Abstract 148: Mst1 Mediates Cell-protective Mechanisms In The Heart Through Phosphorylation Of FoxO1 And C/EBP-beta

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Yasuhiro Maejima ◽  
Peiyong Zhai ◽  
Mitsuaki Isobe ◽  
Junichi Sadoshima
Keyword(s):  
Cell Survival ◽  
Posttranslational Modifications ◽  
Leucine Zipper ◽  
Ischemia Reperfusion ◽  
Protective Effects ◽  
Area At Risk ◽  
Protective Mechanisms ◽  
Antioxidant Genes ◽  
Induced Apoptosis

Forkhead box O1 (FoxO1) regulates both cell survival and death in the heart. The function of FoxO1 is modulated by posttranslational modifications, including phosphorylation. Mst1 (mammalian sterile 20-like kinase 1), a stress-activated pro-apoptotic kinase, phosphorylates FoxO1 at Ser209/216/231/232. We investigated the molecular mechanism by which FoxO1 regulates both cell survival and death in the heart. Overexpression of FoxO1 significantly suppressed, whereas downregulation of endogenous FoxO1 exacerbated, Mst1 activity and Mst1-induced apoptosis. In the presence of Mst1, FoxO1 downregulated pro-apoptotic genes, including FasL, but upregulated antioxidant genes, including catalase. Chromatin immunoprecipitation assays revealed that Mst1 attenuated FoxO1 binding to the FasL promoter, but it enhanced binding to the catalase promoter. Reporter gene assays showed that C/EBP-β binding elements, but not the FoxO binding ones, in the catalase promoter, are critical for Mst1-mediated upregulation of the catalase gene. FoxO1 interacted with C/EBP-β and their interaction was enhanced in the presence of Mst1. Mass spectrometry analyses revealed that Mst1 phosphorylates C/EBP-β at Thr 299 in its leucine zipper domain. Downregulation of endogenous C/EBP-β reversed the cell-protective effects of FoxO1 expression against Mst1-induced apoptosis. We evaluated the role of Mst1-mediated phosphorylation of FoxO1 and C/EBP-β in mediating survival and death of cardiomyocytes in response to ischemia/reperfusion (I/R). The size of myocardial infarction (MI) in left ventricle (LV) after I/R was significantly greater in cardiac-specific FoxO1 knockout ( c-FoxO1 -/- ) mice than in wild-type mice (MI/area at risk: 59±2, 41±1%, p <0.05, n=6). Injection of a C/EBP-β phospho-mimetic mutant adenovirus into the LV of c-FoxO1 -/- significantly reduced the I/R-induced LV infarct size observed in c-FoxO1 -/- (50±2%, p <0.05, n=6). In summary, Mst1-mediated phosphorylation of FoxO1 inhibits the DNA binding of FoxO1 to the cell death promoting genes whereas it enhances FoxO1-C/EBP-β interaction, Mst1-mediated phosphorylation of C/EBP-β, and C/EBP-β-mediated transcription, which activates cell-protective mechanisms in the heart.

Interleukin 35 protects cardiomyocytes following ischemia/reperfusion-induced apoptosis via activation of mitochondrial STAT3

2021 ◽  
Author(s):  
Fengyun Zhou ◽  
Ting Feng ◽  
Xiangqi Lu ◽  
Huicheng Wang ◽  
Yangping Chen ◽  
...  
Keyword(s):  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Protective Role ◽  
Cytochrome C Release ◽  
Neonatal Cardiomyocytes ◽  
Myocardial Ischemia Reperfusion Injury ◽  
Myocardial Ischemia Reperfusion ◽  
Underlying Mechanisms ◽  
Induced Apoptosis

Abstract Mitochondrial reactive oxygen species (mtROS)-induced apoptosis has been suggested to contribute to myocardial ischemia/reperfusion injury. Interleukin 35 (IL-35), a novel anti-inflammatory cytokine, has been shown to protect the myocardium and inhibit mtROS production. However, its effect on cardiomyocytes upon exposure to hypoxia/reoxygenation (H/R) damage has not yet been elucidated. The present study aimed to investigate the potential protective role and underlying mechanisms of IL-35 in H/R-induced mouse neonatal cardiomyocyte injury. Mouse neonatal cardiomyocytes were challenged to H/R in the presence of IL-35, and we found that IL-35 dose dependently promotes cell viability, diminishes mtROS, maintains mitochondrial membrane potential, and decreases the number of apoptotic cardiomyocytes. Meanwhile, IL-35 remarkably activates mitochondrial STAT3 (mitoSTAT3) signaling, inhibits cytochrome c release, and reduces apoptosis signaling. Furthermore, co-treatment of the cardiomyocytes with the STAT3 inhibitor AG490 abrogates the IL-35-induced cardioprotective effects. Our study identified the protective role of IL-35 in cardiomyocytes following H/R damage and revealed that IL-35 protects cardiomyocytes against mtROS-induced apoptosis through the mitoSTAT3 signaling pathway during H/R.

Ischemic preconditioning in rats: role of mitochondrial KATP channel in preservation of mitochondrial function

2000 ◽  
Vol 278 (1) ◽  
pp. H305-H312 ◽  
Author(s):  
Ryan M. Fryer ◽  
Janis T. Eells ◽  
Anna K. Hsu ◽  
Michele M. Henry ◽  
Garrett J. Gross
Keyword(s):  
Infarct Size ◽  
Ischemic Preconditioning ◽  
Mitochondrial Function ◽  
Ischemia Reperfusion ◽  
Mitochondrial Protein ◽  
Atp Synthesis ◽  
Area At Risk ◽  
Selective Antagonist ◽  
Mitochondrial Katp Channel

We examined the role of the sarcolemmal and mitochondrial KATPchannels in a rat model of ischemic preconditioning (IPC). Infarct size was expressed as a percentage of the area at risk (IS/AAR). IPC significantly reduced infarct size (7 ± 1%) versus control (56 ± 1%). The sarcolemmal KATP channel-selective antagonist HMR-1098 administered before IPC did not significantly attenuate cardioprotection. However, pretreatment with the mitochondrial KATP channel-selective antagonist 5-hydroxydecanoic acid (5-HD) 5 min before IPC partially abolished cardioprotection (40 ± 1%). Diazoxide (10 mg/kg iv) also reduced IS/AAR (36.2 ± 4.8%), but this effect was abolished by 5-HD. As an index of mitochondrial bioenergetic function, the rate of ATP synthesis in the AAR was examined. Untreated animals synthesized ATP at 2.12 ± 0.30 μmol ⋅ min−1 ⋅ mg mitochondrial protein−1. Rats subjected to ischemia-reperfusion synthesized ATP at 0.67 ± 0.06 μmol ⋅ min−1 ⋅ mg mitochondrial protein−1. IPC significantly increased ATP synthesis to 1.86 ± 0.23 μmol ⋅ min−1 ⋅ mg mitochondrial protein−1. However, when 5-HD was administered before IPC, the preservation of ATP synthesis was attenuated (1.18 ± 0.15 μmol ⋅ min−1 ⋅ mg mitochondrial protein−1). These data are consistent with the notion that inhibition of mitochondrial KATPchannels attenuates IPC by reducing IPC-induced protection of mitochondrial function.

scholarly journals Dengue activates mTORC2 signaling to counteract apoptosis and maximize viral replication

2020 ◽  
Author(s):  
Christoph C. Carter ◽  
Jean Paul Olivier ◽  
Alexis Kaushansky ◽  
Fred D. Mast ◽  
John D. Aitchison
Keyword(s):  
Viral Replication ◽  
Cell Survival ◽  
Structural Protein ◽  
Protein Protein Interaction ◽  
Mechanistic Target Of Rapamycin ◽  
Promote Cell Survival ◽  
Important Regulator ◽  
Infected Cells ◽  
Induced Apoptosis

ABSTRACTThe mechanistic target of rapamycin (mTOR) functions in at least two distinct complexes: mTORC1, which regulates cellular anabolic-catabolic homeostasis, and mTORC2, which is an important regulator of cell survival and cytoskeletal maintenance. mTORC1 has been implicated in the pathogenesis of flaviviruses including dengue, where it contributes to the establishment of a pro-viral autophagic state. In contrast, the role of mTORC2 in viral pathogenesis is unknown. In this study, we explore the consequences of a physical protein-protein interaction between dengue non-structural protein 5 (NS5) and host cell mTOR proteins during infection. Using shRNA to differentially target mTORC1 and mTORC2 complexes, we show that mTORC2 is required for optimal dengue replication. Furthermore, we show that mTORC2 is activated during viral replication, and that mTORC2 counteracts virus-induced apoptosis, promoting the survival of infected cells. This work reveals a novel mechanism by which the dengue flavivirus can promote cell survival to maximize viral replication.

scholarly journals Ticagrelor Conditioning Effects Are Not Additive to Cardioprotection Induced by Direct NLRP3 Inflammasome Inhibition: Role of RISK, NLRP3, and Redox Cascades

2020 ◽  
Vol 2020 ◽  
pp. 1-12
Author(s):  
Claudia Penna ◽  
Manuela Aragno ◽  
Alessia Sofia Cento ◽  
Saveria Femminò ◽  
Isabella Russo ◽  
...  
Keyword(s):  
Nlrp3 Inflammasome ◽  
Ex Vivo ◽  
Isolated Heart ◽  
Ischemia Reperfusion ◽  
Oral Treatment ◽  
P2y12 Receptor ◽  
Protective Effects ◽  
Area At Risk ◽  
Beneficial Effects ◽  
Pyrin Domain

Inhibition of either P2Y12 receptor or the nucleotide-binding oligomerization domain- (NOD-) like receptor pyrin domain containing 3 (NLRP3) inflammasome provides cardioprotective effects. Here, we investigate whether direct NLRP3 inflammasome inhibition exerts additive effects on myocardial protection induced by the P2Y12 receptor antagonist Ticagrelor. Ticagrelor (150 mg/kg) was orally administered to rats for three consecutive days. Then, isolated hearts underwent an ischemia/reperfusion (30 min ischemia/60 min reperfusion; IR) protocol. The selective NLRP3 inflammasome inhibitor INF (50 μM) was infused before the IR protocol to the hearts from untreated animals or pretreated with Ticagrelor. In parallel experiments, the hearts isolated from untreated animals were perfused with Ticagrelor (3.70 μM) before ischemia and subjected to IR. The hearts of animals pretreated with Ticagrelor showed a significantly reduced infarct size (IS, 49±3% of area at risk, AAR) when compared to control IR group (69±2% of AAR). Similarly, ex vivo administration of INF before the IR injury resulted in significant IS reduction (38±3% of AAR). Myocardial IR induced the NLRP3 inflammasome complex formation, which was attenuated by either INF pretreatment ex vivo, or by repeated oral treatment with Ticagrelor. The beneficial effects induced by either treatment were associated with the protective Reperfusion Injury Salvage Kinase (RISK) pathway activation and redox defence upregulation. In contrast, no protective effects nor NLRP3/RISK modulation were recorded when Ticagrelor was administered before ischemia in isolated heart, indicating that Ticagrelor direct target is not in the myocardium. Our results confirm that Ticagrelor conditioning effects are likely mediated through platelets, but are not additives to the ones achieved by directly inhibiting NLRP3.

Protective effects of 5-(N,N-dimethyl)amiloride on ischemia-reperfusion injury in hearts

1990 ◽  
Vol 258 (5) ◽  
pp. H1615-H1619 ◽  
Author(s):  
H. P. Meng ◽  
G. N. Pierce
Keyword(s):  
Reperfusion Injury ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Significant Rise ◽  
Similar Degree ◽  
Protective Effects ◽  
Tension Development ◽  
Right Ventricular Wall ◽  
Dimethyl Amiloride

An Na(+)-H+ exchange inhibitor, 5-(N,N-dimethyl)amiloride (DMA), was used to probe the possible role of Na(+)-H+ exchange in ischemia-reperfusion injury in coronary perfused isolated rat right ventricular wall. In DMA-untreated hearts, 60 min of ischemia resulted in a significant rise in testing tension (RT: 174 +/- 8% of preischemic level). Thirty minutes of reperfusion further increased RT (273 +/- 12%) and induced a poor recovery in developed tension (DT: 28 +/- 4%). Both the rate of tension development and relaxation (+dT/dt and -dT/dt) recovered to a similar degree. When 1, 5, or 20 microM DMA was included in the perfusate (3 min before ischemia and in the first 3 min of reperfusion), the maximal postischemic RT of the heart was reduced to 204 +/- 21, 166 +/- 15, and 139 +/- 45% of the preischemic levels (P less than 0.05), respectively, and DT was 39 +/- 3, 63 +/- 10, and 79 +/- 8% of the preischemic levels (P less than 0.05), respectively. Similar qualitative recovery of +/- dT/dt was observed. Recovery was similar if DMA was present only during reperfusion. DMA treatment also significantly protected against creatine phosphokinase release during reperfusion. The results demonstrate that DMA can significantly protect the heart during the initial stages of reperfusion. The data suggest that Na(+)-H+ exchange may play an important role in the development of cardiac dysfunction and damage during the first minutes of reperfusion.

scholarly journals Ischemic Postconditioning as a Novel Avenue to Protect against Brain Injury after Stroke

2009 ◽  
Vol 29 (5) ◽  
pp. 873-885 ◽  
Author(s):  
Heng Zhao
Keyword(s):  
Protein Kinase ◽  
Reperfusion Injury ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Ischemic Postconditioning ◽  
Mitogen Activated Protein Kinase ◽  
Protective Effects ◽  
Protective Mechanisms ◽  
Mitogen Activated Protein ◽  
Cerebral Ischemic

Ischemic postconditioning initially referred to a stuttering reperfusion performed immediately after reperfusion, for preventing ischemia/reperfusion injury in both myocardial and cerebral infarction. It has evolved into a concept that can be induced by a broad range of stimuli or triggers, and may even be performed as late as 6 h after focal ischemia and 2 days after transient global ischemia. The concept is thought to be derived from ischemic preconditioning or partial/gradual reperfusion, but in fact the first experiment for postconditioning was carried out much earlier than that of preconditioning or partial/gradual reperfusion, in the research on myocardial ischemia. This review first examines the protective effects and parameters of postconditioning in various cerebral ischemic models. Thereafter, it provides insights into the protective mechanisms of postconditioning associated with reperfusion injury and the Akt, mitogen-activated protein kinase (MAPK), protein kinase C (PKC), and ATP-sensitive K+ (KATP) channel cell signaling pathways. Finally, some open issues and future challenges regarding clinical translation of postconditioning are discussed.

The Regulation of STAT3 and Its Role in the Adhesion and Migration of Chronic Lymphocytic Leukaemia Cells

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 4347-4347
Author(s):  
Sarah Brophy ◽  
Fiona M Quinn ◽  
David O'Brien ◽  
Paul Browne ◽  
Elisabeth A. Vandenberghe ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Cell Adhesion ◽  
Cell Survival ◽  
Lymphocytic Leukaemia ◽  
Serine Phosphorylation ◽  
Pre Treatment ◽  
And Migration ◽  
Induced Apoptosis ◽  
Stat3 Inhibition

Abstract The bone marrow and lymph node microenvironments are important in promoting cell proliferation, survival and protection from drug induced apoptosis in chronic lymphocytic leukaemia (CLL). Chemokine networks, such as the CXCR4/CXCL12 axis, in combination with selectins, such as CD62L and integrins allow the migration of CLL cells to these protective niches. The B-cell receptor (BCR) signalling pathway is the most important pathway involved in micro-environmental crosstalk and CLL cell survival. Further, it has been shown to interact with the signal transducer and activator of transcription 3 (STAT3) signalling pathway. The role of the STAT3 in CLL pathogenesis is unclear; however, it is constitutively phosphorylated on serine residue 727 (serine pSTAT3) in CLL cells. Here, we investigate the role of STAT3 in CLL cell survival and migration, using pharmacological inhibition and siRNA knockdown. Phospho-tyrosine and phospho-serine STAT3 were assessed by flow cytometry and western blotting. Apoptosis was assessed by Annexin V/Propidium Iodide staining by flow cytometry. The expression of cell surface markers involved in cell adhesion and homing was determined by multicolour flow cytometry. Stimulation of the BCR using immunoglobulin F(ab´)2 fragments induced tyrosine phosphorylation of STAT3 in CLL cells with unmutated immunoglobulin (IgVH) genes (n=7) but not mutated IgVH genes (n=5). This induced tyrosine phosphorylation was abrogated by pre-treatment with the Janus kinase (JAK) inhibitor Ruxolitinib and the BCR inhibitors Ibrutinib and Idelalisib (p<0.05, n=5). Gene expression studies using Taqman Assays showed BCR stimulation resulted in an upregulation of STAT3 regulated genes in CLL cells with unmutated IgVH genes. Interestingly, stimulation of BCR resulted in a significant increase in CD62L expression, which was inhibited by pre-treatment with Ibrutinib and Ruxolitinib (p<0.05, n=5). STAT3 inhibition was shown to have a divergent effect on CLL cell survival: In patient samples with >70% positive serine pSTAT3 cells, the STAT3 inhibitor cucurbitacin I induced apoptosis with a concurrent downregulation in serine phosphorylation (n=3); while in patient samples with <70% positive serine pSTAT3 cells, treatment with cucurbitacin I resulted in a decrease in apoptosis and a concurrent increase in serine phosphorylation (n=3). The STAT3 inhibitor S3I-201 had a similar effect but the upstream JAK inhibitor Ruxolitinib had no effect on serine phosphorylation and no effect on the apoptosis of CLL cells. In addition, siRNA mediated STAT3 knockdown and treatment with cucurbitacin I and S31-201, resulted in a significant decrease in CD62L positive CLL cells (p<0.0001, n=29). The role of STAT3 in CLL cell adhesion under shear flow conditions was investigated using a microfluidics system including a neMESYS Low Pressure syringe pump system and Human Umbilical Vein Endothelial Cells (HUVEC) coated biochips. Treatment of CLL cells with cucurbitacin I resulted in a significant decrease in adhesion to endothelial cells (p<0.001, n=4). The effect of STAT3 inhibition on the chemotaxis of CLL cells was investigated using Neuroprobe 96-well ChemoTx plates. Treatment with cucurbitacin I resulted in a significant decrease in CLL cells migrated in response to the chemokine CXCL12 compared to control (p=0.0001, n=8). In addition, treatment of CLL cells with CXCL12 resulted in an increase in serine pSTAT3 that was downregulated by pretreatment with cucurbitacin I. This study has shown: 1. Activation of STAT3 by BCR stimulation occurs in poor prognostic unmutated IgVH genes 2. Serine pSTAT3 has a role in cell survival in response to STAT3 inhibition 3. A role for STAT3 in CLL cell adhesion and migration, in particular in the regulation of the expression of CD62L. In conclusion, this study shows a role for the STAT3 pathway in cell survival and CLL cell-microenvironment crosstalk, suggesting therapeutic potential by interfering with the migration and homing of CLL cells to the lymph node and bone marrow microenvironments. Disclosures No relevant conflicts of interest to declare.

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