, an urban academic pediatric Level 1 trauma center. Injured children ages 1 to 17 years who received a total of >40 mL/kg of LTOWB and/or conventional components over the 24 hours after admission were included. Patient characteristics, blood product utilization and clinical outcomes were analyzed using Kaplan-Meier survival curves, log rank tests and Cox proportional hazards regression analyses. The primary outcome was 28-day survival., Results: Of patients analyzed, 27 of 80 (33%) received LTOWB as part of their hemostatic resuscitation. The LTOWB group was comparable to the component therapy group on baseline demographic and physiologic parameters except older age, higher body weight, and lower red blood cell and plasma transfusion volumes. After adjusting for age, total blood product volume transfused in 24 hours, admission base deficit, international normalized ratio (INR), and injury severity score (ISS), children who received LTOWB as part of their resuscitation had significantly improved survival at both 72 hours and 28 days post-trauma [adjusted odds ratio (AOR) 0.23, P = 0.009 and AOR 0.41, P = 0.02, respectively]; 6-hour survival was not statistically significant (AOR = 0.51, P = 0.30). Survivors at 28 days in the LTOWB group had reduced hospital LOS, ICU LOS, and ventilator days compared to the CT group., Conclusion: Administration of LTOWB during the hemostatic resuscitation of injured children requiring massive transfusion was independently associated with improved 72-hour and 28-day survival., Competing Interests: The following conflicts of interest are relevant to this study: M.H.Y. has given paid lectures and received travel reimbursement from Terumo BCT, the manufacturer of the PLT-sparing leukoreduction filter used in the preparation of the LTOWB units. The authors report no conflicts of interest., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)
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- 2023
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24. Expert consensus on the monitoring and treatment of sepsis-induced immunosuppression.
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Pei F, Yao RQ, Ren C, Bahrami S, Billiar TR, Chaudry IH, Chen DC, Chen XL, Cui N, Fang XM, Kang Y, Li WQ, Li WX, Liang HP, Lin HY, Liu KX, Lu B, Lu ZQ, Maegele M, Peng TQ, Shang Y, Su L, Sun BW, Wang CS, Wang J, Wang JH, Wang P, Xie JF, Xie LX, Zhang LN, Zingarelli B, Guan XD, Wu JF, and Yao YM
- Subjects
- Humans, Consensus, Delphi Technique, Surveys and Questionnaires, Immunosuppression Therapy, Sepsis therapy
- Abstract
Emerged evidence has indicated that immunosuppression is involved in the occurrence and development of sepsis. To provide clinical practice recommendations on the immune function in sepsis, an expert consensus focusing on the monitoring and treatment of sepsis-induced immunosuppression was developed. Literature related to the immune monitoring and treatment of sepsis were retrieved from PubMed, Web of Science, and Chinese National Knowledge Infrastructure to design items and expert opinions were collected through an online questionnaire. Then, the Delphi method was used to form consensus opinions, and RAND appropriateness method was developed to provide consistency evaluation and recommendation levels for consensus opinions. This consensus achieved satisfactory results through two rounds of questionnaire survey, with 2 statements rated as perfect consistency, 13 as very good consistency, and 9 as good consistency. After summarizing the results, a total of 14 strong recommended opinions, 8 weak recommended opinions and 2 non-recommended opinions were produced. Finally, a face-to-face discussion of the consensus opinions was performed through an online meeting, and all judges unanimously agreed on the content of this consensus. In summary, this expert consensus provides a preliminary guidance for the monitoring and treatment of immunosuppression in patients with sepsis., (© 2022. The Author(s).)
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- 2022
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25. Short-Term Administration of HIV Protease Inhibitor Saquinavir Improves Skull Bone Healing with Enhanced Osteoclastogenesis.
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Liu H, Shen Y, Zhao B, Poon EH, Qi S, Ker DFE, Billiar TR, Cooper GM, Xu Y, and Wang D
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- Mice, Animals, Toll-Like Receptor 4 physiology, Osteogenesis, Skull injuries, Saquinavir pharmacology, Saquinavir metabolism, Saquinavir therapeutic use, HIV Protease Inhibitors pharmacology, HIV Protease Inhibitors metabolism, HIV Protease Inhibitors therapeutic use
- Abstract
Background: Using immunomodulatory methods to address the challenging issue of craniofacial bone repair may be a potentially effective approach. The protease inhibitor saquinavir has been shown to inhibit the inflammatory response by targeting the toll-like receptor 4/myeloid differentiation primary response complex. Independently, inhibition of toll-like receptor 4 or myeloid differentiation primary response led to enhanced skull bone repair. Therefore, the authors aimed to investigate the effects of saquinavir on skull bone healing., Methods: The effects of saquinavir on skull bone healing were assessed by means of gene expression, histology, immunohistochemistry, and tomography in a mouse calvarial defect model. Subsequently, the role of saquinavir in cell viability, migration, and osteogenic and osteoclastogenic differentiation was also evaluated in vitro., Results: One-week saquinavir administration improved skull bone healing based on micro-computed tomographic and histomorphometric analyses. Compared to the vehicle control, 1-week saquinavir treatment (1) enhanced osteoclast infiltration (tartrate-resistant acid phosphatase staining) at day 7, but not at days 14 and 28; (2) induced more CD206 + M2 macrophage infiltration, but not F4/80 + M0 macrophages at days 7, 14, and 28; and (3) elevated osteoclastogenic gene RANKL (quantitative polymerase chain reaction) expression and other osteogenic and cytokine expression. Furthermore, in vitro data showed that saquinavir administration did not influence MC3T3-E1 cell migration or mineralization, whereas higher concentrations of saquinavir inhibited cell viability. Saquinavir treatment also enhanced the osteoclastic differentiation of bone marrow-derived precursors, and partially reversed high-mobility group box 1-driven osteoclastogenesis inhibition and elevated proinflammatory cytokine expression., Conclusion: The improved skull bone repair following short-term saquinavir treatment may involve enhanced osteoclastogenesis and modulated inflammatory response following skull injury., Clinical Relevance Statement: The authors' work demonstrates improved skull bone healing by short-term application of saquinavir, a drug traditionally used in the treatment of acquired immunodeficiency syndrome. As such, saquinavir may be repurposed for skeletal repair., (Copyright © 2022 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of the American Society of Plastic Surgeons. All rights reserved.)
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- 2022
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26. Plasma proteomics reveals early, broad release of chemokine, cytokine, TNF, and interferon mediators following trauma with delayed increases in a subset of chemokines and cytokines in patients that remain critically ill.
- Author
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Bonaroti J, Billiar I, Moheimani H, Wu J, Namas R, Li S, Kar UK, Vodovotz Y, Neal MD, Sperry JL, and Billiar TR
- Subjects
- Humans, Critical Illness, Proteomics, Chemokines, Receptors, Cytokine, Cytokines, Interferons
- Abstract
Severe injury is known to cause a systemic cytokine storm that is associated with adverse outcomes. However, a comprehensive assessment of the time-dependent changes in circulating levels of a broad spectrum of protein immune mediators and soluble immune mediator receptors in severely injured trauma patients remains uncharacterized. To address this knowledge gap, we defined the temporal and outcome-based patterns of 184 known immune mediators and soluble cytokine receptors in the circulation of severely injured patients. Proteomics (aptamer-based assay, SomaLogic, Inc) was performed on plasma samples drawn at 0, 24, and 72 hours (h) from time of admission from 150 trauma patients, a representative subset from the Prehospital Plasma during Air Medical Transport in Trauma Patients at Risk for Hemorrhagic Shock (PAMPer) trial. Patients were categorized into outcome groups including Early Non-Survivors (died within 72 h; ENS; n=38), Non-Resolvers (died after 72 h or required ≥7 days of intensive care; NR; n=78), and Resolvers (survivors that required < 7 days of intensive care; R; n=34), with low Injury Severity Score (ISS) patients from the Tranexamic Acid During Prehospital Transport in Patients at Risk for Hemorrhage After Injury (STAAMP) trial as controls. The major findings include an extensive release of immune mediators and cytokine receptors at time 0h that is more pronounced in ENS and NR patients. There was a selective subset of mediators elevated at 24 and 72 h to a greater degree in NR patients, including multiple cytokines and chemokines not previously described in trauma patients. These findings were validated in a quantitative fashion using mesoscale discovery immunoassays (MSD) from an external validation cohort (VC) of samples from 58 trauma patients matched for R and NR status. This comprehensive longitudinal description of immune mediator patterns associated with trauma outcomes provides a new level of characterization of the immune response that follows severe injury., Competing Interests: YV is co-founder of, and stakeholder in, Immunetrics, Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Bonaroti, Billiar, Moheimani, Wu, Namas, Li, Kar, Vodovotz, Neal, Sperry and Billiar.)
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- 2022
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27. Lipidomic signatures align with inflammatory patterns and outcomes in critical illness.
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Wu J, Cyr A, Gruen DS, Lovelace TC, Benos PV, Das J, Kar UK, Chen T, Guyette FX, Yazer MH, Daley BJ, Miller RS, Harbrecht BG, Claridge JA, Phelan HA, Zuckerbraun BS, Neal MD, Johansson PI, Stensballe J, Namas RA, Vodovotz Y, Sperry JL, and Billiar TR
- Subjects
- Humans, Lipidomics, Biomarkers, Inflammation, Critical Illness, COVID-19
- Abstract
Alterations in lipid metabolism have the potential to be markers as well as drivers of pathobiology of acute critical illness. Here, we took advantage of the temporal precision offered by trauma as a common cause of critical illness to identify the dynamic patterns in the circulating lipidome in critically ill humans. The major findings include an early loss of all classes of circulating lipids followed by a delayed and selective lipogenesis in patients destined to remain critically ill. The previously reported survival benefit of early thawed plasma administration was associated with preserved lipid levels that related to favorable changes in coagulation and inflammation biomarkers in causal modelling. Phosphatidylethanolamines (PE) were elevated in patients with persistent critical illness and PE levels were prognostic for worse outcomes not only in trauma but also severe COVID-19 patients. Here we show selective rise in systemic PE as a common prognostic feature of critical illness., (© 2022. The Author(s).)
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- 2022
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28. Intraperitoneal injection of class A TLR9 agonist enhances anti-PD-1 immunotherapy in colorectal peritoneal metastases.
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Jiang T, Zhang H, Li Y, Jayakumar P, Liao H, Huang H, Billiar TR, and Deng M
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- Mice, Animals, Toll-Like Receptor 9 metabolism, Injections, Intraperitoneal, GATA6 Transcription Factor, Immune Checkpoint Inhibitors pharmacology, Immune Checkpoint Inhibitors therapeutic use, Immunotherapy, Adjuvants, Immunologic, Retinoids, Peritoneal Neoplasms drug therapy, Colorectal Neoplasms drug therapy
- Abstract
Peritoneal metastases are associated with a low response rate to immune checkpoint blockade (ICB) therapy. The numbers of peritoneal resident macrophages (PRMs) are reversely correlated with the response rate to ICB therapy. We have previously shown that TLR9 in fibroblastic reticular cells (FRCs) plays a critical role in regulating peritoneal immune cell recruitment. However, the role of TLR9 in FRCs in regulating PRMs is unclear. Here, we demonstrated that the class A TLR9 agonist, ODN1585, markedly enhanced the efficacy of anti-PD-1 therapy in mouse models of colorectal peritoneal metastases. ODN1585 injected i.p. reduced the numbers of Tim4+ PRMs and enhanced CD8+ T cell antitumor immunity. Mechanistically, treatment of ODN1585 suppressed the expression of genes required for retinoid metabolism in FRCs, and this was associated with reduced expression of the PRM lineage-defining transcription factor GATA6. Selective deletion of TLR9 in FRCs diminished the benefit of ODN1585 in anti-PD-1 therapy in reducing peritoneal metastases. The crosstalk between PRMs and FRCs may be utilized to develop new strategies to improve the efficacy of ICB therapy for peritoneal metastases.
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- 2022
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29. High Dimensional Multiomics Reveals Unique Characteristics of Early Plasma Administration in Polytrauma Patients With TBI.
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Wu J, Moheimani H, Li S, Kar UK, Bonaroti J, Miller RS, Daley BJ, Harbrecht BG, Claridge JA, Gruen DS, Phelan HA, Guyette FX, Neal MD, Das J, Sperry JL, and Billiar TR
- Subjects
- Humans, Plasma, Proteomics, Brain Injuries, Traumatic therapy, Emergency Medical Services methods, Multiple Trauma therapy
- Abstract
Objectives: The authors sought to identify causal factors that explain the selective benefit of prehospital administration of thawed plasma (TP) in traumatic brain injury (TBI) patients using mediation analysis of a multiomic database., Background: The Prehospital Air Medical Plasma (PAMPer) Trial showed that patients with TBI and a pronounced systemic response to injury [defined as endotype 2 (E2)], have a survival benefit from prehospital administration of TP. An interrogation of high dimensional proteomics, lipidomics and metabolomics previously demonstrated unique patterns in circulating biomarkers in patients receiving prehospital TP, suggesting that a deeper analysis could reveal causal features specific to TBI patients., Methods: A novel proteomic database (SomaLogic Inc., aptamer-based assay, 7K platform) was generated using admission blood samples from a subset of patients (n=149) from the PAMPer Trial. This proteomic dataset was combined with previously reported metabolomic and lipidomic datasets from these same patients. A 2-step analysis was performed to identify factors that promote survival in E2-TBI patients who had received early TP. First, features were selected using both linear and multivariate-latent-factor regression analyses. Then, the selected features were entered into the causal mediation analysis., Results: Causal mediation analysis of observable features identified 16 proteins and 41 lipids with a high proportion of mediated effect (>50%) to explain the survival benefit of early TP in E2-TBI patients. The multivariate latent-factor regression analyses also uncovered 5 latent clusters of features with a proportion effect >30%, many in common with the observable features. Among the observable and latent features were protease inhibitors known to inhibit activated protein C and block fibrinolysis (SERPINA5 and CPB2), a clotting factor (factor XI), as well as proteins involved in lipid transport and metabolism (APOE3 and sPLA(2)-XIIA)., Conclusions: These findings suggest that severely injured patients with TBI process exogenous plasma differently than those without TBI. The beneficial effects of early TP in E2-TBI patients may be the result of improved blood clotting and the effect of brain protective factors independent of coagulation., Competing Interests: T.R.B. is a stakeholder in Immunetrics Inc. M.D.N holds an equity stake in Haima Therapeutics. He has received research support and/or honoraria from Haemonetics, Instrumentation Laboratories, and Janssen Pharmaceuticals. The remaining authors report no conflicts of interest.
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- 2022
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30. Aconitate decarboxylase 1 is a mediator of polymicrobial sepsis.
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Wu R, Liu J, Wang N, Zeng L, Yu C, Chen F, Wang H, Billiar TR, Jiang J, Tang D, and Kang R
- Subjects
- Animals, Cytokine Release Syndrome, Humans, Mice, Mitogen-Activated Protein Kinases metabolism, Carboxy-Lyases metabolism, Lipopolysaccharides pharmacology, Sepsis metabolism
- Abstract
Sepsis is a challenging clinical syndrome caused by a dysregulated host response to infection. Here, we identified an unexpected proseptic activity of aconitate decarboxylase 1 (ACOD1) in monocytes and macrophages. Previous studies have suggested that ACOD1, also known as immune-responsive gene 1, is an immunometabolic regulator that favors itaconate production to inhibit bacterial lipopolysaccharide-induced innate immunity. We used next-generation sequencing of lipopolysaccharide-activated THP1 cells to demonstrate that ACOD1 accumulation confers a robust proinflammation response by activating a cytokine storm, predominantly through the tumor necrosis factor signaling pathway. We further revealed that the phosphorylation of cyclin-dependent kinase 2 (CDK2) on threonine-160 mediates the activation of mitogen-activated protein kinase 8 through receptor for activated C kinase 1, leading to JUN-dependent transcription of ACOD1 in human and mouse macrophages or monocytes. Genetic deletion of CDK2 or ACOD1 in myeloid cells, or the administration of the CDK inhibitor dinaciclib, protected mice against polymicrobial sepsis and was associated with improved survival and decreased cytokine storm. The expression of the CDK2-ACOD1 axis also correlated with severity of illness in a cohort of 40 patients with bacterial sepsis. Thus, our findings provide evidence for a previously unrecognized function of ACOD1 in innate immunity and suggest it as a potential therapeutic target for the treatment of sepsis.
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- 2022
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31. Multi-Omic Admission-Based Prognostic Biomarkers Identified by Machine Learning Algorithms Predict Patient Recovery and 30-Day Survival in Trauma Patients.
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Abdelhamid SS, Scioscia J, Vodovotz Y, Wu J, Rosengart A, Sung E, Rahman S, Voinchet R, Bonaroti J, Li S, Darby JL, Kar UK, Neal MD, Sperry J, Das J, and Billiar TR
- Abstract
Admission-based circulating biomarkers for the prediction of outcomes in trauma patients could be useful for clinical decision support. It is unknown which molecular classes of biomolecules can contribute biomarkers to predictive modeling. Here, we analyzed a large multi-omic database of over 8500 markers (proteomics, metabolomics, and lipidomics) to identify prognostic biomarkers in the circulating compartment for adverse outcomes, including mortality and slow recovery, in severely injured trauma patients. Admission plasma samples from patients ( n = 129) enrolled in the Prehospital Air Medical Plasma (PAMPer) trial were analyzed using mass spectrometry (metabolomics and lipidomics) and aptamer-based (proteomics) assays. Biomarkers were selected via Least Absolute Shrinkage and Selection Operator (LASSO) regression modeling and machine learning analysis. A combination of five proteins from the proteomic layer was best at discriminating resolvers from non-resolvers from critical illness with an Area Under the Receiver Operating Characteristic curve (AUC) of 0.74, while 26 multi-omic features predicted 30-day survival with an AUC of 0.77. Patients with traumatic brain injury as part of their injury complex had a unique subset of features that predicted 30-day survival. Our findings indicate that multi-omic analyses can identify novel admission-based prognostic biomarkers for outcomes in trauma patients. Unique biomarker discovery also has the potential to provide biologic insights.
- Published
- 2022
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32. LONGITUDINAL ANALYSIS OF TRANSCRIPTOMIC SUBTYPES IN TRAUMA PATIENTS.
- Author
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Chen T, Wei Y, Vodovotz Y, Chen W, and Billiar TR
- Subjects
- Humans, Prognosis, Transcriptome genetics, Wounds, Nonpenetrating
- Abstract
Abstract: Objective: We previously identified two transcriptomic subtypes (Signature Groups: SG1 vs. SG2) in trauma patients at 12 hours postinjury, with SG1 associated with worse outcomes. In this study, we aimed to further characterize the changes in SG subtype categorization of trauma patients over time after injury and define the corresponding association with outcomes based on the timing of the subtype designation. Methods and Results: This study was a secondary analysis of published data of whole-blood leukocyte transcriptomics, a longitudinal data from 167 severe blunt trauma patients. We assigned trauma patients to SG1 or SG2 subtype for time points between 12 hours and 28 days, inclusive, postinjury and characterized their longitudinal outcomes. SG1 assignment, regardless of time point, was associated consistently with slower recovery. Further analysis revealed that additional prognostic information could be obtained by assessing SG subtype at both 12 hours and 1 day. Conclusions: This study provides a proof of concept that immune status can worsen after admission and highlights the benefit of longitudinally monitoring SG subtypes in trauma patients., Competing Interests: Y.V. is a field chief editor of Frontiers in Systems Biology . Y.V. is also a cofounder of and stakeholder in Immunetrics, Inc. The other authors report no conflict of interest., (Copyright © 2022 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the Shock Society.)
- Published
- 2022
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33. Impaired interferon-γ signaling promotes the development of silicosis.
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Peng Z, Duan M, Tang Y, Wu J, Zhao K, Zhong Y, He Z, Meng J, Chen F, Xiao X, Wang H, Billiar TR, Lu B, and Liang F
- Abstract
Silicosis is caused by inhalation of crystalline silica dust particles and known as one of the most serious occupational diseases worldwide. However, little is known about intrinsic factors leading to disease susceptibility. Single-cell sequencing of bronchoalveolar lavage fluid cells of mine workers with silicosis and their co-workers who did not develop silicosis revealed that the impaired interferon (IFN)-γ signaling in myeloid cells was strongly associated with the occurrence of silicosis. Global or myeloid cell-specific deletion of interferon γ receptor (IFN-γR) markedly enhanced the crystalline silica-induced pulmonary injury in wild-type but not in NLRP3 deficient mice. In vitro , IFN-γ priming of macrophages suppressed the crystalline silica-induced NLRP3 inflammasome activation partly by inducing the formation of spacious phagosomes with relatively reduced ratio of crystalline silica/phagosomal areas volumes to resistant crystalline silica-induced lysosomal membrane damage. Thus, these findings provide molecular insights into the intricate mechanisms underlying innate immunity-mediated host responses to environmental irritants., Competing Interests: The authors declare no competing interests., (© 2022.)
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- 2022
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34. Single-cell transcriptome profiling of the immune space-time landscape reveals dendritic cell regulatory program in polymicrobial sepsis.
- Author
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Yao RQ, Li ZX, Wang LX, Li YX, Zheng LY, Dong N, Wu Y, Xia ZF, Billiar TR, Ren C, and Yao YM
- Subjects
- Animals, Dendritic Cells, Gene Expression Profiling, Humans, Mice, T-Lymphocytes, Regulatory, COVID-19, Sepsis
- Abstract
Rationale: Evident immunosuppression has been commonly seen among septic patients, and it is demonstrated to be a major driver of morbidity. Nevertheless, a comprehensive view of the host immune response to sepsis is lacking as the majority of studies on immunosuppression have focused on a specific type of immune cells. Methods: We applied multi-compartment, single-cell RNA sequencing (scRNA-seq) to dissect heterogeneity within immune cell subsets during sepsis progression on cecal ligation and puncture (CLP) mouse model. Flow cytometry and multiplex immunofluorescence tissue staining were adopted to identify the presence of 'mature DCs enriched in immunoregulatory molecules' (mregDC) upon septic challenge. To explore the function of mregDC, sorted mregDC were co-cultured with naïve CD4
+ T cells. Intracellular signaling pathways that drove mregDC program were determined by integrating scRNA-seq and bulk-seq data, combined with inhibitory experiments. Results: ScRNA-seq analysis revealed that sepsis induction was associated with substantial alterations and heterogeneity of canonical immune cell types, including T, B, natural killer (NK), and myeloid cells, across three immune-relevant tissue sites. We found a unique subcluster of conventional dendritic cells (cDCs) that was characterized by specific expression of maturation- and migration-related genes, along with upregulation of immunoregulatory molecules, corresponding to the previously described 'mregDCs' in cancer. Flow cytometry and in stiu immunofluorescence staining confirmed the presence of sepsis-induced mregDC at protein level. Functional experiments showed that sepsis-induced mregDCs potently activated naive CD4+ T cells, while promoted CD4+ T cell conversion to regulatory T cells. Further observations indicated that the mregDC program was initiated via TNFRSF-NF-κB- and IFNGR2-JAK-STAT3-dependent pathways within 24 h of septic challenge. Additionally, we confirmed the detection of mregDC in human sepsis using publicly available data from a recently published single-cell study of COVID-19 patients. Conclusions: Our study generates a comprehensive single-cell immune landscape for polymicrobial sepsis, in which we identify the significant alterations and heterogeneity in immune cell subsets that take place during sepsis. Moreover, we find a conserved and potentially targetable immunoregulatory program within DCs that associates with hyperinflammation and organ dysfunction early following sepsis induction., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)- Published
- 2022
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35. Inferring Tissue-Specific, TLR4-Dependent Type 17 Immune Interactions in Experimental Trauma/Hemorrhagic Shock and Resuscitation Using Computational Modeling.
- Author
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Shah AM, Zamora R, Korff S, Barclay D, Yin J, El-Dehaibi F, Billiar TR, and Vodovotz Y
- Subjects
- Animals, Computer Simulation, Granulocyte-Macrophage Colony-Stimulating Factor, Inflammation Mediators, Interleukin-10, Interleukin-17, Mice, Mice, Inbred C57BL, Signal Transduction, Toll-Like Receptor 4 genetics, Toll-Like Receptor 4 metabolism, Shock, Hemorrhagic
- Abstract
Trauma/hemorrhagic shock followed by resuscitation (T/HS-R) results in multi-system inflammation and organ dysfunction, in part driven by binding of damage-associated molecular pattern molecules to Toll-like Receptor 4 (TLR4). We carried out experimental T/HS-R (pseudo-fracture plus 2 h of shock followed by 0-22 h of resuscitation) in C57BL/6 (wild type [WT]) and TLR4-null (TLR4
-/- ) mice, and then defined the dynamics of 20 protein-level inflammatory mediators in the heart, gut, lung, liver, spleen, kidney, and systemic circulation. Cross-correlation and Principal Component Analysis (PCA) on data from the 7 tissues sampled suggested that TLR4-/- samples express multiple inflammatory mediators in a small subset of tissue compartments as compared to the WT samples, in which many inflammatory mediators were localized non-specifically to nearly all compartments. We and others have previously defined a central role for type 17 immune cells in human trauma. Accordingly, correlations between IL-17A and GM-CSF (indicative of pathogenic Th17 cells); between IL-17A and IL-10 (indicative of non-pathogenic Th17 cells); and IL-17A and TNF (indicative of memory/effector T cells) were assessed across all tissues studied. In both WT and TLR4-/- mice, positive correlations were observed between IL-17A and GM-CSF, IL-10, and TNF in the kidney and gut. In contrast, the variable and dynamic presence of both pathogenic and non-pathogenic Th17 cells was inferred in the systemic circulation of TLR4-/- mice over time, suggesting a role for TLR4 in efflux of these cells into peripheral tissues. Hypergraph analysis - used to define dynamic, cross compartment networks - in concert with PCA-suggested that IL-17A was present persistently in all tissues at all sampled time points except for its absence in the plasma at 0.5h in the WT group, supporting the hypothesis that T/HS-R induces efflux of Th17 cells from the circulation and into specific tissues. These analyses suggest a complex, context-specific role for TLR4 and type 17 immunity following T/HS-R., Competing Interests: YV is a co-founder of, and stakeholder in, Immunetrics, Inc. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Shah, Zamora, Korff, Barclay, Yin, El-Dehaibi, Billiar and Vodovotz.)- Published
- 2022
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36. Z-DNA binding protein 1 promotes heatstroke-induced cell death.
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Yuan F, Cai J, Wu J, Tang Y, Zhao K, Liang F, Li F, Yang X, He Z, Billiar TR, Wang H, Su L, and Lu B
- Subjects
- Cell Death, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Humans, RNA-Binding Proteins genetics, RNA-Binding Proteins metabolism, Receptor-Interacting Protein Serine-Threonine Kinases genetics, Heat Stroke, Nucleic Acids
- Abstract
Heatstroke is a heat stress-induced, life-threatening condition associated with circulatory failure and multiple organ dysfunctions. If global warming continues, heatstroke might become a more prominent cause of mortality worldwide, but its pathogenic mechanism is not well understood. We found that Z-DNA binding protein 1 (ZBP1), a Z-nucleic acid receptor, mediated heatstroke by triggering receptor-interacting protein kinase 3 (RIPK3)-dependent cell death. Heat stress increased the expression of ZBP1 through heat shock transcription factor 1 (HSF1) and activated ZBP1 through a mechanism independent of the nucleic acid sensing action. Deletion of ZBP1, RIPK3, or both mixed lineage kinase domain-like (MLKL) and caspase-8 decreased heat stress-induced circulatory failure, organ injury, and lethality. Thus, ZBP1 appears to have a second function that orchestrates host responses to heat stress.
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- 2022
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37. Prehospital low titer group O whole blood is feasible and safe: Results of a prospective randomized pilot trial.
- Author
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Guyette FX, Zenati M, Triulzi DJ, Yazer MH, Skroczky H, Early BJ, Adams PW, Brown JB, Alarcon L, Neal MD, Forsythe RM, Zuckerbraun BS, Peitzman AB, Billiar TR, and Sperry JL
- Subjects
- ABO Blood-Group System, Blood Transfusion, Humans, Pilot Projects, Prospective Studies, Emergency Medical Services, Resuscitation methods
- Abstract
Introduction: Low titer group O whole blood (LTOWB) resuscitation is increasingly common in both military and civilian settings. Data regarding the safety and efficacy of prehospital LTOWB remain limited., Methods: We performed a single-center, prospective, cluster randomized, prehospital through in-hospital whole blood pilot trial for injured air medical patients. We compared standard prehospital air medical care including red cell transfusion and crystalloids followed by in-hospital component transfusion to prehospital and in-hospital LTOWB resuscitation. Prehospital vital signs were used as inclusion criteria (systolic blood pressure ≤90 mm Hg and heart rate ≥108 beats per minute or systolic blood pressure ≤70 mm Hg for patients at risk of hemorrhage). Primary outcome was feasibility. Secondary outcomes included 28-day and 24-hour mortality, multiple organ failure, nosocomial infection, 24-hour transfusion requirements, and arrival coagulation parameters., Results: Between November 2018 and October 2020, 86 injured patients were cluster randomized by helicopter base. The trial has halted early at 77% enrollment. Overall, 28-day mortality for the cohort was 26%. Injured patients randomized to prehospital LTOWB (n = 40) relative to standard care (n = 46) were similar in demographics and injury characteristics. Intent-to-treat Kaplan-Meier survival analysis demonstrated no statistical mortality benefit at 28 days (25.0% vs. 26.1%, p = 0.85). Patients randomized to prehospital LTOWB relative to standard care had lower red cell transfusion requirements at 24 hours (p < 0.01) and a lower incidence of abnormal thromboelastographic measurements. No transfusion reactions during the prehospital or in-hospital phase of care were documented., Conclusion: Prehospital through in-hospital LTOWB resuscitation is safe and may be associated with hemostatic benefits. A large-scale clinical trial is feasible with protocol adjustment and would allow the effects of prehospital LTOWB on survival and other pertinent clinical outcomes to be appropriately characterized., Level of Evidence: Therapeutic/Care Management, Level II., (Copyright © 2022 American Association for the Surgery of Trauma.)
- Published
- 2022
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38. GRK2 regulates group 2 innate lymphoid cell mobilization in sepsis.
- Author
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Lai D, Chen W, Zhang K, Scott MJ, Li Y, Billiar TR, Wilson MA, and Fan J
- Subjects
- Animals, G-Protein-Coupled Receptor Kinase 2, Immunity, Innate, Interleukin-1 Receptor-Like 1 Protein, Lung metabolism, Lymphocytes, Mice, Mice, Inbred C57BL, Interleukin-33, Sepsis metabolism
- Abstract
Background: Sepsis induces group 2 innate lymphoid cell (ILC2) expansion in the lung. However, the origin of these lung-recruited ILC2 and the mechanism of ILC2 expansion are unclear. This study aims to determine the origin of lung-recruited ILC2 and its underlying mechanism in sepsis., Methods: Sepsis was induced by cecal ligation and puncture (CLP) model in wild-type, IL-33-deficient and ST2-deficient mice. The frequency, cell number and C-X-C chemokine receptor 4 (CXCR4) expression of ILC2 in bone marrow (BM), blood and lung were measured by flow cytometry. In the in vitro studies, purified ILC2 progenitor (ILC2p) were challenged with IL-33 or G protein-coupled receptor kinase 2 (GRK2) inhibitor, the CXCR4 expression and GRK2 activity were detected by confocal microscopy or flow cytometry., Results: We show that IL-33 acts through its receptor, ST2, on BM ILC2p to induce GRK2 expression and subsequent downregulation of cell surface expression of CXCR4, which results in decreasing retention of ILC2p in the BM and promoting expansion of ILC2 in the lung. Importantly, we demonstrate that reduced IL-33 level in aging mice contributes to impaired ILC2 mobilization from BM and accumulation in the lung following sepsis., Conclusion: This study identifies a novel pathway in regulating ILC2p mobilization and expansion during sepsis and indicates BM as the main source of ILC2 in the lung following sepsis., (© 2022. The Author(s).)
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- 2022
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39. The independent prognostic value of global epigenetic alterations: An analysis of single-cell ATAC-seq of circulating leukocytes from trauma patients followed by validation in whole blood leukocyte transcriptomes across three etiologies of critical illness.
- Author
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Chen T, Conroy J, Wang X, Situ M, Namas RA, Vodovotz Y, Chen W, Singh H, and Billiar TR
- Subjects
- Chromatin Immunoprecipitation Sequencing, Epigenesis, Genetic, Humans, Leukocytes, Leukocytes, Mononuclear, Prognosis, Critical Illness, Transcriptome
- Abstract
Background: While bulk and single cell transcriptomic patterns in circulating leukocytes from trauma patients have been reported, how these relate to changes in open chromatin patterns remain unstudied. Here, we investigated whether single-cell ATAC-seq would provide further resolution of transcriptomic patterns that align with patient outcomes., Methods: We performed scATAC-seq on peripheral blood mononuclear cells from four trauma patients at <4 h, 24 h, 72 h post-injury and four matched healthy controls, and extracted the features associated with the global epigenetic alterations. Three large-scale bulk transcriptomic datasets from trauma, burn and sepsis patients were used to validate the scATAC-seq derived signature, explore patient epigenetic heterogeneity (Epigenetic Groups: EG_hi vs. EG_lo), and associate patterns with clinical outcomes in critical illness., Findings: Patient subsets with gene expression patterns in blood leukocytes representative of a high global epigenetic signature (EG_hi) had worse outcomes across three etiologies of critical illness. EG_hi designation contributed independent of the known immune leukocyte transcriptomic responses to patient prognosis (Trauma: HR=0.62 [95% CI: 0.43-0.89, event set as recovery], p=0.01, n=167; Burns: HR=4.35 [95% CI: 0.816-23.2, event set as death], p=0.085, n=121; Sepsis: HR=1.60 [95% CI: 1.10-2.33, event set as death], p=0.013, n=479; Cox proportional hazards regression)., Interpretation: The inclusion of gene expression patterns that associate with global epigenetic changes in circulating leukocytes improves the resolution of transcriptome-based patient classification in acute critical illnesses. Early detection of both the global epigenetic signature and the known immune transcriptomic patterns associates with the worse prognosis in trauma, burns and sepsis., Funding: This project was supported by an R35 grant from National Institutes of Health: 1R35GM127027-01 (T.B.)., Competing Interests: Declaration of interests Y.V. is a field chief editor of Frontiers in Systems Biology. Y.V. is also a co-founder of, and stakeholder in Immunetrics, Inc. The other authors declare no competing interests., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)
- Published
- 2022
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40. Analysis of glutathione mediated S-(de)nitrosylation in complex biological matrices by immuno-spin trapping and identification of two novel substrates.
- Author
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Sircar E, Stoyanovsky DA, Billiar TR, Holmgren A, and Sengupta R
- Subjects
- Cyclic N-Oxides chemistry, Glutaredoxins chemistry, Hep G2 Cells, Humans, Ribonucleoside Diphosphate Reductase chemistry, S-Nitrosothiols chemistry, Spin Labels, Spin Trapping, Thioredoxins chemistry, Thioredoxins metabolism, Glutaredoxins metabolism, Glutathione metabolism, Ribonucleoside Diphosphate Reductase metabolism, S-Nitrosothiols metabolism
- Abstract
The intracellular concentration of reduced glutathione (GSH) lies in the range of 1-10 mM, thereby indisputably making it the most abundant intracellular thiol. Such a copious amount of GSH makes it the most potent and robust cellular antioxidant that plays a crucial role in cellular defence against redox stress. The role of GSH as a denitrosylating agent is well established; in this study, we demonstrate GSH mediated denitrosylation of HepG2 cell-derived protein nitrosothiols (PSNOs), by a unique spin-trapping mechanism, using 5,5-dimethyl-1-pyrroline N-oxide (DMPO) as the spin trapping agent, followed by a western blot analysis. We also report our findings of two, hitherto unidentified substrates of GSH mediated S-denitrosylation, namely S-nitrosoglutaredoxin 1 (Grx1-SNO) and S-nitrosylated R1 subunit of ribonucleotide reductase (R1-SNO)., (Copyright © 2021 Elsevier Inc. All rights reserved.)
- Published
- 2022
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41. Hepatocytes Are Resistant to Cell Death From Canonical and Non-Canonical Inflammasome-Activated Pyroptosis.
- Author
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Sun P, Zhong J, Liao H, Loughran P, Mulla J, Fu G, Tang D, Fan J, Billiar TR, Gao W, and Scott MJ
- Subjects
- Hepatocytes metabolism, Intracellular Signaling Peptides and Proteins metabolism, Phosphate-Binding Proteins metabolism, Inflammasomes metabolism, Pyroptosis
- Abstract
Background: Pyroptosis, gasdermin-mediated programmed cell death, is readily induced in macrophages by activation of the canonical inflammasome (caspase-1) or by intracellular lipopolysaccharide (LPS)-mediated non-canonical inflammasome (caspase-11) activation. However, whether pyroptosis is induced similarly in hepatocytes is still largely controversial but highly relevant to liver pathologies such as alcoholic/nonalcoholic liver disease, drug-induced liver injury, ischemia-reperfusion and liver transplant injury, or organ damage secondary to sepsis., Methods and Results: In this study we found that hepatocytes activate and cleave gasdermin-D (GSDMD) at low levels after treatment with LPS. Overexpression of caspase-1 or caspase-11 p10/p20 activated domains was able to induce typical GSDMD-dependent pyroptosis in hepatocytes both in vitro and in vivo. However, morphologic features of pyroptosis in macrophages (eg, pyroptotic bodies, cell flattening, loss of cell structure) did not occur in pyroptotic hepatocytes, with cell structure remaining relatively intact despite the cell membrane being breached. Our results suggest that hepatocytes activate pyroptosis pathways and cleave GSDMD, but this does not result in cell rupture and confer the same pyroptotic morphologic changes as previously reported in macrophages. This is true even with caspase-1 or caspase-11 artificial overexpression way above levels seen endogenously even after priming or in pathologic conditions., Conclusions: Our novel findings characterize hepatocyte morphology in pyroptosis and suggest alternative use for canonical/non-canonical inflammasome activation/signaling and subsequent GSDMD cleavage because there is no rapid cell death as in macrophages. Improved understanding and recognition of the role of these pathways in hepatocytes may result in novel therapeutics for a range of liver diseases., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)
- Published
- 2022
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42. Patient-Specific Precision Injury Signatures to Optimize Orthopaedic Interventions in Multiply Injured Patients (PRECISE STUDY).
- Author
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McKinley TO, Gaski GE, Billiar TR, Vodovotz Y, Brown KM, Elster EA, Constantine GM, Schobel SA, Robertson HT, Meagher AD, Firoozabadi R, Gary JL, O'Toole RV, Aneja A, Trochez KM, Kempton LB, Steenburg SD, Collins SC, Frey KP, and Castillo RC
- Subjects
- Humans, Precision Medicine, Prospective Studies, Multiple Trauma surgery, Orthopedic Procedures, Orthopedics
- Abstract
Summary: Optimal timing and procedure selection that define staged treatment strategies can affect outcomes dramatically and remain an area of major debate in the treatment of multiply injured orthopaedic trauma patients. Decisions regarding timing and choice of orthopaedic procedure(s) are currently based on the physiologic condition of the patient, resource availability, and the expected magnitude of the intervention. Surgical decision-making algorithms rarely rely on precision-type data that account for demographics, magnitude of injury, and the physiologic/immunologic response to injury on a patient-specific basis. This study is a multicenter prospective investigation that will work toward developing a precision medicine approach to managing multiply injured patients by incorporating patient-specific indices that quantify (1) mechanical tissue damage volume; (2) cumulative hypoperfusion; (3) immunologic response; and (4) demographics. These indices will formulate a precision injury signature, unique to each patient, which will be explored for correspondence to outcomes and response to surgical interventions. The impact of the timing and magnitude of initial and staged surgical interventions on patient-specific physiologic and immunologic responses will be evaluated and described. The primary goal of the study will be the development of data-driven models that will inform clinical decision-making tools that can be used to predict outcomes and guide intervention decisions., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)
- Published
- 2022
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43. ADAR1 RNA editing regulates endothelial cell functions via the MDA-5 RNA sensing signaling pathway.
- Author
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Guo X, Liu S, Yan R, Nguyen V, Zenati M, Billiar TR, and Wang Q
- Subjects
- Animals, Biomarkers, Gene Deletion, Genes, Lethal, Immunity, Innate, Mice, Mice, Knockout, Mice, Transgenic, Repetitive Sequences, Nucleic Acid, Transcription, Genetic, Adenosine Deaminase genetics, Endothelial Cells metabolism, Gene Expression Regulation, Interferon-Induced Helicase, IFIH1 genetics, RNA Editing, Signal Transduction
- Abstract
The RNA-sensing signaling pathway has been well studied as an essential antiviral mechanism of innate immunity. However, its role in non-infected cells is yet to be thoroughly characterized. Here, we demonstrated that the RNA sensing signaling pathway also reacts to the endogenous cellular RNAs in endothelial cells (ECs), and this reaction is regulated by the RNA-editing enzyme ADAR1. Cellular RNA sequencing analysis showed that EC RNAs endure extensive RNA editing, especially in the RNA transcripts of short interspersed nuclear elements. The EC-specific deletion of ADAR1 dramatically reduced the editing level on short interspersed nuclear element RNAs, resulting in newborn death in mice with damage evident in multiple organs. Genome-wide gene expression analysis revealed a prominent innate immune activation with a dramatically elevated expression of interferon-stimulated genes. However, blocking the RNA sensing signaling pathway by deletion of the cellular RNA receptor MDA-5 prevented interferon-stimulated gene expression and rescued the newborn mice from death. This evidence demonstrated that the RNA-editing/RNA-sensing signaling pathway dramatically modulates EC function, representing a novel molecular mechanism for the regulation of EC functions., (© 2021 Guo et al.)
- Published
- 2021
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44. Multi-omic analysis in injured humans: Patterns align with outcomes and treatment responses.
- Author
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Wu J, Vodovotz Y, Abdelhamid S, Guyette FX, Yaffe MB, Gruen DS, Cyr A, Okonkwo DO, Kar UK, Krishnamoorthi N, Voinchet RG, Billiar IM, Yazer MH, Namas RA, Daley BJ, Miller RS, Harbrecht BG, Claridge JA, Phelan HA, Zuckerbraun BS, Johansson PI, Stensballe J, Morrissey JH, Tracy RP, Wisniewski SR, Neal MD, Sperry JL, and Billiar TR
- Subjects
- Cluster Analysis, Cohort Studies, Humans, Metabolome, Plasma, Proteome metabolism, Time Factors, Treatment Outcome, Brain Injuries, Traumatic genetics, Brain Injuries, Traumatic therapy, Genomics
- Abstract
Trauma is a leading cause of death and morbidity worldwide. Here, we present the analysis of a longitudinal multi-omic dataset comprising clinical, cytokine, endotheliopathy biomarker, lipidome, metabolome, and proteome data from severely injured humans. A "systemic storm" pattern with release of 1,061 markers, together with a pattern suggestive of the "massive consumption" of 892 constitutive circulating markers, is identified in the acute phase post-trauma. Data integration reveals two human injury response endotypes, which align with clinical trajectory. Prehospital thawed plasma rescues only endotype 2 patients with traumatic brain injury (30-day mortality: 30.3 versus 75.0%; p = 0.0015). Ubiquitin carboxy-terminal hydrolase L1 (UCHL1) was identified as the most predictive circulating biomarker to identify endotype 2-traumatic brain injury (TBI) patients. These response patterns refine the paradigm for human injury, while the datasets provide a resource for the study of critical illness, trauma, and human stress responses., Competing Interests: Y.V. is co-founder and a stakeholder in Immunetrics. H.A.P. is a consultant for Avita Medical and Spectral MD. M.D.N. holds an equity stake in Haima Therapeutics. He has received research support and/or honoraria from Haemonetics, Instrumentation Laboratories, and Janssen Pharmaceuticals. T.R.B. is a stakeholder in Immunetrics. Other authors declare no conflict of interests.
- Published
- 2021
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45. Single-Cell Transcriptomics Reveals Compartment-Specific Differences in Immune Responses and Contributions for Complement Factor 3 in Hemorrhagic Shock Plus Tissue Trauma.
- Author
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Fu G, Chen T, Wu J, Jiang T, Tang D, Bonaroti J, Conroy J, Scott MJ, Deng M, and Billiar TR
- Subjects
- Animals, Male, Mice, Mice, Inbred C57BL, Shock, Hemorrhagic complications, Wounds and Injuries complications, Complement C3 immunology, Immunity, Cellular, Shock, Hemorrhagic immunology, Transcriptome immunology, Wounds and Injuries immunology
- Abstract
Abstract: Hemorrhagic shock with tissue trauma (HS/T) leads to the activation of a system-wide immune-inflammatory response that involves all organs and body compartments. Recent advances in single-cell analysis permit the simultaneous assessment of transcriptomic patterns in a large number of cells making it feasible to survey the landscape of immune cell responses across numerous anatomic sites. Here, we used single-cell RNA sequencing of leukocytes from the blood, liver, and spleen to identify the major shifts in gene expression by cell type and compartment in a mouse HS/T model. At 6 h, dramatic changes in gene expression were observed across multiple-cell types and in all compartments in wild-type mice. Monocytes from circulation and liver exhibited a significant upregulation of genes associated with chemotaxis and migration and a simultaneous suppression of genes associated with interferon signaling and antigen presentation. In contrast, liver conventional DC exhibited a unique pattern compared with other myeloid cells that included a pronounced increase in major histocompatibility complex class II (MHCII) gene expression. The dominant pattern across all compartments for B and T cells was a suppression of genes associated with cell activation and signaling after HS/T. Using complement factor 3 (C3) knockout mice we unveiled a role for C3 in the suppression of monocyte Major Histocompatibility Complex class II expression and activation of gene expression associated with migration, phagocytosis and cytokine upregulation, and an unexpected role in promoting interferon-signaling in a subset of B and T cells across all three compartments after HS/T. This transcriptomic landscape study of immune cells provides new insights into the host immune response to trauma, as well as a rich resource for further investigation of trauma-induced immune responses and complement in driving interferon signaling., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 by the Shock Society.)
- Published
- 2021
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46. Immuno-Spin Trapping Method for the Analysis of S-Nitrosylated Proteins.
- Author
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Sircar E, Stoyanovsky DA, Billiar TR, and Sengupta R
- Subjects
- Free Radicals, Spin Trapping, Sulfhydryl Compounds, Proteins, Proteomics
- Abstract
The dynamic and unstable nature of protein nitrosothiols (PSNOs) derived from complex biological matrices (like cell lysates) make them unsuitable for proteomic/biochemical analysis in vitro. In an attempt to increase the stability of cell-derived PSNOs, scientists have devised methods to derivatize thiols undergoing nitrosylation, with a suitable molecule, to yield a stable adduct that could easily be detected using appropriate antibodies. The Biotin Switch Assay (BTSA) is currently the most widely used method for tagging PSNOs; however, the error-prone and cumbersome nature of the BTSA protocol prompted the development of alternative mechanisms of tagging cell-derived PSNOs. One such method is the immuno-spin trapping method using 5,5-dimethyl-1-pyrroline N-oxide (DMPO), which effectively overcomes the shortcomings of the BTSA and proves to be a promising alternative. Here we describe the protocol for DMPO-based PSNO labeling and subsequent proteomic analysis by western blotting with an anti-DMPO antibody. © 2021 Wiley Periodicals LLC. Basic Protocol: Labeling of cell-derived PSNOs by DMPO-based immuno-spin trapping and their subsequent analysis by immunostaining., (© 2021 Wiley Periodicals LLC.)
- Published
- 2021
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47. Mechanical Ventilation with Moderate Tidal Volume Exacerbates Extrapulmonary Sepsis-Induced Lung Injury via IL33-WISP1 Signaling Pathway.
- Author
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Liu S, Deng M, Pan P, Turnquist HR, Pitt BR, Billiar TR, and Zhang LM
- Subjects
- Animals, Disease Models, Animal, Male, Mice, Mice, Inbred C57BL, Sepsis therapy, Signal Transduction physiology, Ventilator-Induced Lung Injury metabolism, CCN Intercellular Signaling Proteins physiology, Interleukin-33 physiology, Proto-Oncogene Proteins physiology, Respiration, Artificial adverse effects, Sepsis complications, Tidal Volume physiology, Ventilator-Induced Lung Injury etiology
- Abstract
Abstract: IL-33 and WNT1-inducible secreted protein (WISP1) play central roles in acute lung injury (ALI) induced by mechanical ventilation with moderate tidal volume (MTV) in the setting of sepsis. Here, we sought to determine the inter-relationship between IL-33 and WISP1 and the associated signaling pathways in this process.We used a two-hit model of cecal ligation puncture (CLP) followed by MTV ventilation (4 h 10 mL/kg) in wild-type, IL-33-/- or ST2-/- mice or wild-type mice treated with intratracheal antibodies to WISP1. Macrophages (Raw 264.7 and alveolar macrophages from wild-type or ST2-/- mice) were used to identify specific signaling components.CLP + MTV resulted in ALI that was partially sensitive to genetic ablation of IL-33 or ST2 or antibody neutralization of WISP1. Genetic ablation of IL-33 or ST2 significantly prevented ALI after CLP + MTV and reduced levels of WISP1 in the circulation and bronchoalveolar lung fluid. rIL-33 increased WISP1 in alveolar macrophages in an ST2, PI3K/AKT, and ERK dependent manner. This WISP1 upregulation and WNT β-catenin activation were sensitive to inhibition of the β-catenin/TCF/CBP/P300 nuclear pathway.We show that IL-33 drives WISP1 upregulation and ALI during MTV in CLP sepsis. The identification of this relationship and the associated signaling pathways reveals a number of possible therapeutic targets to prevent ALI in ventilated sepsis patients., Competing Interests: The authors report no conflicts of interest., (Copyright © 2021 by the Shock Society.)
- Published
- 2021
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