scholarly journals ATF3 Protects against LPS-Induced Inflammation in Mice via Inhibiting HMGB1 Expression

2013 ◽  
Vol 2013 ◽  
pp. 1-14 ◽  
Author(s):  
Pei-Fang Lai ◽  
Ching-Feng Cheng ◽  
Heng Lin ◽  
Tzu-Ling Tseng ◽  
Hsi-Hsien Chen ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Gene Knockout ◽  
Critical Role ◽  
Negative Regulator ◽  
Tlr4 Signaling ◽  
Lps Challenge ◽  
Hmgb1 Expression ◽  
Elevated Expression ◽  
Sepsis Therapy

Lipopolysaccharide (LPS) triggers innate immunity mainly via TLR4 signaling. ATF3 is a negative regulator of TLR4 signaling. HMGB1 plays a critical role in the final step of sepsis. However, the mechanisms of ATF3 and the role of HMGB1 in regulating innate immunity-induced sepsis are incompletely understood. In this study, we found that serum HMGB1 levels were 10-fold higher in patients with sepsis than normal controls. We further demonstrated that ATF3 gene knockout in mice subjected to LPS-induced endotoxemia correlates with an increase in the mortality rate and the elevated expression of IL-6, TNF-α, NO, MCP-1, and HMGB1 in the lung tissues or serum. The biochemical effects of ATF3 were observed inin vitromacrophages and blocked by ATF3 siRNA treatment. We have also shown that adeno-associated virus-mediated ATF3 gene transfer protected ATF3 knockout mice from LPS-induced mortality. In addition, ATF3 knockdown increased LPS-induced release of HMGB1. In conclusion, upregulation of ATF3 contributes to the reduced release of inflammatory molecules, especially HMGB1, which induced lung injury and increased the survival rate of mice after LPS challenge. Therefore, suppressing LPS-induced inflammation with ATF3 induction or ATF3 mimetics may be an important strategy for sepsis therapy.

Expression of ATF3 in mouse protects the liver against sepsis via inhibiting HMGB expression

2014 ◽  
Vol 2 (4) ◽  
pp. 288-299 ◽  
Keyword(s):  
Innate Immunity ◽  
Liver Tissue ◽  
Mononuclear Cells ◽  
Critical Role ◽  
Negative Regulator ◽  
Immunofluorescent Staining ◽  
Protective Effects ◽  
Tlr4 Signaling ◽  
Tnf Α ◽  
Sepsis Therapy

Lipopolysaccharides are components of Gram-negative enterobacteria that cause septic shock in mammals and triggers innate immunity mainly via TLR4 signaling. HMGB1 play a critical role in regulating innate immunity-induced sepsis. ATF3 is a negative regulator of TLR4 signaling and the mechanism of HMGB1 induced liver injury after sepsis are incompletely understood. In this study, we investigated the protective effects of ATF3 after LPS injection. Adult (4-6 months) C57/BL6 mice and ATF3 knockout mice were treated with a low dose of LPS (0.5 mg/kg, iv) for 6, 12 hrs. Liver enzymes and cytokines (TNF-α, IL-1β and IL-6) are assessed. The neutrophil and mononuclear cells in the liver tissue were examined using immunofluorescent staining. We found that serum HMGB1 levels were 8-fold higher in C57/BL6 mice with sepsis than ATF3 knockout with greater densities of neutrophils and mononuclear cells in the liver tissue, and higher levels of TNF-α, IL-1β and IL-6 in the circulation and liver tissue as well as associated with an increase in the mortality rate. In conclusion, upregulation of ATF3 contributes to the reduced release of HMGB1, and increased the survival rate of mice after LPS treated. Therefore, suppressing LPS-induced inflammation with ATF3 induction may be an important strategy for sepsis therapy.

Abstract P317: Cardiac Fibroblast GSK3α Promotes Myocardial Fibrotic Remodeling Through GSK3α-ERK-IL11 Signaling Circuit

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Prachi Umbarkar ◽  
Sultan Tousif ◽  
Anand P Singh ◽  
Joshua C Anderson ◽  
qinkun zhang ◽  
...  
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Critical Role ◽  
Flow Cytometric Analysis ◽  
Collagen Gel ◽  
Pressure Overload ◽  
Negative Regulator ◽  
Post Injury

Background: Myocardial fibrosis contributes significantly to heart failure (HF). Fibroblasts are among the predominant cell type in the heart and are primary drivers of fibrosis. To identify the kinases involved in fibrosis, we analyzed the kinome of mouse cardiac fibroblasts (CF) isolated from normal and failing hearts. This unbiased screening revealed the critical role of the GSK-3 family-centric pathways in fibrosis. Previously we have shown that among two isoforms of GSK3, CF-GSK3β acts as a negative regulator of fibrosis in the injured heart. However, the role of CF-GSK3α in the pathogenesis of cardiac diseases is completely unknown. Methods and Results: To define the role of CF-GSK3α in HF, we employed two novel fibroblast-specific KO mouse models. Specifically, GSK3α was deleted from fibroblasts or myofibroblasts with tamoxifen-inducible Tcf21- or periostin- promoter-driven Cre recombinase. In both models, GSK3α deletion restricted pressure overload-induced cardiac fibrosis and preserved cardiac function. We examined the effect of GSK3α deletion on myofibroblast transformation and pro-fibrotic TGFβ1-SMAD3 signaling in vitro . A significant reduction in cell migration, collagen gel contraction, and α-SMA expression in TGFβ1-treated KO CFs confirmed that GSK3α is required for myofibroblast transformation. Surprisingly, GSK3α deletion did not affect SMAD3 activation, indicating the pro-fibrotic role of GSK3α is SMAD3 independent. To further delineate the underlying mechanisms, proteins were isolated from CFs of WT and KO mice at 4 weeks post-injury, and kinome profiling was performed. The kinome analysis identified the downregulation of RAF family kinase activity in KO CFs. Moreover, mapping of significantly altered kinases against literature annotated interactions generated ERK-centric networks. Consistently, flow cytometric analysis of CFs confirmed significantly low levels of pERK in KO mice. Additionally, our in vitro studies demonstrated that GSK3α deletion prevents TGFβ1-induced ERK activation. Interestingly, IL-11, a pro-fibrotic downstream effector of TGFβ1, was remarkably reduced in KO CFs and ERK inhibition further decreased IL-11 expression. Taken together, herein, we discovered the GSK3α-ERK-IL-11 signaling as a critical pro-fibrotic pathway in the heart. Strategies to inhibit this pro-fibrotic network could prevent adverse fibrosis and HF. Conclusion: CF-GSK3α plays a causal role in myocardial fibrosis that could be therapeutically targeted for future clinical applications.

scholarly journals NG2 glia regulate brain innate immunity via TGF-β2/TGFBR2 axis

BMC Medicine ◽  
2019 ◽  
Vol 17 (1) ◽  
Author(s):  
Shu-zhen Zhang ◽  
Qin-qin Wang ◽  
Qiao-qiao Yang ◽  
Huan-yu Gu ◽  
Yan-qing Yin ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Diphtheria Toxin ◽  
Transforming Growth Factor ◽  
Critical Role ◽  
Normal Brain ◽  
Sequencing Analysis ◽  
Brain Functions ◽  
Lps Challenge ◽  
The Impact ◽  
The Brain

Abstract Background Brain innate immunity is vital for maintaining normal brain functions. Immune homeostatic imbalances play pivotal roles in the pathogenesis of neurological diseases including Parkinson’s disease (PD). However, the molecular and cellular mechanisms underlying the regulation of brain innate immunity and their significance in PD pathogenesis are still largely unknown. Methods Cre-inducible diphtheria toxin receptor (iDTR) and diphtheria toxin-mediated cell ablation was performed to investigate the impact of neuron-glial antigen 2 (NG2) glia on the brain innate immunity. RNA sequencing analysis was carried out to identify differentially expressed genes in mouse brain with ablated NG2 glia and lipopolysaccharide (LPS) challenge. Neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice were used to evaluate neuroinflammatory response in the presence or absence of NG2 glia. The survival of dopaminergic neurons or glial cell activation was evaluated by immunohistochemistry. Co-cultures of NG2 glia and microglia were used to examine the influence of NG2 glia to microglial activation. Results We show that NG2 glia are required for the maintenance of immune homeostasis in the brain via transforming growth factor-β2 (TGF-β2)-TGF-β type II receptor (TGFBR2)-CX3C chemokine receptor 1 (CX3CR1) signaling, which suppresses the activation of microglia. We demonstrate that mice with ablated NG2 glia display a profound downregulation of the expression of microglia-specific signature genes and remarkable inflammatory response in the brain following exposure to endotoxin lipopolysaccharides. Gain- or loss-of-function studies show that NG2 glia-derived TGF-β2 and its receptor TGFBR2 in microglia are key regulators of the CX3CR1-modulated immune response. Furthermore, deficiency of NG2 glia contributes to neuroinflammation and nigral dopaminergic neuron loss in MPTP-induced mouse PD model. Conclusions These findings suggest that NG2 glia play a critical role in modulation of neuroinflammation and provide a compelling rationale for the development of new therapeutics for neurological disorders.

scholarly journals B7-H4–deficient mice display augmented neutrophil-mediated innate immunity

Blood ◽  
2009 ◽  
Vol 113 (8) ◽  
pp. 1759-1767 ◽  
Author(s):  
Gefeng Zhu ◽  
Mathew M. Augustine ◽  
Takeshi Azuma ◽  
Liqun Luo ◽  
Sheng Yao ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Negative Regulator ◽  
Immunoglobulin Superfamily ◽  
Cell Responses ◽  
Inhibitory Function ◽  
Genetic Targeting ◽  
Neutrophil Response ◽  
Deficient Mice

Abstract B7-H4 is an immunoglobulin superfamily molecule and shown to be inhibitory for T-cell responses. To explore physiologic roles of B7-H4, we created B7-H4–deficient (KO) mice by genetic targeting. B7-H4KO mice are healthy and their T- and B-cell responses to polyclonal antigens are in normal range. However, B7-H4KO mice are more resistant to infection by Listeria monocytogenes than their littermates. Within 3 days after infection, bacterial colonies in livers and spleens are significantly lower than the controls, suggesting a role of B7-H4 in enhancing innate immunity. Further studies demonstrate that neutrophils increase in peripheral organs of B7-H4KO mice more so than their littermates but their bactericidal functions remain unchanged. Augmented innate resistance is completely dependent on neutrophils, even in the absence of adaptive immunity. In vitro B7-H4 inhibits the growth of bone marrow–derived neutrophil progenitors, suggesting an inhibitory function of B7-H4 in neutrophil expansion. Our results identify B7-H4 as a negative regulator of the neutrophil response to infection and provide a new target for manipulation of innate immunity.

Interaction and Regulation of HSC with Osteopontin Is Mediated through α4β1 and α9β1Integrins, Which Are Differentially Expressed in the HSC Pool.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1330-1330
Author(s):  
David N. Haylock ◽  
Genevieve A. Whitty ◽  
Brenda Williams ◽  
Melonie J. Storan ◽  
Susie K. Nilsson
Keyword(s):  
Cord Blood ◽  
Critical Role ◽  
Negative Regulator ◽  
Hemopoietic Stem Cell ◽  
Null Mouse ◽  
Dependent Manner ◽  
Independent Manner ◽  
Hsc Niche

Abstract Osteoblasts are a key cellular component of the hemopoietic stem cell (HSC) niche and directly regulate the HSC pool. Molecules synthesised by osteoblasts both promote or inhibit HSC proliferation. Osteopontin (Opn) is an osteoblast produced, RGD containing protein with roles in cell adhesion and migration. Until recently, the role of Opn in hemopoiesis was seen as restricted to the regulation of bone turnover. However, from analysis of hemopoiesis in the Opn null mouse, we have demonstrated that Opn plays a critical role in regulating the HSC pool. Furthermore Opn is critical in trans-marrow migration and lodgement of HSC within the BM after transplantation. When added to in vitro HSC cultures, exogenous thrombin-cleaved Opn also inhibits cell proliferation and potently suppresses HSC differentiation. We have now demonstrated that this interaction occurs in an RGD-independent manner via the cryptic SVVYGLR epitope revealed on the N-terminal fragment of Opn following thrombin cleavage. This epitope has previously been shown to bind to α4β1 and α9β1. HSC are known to express α4β1, but we have now shown that within the HSC pool this occurs in a differential manner, mimicking that of CD38, with more committed CD34+CD38+ cord blood progenitors having the highest levels of expression. In addition, we have shown the previously unrecognised characteristic of human marrow and cord blood HSC, the expression of α9β1, which also occurs in a differential manner, but mimicking CD34. Expression of α9β1 is highest on cord blood CD34+CD38− cells, a population highly enriched for HSC. Using the synthetic SVVYGLR peptide in culture, we re-capitulated the thrombin-cleaved Opn induced suppression of HSC differentiation in a dose dependent manner. Antibody blocking experiments demonstrated that binding to this peptide was occurring through both α4β1 and α9β1. In contrast, suppression of HSC proliferation and differentiation did not occur through the upstream alternate α4β1 binding site. Furthermore, we have now demonstrated endogenous binding of Opn to α4β1 and α9β1 to cord blood HSC in vivo. Together, these data provide strong evidence that Opn is an important component of the HSC niche which acts as a physiological negative regulator. Furthermore, our studies identify the previously unrecognised characteristic of HSC, the expression of α9β1, which together with α4β1 provides two receptors on HSC with differing expression signatures and potentially a mechanism for fine tunning the physiological effects of Opn.

scholarly journals Wnt Modulating Agents Inhibit Human Cytomegalovirus Replication

2013 ◽  
Vol 57 (6) ◽  
pp. 2761-2767 ◽  
Author(s):  
Arun Kapoor ◽  
Ran He ◽  
Rajkumar Venkatadri ◽  
Michael Forman ◽  
Ravit Arav-Boger
Keyword(s):  
Stem Cell ◽  
Cancer Stem Cell ◽  
Human Cytomegalovirus ◽  
Wnt Pathway ◽  
Critical Role ◽  
Wnt Signaling Pathway ◽  
Negative Regulator ◽  
Infected Cells ◽  
Hcmv Replication

ABSTRACTInfection with human cytomegalovirus (HCMV) continues to be a threat for pregnant women and immunocompromised hosts. Although limited anti-HCMV therapies are available, development of new agents is desired. The Wnt signaling pathway plays a critical role in embryonic and cancer stem cell development and is targeted by gammaherpesviruses, Epstein-Barr virus (EBV), and Kaposi's sarcoma-associated herpesvirus (KSHV). HCMV infects stem cells, including neural progenitor cells, during embryogenesis. To investigate the role of Wnt in HCMV replicationin vitro, we tested monensin, nigericin, and salinomycin, compounds that inhibit cancer stem cell growth by modulating the Wnt pathway. These compounds inhibited the replication of HCMV Towne and a clinical isolate. Inhibition occurred prior to DNA replication but persisted throughout the full replication cycle. There was a significant decrease in expression of IE2, UL44, and pp65 proteins. HCMV infection resulted in a significant and sustained decrease in expression of phosphorylated and total lipoprotein receptor-related protein 6 (pLRP6 and LRP6, respectively), Wnt 5a/b, and β-catenin and a modest decrease in Dvl2/3, while levels of the negative regulator axin 1 were increased. Nigericin decreased the expression of pLRP6, LRP6, axin 1, and Wnt 5a/b in noninfected and HCMV-infected cells. For all three compounds, a correlation was found between expression levels of Wnt 5a/b and axin 1 and HCMV inhibition. The decrease in Wnt 5a/b and axin 1 expression was more significant in HCMV-infected cells than noninfected cells. These data illustrate the complex effects of HCMV on the Wnt pathway and the fine balance between Wnt and HCMV, resulting in abrogation of HCMV replication. Additional studies are required to elucidate how HCMV targets Wnt for its benefit.

scholarly journals Role of the CBP catalytic core in intramolecular SUMOylation and control of histone H3 acetylation

2017 ◽  
Vol 114 (27) ◽  
pp. E5335-E5342 ◽  
Author(s):  
Sangho Park ◽  
Robyn L. Stanfield ◽  
Maria A. Martinez-Yamout ◽  
H. Jane Dyson ◽  
Ian A. Wilson ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Critical Role ◽  
Histone H3 ◽  
Negative Regulator ◽  
Creb Binding Protein ◽  
Catalytic Core ◽  
Intrinsically Disordered ◽  
Histone H3 Acetylation ◽  
H3 Acetylation

The histone acetyl transferases CREB-binding protein (CBP) and its paralog p300 play a critical role in numerous cellular processes. Dysregulation of their catalytic activity is associated with several human diseases. Previous work has elucidated the regulatory mechanisms of p300 acetyltransferase activity, but it is not known whether CBP activity is controlled similarly. Here, we present the crystal structure of the CBP catalytic core encompassing the bromodomain (BRD), CH2 (comprising PHD and RING), HAT, and ZZ domains at 2.4-Å resolution. The BRD, PHD, and HAT domains form an integral structural unit to which the RING and ZZ domains are flexibly attached. The structure of the apo-CBP HAT domain is similar to that of acyl-CoA–bound p300 HAT complexes and shows that the acetyl-CoA binding site is stably formed in the absence of cofactor. The BRD, PHD, and ZZ domains interact with small ubiquitin-like modifier 1 (SUMO-1) and Ubc9, and function as an intramolecular E3 ligase for SUMOylation of the cell cycle regulatory domain 1 (CRD1) of CBP, which is located adjacent to the BRD. In vitro HAT assays suggest that the RING domain, the autoregulatory loop (AL) within the HAT domain, and the ZZ domain do not directly influence catalytic activity, whereas the BRD is essential for histone H3 acetylation in nucleosomal substrates. Several lysine residues in the intrinsically disordered AL are autoacetylated by the HAT domain. Upon autoacetylation, acetyl-K1596 (Ac-K1596) binds intramolecularly to the BRD, competing with histones for binding to the BRD and acting as a negative regulator that inhibits histone H3 acetylation.

Abstract 330: Metallothionein Preservation Of Cardiac Akt2 Signaling By Down-regulating Trb3 Prevents Diabetic Cardiomyopathy

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
YI TAN ◽  
Xiaoqing Yan ◽  
Shanshan Zhou ◽  
Yong Li ◽  
Yan Li ◽  
...  
Keyword(s):  
Diabetic Cardiomyopathy ◽  
Insulin Signaling ◽  
Glycogen Synthase ◽  
Critical Role ◽  
Negative Regulator ◽  
H9c2 Cells ◽  
Akt Phosphorylation ◽  
Gsk 3Β

Cardiac insulin resistance is a key pathogenic factor for diabetic cardiomyopathy, but its mechanism remains largely unclear. Here we demonstrated that diabetes significantly inhibited cardiac Akt phosphorylation from 2 weeks to 2 months in wide-type (WT) mice, but not in cardiac-specific metallothionein-transgenic (MT-TG) mice. Cardiac Akt2 expression and phosphorylation was decreased and insulin-induced cardiac Akt2 and GSK-3β phosphorylation and glycogen synthase dephosphorylation were also decreased in WT, but not MT-TG, diabetic mice. Deletion of the Akt2 gene either in vitro H9c2 cells or in vivo significantly impaired cardiac glucose metabolic signaling. In addition, diabetes significantly increased cardiac Akt negative regulator tribbles (TRB)3 expression only in WT mice, suggesting the possible contribution of MT inhibition of diabetic up-regulation of TRB3 to Akt2 function preservation. Cardiac H9c2 cells with and without forced MT-overexpression (MT-H9c2) were treated with tert-butyl hydroperoxide (tBHP), which significantly reduced Akt2 phosphorylation in both basal and insulin-stimulating conditions only in H9c2 cells. Silencing TRB3 expression with SiRNA completely prevented tBHP’s inhibition of insulin-stimulated Akt2 phosphorylation in H9c2 cells, while overexpression of TRB3 in MT-H9c2 cells completely abolished MT preservation of insulin-stimulated Akt2 phosphorylation. Forced-overexpression of TRB3 by adenovirus-mediated gene delivery in MT-TG hearts also abolished MT’s preservation of cardiac insulin signaling and prevention of diabetic cardiomyopathy. These results suggest that diabetes-attenuated cardiac Akt2 function via up-regulating TRB3 plays a critical role in diabetic inhibition of insulin signaling in the heart. MT preserved cardiac Akt2-mediated insulin signaling by inhibiting TRB3, leading to the prevention of diabetic cardiomyopathy.

scholarly journals CRISPR-Cas9 mediated efficient PD-1 disruption on human primary T cells from cancer patients

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Shu Su ◽  
Bian Hu ◽  
Jie Shao ◽  
Bin Shen ◽  
Juan Du ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Checkpoint Inhibitors ◽  
Gene Knockout ◽  
Cell Activity ◽  
Negative Regulator ◽  
Activated T Cells ◽  
Human T Cells ◽  
New Strategy

Abstract Strategies that enhance the function of T cells are critical for immunotherapy. One negative regulator of T-cell activity is ligand PD-L1, which is expressed on dentritic cells (DCs) or some tumor cells and functions through binding of programmed death-1 (PD-1) receptor on activated T cells. Here we described for the first time a non-viral mediated approach to reprogram primary human T cells by disruption of PD-1. We showed that the gene knockout of PD-1 by electroporation of plasmids encoding sgRNA and Cas9 was technically feasible. The disruption of inhibitory checkpoint gene PD-1 resulted in significant reduction of PD-1 expression but didn’t affect the viability of primary human T cells during the prolonged in vitro culture. Cellular immune response of the gene modified T cells was characterized by up-regulated IFN-γ production and enhanced cytotoxicity. These results suggest that we have demonstrated an approach for efficient checkpoint inhibitor disruption in T cells, providing a new strategy for targeting checkpoint inhibitors, which could potentialy be useful to improve the efficacy of T-cell based adoptive therapies.

Osteopontin, a key component of the hematopoietic stem cell niche and regulator of primitive hematopoietic progenitor cells

Blood ◽  
2005 ◽  
Vol 106 (4) ◽  
pp. 1232-1239 ◽  
Author(s):  
Susan K. Nilsson ◽  
Hayley M. Johnston ◽  
Genevieve A. Whitty ◽  
Brenda Williams ◽  
Ryan J. Webb ◽  
...  
Keyword(s):  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Inflammatory Responses ◽  
Critical Role ◽  
Negative Regulator ◽  
Hematopoietic Stem ◽  
Hematopoietic Stem Cell Niche ◽  
Hsc Niche ◽  
Cell Niche

Abstract Although recent data suggests that osteoblasts play a key role within the hematopoietic stem cell (HSC) niche, the mechanisms underpinning this remain to be fully defined. The studies described herein examine the role in hematopoiesis of Osteopontin (Opn), a multidomain, phosphorylated glycoprotein, synthesized by osteoblasts, with well-described roles in cell adhesion, inflammatory responses, angiogenesis, and tumor metastasis. We demonstrate a previously unrecognized critical role for Opn in regulation of the physical location and proliferation of HSCs. Within marrow, Opn expression is restricted to the endosteal bone surface and contributes to HSC transmarrow migration toward the endosteal region, as demonstrated by the markedly aberrant distribution of HSCs in Opn–/– mice after transplantation. Primitive hematopoietic cells demonstrate specific adhesion to Opn in vitro via β1 integrin. Furthermore, exogenous Opn potently suppresses the proliferation of primitive HPCs in vitro, the physiologic relevance of which is demonstrated by the markedly enhanced cycling of HSC in Opn–/– mice. These data therefore provide strong evidence that Opn is an important component of the HSC niche which participates in HSC location and as a physiologic-negative regulator of HSC proliferation.

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