scholarly journals Molecular Dissection of Pro-Fibrotic IL11 Signaling in Cardiac and Pulmonary Fibroblasts

2021 ◽  
Vol 8 ◽  
Author(s):  
Anissa A. Widjaja ◽  
Sivakumar Viswanathan ◽  
Dong Jinrui ◽  
Brijesh K. Singh ◽  
Jessie Tan ◽  
...  
Keyword(s):  
Er Stress ◽  
Signaling Pathways ◽  
Protein Translation ◽  
Null Mouse ◽  
Autocrine Loop ◽  
Erk Activation ◽  
Fibroblast Activation ◽  
Stat3 Phosphorylation ◽  
Dependent Protein ◽  
Dose Dependent

In fibroblasts, TGFβ1 stimulates IL11 upregulation that leads to an autocrine loop of IL11-dependent pro-fibrotic protein translation. The signaling pathways downstream of IL11, which acts via IL6ST, are contentious with both STAT3 and ERK implicated. Here we dissect IL11 signaling in fibroblasts and study IL11-dependent protein synthesis pathways in the context of approved anti-fibrotic drug mechanisms of action. We show that IL11-induced ERK activation drives fibrogenesis and while STAT3 phosphorylation (pSTAT3) is also seen, this appears unrelated to fibroblast activation. Ironically, recombinant human IL11, which has been used extensively in mouse experiments to infer STAT3 activity downstream of IL11, increases pSTAT3 in Il11ra1 null mouse fibroblasts. Unexpectedly, inhibition of STAT3 was found to induce severe proteotoxic ER stress, generalized fibroblast dysfunction and cell death. In contrast, inhibition of ERK prevented fibroblast activation in the absence of ER stress. IL11 stimulated an axis of ERK/mTOR/P70RSK protein translation and its selectivity for Collagen 1 synthesis was ascribed to an EPRS-regulated, ribosome stalling mechanism. Surprisingly, the anti-fibrotic drug nintedanib caused dose-dependent ER stress and lesser pSTAT3 expression. Pirfenidone had no effect on ER stress whereas anti-IL11 specifically inhibited the ERK/mTOR axis while reducing ER stress. These studies define the translation-specific signaling pathways downstream of IL11, intersect immune and metabolic signaling and reveal unappreciated effects of nintedanib.

scholarly journals Molecular dissection of pro-fibrotic signaling identifies the mechanism underlying IL11-driven fibrosis gene translation, reveals non-specific effects of STAT3 and suggests a new mechanism of action for nintedanib

2021 ◽  
Author(s):  
Anissa A Widjaja ◽  
Sivakumar Viswanathan ◽  
Jinrui Dong ◽  
Brijesh K Singh ◽  
Jessie Tan ◽  
...  
Keyword(s):  
Er Stress ◽  
Mechanism Of Action ◽  
Protein Translation ◽  
Null Mouse ◽  
Autocrine Loop ◽  
Erk Activation ◽  
Specific Effects ◽  
Stat3 Phosphorylation ◽  
Dose Dependent ◽  
Pharmacologic Inhibition

In fibroblasts, TGFβ1 stimulates IL11 upregulation that leads to an autocrine loop of IL11-dependent pro-fibrotic protein translation. The signalling pathways downstream of IL11 are contentious and both STAT3 and ERK have been implicated. Here we show that TGFβ1- or IL11- induced ERK activation is consistently associated with fibrogenesis whereas STAT3 phosphorylation (pSTAT3) is unrelated to fibroblast activation. Surprisingly, recombinant human IL11, which has been used extensively in mouse experiments to infer STAT3 activity downstream of IL11, non-specifically increases pSTAT3 in Il11ra1 null mouse fibroblasts. Pharmacologic inhibition of STAT3 prevents TGFβ1-induced fibrogenesis but this effect was found to reflect fibroblast dysfunction due to severe proteotoxic ER stress. In contrast, inhibition of MEK/ERK prevented fibrosis in the absence of ER stress. TGFβ1-stimulated ERK/mTOR/P70RSK-driven protein translation was IL11-dependent and selectivity for pro-fibrotic protein synthesis was ascribed to an EPRS-related mechanism. In TGFβ1-stimulated fibroblasts, the anti-fibrotic drug nintedanib caused dose-dependent ER stress, reduced pSTAT/pERK and inhibited pro-fibrotic protein translation, similarly to generic STAT3 inhibitors or ER stressors. Pirfenidone, while anti-fibrotic, had no effect on ER stress whereas anti-IL11 inhibited the ERK/mTOR axis while reducing ER stress. These studies discount a specific role for STAT3 in pro-fibrotic signaling, suggest a novel mechanism of action for nintedanib and prioritise further the IL11 pathway as a therapeutic target for fibrosis.

scholarly journals Endoplasmic Reticulum Stress/Ca2+-Calmodulin-Dependent Protein Kinase/Signal Transducer and Activator of Transcription 3 Pathway Plays a Role in the Regulation of Cellular Zinc Deficiency in Myocardial Ischemia/Reperfusion Injury

2022 ◽  
Vol 12 ◽  
Author(s):  
Huanhuan Zhao ◽  
Dan Liu ◽  
Qiumei Yan ◽  
Xiyun Bian ◽  
Jing Yu ◽  
...  
Keyword(s):  
Er Stress ◽  
Zinc Deficiency ◽  
Calcium Concentration ◽  
Ischemia Reperfusion ◽  
Zinc Homeostasis ◽  
Dependent Protein Kinase ◽  
Stat3 Phosphorylation ◽  
Dependent Protein ◽  
Myocardial Ischemia Reperfusion ◽  
Cellular Zinc

Zinc homeostasis has been known to play a role in myocardial ischemia/reperfusion (I/R) injury, but the precise molecular mechanisms regulating the expression of ZIP transporters during reperfusion are still unclear. The aim of this study was to determine whether ER Stress/CaMKII/STAT3 pathway plays a role in the regulation of cellular zinc homeostasis. Zinc deficiency increased mRNA and protein expressions of the ER stress relevant markers Chop and Bip, and STAT3 phosphorylation in H9c2 or HL-1 cells, an effect that was abolished by ZnCl2. ER calcium concentration [(Ca2+)ER] was decreased and cytosolic calcium concentration [(Ca2+)I] was increased at the condition of normoxia or ischemia/reperfusion, indicating that zinc deficiency triggers ER stress and Ca2+ leak. Further studies showed that upregulation of STAT3 phosphorylation was reversed by Ca2+ chelator, indicating that intracellular Ca2+ is important for zinc deficiency-induced STAT3 activation. In support, zinc deficiency enhanced ryanodine receptors (RyR), a channel in the ER that mediate Ca2+ release, and Ca2+-calmodulin-dependent protein kinase (CaMKII) phosphorylation, implying that zinc deficiency provoked Ca2+ leak from ER via RyR and p-CaMKII is involved in STAT3 activation. Moreover, inhibition of STAT3 activation blocked zinc deficiency induced ZIP9 expression, and resulted in increased Zn2+ loss in cardiomyocytes, further confirming that STAT3 activation during reperfusion promotes the expression of ZIP9 zinc transporter to correct the imbalance in zinc homeostasis. In addition, suppressed STAT3 activation aggravated reperfusion injury. These data suggest that the ER Stress/CaMKII/STAT3 axis may be an endogenous protective mechanism, which increases the resistance of the heart to I/R.

Apelin-13 protects the heart against ischemia-reperfusion injury through inhibition of ER-dependent apoptotic pathways in a time-dependent fashion

2011 ◽  
Vol 301 (4) ◽  
pp. H1471-H1486 ◽  
Author(s):  
Jianping Tao ◽  
Wei Zhu ◽  
Yapeng Li ◽  
Ping Xin ◽  
Jing Li ◽  
...  
Keyword(s):  
Er Stress ◽  
Signaling Pathways ◽  
Time Course ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Fold Increase ◽  
Erk Activation ◽  
Caspase 12 ◽  
Induced Apoptosis ◽  
Stress Dependent

Endoplasmic reticulum (ER) stress is activated during and contributes to ischemia-reperfusion (I/R) injury. Attenuation of ER stress-induced apoptosis protects the heart against I/R injury. Using apelin, a ligand used to activate the apelin APJ receptor, which is known to be cardioprotective, this study was designed to investigate 1) the time course of changes in I/R injury after ER stress; 2) whether apelin infusion protects the heart against I/R injury via modulation of ER stress-dependent apoptosis signaling pathways; and 3) how phosphatidylinositol 3-kinase (PI3K)/Akt, endothelial nitric oxide synthase (eNOS), AMP-activated protein kinase (AMPK), and ERK activation are involved in the protection offered by apelin treatment. The results showed that, using an in vivo rat I/R model induced by 30 min of ischemia followed by reperfusion, infarct size (IS) increased from 2 h of reperfusion (34.85 ± 2.14%) to 12 h of reperfusion (48.98 ± 3.35, P < 0.05), which was associated with an abrupt increase in ER stress-dependent apoptosis activation, as evidenced by increased CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, and JNK activation (CHOP: 2.49-fold increase, caspase-12: 2.09-fold increase, and JNK: 3.38-fold increase, P < 0.05, respectively). Administration of apelin at 1 μg/kg not only completely abolished the activation of ER stress-induced apoptosis signaling pathways at 2 h of reperfusion but also significantly attenuated time-related changes at 24 h of reperfusion. Using pharmacological inhibition, we also demonstrated that PI3K/Akt, AMPK, and ERK activation were involved in the protection against I/R injury via inhibition of ER stress-dependent apoptosis activation. In contrast, although eNOS activation played a role in decreasing IS at 2 h of reperfusion, it failed to modify either IS or ER stress-induced apoptosis signaling pathways at 24 h after reperfusion.

Role of extracellular signal-regulated kinase (ERK) signaling in nucleotide excision repair and genotoxicity in response to As(III) and Pb(II)

2008 ◽  
Vol 80 (12) ◽  
pp. 2735-2750
Author(s):  
Ju-Pi Li ◽  
Chun-Yu Wang ◽  
Yen-An Tang ◽  
Yun-Wei Lin ◽  
Jia-Ling Yang
Keyword(s):  
Signaling Pathways ◽  
Nucleotide Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Erk Signaling ◽  
Erk Activation ◽  
Extracellular Signal Regulated Kinase ◽  
Extracellular Signal ◽  
Nucleotide Excision

Arsenic and lead can induce genetic injuries and epigenetic signaling pathways in cultured mammalian cells. To test whether signaling pathways affect the extent of genetic injuries, we explored the impacts of extracellular signal-regulated kinase 1 and 2 (ERK) on nucleotide excision repair (NER), cytotoxicity, and genotoxicity following sodium arsenite [As(III)] and lead acetate [Pb(II)]. Sustained ERK activation was observed in human cells exposed to As(III) and Pb(II). As(III) inhibited the cellular NER synthesis capability; conversely, Pb(II) stimulated it. ERK activation contributed to the As(III)-induced NER inhibition and micronucleus formation. In contrast, this signal was required for inducing cellular NER activity and preventing mutagenesis following Pb(II). ERK activation by Pb(II) was dependent on protein kinase C (PKCα) that also exhibited anti-mutagenicity. Enforced expression of ERK signaling markedly elevated the cellular NER activity, which was suppressed by As(III). Nonetheless, ERK activation could counteract the cytotoxicity caused by these two metals. Together, the results indicate that pro-survival ERK signaling exhibits dual and opposing impacts on NER process following As(III) and Pb(II) exposures. The findings also suggest that ERK is an important epigenetic signaling in the determination of metal genotoxicity.

scholarly journals Ca2+ and phospholipid-dependent protein kinase (protein kinase C) activity is not necessarily required for secretion by human neutrophils

Blood ◽  
1986 ◽  
Vol 68 (4) ◽  
pp. 810-817
Author(s):  
KJ Balazovich ◽  
JE Smolen ◽  
LA Boxer
Keyword(s):  
Protein Kinase ◽  
Phorbol Esters ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Soluble Form ◽  
Human Neutrophils ◽  
Dependent Manner ◽  
Dependent Protein Kinase ◽  
Dependent Protein ◽  
Dose Dependent

Ca2+-dependent and phospholipid-dependent protein kinase (PKC) is a receptor for and is activated by phorbol esters. This enzyme is reportedly involved in the mechanism of superoxide anion (O2-) production and the release of intracellular granule contents from human neutrophils. As previously reported by others, we found that greater than 75% of the total cellular PKC activity existed in a soluble form in untreated neutrophils and that this activity was enhanced in a dose- dependent manner by phorbol 12-myristate 13-acetate (PMA) and by phorbol 12,13-dibutyrate (PDBu). Furthermore, mezerein, an analogue of PMA that is thought to be a competitive inhibitor, did not activate PKC, and on the contrary, inhibited PMA-stimulated activity in a dose- dependent manner. Pretreatment of intact neutrophils with PMA or PDBu caused the “translocation” of PKC activity to the insoluble cell fraction; PKC translocation was not detected after mezerein stimulation at any of the tested concentrations. Neither did mezerein cause an increase in intracellular Ca2+, as monitored by Quin 2 fluorescence. Both phorbol esters and mezerein stimulated intact neutrophils to generate O2- and release lysosomal enzymes into the extracellular medium. Finally sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis demonstrated key differences in the patterns of endogenous phosphoproteins of neutrophils stimulated with phorbol as compared with mezerein. We therefore suggest that PKC activation may not be the only pathway required to elicit neutrophil responses.

scholarly journals The Dose-dependent Dual Effects of Corticosterone on Spatial Memory Ability After Traumatic Brain Injury Are Mediated by Two Corticosteroid Receptor Systems

2021 ◽  
Author(s):  
Bin Zhang ◽  
Mengshi Yang ◽  
Qiongyu Yan ◽  
Xiaojian Xu ◽  
Fei Niu ◽  
...  
Keyword(s):  
Traumatic Brain Injury ◽  
Brain Injury ◽  
Spatial Memory ◽  
Signaling Pathways ◽  
Hippocampal Neurons ◽  
High Dose ◽  
Short Term ◽  
Memory Ability ◽  
Neurotoxic Effects ◽  
Dose Dependent

Abstract Background: Our recent studies reported the opposite effects of mineralocorticoid receptor (MR) and glucocorticoid receptor (GR) on neuron survival after traumatic brain injury (TBI). However, as a mixed agonist for MR and GR, whether short term use of high-dose endogenous glucocorticoids exerts neurotoxic effects by excessive activation of GR, what is the set-point, and the possible signaling pathways remain unclear. This study examined the dose-dependent dual effects of corticosterone (CORT) on the spatial memory, the survival of hippocampal neurons and the possible receptor-mediated downstream signaling pathways after TBI.Methods: Based on controlled cortical impact (CCI) and CORT treatments, Sprague-Dawley rats (n=168) were randomly divided into the sham, CCI, CCI + CORT1 (0.3 mg/kg), CCI + CORT2 (3 mg/kg), CCI + CORT3 (30 mg/kg), CCI + CORT1 + spirolactone (spirolactone: 50 mg/kg/d), and CCI + CORT3 + RU486 (RU486: 50 mg/kg/d) groups. Brain tissues were collected on postinjury day 3 and processed for histology and western blot analysis.Results: On postinjury day 3, we tested the learning and memory ability, neuronal apoptosis in the hippocampus, activation levels of MR and GR, Bcl-2 family proteins, and apoptosis-related intracellular signaling pathways. We found that different doses of CORT exhibited dual effects on the survival of hippocampal neurons and the spatial memory. Lower doses of CORT (0.3, 3 mg/kg) significantly increased the activation of MR, upregulated the phosphorylation of Akt/CREB/Bad and the Bcl-2 expression, reduced the number of apoptotic neurons, and subsequently improved the spatial memory. In contrast, higher dose of CORT (30 mg/kg) exerted opposite effect by over activating GR, upregulating the expressions of P53/Bax, and inhibiting the Erk/CREB activities. Conclusion: The results suggest that there is a threshold between the neuroprotective and neurotoxic effects of endogenous GC, higher dose of which, even for short-term use, should also be avoided after TBI.

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