Abstract 356: Sam68 Promotes NF-kB Signaling and Inflammation in Macrophages and Impedes the Recovery of Arterial Injury

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Shuling Han ◽  
Junlan Zhou ◽  
Gangjian Qin
Keyword(s):  
Inflammatory Cytokines ◽  
Vascular Inflammation ◽  
Neointimal Hyperplasia ◽  
Critical Role ◽  
Inflammatory Cells ◽  
Arterial Injury ◽  
Filamin A ◽  
Interacting Protein ◽  
Raw264.7 Macrophages ◽  
Pro Inflammatory Cytokines

Background: The role of Src-associated-in-mitosis-of-68-kDa (Sam68) protein in vascular biology has not been studied. A recent report suggests that Sam68 suppresses TNF-a-mediated NF-kB activation in embryonic fibroblasts. Since NF-kB plays a critical role in vascular inflammation and injury, we sought to dissect the molecular mechanism by which Sam68 regulates NF-kB signaling and its functional significance in vascular injury. Methods & Results: The endothelial denudation injury was induced in the carotid arteries of Sam68 -/- mice and WT littermates. Sam68 -/- mice displayed a significantly accelerated re-endothelialization and attenuated neointimal hyperplasia, which was associated with reduced macrophage infiltration and lowered expression of pro-inflammatory cytokines (i.e., TNF-a, MCP-1, IL-1, and IL-6) in the injured vessels. The improved carotid recovery in Sam68 -/- mice was recapitulated in WT mice that had received Sam68 -/- bone-marrow (BM) transplantation, suggesting that Sam68 impedes vascular recovery primarily by its function in BM cells, likely pro-inflammatory cells. In cultured Raw264.7 macrophages, knockdown of Sam68 resulted in a significant reduction in the TNF-a-induced expression of pro-inflammatory cytokines and in the level of P-IKKαβ, P-IkBa (in cytosol) and P-P65 (in nucleus), indicating attenuated NF-kB activation. These results were confirmed in peritoneal and BM-derived macrophages from Sam68 -/- and WT mice. Furthermore, Sam68 co-immunoprecipitation and mass-spectrometric analyses identified cytoskeleton protein Filamin A (FLNA) as a novel Sam68-interacting protein in response to TNF-a stimulation. Reverse co-immunoprecipitation and truncational mutagenesis confirmed that Sam68-FLNA interactions require the N-terminus of Sam68. Finally, analyses in Raw264.7 cells with FLNA knockdown revealed that the effects of Sam68 on TNF-a[[Unsupported Character - Codename ­]]-induced NF-kB signaling and pro-inflammatory cytokine expression were dependent on FLNA. Conclusions: Our data suggest that Sam68 promotes pro-inflammatory response in injured arteries and impedes recovery, and this effect may be attributable, at least partially, to the exaggerated NF-kB activity in macrophages.

Abstract 14810: Sam68 Promotes Nf-kb Signaling and Inflammation and Impedes the Recovery of Arterial Injury

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Shuling Han ◽  
Junlan Zhou ◽  
Gangjian Qin
Keyword(s):  
Inflammatory Cytokines ◽  
Molecular Mechanisms ◽  
Mononuclear Cells ◽  
Vascular Inflammation ◽  
Critical Role ◽  
Inflammatory Cells ◽  
Peritoneal Macrophages ◽  
Post Injury ◽  
Raw264.7 Macrophages ◽  
Tnf Α

Background: The role of Src-associated in mitosis 68 kDa (Sam68) protein in cardiovascular biology has not been studied. A recent report suggests that Sam68 suppresses TNF-α-mediated NF-kB activation. Since NF-kB plays a critical role in vascular inflammation and injury via generation of inflammatory cytokines and recruitment of inflammatory cells, we sought to dissect the molecular mechanism by which Sam68 regulates NF-kB signaling and its functional significance during vascular injury. Methods & Results: The endothelial denudation injury was induced in the carotid arteries of Sam68-null (Sam68 -/- ) and WT mice. Sam68 -/- mice displayed an accelerated re-endothelialization ( P <0.05 at day 5 post-injury) and attenuated neointima formation (by 2.2 folds, P <0.05, at day 14), which was associated with a reduced number of macrophages and lowered expression of pro-inflammatory cytokines (i.e., TNF-alpha, MCP-1 and IL-6) in the injured vessels. In cultured Raw264.7 macrophages, knockdown of Sam68 resulted in a significant reduction in the TNF-α-induced expression of TNF-α, MCP-1, and IL-6 and in the level of nuclear phospho-p65, which indicates attenuated NF-kB activation. These results were confirmed in peritoneal macrophages and macrophages differentiated from bone-marrow mononuclear cells of Sam68 -/- and WT mice. To identify molecular mechanisms, Raw264.7 cells were treated with TNF-α and Vehicle, followed by Sam68 co-immunoprecipitation and mass-spectrometric identification of the Sam68-interacting proteins. We found that TNF-α treatment results in altered interactions of Sam68 with 22 cytosolic, cytoskeletal, and nuclear proteins. Further experiments are under way to validate their involvement in the NF-kB signaling. Conclusions: Our results for the first time suggest that Sam68 promotes pro-inflammatory response in injured arteries and impedes recovery, and this effect may be partially attributable to the exaggerated NF-kB activity.

Abstract 124: Sam68 Impedes the Recovery of Arterial Injury by Augmenting Inflammatory Response

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Shuling Han ◽  
Junlan Zhou ◽  
Baron T Arnone ◽  
Dauren Biyashev ◽  
Chan Boriboun ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Molecular Mechanisms ◽  
Mononuclear Cells ◽  
Vascular Inflammation ◽  
Critical Role ◽  
Inflammatory Cells ◽  
Peritoneal Macrophages ◽  
Raw264.7 Macrophages ◽  
Tnf Α

Background: The role of Src-associated in mitosis 68 kDa (Sam68) in cardiovascular biology has not been studied. A recent report suggests that Sam68 suppresses TNF-α-induced NF-κB activation. Since NF-κB plays a critical role in vascular inflammation and injury via generation of inflammatory cytokines and recruitment of inflammatory cells, we sought to dissect the mechanism by which Sam68 regulates NF-κB signaling and its functional significance during vascular injury. Methods & Results: The endothelial denudation injury was induced in the carotid arteries of Sam68-/- and WT mice. Sam68-/- mice displayed an accelerated re-endothelialization and attenuated neointima hyperplasia, which was associated with a reduced number of macrophages and lowered expression of pro-inflammatory cytokines (i.e., TNF-α, IL-1β and IL-6) in the injured vessels. Importantly, the ameliorated vascular remodeling was recapitulated in WT mice after transplantation of bone marrow (BM) from Sam68-/- mice, suggesting beneficial role was attributed largely to BM-derived inflammatory cells. In cultured Raw264.7 macrophages, knockdown of Sam68 resulted in a significant reduction in the TNF-α-induced expression of TNF-α, IL-1β, and IL-6 and in the level of nuclear phospho-p65, indicating an attenuated NF-κB activation. These results were confirmed in peritoneal macrophages and macrophages differentiated from BM mononuclear cells of Sam68-/- and WT mice. To identify molecular mechanisms, Raw264.7 cells were treated with TNF-α and Vehicle, followed by Sam68 co-immunoprecipitation and mass-spec identification of Sam68-interacting proteins. Specifically, TNF-α treatment results in altered interactions of Sam68 with Filamin A (FLNA), a cytoskeleton protein known to be involved in NF-κB activation. Loss- and gain-of-function of Sam68 and FLNA suggest their mutual dependence in NF-κB activation and pro-inflammatory cytokine expression, and Sam68 is required for TRAF2-FLNA interaction. Conclusions: Our results for the first time suggest that Sam68 promotes pro-inflammatory response in injured arteries and impedes recovery, and this effect is attributed, in part, to the exaggerated NF-κB activity via Sam68-FLNA interaction and consequent TRAF2 stabilization.

scholarly journals Punicalagin Prevents Inflammation in LPS-Induced RAW264.7 Macrophages by Inhibiting FoxO3a/Autophagy Signaling Pathway

Nutrients ◽  
2019 ◽  
Vol 11 (11) ◽  
pp. 2794 ◽  
Author(s):  
Cao ◽  
Chen ◽  
Ren ◽  
Zhang ◽  
Tan ◽  
...  
Keyword(s):  
Western Blot ◽  
Signaling Pathway ◽  
Inflammatory Cytokines ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Inflammatory Responses ◽  
Raw264.7 Macrophages ◽  
Tnf Α ◽  
Autophagy Inhibition ◽  
Pro Inflammatory Cytokines

Punicalagin, a hydrolysable tannin of pomegranate juice, exhibits multiple biological effects, including inhibiting production of pro-inflammatory cytokines in macrophages. Autophagy, an intracellular self-digestion process, has been recently shown to regulate inflammatory responses. In this study, we investigated the anti-inflammatory potential of punicalagin in lipopolysaccharide (LPS) induced RAW264.7 macrophages and uncovered the underlying mechanisms. Punicalagin significantly attenuated, in a concentration-dependent manner, LPS-induced release of NO and decreased pro-inflammatory cytokines TNF-α and IL-6 release at the highest concentration. We found that punicalagin inhibited NF-κB and MAPK activation in LPS-induced RAW264.7 macrophages. Western blot analysis revealed that punicalagin pre-treatment enhanced LC3II, p62 expression, and decreased Beclin1 expression in LPS-induced macrophages. MDC assays were used to determine the autophagic process and the results worked in concert with Western blot analysis. In addition, our observations indicated that LPS-induced releases of NO, TNF-α, and IL-6 were attenuated by treatment with autophagy inhibitor chloroquine, suggesting that autophagy inhibition participated in anti-inflammatory effect. We also found that punicalagin downregulated FoxO3a expression, resulting in autophagy inhibition. Overall these results suggested that punicalagin played an important role in the attenuation of LPS-induced inflammatory responses in RAW264.7 macrophages and that the mechanisms involved downregulation of the FoxO3a/autophagy signaling pathway.

Abstract 15691: The Novel High Mobility Group Protein Hmgxb4 Promotes Neointimal Formation in Response to Arterial Injury

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
xiangqin he ◽  
Kunzhe Dong ◽  
Jian Shen ◽  
Islam Osman ◽  
Guoqing Hu ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Neointimal Hyperplasia ◽  
Critical Role ◽  
High Mobility ◽  
Arterial Injury ◽  
The Novel ◽  
Neointimal Formation ◽  
Vsmc Proliferation ◽  
And Migration

Introduction: Restenosis after percutaneous intervention is predominantly attributed to proliferation and migration of vascular smooth muscle cells (VSMCs). However, the key regulators responsible for VSMC proliferation and migration remain to be identified. Hypothesis: We previously reported that the novel high mobility group (HMG) nuclear protein HMGXB4 (HMG-Box containing 4) plays a critical role in the de-differentiation of vascular smooth muscle cells in vitro and in acute inflammatory response to septic shock. We hypothesize that HMGXB4 is critical for neointimal hyperplasia in response to inflammatory stimuli. Methods and Results: We found that the expression of HMGXB4 is dramatically induced in ligation or wire injury-induced neointimal hyperplasia and correlated with the activation of inflammatory signaling in mice. Using an inducible smooth muscle-specific Hmgxb4 KO (knockout) mice model, we found specific KO of Hmgxb4 in VSMCs ameliorates ligation- or wire- injury induced neointimal formation. Among an array of growth factors and inflammation cytokines, we found that TNFα and INFγ effectively induces the expression of HMGXB4 in VSMCs and correlates with the VSMC proliferation in vitro. Furthermore, we found deletion of HMGXB4 attenuates while over-expression of HMGXB4 promotes inflammation cytokines-induced VSMC proliferation in vitro. These results suggest injury-induced inflammatory signal triggers HMGXB4 induction, which, in turn, promotes the VSMC proliferation and neointimal formation. Conclusions: Our study not only demonstrates a critical role of HMGXB4 in promoting neointimal hyperplasia in response the arterial injury, but also suggests HMGXB4 is a potential novel target for the management of restenosis in human.

Pro-inflammatory cytokines reduce human TAFI expression via tristetraprolin-mediated mRNA destabilisation and decreased binding of HuR

2015 ◽  
Vol 114 (08) ◽  
pp. 337-349 ◽  
Author(s):  
Dragana Komnenov ◽  
Corey Scipione ◽  
Zainab Bazzi ◽  
Justin Garabon ◽  
Marlys Koschinsky ◽  
...  
Keyword(s):  
Inflammatory Cytokines ◽  
Inflammatory Mediators ◽  
Hepg2 Cells ◽  
Inflammatory Cells ◽  
Gene Encoding ◽  
Protein Levels ◽  
Anti Inflammatory ◽  
Mechanistic Basis ◽  
Pro Inflammatory Cytokines

SummaryThrombin activatable fibrinolysis inhibitor (TAFI) is the zymogen form of a basic carboxypeptidase (TAFIa) with both anti-fibrinolytic and anti-inflammatory properties. The role of TAFI in inflammatory disease is multifaceted and involves modulation both of specific inflammatory mediators as well as of the behaviour of inflammatory cells. Moreover, as suggested by in vitro studies, inflammatory mediators are capable of regulating the expression of CPB2, the gene encoding TAFI. In this study we addressed the hypothesis that decreased TAFI levels observed in inflammation are due to post-transcriptional mechanisms. Treatment of human HepG2 cells with pro-inflammatory cytokines TNFα, IL-6 in combination with IL-1β, or with bacterial lipopolysaccharide (LPS) decreased TAFI protein levels by approximately two-fold over 24 to 48 hours of treatment. Conversely, treatment of HepG2 cells with the anti-inflammatory cytokine IL-10 increased TAFI protein levels by two-fold at both time points. We found that the mechanistic basis for this modulation of TAFI levels involves binding of tristetraprolin (TTP) to the CPB2 3′-UTR, which mediates CPB2 mRNA destabilisation. In this report we also identified that HuR, another ARE-binding protein but one that stabilises transcripts, is capable of binding the CBP2 3’UTR. We found that pro-inflammatory mediators reduce the occupancy of HuR on the CPB2 3’-UTR and that the mutation of the TTP binding site in this context abolishes this effect, although TTP and HuR appear to contact discrete binding sites. Interestingly, all of the mediators tested appear to increase TAFI protein expression in THP-1 macrophages, likewise through effects on CPB2 mRNA stability.

scholarly journals Alzheimer's disease and periodontitis - an elusive link

2014 ◽  
Vol 60 (2) ◽  
pp. 173-180 ◽  
Author(s):  
Abhijit N. Gurav
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Inflammatory Cytokines ◽  
Critical Role ◽  
Anaerobic Bacteria ◽  
Systemic Infection ◽  
Low Grade ◽  
Inflammatory Status ◽  
Pro Inflammatory Cytokines

Alzheimer's disease is the preeminent cause and commonest form of dementia. It is clinically characterized by a progressive descent in the cognitive function, which commences with deterioration in memory. The exact etiology and pathophysiologic mechanism of Alzheimer's disease is still not fully understood. However it is hypothesized that, neuroinflammation plays a critical role in the pathogenesis of Alzheimer's disease. Alzheimer's disease is marked by salient inflammatory features, characterized by microglial activation and escalation in the levels of pro-inflammatory cytokines in the affected regions. Studies have suggested a probable role of systemic infection conducing to inflammatory status of the central nervous system. Periodontitis is common oral infection affiliated with gram negative, anaerobic bacteria, capable of orchestrating localized and systemic infections in the subject. Periodontitis is known to elicit a "low grade systemic inflammation" by release of pro-inflammatory cytokines into systemic circulation. This review elucidates the possible role of periodontitis in exacerbating Alzheimer's disease. Periodontitis may bear the potential to affect the onset and progression of Alzheimer's disease. Periodontitis shares the two important features of Alzheimer's disease namely oxidative damage and inflammation, which are exhibited in the brain pathology of Alzheimer's disease. Periodontitis can be treated and hence it is a modifiable risk factor for Alzheimer's disease.

scholarly journals Filamin A Links Sphingosine Kinase 1 and Sphingosine-1-Phosphate Receptor 1 at Lamellipodia To Orchestrate Cell Migration

2008 ◽  
Vol 28 (18) ◽  
pp. 5687-5697 ◽  
Author(s):  
Michael Maceyka ◽  
Sergio E. Alvarez ◽  
Sheldon Milstien ◽  
Sarah Spiegel
Keyword(s):  
Critical Role ◽  
Cell Movement ◽  
Sphingosine Kinase ◽  
Lipid Mediator ◽  
Sphingosine Kinase 1 ◽  
Filamin A ◽  
Interacting Protein ◽  
Surface Receptors ◽  
Sphingosine 1 Phosphate ◽  
Lamellipodia Formation

ABSTRACT Sphingosine kinase 1 (SphK1) catalyzes the phosphorylation of sphingosine to produce the potent lipid mediator sphingosine-1-phosphate (S1P), which plays a critical role in cell motility via its cell surface receptors. Here, we have identified filamin A (FLNa), an actin-cross-linking protein involved in cell movement, as a bona fide SphK1-interacting protein. Heregulin stimulated SphK1 activity only in FLNa-expressing A7 melanoma cells but not in FLNa-deficient cells and induced its translocation and colocalization with FLNa at lamellipodia. SphK1 was required for heregulin-induced migration, lamellipodia formation, activation of PAK1, and subsequent FLNa phosphorylation. S1P directly stimulated PAK1 kinase, suggesting that it may be a target of intracellularly generated S1P. Heregulin also induced colocalization of S1P1 (promotility S1P receptor) but not S1P2, with SphK1 and FLNa at membrane ruffles. Moreover, an S1P1 antagonist inhibited the lamellipodia formation induced by heregulin. Hence, FLNa links SphK1 and S1P1 to locally influence the dynamics of actin cytoskeletal structures by orchestrating the concerted actions of the triumvirate of SphK1, FLNa, and PAK1, each of which requires and/or regulates the actions of the others, at lamellipodia to promote cell movement.

scholarly journals Anti-Inflammatory Activity of Sonchus oleraceus Extract in Lipopoly saccharide-Stimulated RAW264.7 Cells

2018 ◽  
Vol 11 (4) ◽  
pp. 1755-1761
Author(s):  
Eun-Jin Yang ◽  
Sungchan Jang ◽  
Kwang Hee Hyun ◽  
Eun-Young Jung ◽  
Seung-Young Kim ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Inflammatory Cytokines ◽  
Inflammatory Activity ◽  
Raw264.7 Cells ◽  
Supporting Evidence ◽  
Dependent Manner ◽  
Sonchus Oleraceus ◽  
Raw264.7 Macrophages ◽  
Anti Inflammatory ◽  
Pro Inflammatory Cytokines

The anti-inflammatory activity and non-toxicity of Sonchus oleraceus extract (J6) were tested by measuring its effect on the levels of nitric oxide (NO), prostaglandin E2 (PGE2), and the pro-inflammatory cytokines, interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α), in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. We treated the RAW264.7 cells with various concentrations (50, 100, or 200 μg/mL) of J6. Our results showed that J6 inhibited the production of NO, PGE2, and pro-inflammatory cytokines in a concentration-dependent manner, without compromising cell viability. In addition, we provided supporting evidence that the inhibitory activity of J6 on the production of NO and PGE2 occurred via the downregulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively. Our findings suggest that J6 is a new source for anti-inflammatory drugs and ingredients for healthcare products that include functional cosmetics.

Abstract 1123: Critical Role Of Adipose Senescence In Insulin Resistance And Vascular Dysfunction

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Masayuki Orimo ◽  
Tohru Minamino ◽  
Hideyuki Miyauchi ◽  
Kaoru Tateno ◽  
Sho Okada ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Adipose Tissue ◽  
Inflammatory Cytokines ◽  
Cellular Senescence ◽  
Vascular Dysfunction ◽  
Critical Role ◽  
Serum Levels ◽  
P53 Expression ◽  
Deficient Mice ◽  
Pro Inflammatory Cytokines

Cellular senescence is originally described as the finite replicative lifespan of human somatic cells in culture. As a consequence of semi-conservative DNA replication, the extreme terminals of the chromosomes are not duplicated completely, resulting in successive shortening of telomeres with each cell division. Telomerase is a ribonucleoprotein that adds telomeres to the ends of chromosomes. Critically short telomeres are thought to trigger DNA damage response, thereby inducing cellular senescence. Accumulating evidence has suggested that senescent cells promote aging phenotypes or age-related pathologies. Here we show that adipose senescence is critically involved in the regulation of insulin resistance that underlies age-associated cardiovascular disease. The later generation of telomerase-deficient mice with short telomeres exhibited insulin resistance and vascular dysfunction when fed on a high-calorie diet. Adipose tissue of these mice revealed senescence-like phenotypes such as an increase in neutral β galactosidase activity and upregulation of p53 and pro-inflammatory cytokines. Serum levels of pro-inflammatory cytokines were markedly elevated in telomerase-deficient mice and treatment of these mice with a neutralizing antibody against TNF-α significantly improved insulin and glucose intolerance. Removal of senescent adipose tissue reduced serum levels of pro-inflammatory cytokines and thereby improved insulin resistance in telomerase-deficient mice. Conversely, implantation of senescent adipose tissue to wild-type mice impaired insulin sensitivity and glucose tolerance in recipients. Introduction of telomere dysfunction to young adipose tissue markedly upregulated p53 expression and increased the production of pro-inflammatory cytokines. Inhibition of p53 activity significantly improved senescence-like phenotypes of adipose tissue, insulin resistance, and vascular dysfunction in telomerase-deficient mice. These results disclose a novel mechanism of insulin resistance and suggest that adipose senescence is a potential therapeutic target for the treatment of diabetes and diabetic vasculopathy.

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