Dysregulated proinflammatory cytokine release has been implicated in the pathogenesis of several life-threatening acute lung illnesses such as pneumonia, sepsis, and acute respiratory distress syndrome. Suppressors of cytokine signaling proteins, particularly SOCS2, have recently been described as antiinflammatory mediators. However, the regulation of SOCS2 protein has not been described. Here we describe a mechanism of SOCS2 regulation by the action of the ubiquitin E3 ligase KIAA0317. KIAA0317-mediated degradation of SOCS2 exacerbated inflammation in vitro, and depletion of KIAA0317 in vivo ameliorated pulmonary inflammation. KIAA0317-knockout mice exhibited resistance to LPS-induced pulmonary inflammation, while KIAA03017 reexpression mitigated this effect. We uncovered a small molecule inhibitor of KIAA0317 protein (BC-1365) that prevented SOCS2 degradation and attenuated LPS- and P. aeruginosa–induced lung inflammation in vivo. These studies show KIAA0317 to be a critical mediator of pulmonary inflammation through its degradation of SOCS2 and a potential candidate target for therapeutic inhibition.
Travis B. Lear, Alison C. McKelvey, John W. Evankovich, Shristi Rajbhandari, Tiffany A. Coon, Sarah R. Dunn, James D. Londino, Bryan J. McVerry, Yingze Zhang, Eleanor Valenzi, Christine L. Burton, Rachael Gordon, Sebastien Gingras, Karina C. Lockwood, Michael J. Jurczak, Robert Lafyatis, Mark J. Shlomchik, Yuan Liu, Bill B. Chen
miR-511-3p, encoded by CD206/Mrc1, was demonstrated to reduce allergic inflammation and promote alternative (M2) macrophage polarization. Here, we sought to elucidate the fundamental mechanism by which miR-511-3p attenuates allergic inflammation and promotes macrophage polarization. Compared with wild-type mice, the allergen-challenged Mrc1-/- mice showed increased airway hyper-responsiveness (AHR) and inflammation. However, this increased AHR and inflammation were significantly attenuated when these mice were pre-transduced with adeno-associated virus (AAV)-miR-511-3p. Gene expression profiling of macrophages identified Ccl2 as one of the major genes that was highly expressed in M2 macrophages but antagonized by miR-511-3p. The interaction between miR-511-3p and Ccl2 was confirmed by in silico analysis and mRNA-miRNA pull-down assay. Further evidence for the inhibition of Ccl2 by miR-511-3p was given by reduced levels of Ccl2 in supernatants of miR-511-3p transduced macrophages and in bronchoalveolar lavage fluids of AAV-miR-511-3p-infected Mrc1-/- mice. Mechanistically, we demonstrated that Ccl2 promotes M1 macrophage polarization by activating RhoA signaling through Ccr2. The interaction between Ccr2 and RhoA was also supported by co-immunoprecipitation assay. Importantly, inhibition of RhoA signaling suppressed cockroach allergen-induced AHR and lung inflammation. These findings suggest a novel mechanism by which miR-511-3p regulates allergic inflammation and macrophage polarization by targeting Ccl2 and its downstream Ccr2/RhoA axis.
Danh C. Do, Jie Mu, Xia Ke, Karan Sachdeva, Zili Qin, Mei Wan, Faoud T. Ishmael, Peisong Gao
Osteoarthritis (OA) is the leading cause of joint failure, yet the underlying mechanisms remain elusive, and no approved therapies that slow progression exist. Dysregulated integrin function was previously implicated in OA pathogenesis. However, the roles of integrin αVβ3 and the integrin-associated receptor CD47 in OA remain largely unknown. Here, transcriptomic and proteomic analyses of human and murine osteoarthritic tissues revealed dysregulated expression of αVβ3, CD47, and their ligands. Using genetically deficient mice and pharmacologic inhibitors, we showed that αVβ3, CD47, and the downstream signaling molecules Fyn and FAK are crucial to OA pathogenesis. MicroPET/CT imaging of a mouse model showed elevated ligand-binding capacities of integrin αVβ3 and CD47 in osteoarthritic joints. Further, our in vitro studies demonstrated that chondrocyte breakdown products, derived from articular cartilage of individuals with OA, induced αVβ3/CD47-dependent expression of inflammatory and degradative mediators, and revealed the downstream signaling network. Our findings identify a central role for dysregulated αVβ3 and CD47 signaling in OA pathogenesis and suggest that activation of αVβ3 and CD47 signaling in many articular cell types contributes to inflammation and joint destruction in OA. Thus, the data presented here provide a rationale for targeting αVβ3, CD47, and their signaling pathways as a disease-modifying therapy.
Qian Wang, Kazuhiro Onuma, Changhao Liu, Heidi Wong, Michelle S. Bloom, Eileen E. Elliott, Richard R.L. Cao, Nick Hu, Nithya Lingampalli, Orr Sharpe, Xiaoyan Zhao, Dong Hyun Sohn, Christin M. Lepus, Jeremy Sokolove, Rong Mao, Cecilia T. Cisar, Harini Raghu, Constance R. Chu, Nicholas J. Giori, Stephen B. Willingham, Susan S. Prohaska, Zhen Cheng, Irving L. Weissman, William H. Robinson
Dendritic cells (DCs) are crucial to balance protective immunity and autoimmune inflammatory processes. Expression of CD83 is a well-established marker for mature DCs although its physiological role is still not completely understood. Using a DC-specific CD83 conditional KO mouse (CD83ΔDC) we provide new insights into the function of CD83 within this cell type. Interestingly, CD83-deficient DCs produced drastically increased IL-2 levels and displayed higher expression of the co-stimulatory molecules CD25 and OX40L, which causes superior induction of antigen-specific T cell responses and compromises Treg suppressive functions. This also directly translates into accelerated immune responses in vivo. Upon Salmonella typhimurium and Listeria monocytogenes infection, CD83ΔDC mice cleared both pathogens more efficiently, and CD83-deficient DCs expressed increased IL-12 levels after bacterial encounter. Using the experimental autoimmune encephalomyelitis (EAE) model, autoimmune inflammation was dramatically aggravated in CD83ΔDC mice, while resolution of inflammation was strongly reduced. This phenotype was associated with increased cell influx into the CNS accompanied by elevated Th17 cell numbers. Concomitantly, CD83ΔDC mice had reduced Treg numbers in peripheral lymphoid organs. In summary, we show that CD83 ablation on DCs results in enhanced immune responses by dysregulating tolerance mechanisms and thereby impairing resolution of inflammation, which also demonstrates high clinical relevance.
Andreas B. Wild, Lena Krzyzak, Katrin Peckert, Lena Stich, Christine Kuhnt, Alina Butterhof, Christine Seitz, Jochen Mattner, Niklas Grüner, Maximilian Gänsbauer, Martin Purtak, Didier Soulat, Thomas H. Winkler, Lars Nitschke, Elisabeth Zinser, Alexander Steinkasserer
Background: Epicardial adipose tissue (EAT) is the visceral fat depot of the heart. Inflammation of EAT is thought to contribute to coronary artery disease (CAD). Therefore, we hypothesized that the EAT of patients with CAD would have increased inflammatory gene expression compared to controls without CAD. Methods: 26 patients referred for cardiac surgery with (n=13) or without CAD (n=13) were consented. Samples of EAT and subcutaneous adipose tissue (SAT) were obtained at the time of surgery. Gene expression analysis was performed using Affymetrix Human Gene 1.0 ST arrays. Differential regulation was defined as a 1.5 fold change (ANOVA p<0.05). Results: When comparing SAT and EAT of controls, 693 genes were differentially expressed. 805 genes were differentially expressed between SAT and EAT in cases. Expression of 326 genes was different between EAT of cases and controls; expression of 14 genes was increased in cases, while 312 were increased in controls. qRT-PCR confirmed that there was no difference in expression of major inflammatory genes (CCL2, CCR2, TNFα, IL6, IL8, PAI1) between cases and controls. Immunohistochemistry demonstrated that there were more macrophages in EAT than SAT, but that there was no difference in the number or activation state between cases and controls. Conclusion: In contrast to prior studies, we did not find increased inflammatory gene expression in the EAT of patients with CAD in comparison to controls without CAD. We conclude that specific adipose tissue organ, rather than CAD status, is responsible for the majority of differential gene expression.
Timothy P. Fitzgibbons, Nancy Lee, Khanh-Van Tran, Sara Nicoloro, Mark Kelly, Stanley K.C. Tam, Michael P. Czech
Although human cytomegalovirus (HCMV) is a known cause of sensorineural hearing loss in infants with congenital HCMV (cCMV) infections, mechanisms that contribute to sensorineural hearing loss (SNHL) in infants with cCMV infection are not well defined. Using a murine model of CMV infection during auditory development, we have shown that peripheral infection of newborn mice with murine CMV (MCMV) results in focal infection of the cochlea and virus-induced cochlear inflammation. Approximately 50%–60% of infected mice exhibited increased auditory brainstem response (ABR) thresholds across a range of sound frequencies. Histological analyses of the cochlea in MCMV-infected mice with elevated ABR thresholds revealed preservation of hair cell (HC) number and morphology in the organ of Corti. In contrast, the number of spiral ganglion neurons (SGN), synapses, and neurites connecting the cochlear HC and SGN nerve terminals were decreased. Decreasing cochlear inflammation by corticosteroid treatment of MCMV-infected mice resulted in preservation of SGN and improved auditory function. These findings show that virus-induced cochlear inflammation during early auditory development, rather than direct virus-mediated damage, could contribute to histopathology in the cochlea and altered auditory function without significant loss of HCs in the sensory epithelium.
Cathy Yea Won Sung, Maria C. Seleme, Shelby Payne, Stipan Jonjic, Keiko Hirose, William Britt
Long-term survivors post hematopoietic stem cell transplantation are at high risk of infection which accounts for one-third of all deaths. Little is known about the cause of inferior host defense after immune cell reconstitution. Here, we exploited a murine syngeneic bone marrow transplantation (BMT) model of late infection with gammaherpesvirus 68 (MHV-68) to determine the role of conventional dendritic cell (cDC) trafficking in adaptive immunity in BMT mice. Post infection, the expression of chemokine Ccl21 in the lung is reduced and the migration of cDCs into lung draining lymph nodes (dLNs) is impaired in BMT mice, limiting the opportunity for cDCs to prime Th cells in the dLNs. While cDC subsets are redundant in priming Th1 cells, Notch2 functions in cDC2s are required for priming increased Th17 responses in BMT mice and cDC1s can lessen this activity. Importantly, Th17 cells can be primed both in the lungs and dLNs, allowing for increased Th17 responses without optimum cDC trafficking in BMT mice. Taken together, impaired cDC trafficking in BMT mice reduces protective Th1 responses and allows increased pathogenic Th17 responses. Thus, we have revealed a previously unknown mechanism for BMT procedures to cause long-term inferior immune responses to herpes viral infection.
Carol A. Wilke, Matthew M. Chadwick, Paul R. Chan, Bethany B. Moore, Xiaofeng Zhou
Mesenchymal stem cells (MSCs) can suppress pathological inflammation. However, the mechanisms underlying the association between MSCs and inflammation remain unclear. Under coculture conditions with macrophages, MSCs highly expressed angiopoietin-like 4 (ANGPTL4) to blunt the polarization of macrophages toward the proinflammatory phenotype. ANGPTL4-deficient MSCs failed to inhibit the inflammatory macrophage phenotype. In inflammation-related animal models, the injection of coculture medium or ANGPTL4 protein increased the antiinflammatory macrophages in both peritonitis and myocardial infarction. In particular, cardiac function and pathology were markedly improved by ANGPTL4 treatment. We found that retinoic acid–related orphan receptor α (RORα) was increased by inflammatory mediators, such as IL-1β, and bound to ANGPTL4 promoter in MSCs. Collectively, RORα-mediated ANGPTL4 induction was shown to contribute to the antiinflammatory activity of MSCs against macrophages under pathological conditions. This study suggests that the capability of ANGPTL4 to induce tissue repair is a promising opportunity for safe stem cell–free regeneration therapy from a translational perspective.
Dong Im Cho, Hye-jin Kang, Ju Hee Jeon, Gwang Hyeon Eom, Hyang Hee Cho, Mi Ra Kim, Meeyoung Cho, Hye-yun Jeong, Hyen Chung Cho, Moon Hwa Hong, Yong Sook Kim, Youngkeun Ahn
It has been hypothesized that interleukin-1alpha (IL-1α) is released from damaged cardiomyocytes following myocardial infarction (MI) and activates cardiac fibroblasts via its receptor (IL-1R1) to drive the early stages of cardiac remodeling. This study aimed to definitively test this hypothesis using cell type-specific IL-1α and IL-1R1 knockout (KO) mouse models. A floxed Il1α mouse was created and used to generate a cardiomyocyte-specific IL-1α KO mouse line (MIL1AKO). A tamoxifen-inducible fibroblast-specific IL-1R1 hemizygous KO mouse line (FIL1R1KO) was also generated. Mice underwent experimental MI (permanent left anterior descending coronary artery ligation) and cardiac function was determined 4 weeks later by conductance pressure-volume catheter analysis. Molecular markers of remodeling were evaluated at various time points by real-time RT-PCR and histology. MIL1AKO mice showed no difference in cardiac function or molecular markers of remodeling post-MI compared with littermate controls. In contrast, FIL1R1KO mice showed improved cardiac function and reduced remodeling markers post-MI compared with littermate controls. In conclusion, these data highlight a key role for the IL-1R1/cardiac fibroblast signaling axis in regulating post-MI remodeling and provide support for the continued development of anti-IL-1 therapies for improving cardiac function after MI. Cardiomyocyte-derived IL-1α was not an important contributor to post-MI remodeling in this model.
Sumia A. Bageghni, Karen E. Hemmings, Nadira Y. Yuldasheva, Azhar Maqbool, Filomena O. Gamboa-Esteves, Neil E. Humphreys, Maj Simonsen Jackson, Christopher P. Denton, Sheila Francis, Karen E. Porter, Justin F. X. Ainscough, Emmanuel Pinteaux, Mark J. Drinkhill, Neil A. Turner
Steroid-refractory intestinal acute graft-versus-host disease (aGVHD) is a frequently fatal condition with little known about mechanisms driving failed steroid responses in gut mucosa. To uncover novel molecular insights in steroid-refractory aGVHD, we compared gene expression profiles of rectosigmoid biopsies from patients at diagnosis of clinical stage 3-4 lower intestinal aGVHD (N=22), to repeat biopsies when the patients became steroid refractory (N=22), and normal controls (N=10). We also performed single gene analyses of factors associated with tolerance (programmed death ligand-1 [PDL1], indoleamine 2,3 dioxygenase [IDO1], and T cell immunoreceptor with Ig and ITIM domains [TIGIT]) and found that significantly higher expression levels of these aGVHD inhibitory genes (PDL1, IDO1, TIGIT) at aGVHD onset became decreased in the steroid-refractory state. We examined genes triggered by microbial ligands to stimulate gut repair, amphiregulin (AREG) and the aryl hydrocarbon receptor (AhR), and found that both AREG and AhR gene expression levels were increased at aGVHD onset and remained elevated in steroid-refractory aGVHD. We also identified higher expression levels of metallothioneines, metal-binding enzymes induced in stress responses, and M2 macrophage genes in steroid-refractory aGVHD. We observed no differences in T-cell subsets between onset and steroid-refractory aGVHD. Patients with a rapidly fatal course showed greater DNA damage and a distinct microbial signature at aGVHD onset, whereas patients with more prolonged survival exhibited a gene expression profile consistent with activation of Smoothened. Our results extend the paradigm beyond T cell-centric therapies for steroid-refractory GI aGVHD and highlight new mechanisms for therapeutic exploration.
Shernan G. Holtan, Ashraf Shabaneh, Brian C. Betts, Armin Rashidi, Margaret L. MacMillan, Celalettin Ustun, Khalid Amin, Byron P. Vaughn, Justin Howard, Alexander Khoruts, Mukta Arora, Todd E. DeFor, Darrell Johnson, Bruce R. Blazar, Daniel J. Weisdorf, Jinhua Wang
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