scholarly journals Endogenous HMGB1 contributes to ischemia-reperfusion-induced myocardial apoptosis by potentiating the effect of TNF-α/JNK

2011 ◽  
Vol 300 (3) ◽  
pp. H913-H921 ◽  
Author(s):  
Hu Xu ◽  
Yongwei Yao ◽  
Zhaoliang Su ◽  
Yunbo Yang ◽  
Raymond Kao ◽  
...  
Keyword(s):  
Ischemia Reperfusion ◽  
Myocardial Inflammation ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Myocardial Apoptosis ◽  
Apoptosis Inhibition ◽  
Tnf Α ◽  
Myocyte Apoptosis ◽  
Jnk Activation

High-mobility group box 1 (HMGB1) is a nuclear protein that has been implicated in the myocardial inflammation and injury induced by ischemia-reperfusion (I/R). The purpose of the present study was to assess the role of HMGB1 in myocardial apoptosis induced by I/R. In vivo, myocardial I/R induced an increase in myocardial HMGB1 expression and apoptosis. Inhibition of HMGB1 (A-box) ameliorated the I/R-induced myocardial apoptosis. In vitro, isolated cardiac myocytes were challenged with anoxia-reoxygenation (A/R; in vitro correlate to I/R). A/R-challenged myocytes also generated HMGB1 and underwent apoptosis. Inhibition of HMGB1 attenuated the A/R-induced myocyte apoptosis. Exogenous HMGB1 had no effect on myocyte apoptosis. However, inhibition of HMGB1 attenuated myocyte TNF-α production after the A/R was challenged; surprisingly, HMGB1 itself did not induce myocyte TNF-α production. Exogenous TNF-α induced a moderate proapoptotic effect on the myocytes, an effect substantially potentiated by coadministration of HMGB1. It is generally accepted that apoptosis induced by TNF-α is regulated by the balance of activation of c-Jun NH2-terminal kinase (JNK) and NF-κB. Indeed, in the present study, TNF-α increased the phosphorylation status of JNK and p65, a subunit of NF-κB; HMGB1 greatly potentiated TNF-α-induced JNK phosphorylation. Furthermore, inhibition of JNK (SP-600125) prevented the myocyte apoptosis induced by a TNF-α/HMGB1 cocktail. Finally, A/R increased HMGB1 production in both wild-type and toll-like receptor 4-deficient myocytes; however, deficiency in toll-like receptor 4 diminished A/R-induced myocyte apoptosis, TNF-α, and JNK activation. Our results indicate that myocyte-derived HMGB1 and TNF-α work in concert to promote I/R-induced myocardial apoptosis through JNK activation.

scholarly journals Micrococcus luteus Teichuronic Acids Activate Human and Murine Monocytic Cells in a CD14- and Toll-Like Receptor 4-Dependent Manner

2001 ◽  
Vol 69 (4) ◽  
pp. 2025-2030 ◽  
Author(s):  
Shuhua Yang ◽  
Shunji Sugawara ◽  
Toshihiko Monodane ◽  
Masahiro Nishijima ◽  
Yoshiyuki Adachi ◽  
...  
Keyword(s):  
Lipid A ◽  
Micrococcus Luteus ◽  
Dependent Manner ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Necrosis Factor Alpha ◽  
Monocytic Cells ◽  
Tnf Α

ABSTRACT Teichuronic acid (TUA), a component of the cell walls of the gram-positive organism Micrococcus luteus (formerlyMicrococcus lysodeikticus), induced inflammatory cytokines in C3H/HeN mice but not in lipopolysaccharide (LPS)-resistant C3H/HeJ mice that have a defect in the Toll-like receptor 4 (TLR4) gene, both in vivo and in vitro, similarly to LPS (T. Monodane, Y. Kawabata, S. Yang, S. Hase, and H. Takada, J. Med. Microbiol. 50:4–12, 2001). In this study, we found that purified TUA (p-TUA) induced tumor necrosis factor alpha (TNF-α) in murine monocytic J774.1 cells but not in mutant LR-9 cells expressing membrane CD14 at a lower level than the parent J774.1 cells. The TNF-α-inducing activity of p-TUA in J774.1 cells was completely inhibited by anti-mouse CD14 monoclonal antibody (MAb). p-TUA also induced interleukin-8 (IL-8) in human monocytic THP-1 cells differentiated to macrophage-like cells expressing CD14. Anti-human CD14 MAb, anti-human TLR4 MAb, and synthetic lipid A precursor IVA, an LPS antagonist, almost completely inhibited the IL-8-inducing ability of p-TUA, as well as LPS, in the differentiated THP-1 cells. Reduced p-TUA did not exhibit any activities in J774.1 or THP-1 cells. These findings strongly suggested that M. luteus TUA activates murine and human monocytic cells in a CD14- and TLR4-dependent manner, similar to LPS.

scholarly journals Toll-like receptor 2 dominance over Toll-like receptor 4 in stressful conditions for its detrimental role in the heart

2017 ◽  
Vol 312 (6) ◽  
pp. H1238-H1247 ◽  
Author(s):  
Ashim K. Bagchi ◽  
Gauri Akolkar ◽  
Soma Mandal ◽  
Prathapan Ayyappan ◽  
Xi Yang ◽  
...  
Keyword(s):  
Transforming Growth Factor ◽  
Heart Function ◽  
Ischemia Reperfusion ◽  
Myeloid Differentiation ◽  
Type I ◽  
Toll Like Receptor ◽  
Tlr4 Signaling ◽  
Tnf Α ◽  
Differentiation Gene

It has been suggested that Toll-like receptor (TLR)4 promotes IL-10-mediated cardiac cell survival, whereas another receptor, TLR2, from the same family, is detrimental. Here, we examined the interactive role of these two innate signaling molecules under stressful conditions, including IL-10 knockout (IL-10−/−) mice, global ischemia-reperfusion (I/R) injury in rat hearts, and in vitro short hairpin RNA experimental models in the presence or absence of IL-10 (10 ng/ml). Circulating and myocardial levels of TNF-α as well as apoptosis and fibrosis were higher in IL-10−/− mice. The increase in TLR2 in IL-10−/− hearts indicated its negative regulation by IL-10. Ex vivo I/R also caused a marked upregulation of TLR2 and TNF-α as well as apoptotic and fibrotic signals. However, a 40-min reperfusion with IL-10 triggered an increase in TLR4 expression and improved recovery of cardiac function. The increase in IL-1 receptor-associated kinase (IRAK)-M and IRAK-2 activity during I/R injury suggested their role in TLR2 signaling. In vitro inhibition of TLR4 activity as a consequence of RNA inhibition-mediated suppression of myeloid differentiation gene (MyD)88 suggested MyD88-dependent activation of TLR4. The inclusion of IL-10 during reperfusion also downregulated the expression of IRAK-2, TNF-α receptor-associated factor 1-interacting protein (TRAIP) and apoptotic signals, caspase-3, and the Bax-to-Bcl-xL ratio. IL-10 reduced the TNF-α receptor-associated increase in TRAIP-induced apoptosis during I/R injury, which led to an increase in IL-1β to mitigate transforming growth factor-β receptor type I-mediated fibrosis. The IL-10 mitigation of these changes suggests that the stimulation through TLR4 signaling promotes IRAK-4 and phosphorylates IRAK-1 instead of IRAK-2 and may be an important therapeutic approach in restoring heart health in stress. NEW & NOTEWORTHY Under stress conditions such as downregulation of the IL-10 gene or ischemia-reperfusion injury, Toll-like receptor (TLR)4 and IL-1 receptor-associated kinase (IRAK)-1 activation is suppressed, along with the upregulation of TLR-2 and IRAK-2, resulting in fibrosis and apoptosis. It is suggested that IL-10 helps to maintain heart function during stress via myeloid differentiation gene 88/IRAK-4/IRAK-1-dependent TLR4 signaling.

scholarly journals Oxidative stress generated by hemorrhagic shock recruits Toll-like receptor 4 to the plasma membrane in macrophages

2006 ◽  
Vol 203 (8) ◽  
pp. 1951-1961 ◽  
Author(s):  
Kinga A. Powers ◽  
Katalin Szászi ◽  
Rachel G. Khadaroo ◽  
Patrick S. Tawadros ◽  
John C. Marshall ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Plasma Membrane ◽  
Hemorrhagic Shock ◽  
Lipid Rafts ◽  
Ischemia Reperfusion ◽  
Resonance Energy ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
H2o2 Treatment

Oxidative stress generated by ischemia/reperfusion is known to prime inflammatory cells for increased responsiveness to subsequent stimuli, such as lipopolysaccharide (LPS). The mechanism(s) underlying this effect remains poorly elucidated. These studies show that alveolar macrophages recovered from rodents subjected to hemorrhagic shock/resuscitation expressed increased surface levels of Toll-like receptor 4 (TLR4), an effect inhibited by adding the antioxidant N-acetylcysteine to the resuscitation fluid. Consistent with a role for oxidative stress in this effect, in vitro H2O2 treatment of RAW 264.7 macrophages similarly caused an increase in surface TLR4. The H2O2-induced increase in surface TLR4 was prevented by depleting intracellular calcium or disrupting the cytoskeleton, suggesting the involvement of receptor exocytosis. Further, fluorescent resonance energy transfer between TLR4 and the raft marker GM1 as well as biochemical analysis of the raft components demonstrated that oxidative stress redistributes TLR4 to lipid rafts in the plasma membrane. Preventing the oxidant-induced movement of TLR4 to lipid rafts using methyl-β-cyclodextrin precluded the increased responsiveness of cells to LPS after H2O2 treatment. Collectively, these studies suggest a novel mechanism whereby oxidative stress might prime the responsiveness of cells of the innate immune system.

scholarly journals Alveolar Macrophages and Toll-like Receptor 4 Mediate Ventilated Lung Ischemia Reperfusion Injury in Mice

2012 ◽  
Vol 117 (4) ◽  
pp. 822-835 ◽  
Author(s):  
Arun Prakash ◽  
Kailin R. Mesa ◽  
Kevin Wilhelmsen ◽  
Fengyun Xu ◽  
Jeffrey M. Dodd-o ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Alveolar Macrophages ◽  
Interleukin 6 ◽  
Ischemia Reperfusion ◽  
Sterile Inflammation ◽  
Interleukin 1Β ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Lung Dysfunction

Background Ischemia-reperfusion (I-R) injury is a sterile inflammatory process that is commonly associated with diverse clinical situations such as hemorrhage followed by resuscitation, transient embolic events, and organ transplantation. I-R injury can induce lung dysfunction whether the I-R occurs in the lung or in a remote organ. Recently, evidence has emerged that receptors and pathways of the innate immune system are involved in recognizing sterile inflammation and overlap considerably with those involved in the recognition of and response to pathogens. Methods The authors used a mouse surgical model of transient unilateral left pulmonary artery occlusion without bronchial involvement to create ventilated lung I-R injury. In addition, they mimicked nutritional I-R injury in vitro by transiently depriving cells of all nutrients. Results Compared with sham-operated mice, mice subjected to ventilated lung I-R injury had up-regulated lung expression of inflammatory mediator messenger RNA for interleukin-1β, interleukin-6, and chemokine (C-X-C motif) ligand-1 and -2, paralleled by histologic evidence of lung neutrophil recruitment and increased plasma concentrations of interleukin-1β, interleukin-6, and high-mobility group protein B1 proteins. This inflammatory response to I-R required toll-like receptor-4 (TLR4). In addition, the authors demonstrated in vitro cooperativity and cross-talk between human macrophages and endothelial cells, resulting in augmented inflammatory responses to I-R. Remarkably, the authors found that selective depletion of alveolar macrophages rendered mice resistant to ventilated lung I-R injury. Conclusions The data reveal that alveolar macrophages and the pattern recognition receptor toll-like receptor-4 are involved in the generation of the early inflammatory response to lung I-R injury.

Abstract 197: Emerging Role of Toll-like Receptor 4 and Heat Shock Protein 60&70 in Ischaemia-Induced Skeletal Muscle Damage In Vitro

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Ali Navi ◽  
Rebekah Yu ◽  
Xu Shi-Wen ◽  
Sidney Shaw ◽  
Daryll Baker ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Inflammatory Cytokine ◽  
Caspase 3 ◽  
Neutralizing Antibody ◽  
Cytokine Release ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Skeletal Muscle Damage ◽  
Tnf Α

OBJECTIVES The innate immune response contributes to the skeletal muscle damage in patients with critical limb ischaemia (CLI); however, the detailed signaling mechanisms are not fully understood. We hypothesized that simulated ischaemia induces inflammatory cytokine release from skeletal myotubes, via a mechanism that involves heat shock protein (HSP) 60&70, known endogenous ligands of Toll-like receptor 4 (TLR4), in vitro. METHODS Human gastrocnemius muscle biopsies were taken from patients with CLI undergoing major lower limb amputation and from patients with no peripheral arterial disease (PAD). Human myoblasts were isolated, cultured to myotubes and then pre-treated with TLR4 neutralizing antibody prior to exposure to simulated ischaemia. Fluorescent immunostaining was carried out to confirm cell differentiation; ELISA analysis were carried out to quantify IL6 and TNF-α release; and Western blot was used to assess expression of HSP60&70, TLR4 and cleaved caspase-3 as a marker of apoptosis. RESULTS Myotubes from patients with CLI expressed greater levels of cleaved caspase-3 and TLR4 as compared to those from patients with no PAD. When exposed to ischaemic conditions, increased IL6 and TNF-α release and upregulation of HSP60&70, cleaved caspase-3 and TLR4 were observed in myotubes from both groups of patients compared to culturing in normoxic conditions (P<0.05). Pre-treatment of myotubes from patients with CLI with TLR4 neutralizing antibody prior to simulated ischaemia was associated with reduced expression of HSP60&70, IL6, TNF-α and cleaved caspase-3 (P<0.05). CONCLUSIONS Increased cytokine release, apoptosis and expression of HSP60&70 and TLR4 occur in ischaemic skeletal muscle in vitro. TLR4 antagonism was associated with reduced apoptosis and inflammatory cytokine release and down-regulation of HSP60&70 expression. This suggests a potential pathway where TLR4 and its endogenous ligands contribute to a positive feedback loop to maintain a proinflammatory environment during ischaemia.

scholarly journals MicroRNA-421 improves ischemia/reperfusion injury via regulation toll-like receptor 4 pathway

2019 ◽  
Vol 48 (3) ◽  
pp. 030006051987186
Author(s):  
Lin-lin Guo ◽  
Ming-lei Guo ◽  
Jian Yao ◽  
Yun-qi Weng ◽  
Xue-zhi Zhang
Keyword(s):  
Reperfusion Injury ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Janus Kinase ◽  
Myocardial Infarct Size ◽  
Serum Concentrations ◽  
Toll Like Receptor ◽  
Model Group ◽  
Toll Like Receptor 4 ◽  
Tnf Α

Objectives The objective was to investigate the effects of microRNA-421 against myocardial ischemia/reperfusion injury in C57BL/6 mice. Methods Male C57BL/6 mice (n = 27) were randomly divided into three groups: normal control (NC) group (sham-treated); I/R model group, which underwent the I30min/R24h model (ischemia for 30 minutes followed by reperfusion for 24 hours); and the miRNA group, which were injected with miR-421. Pathology was assessed by hematoxylin and eosin staining and myocardial infarct size was measured by triphenyltetrazolium chloride staining. The apoptosis rate was measured by TUNEL assay, and relative expression of toll-like receptor-4 (TLR4), Janus kinase 2 (JAK2), and signal transducer and activator of translation 3 (STAT3) was evaluated by immunohistochemistry. Interleukin (IL)-6, tumor necrosis factor (TNF)-α, IL-10, and high mobility group protein B1 (HMGB1) serum concentrations were measured by ELISA. Results Compared with the NC group, in the model group, the myocardial infarction was large; inflammatory cell infiltration was severe; apoptosis was enhanced; expression of TLR4, JAK2, and STAT3 was increased; and serum concentrations of IL-6, TNF-α, IL-10, and HMGB1 were significantly increased. In the miRNA group, the ischemia/reperfusion injury was significantly improved. Conclusions Overexpression of miRNA-421 could reduce ischemia/reperfusion inflammatory response, perhaps via inactivation of TLR4, JAK2, and STAT3.

scholarly journals Toll-Like Receptor 4 Activation Prevents Rat Cardiac Fibroblast Death Induced by Simulated Ischemia/Reperfusion

2021 ◽  
Vol 8 ◽  
Author(s):  
Pablo Parra-Flores ◽  
Jenaro Espitia-Corredor ◽  
Claudio Espinoza-Pérez ◽  
Cristian Queirolo ◽  
Pedro Ayala ◽  
...  
Keyword(s):  
Cell Viability ◽  
Apoptotic Cell ◽  
Ischemia Reperfusion ◽  
Protective Effects ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Trypan Blue Exclusion ◽  
Simulated Ischemia ◽  
Tlr4 Activation

Death of cardiac fibroblasts (CFs) by ischemia/reperfusion (I/R) has major implications for cardiac wound healing. In in vivo models of myocardial infarction, toll-like receptor 4 (TLR4) activation has been reported as a cardioprotector; however, it remains unknown whether TLR4 activation can prevent CF death triggered by simulated I/R (sI/R). In this study, we analyzed TLR4 activation in neonate CFs exposed to an in vitro model of sI/R and explored the participation of the pro-survival kinases Akt and ERK1/2. Simulated ischemia was performed in a free oxygen chamber in an ischemic medium, whereas reperfusion was carried out in normal culture conditions. Cell viability was analyzed by trypan blue exclusion and the MTT assay. Necrotic and apoptotic cell populations were evaluated by flow cytometry. Protein levels of phosphorylated forms of Akt and ERK1/2 were analyzed by Western blot. We showed that sI/R triggers CF death by necrosis and apoptosis. In CFs exposed only to simulated ischemia or only to sI/R, blockade of the TLR4 with TAK-242 further reduced cell viability and the activation of Akt and ERK1/2. Preconditioning with lipopolysaccharide (LPS) or treatment with LPS in ischemia or reperfusion was not protective. However, LPS incubation during both ischemia and reperfusion periods prevented CF viability loss induced by sI/R. Furthermore, LPS treatment reduced the sub-G1 population, but not necrosis of CFs exposed to sI/R. On the other hand, the protective effects exhibited by LPS were abolished when TLR4 was blocked and Akt and ERK1/2 were inhibited. In conclusion, our results suggest that TLR4 activation protects CFs from apoptosis induced by sI/R through the activation of Akt and ERK1/2 signaling pathways.

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