Pulmonology
Abstract
Lung allograft rejection results in the accumulation of low molecular weight hyaluronic acid (LMW-HA), which further propagates inflammation and tissue injury. We have previously shown that therapeutic lymphangiogenesis in a murine model of lung allograft rejection reduced tissue LMW-HA and was associated with improved transplant outcomes. Herein we investigated the use of 4-Methylumbelliferone, a known inhibitor of HA synthesis, to alleviate acute allograft rejection in a murine model of lung transplantation. We found that treating mice with 4MU from day 20-30 post-transplant was sufficient to significantly improve outcomes, characterized by a reduction in T-cell mediated lung inflammation, LMW-HA content, and improved pathology scores. In vitro, 4MU directly attenuated activation, proliferation, and differentiation of naïve CD4+ T-cells into Th1 cells. As 4MU has already been demonstrated to be safe for human use, we believe examining 4MU for the treatment of acute lung allograft rejection may be of clinical significance.
Authors
Jewel Imani, Kaifeng Liu, Ye Cui, Jean-Pierre Assaker, Junwen Han, Auyon J. Ghosh, Julie Ng, Shikshya Shrestha, Anthony M. Lamattina, Pierce H. Louis, Anne Hentschel, Anthony J. Esposito, Ivan O. Rosas, Xiaoli Liu, Mark A. Perrella, Jamil Azzi, Gary Visner, Souheil El-Chemaly
Abstract
Asthma is a common disease with profoundly variable natural history and patient morbidity. Heterogeneity has long been appreciated and much work has focused on identifying subgroups of patients with similar pathobiological underpinnings. Previous studies of the Severe Asthma Research Program (SARP) cohort linked gene expression changes to specific clinical and physiologic characteristics. While invaluable for hypothesis generation, these data include extensive candidate gene lists that complicate target identification and validation. In this analysis, we performed unsupervised clustering of the SARP cohort using bronchial epithelial cell gene expression data, identifying a transcriptional signature for participants suffering exacerbation prone asthma with impaired lung function. Clinically, participants in this asthma cluster exhibited a mixed inflammatory process and bore transcriptional hallmarks of nuclear factor kappa B (NF-κB) and activator protein 1 (AP-1) activation despite high corticosteroid exposure. Using supervised machine learning, we found a set of 31 genes that classified patients with high accuracy and could reconstitute clinical and transcriptional hallmarks of our patient clustering in an external cohort. Of these genes, IL18R1 (IL-18 Receptor 1) negatively associated with lung function and was highly expressed in the most severe patient cluster. We validated IL18R1 protein expression in lung tissue and identified downstream NF-κB and AP-1 activity, supporting IL-18 signaling in severe asthma pathogenesis and highlighting this approach for gene/pathway discovery.
Authors
Matthew J. Camiolo, Xiuxia Zhou, Qi Wei, Humberto E. Trejo Bittar, Naftali Kaminski, Anuradha Ray, Sally Wenzel
Abstract
Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease associated with unremitting fibroblast activation including fibroblast-to-myofibroblast transformation (FMT), migration, resistance to apoptotic clearance, and excessive deposition of extracellular matrix (ECM) proteins in the distal lung parenchyma. Aberrant activation of lung-developmental pathways is associated with severe fibrotic lung disease; however, the mechanisms through which these pathways activate fibroblasts in IPF remain unclear. Sox9 is a member of the HMG box family of DNA-binding transcription factors that are selectively expressed by epithelial cell progenitors to modulate branching morphogenesis during lung development. We demonstrate that Sox9 is upregulated via MAPK/PI3K-dependent signaling and by the transcription factor Wilms’ tumor 1 in distal lung-resident fibroblasts in IPF. Mechanistically, using fibroblast activation assays, we demonstrate that Sox9 functions as a positive regulator of FMT, migration, survival, and ECM production. Importantly, our in vivo studies demonstrate that fibroblast-specific deletion of Sox9 is sufficient to attenuate collagen deposition and improve lung function during TGFα-induced pulmonary fibrosis. Using a mouse model of bleomycin-induced pulmonary fibrosis, we show that myofibroblast-specific Sox9 overexpression augments fibroblast activation and pulmonary fibrosis. Thus, Sox9 functions as a profibrotic transcription factor in activating fibroblasts, illustrating the potential utility of targeting Sox9 in IPF treatment.
Authors
Prathibha R. Gajjala, Rajesh K. Kasam, Divyalakshmi Soundararajan, Debora Sinner, Steven K. Huang, Anil G. Jegga, Satish K. Madala
Abstract
Pulmonary hypertension (PH) is a severe cardiopulmonary disease characterized by complement-dependent, fibroblast-induced perivascular accumulation and pro-inflammatory activation of macrophages. We hypothesized that, in PH, nanoscale-sized small extracellular vesicles (sEVs), released by perivascular/adventitial fibroblasts, are critical mediators of complement-dependent pro-inflammatory activation of macrophages. Pulmonary adventitial fibroblasts were isolated from calves with severe PH (PH-Fibs) and age-matched controls (CO-Fibs). PH-Fibs exhibited increased secretion of sEVs, compared to CO-Fibs, and sEV biological activity was tested on mouse and bovine bone marrow-derived macrophages (BMDMs) and showed similar responses. PH-Fib-sEVs induced augmented expression of pro-inflammatory cytokines/chemokines and metabolic genes in BMDMs, compared to CO-Fib-sEVs. Pharmacological blockade of exosome release from PH-Fibs resulted in significant attenuation of pro-inflammatory activation of BMDMs. “Bottom-up” proteomic analyses revealed significant enrichment of complement and Coagulation cascades in PH-Fib-sEVs, including augmented expression of complement component C3. We therefore examined whether PH-Fib-sEVs-mediated pro-inflammatory activation of BMDMs was complement C3-dependent. Treatment of PH-Fibs with siC3-RNA significantly attenuated the capacity of PH-Fib-sEVs for pro-inflammatory activation of BMDMs. PH-Fib-sEVs mediated pro-glycolytic alterations and complement-dependent activation of macrophages toward a pro-inflammatory phenotype, as confirmed by metabolomic studies. Thus, fibroblast-released sEVs can serve as critical mediators of complement-induced perivascular/microenvironmental inflammation in PH.
Authors
Sushil Kumar, Maria G. Frid, Hui Zhang, Min Li, Suzette Riddle, R. Dale Brown, Subhash Chandra Yadav, Micaela K. Roy, Monika E. Dzieciatkowska, Angelo D’Alessandro, Kirk C. Hansen, Kurt R. Stenmark
Abstract
A role for hereditary influences in the susceptibility for chronic obstructive pulmonary disease (COPD) is widely recognized. Cytotoxic lymphocytes are implicated in COPD pathogenesis, and functions of these leukocytes are modulated by interactions between their killer-cell immunoglobulin-like receptors (KIR) and human leukocyte antigen (HLA)-Class I molecules on target cells. We hypothesized HLA-Class I and KIR inheritance affect risks for COPD. HLA-Class I alleles and KIR genotypes were defined by candidate gene analyses in multiple cohorts of COPD patients (total n=392) and control smokers with normal spirometry (total n=342). Compared to controls, COPD patients had over-representations of HLA-C*07 and activating KIR2DS1, with under-representations of HLA-C*12. Particular HLA-KIR permutations were synergistic; e.g. the presence of HLA-C*07 + KIR2DS1 + HLA-C12null vs. HLAC*07null + KIR2DS1null + HLA-C12 was associated with COPD, especially among HLA-C1 allotype homozygotes (OR=18.5, 95%CI=3.7-90.9, p<0.0001). Cytotoxicity of COPD lymphocytes was more enhanced by KIR stimulation than those of controls (p=0.005) and was correlated with lung function (r=0.44, p=0.004). These data show HLA-C and KIR polymorphisms strongly influence COPD susceptibility and highlight the importance of lymphocyte-mediated cytotoxicity in COPD pathogenesis. Findings here also indicate HLA-KIR typing could stratify at-risk patients and raise possibilities HLA-KIR axis modulation may have therapeutic potential.
Authors
Takudzwa Mkorombindo, Thi Tran-Nguyen, Kaiyu Yuan, Yingze Zhang, Jianmin Xue, Gerard J. Criner, Young-il Kim, Joseph M. Pilewski, Amit Gaggar, Michael H. Cho, Frank C. Sciurba, Steven R. Duncan
Abstract
Chronic obstructive pulmonary disease (COPD) is mainly caused by cigarette smoking, characterized by chronic inflammation in vulnerable individuals. However, it is unknown how genetic factors may shape chronic inflammation in COPD. To understand how hedgehog interacting protein, encoded by HHIP gene identified in the genome-wide association study in COPD, plays a role in inflammation, we utilized Hhip+/- mice that resemble persistent inflammation and emphysema observed in human COPD upon aging. By performing single cell RNA sequencing of the whole lung from mice at different ages, we found that Hhip+/- mice develop a cytotoxic immune response with a specific increase in Klrg1+CD8+T cells with upregulated Ifnγ expression recapitulating human COPD. Hhip expression was restricted to a lung fibroblast subpopulation that has increased interaction with CD8+T lymphocytes in Hhip+/- compared to Hhip+/+ during aging. Hhip-expressing lung fibroblasts have upregulated IL-18 pathway genes in Hhip+/- lung fibroblasts, which was sufficient to drive increased levels of IFNγ in CD8+T cells ex vivo. Our finding provides insight into how a common genetic variation contributes to the amplified lymphocytic inflammation in COPD.
Authors
Jeong H. Yun, ChangHee Lee, Tao Liu, Siqi Liu, Edy Y. Kim, Shuang Xu, Jeffrey L. Curtis, Luca Pinello, Russell P. Bowler, Edwin K. Silverman, Craig P. Hersh, Xiaobo Zhou
Abstract
Macrophage activation syndrome (MAS) is a life-threatening cytokine storm complicating systemic juvenile idiopathic arthritis (SJIA) driven by IFNγ. SJIA and MAS are associated with an unexplained emerging lung disease (SJIA-LD), with our recent work supporting pulmonary activation of IFNγ pathways pathologically linking SJIA-LD and MAS. Our objective was to mechanistically define the novel observation of pulmonary inflammation in the TLR9 mouse model of MAS. In acute MAS, lungs exhibit mild but diffuse CD4-predominant, perivascular interstitial inflammation with elevated IFNγ, IFN-induced chemokines, and AMΦ expression of IFNγ-induced genes. Single-cell RNA-sequencing confirmed IFN-driven transcriptional changes across lung cell types with myeloid expansion and detection of MAS-specific macrophage populations. Systemic MAS resolution was associated with increased AMΦ and interstitial lymphocytic infiltration. AMΦ transcriptomic analysis confirmed IFNγ-induced proinflammatory polarization during acute MAS, which switches towards an anti-inflammatory phenotype after systemic MAS resolution. Interestingly, recurrent MAS led to increased alveolar inflammation and lung injury, and reset AMΦ polarization towards a proinflammatory state. Furthermore, in mice bearing macrophages insensitive to IFNγ, both systemic feature of MAS and pulmonary inflammation were attenuated. These findings demonstrate that experimental MAS induces IFNγ-driven pulmonary inflammation replicating key features of SJIA-LD, and provides a model system for testing novel treatments directed towards SJIA-LD.
Authors
Denny K. Gao, Nathan Salomonis, Maggie Henderlight, Christopher Woods, Kairavee Thakkar, Alexei A. Grom, Sherry Thornton, Michael B. Jordan, Kathryn A. Wikenheiser-Brokamp, Grant S. Schulert
Abstract
In the current study, we followed 839 household contacts (HHCs) of tuberculosis (TB) patients for 2 years and identified the factors that enhanced the development of TB. Fourteen of the 17 HHCs who progressed to TB were in the 15- to 30-year-old age group. At baseline (the “0“ time point, when all the individuals were healthy), the concentration of the thyroid hormone thyroxine (T4) was lower, and there were increased numbers of Tregs in PBMCs of TB progressors. At baseline, PBMCs from TB progressors stimulated with early secretory antigenic target 6 (ESAT-6) and 10 kDa culture filtrate antigen (CFP-10) produced less IL-1α. Thyroid hormones inhibited Mycobacterium tuberculosis (Mtb) growth in macrophages in an IL-1α–dependent manner. Mtb-infected Thra1PV/+ (mutant thyroid hormone receptor) mice had increased mortality and reduced IL-1α production. Our findings suggest that young HHCs who exhibit decreased production of thyroid hormones are at high risk of developing active TB disease.
Authors
Kamakshi Prudhula Devalraju, Deepak Tripathi, Venkata Sanjeev Kumar Neela, Padmaja Paidipally, Rajesh Kumar Radhakrishnan, Karan P. Singh, Mohammad Soheb Ansari, Martin Jaeger, Romana T. Netea-Maier, Mihai G. Netea, Sunmi Park, Sheue-yann Cheng, Vijaya Lakshmi Valluri, Ramakrishna Vankayalapati
Abstract
Sepsis is a critical illness characterized by dysregulated inflammatory responses lacking counter-regulation. Specialized pro-resolving mediators are agonists for anti-inflammation and promoting resolution and are protective in preclinical sepsis models. Here, in human sepsis, we mapped resolution circuits for the specialized pro-resolving mediators resolvin D1 and resolvin D2 in peripheral blood neutrophils and monocytes, their regulation of leukocyte activation and function ex vivo, and their relationships to measures of clinical severity. Neutrophils and monocytes were isolated from healthy subjects and sepsis patients by inertial microfluidics and resolvin D1 and resolvin D2 receptor expression determined by flow cytometry. The impact of these resolvins on leukocyte activation was determined by isodielectric separation and leukocyte function by stimulated phagolysosome formation. Leukocyte pro-resolving receptor expression was significantly higher in sepsis. In nanomolar concentrations, resolvin D1 and resolvin D2 partially reversed sepsis-induced changes in leukocyte activation and function. Principal component analyses of leukocyte resolvin receptor expression and responses differentiated sepsis from health and were associated with measures of sepsis severity. These findings indicate that resolvin D1 and resolvin D2 signaling for anti-inflammation and resolution are uncoupled from leukocyte activation in early sepsis and suggest that indicators of diminished resolution signaling correlate with clinical disease severity.
Authors
Bakr Jundi, Do-Hyun Lee, Hyungkook Jeon, Melody G. Duvall, Julie Nijmeh, Raja-Elie E. Abdulnour, Mayra Pinilla-Vera, Rebecca M. Baron, Jongyoon Han, Joel Voldman, Bruce D. Levy
Abstract
Necroptosis has emerged as a potential mechanism in the pathogenesis of chronic obstructive pulmonary disease (COPD). Here, we found that markers of necroptosis, including high mobility group box 1 release and phosphorylation of mixed lineage kinase domain-like protein (p-MLKL), were markedly induced in the late stage of cigarette smoking–induced (CS-induced) emphysema in mouse lung tissue as well as in lung epithelial cells and organoids with higher dosage of or more prolonged exposure to cigarette smoking extract (CSE). Apoptotic signals were also detected and maximally induced in the early stage of CS-exposed mice and CSE-treated epithelial cells. Inhibition of apoptosis by Z-VAD, a pan-caspase inhibitor, switched the cellular stress to enhanced necroptosis in lung epithelial cells and organoids treated with CSE. Depletion or inhibition of receptor-interacting protein kinase 3 (RIP3) or MLKL attenuated the CSE-induced cell death, suggesting that necroptosis contributes to CSE-induced cell death. Silencing or inhibition of RIP1 had no protective effect, indicating a RIP1-independent RIP3 activation pathway. CSE-induced necroptosis released more damage-associated molecular patterns and evoked greater engulfment but slower clearance by bone marrow–derived macrophages, leading to enhanced expression of proinflammatory cytokines Tnfα and Il6. Finally, our in vivo data verified that inhibition of necroptosis by RIP3 inhibitor GSK’872 protected mice from CS-induced emphysema and suppressed the lung inflammation. In conclusion, we provide evidence that necroptosis contributes to the pathogenesis of COPD. Targeting RIP3 and its downstream pathway may be an effective therapy for COPD.
Authors
Dongshi Chen, Alyssa D. Gregory, Xiaoyun Li, Jianxin Wei, Christine L. Burton, Gregory Gibson, Stephen J. Scott, Claudette M. St. Croix, Yingze Zhang, Steven D. Shapiro
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