scholarly journals WNT3a and WNT5a Transported by Exosomes Activate WNT Signaling Pathways in Human Cardiac Fibroblasts

2019 ◽  
Vol 20 (6) ◽  
pp. 1436 ◽  
Author(s):  
Edyta Działo ◽  
Michał Rudnik ◽  
Roman Koning ◽  
Marcin Czepiel ◽  
Karolina Tkacz ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathways ◽  
Cardiac Fibrosis ◽  
Nuclear Translocation ◽  
Glycogen Synthase ◽  
Cardiac Fibroblasts ◽  
Interacting Protein ◽  
L Cells ◽  
Wnt Proteins ◽  
Human Cardiac Fibroblasts

WNT signaling plays an important role in fibrotic processes in the heart. Recently, exosomes have been proposed as novel extracellular transporters for WNT proteins. In this study, we analyzed whether WNT3a and WNT5a carried by exosomes could activate downstream molecular pathways in human cardiac fibroblasts. Exosomes were isolated from conditioned medium of control, WNT3a- and WNT5a-producing L cells by differential ultracentrifugations. Obtained exosomes showed size ranging between 20–150 nm and expressed exosomal markers ALG-2-interacting protein X (ALIX) and CD63. Treatment with WNT3a-rich exosomes inhibited activity of glycogen synthase kinase 3β (GSK3β), induced nuclear translocation of β-catenin, and activated T-cell factor (TCF)/lymphoid enhancer factor (LEF) transcription factors as well as expression of WNT/β-catenin responsive genes in cardiac fibroblasts, but did not coactivate extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and activator protein 1 (AP-1) signaling pathways. In contrast, exosomes produced by WNT5a-producing L cells failed to activate β-catenin-dependent response, but successfully triggered phosphorylation of ERK1/2 and JNK and stimulated IL-6 production. In conclusion, exosomes containing WNT proteins can functionally contribute to cardiac fibrosis by activating profibrotic WNT pathways on cardiac fibroblasts and may represent a novel mechanism of spreading profibrotic signals in the heart.

scholarly journals Metformin enhances glucagon-like peptide 1 via cooperation between insulin and Wnt signaling

2013 ◽  
Vol 220 (2) ◽  
pp. 117-128 ◽  
Author(s):  
Mi-Hyun Kim ◽  
Jae-Hwan Jee ◽  
Sunyoung Park ◽  
Myung-Shik Lee ◽  
Kwang-Won Kim ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Cross Talk ◽  
Insulin Signaling ◽  
Nuclear Translocation ◽  
Glycogen Synthase ◽  
Oral Glucose ◽  
High Blood Glucose ◽  
Insulin Sensitizer ◽  
L Cells ◽  
Glucagon Like Peptide

One aspect of the effects of metformin on glucagon-like peptide (GLP)-1 might be associated with the mechanism by which the cross talk between insulin and Wnt signaling enhances GLP1 secretion, due to the action of metformin as an insulin sensitizer. However, this remains completely unknown. In this study, we have investigated the mechanisms of the action of metformin on cross talk between insulin and Wnt signaling. GLP1 enhancement by meformin was determined in human NCI-H716 intestinal L-cells and hyperglycemic db/db mice treated with metformin (0.25 and 0.5 mM and/or 12.5 mg/kg body weight) for 24 h and 2 months. Metformin increased GLP1 secretion in L-cells and db/db mice. Metformin stimulated the nuclear translocation of β-catenin and TOPflash reporter activity, and gene depletion of β-catenin or enhancement of mutation of transcription factor 7-like 2 binding site offset GLP1. In addition, insulin receptor substrate 2 gene depletion blocked metformin-enhanced β-catenin translocation. These effects were preceded by an increase in glucose utilization and calcium influx, the activation of calcium-dependent protein kinase, and, in turn, the activation of insulin signaling, and the inhibition of glycogen synthase kinase 3β, a potent inhibitor of β-catenin. Furthermore, high blood glucose levels were controlled via GLP1 receptor-dependent insulinotropic pathways in db/db mice, which were evidenced by the increase in GLP1 and insulin levels at 30 min after oral glucose loading and pancreatic insulinotropic gene expression. Our findings indicate that the cooperation between Wnt and its upstream insulin signaling pathways might be a novel and important mechanism underlying the effects of metformin on GLP1 production.

Abstract 121: Interleukin 11 Mediates Wnt/β-catenin-dependent Fibrotic Response Of Human Cardiac Fibroblasts

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Edyta Dzialo ◽  
Marcin Czepiel ◽  
Maciej Siedlar ◽  
Gabriela Kania ◽  
Przemyslaw Blyszczuk
Keyword(s):  
Smooth Muscle ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Smooth Muscle Actin ◽  
Neutralizing Antibody ◽  
Cell Culture Supernatant ◽  
Muscle Actin ◽  
Alpha Smooth Muscle Actin ◽  
Wnt Proteins ◽  
Human Cardiac Fibroblasts

Wnt proteins family represents secreted glycoproteins implicated in the number of fibrotic cardiac pathologies. The transcriptional activity of Wnts is broad and involves β-catenin-dependent or β-catenin-independent responses. In this study, we examined the effect of exogenous Wnt3a (β-catenin-dependent) and Wnt5a (β-catenin-independent) in TGF-β-activated human cardiac fibroblasts. Furthermore, we assessed the hypothesis that Wnt3a could regulate IL-11 production and analyzed its contribution to profibrotic response in cardiac fibroblasts.By employing a full genome transcriptomics, we analyzed transformation of human cardiac fibroblasts induced by TGF-β in the presence of Wnt3a or Wnt5a produced by cell culture supernatant of L-Wnt3a, L-Wnt5a or control L-cells. Stimulation with Wnt3a of TGF-β-activated fibroblasts resulted in induction of 66 genes, specifically involved in myofibroblast differentiation including ACTA2 (encoding alpha smooth muscle actin; αSMA) ACTG2 (encoding gamma smooth muscle actin; γSMA) and VCL (encoding vinculin). In contrast to Wnt3a, treatment with Wnt5a upregulated expression of only 2 genes in TGF-β-activated cells. Additionally, in the presence of TGF-β, Wnt3a enhanced phosphorylation of TAK1 and production and secretion of IL-11. Importantly, in the absence of TGF-β, Wnt3a did not promote fibroblast-to-myofibroblast transition, TAK1 phosphorylation and IL-11 production. To determine, if Wnt3a-dependent production of IL-11 could contribute to profibrotic response we blocked IL-11 activity with anti-IL-11 neutralizing antibody in cardiac fibroblasts activated with TGF-β and Wnt3a. We found that neutralizing anti-IL11 antibody effectively suppressed production of αSMA, γSMA, fibronectin and pro-collagen I alpha 1, both on mRNA and protein levels. In line with these findings, blockade of IL-11 suppressed contractile properties of TGF-β/Wnt3a-activated cardiac fibroblasts. In conclusion, Wnt3a and Wnt5a differentially regulate gene expression of TGF-β-activated cardiac. Activation of the Wnt/β-catenin pathway promotes fibroblast-to-myofibroblast transition by enhancing production of profibrotic IL-11. It seems that identifying the profibrotic Wnt/β-catenin-IL11 mechanism in cardiac fibroblasts might represent a promising strategy in development of new therapies against cardiac fibrosis.

scholarly journals Ykt6 membrane-to-cytosol cycling regulates exosomal Wnt secretion

2018 ◽  
Author(s):  
Karen Linnemannstöns ◽  
Pradhipa Karuna M ◽  
Leonie Witte ◽  
Jeanette Clarissa Kittel ◽  
Adi Danieli ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Wnt Signaling ◽  
Signaling Pathways ◽  
Long Range ◽  
Protein Trafficking ◽  
Secretory Pathway ◽  
Multivesicular Bodies ◽  
Phosphorylation Sites ◽  
Wnt Proteins ◽  
C Terminus

Protein trafficking in the secretory pathway, for example the secretion of Wnt proteins, requires tight regulation. These ligands activate Wnt signaling pathways and are crucially involved in development and disease. Wnt is transported to the plasma membrane by its cargo receptor Evi, where Wnt/Evi complexes are endocytosed and sorted onto exosomes for long-range secretion. However, the trafficking steps within the endosomal compartment are not fully understood. The promiscuous SNARE Ykt6 folds into an auto-inhibiting conformation in the cytosol, but a portion associates with membranes by its farnesylated and palmitoylated C-terminus. Here, we demonstrate that membrane detachment of Ykt6 is essential for exosomal Wnt secretion. We identified conserved phosphorylation sites within the SNARE domain of Ykt6, which block Ykt6 cycling from the membrane to the cytosol. In Drosophila, Ykt6-RNAi mediated block of Wg secretion is rescued by wildtype but not phosphomimicking Ykt6. The latter accumulates at membranes, while wildtype Ykt6 regulates Wnt trafficking between the plasma membrane and multivesicular bodies. Taken together, we show that a regulatory switch in Ykt6 fine-tunes sorting of Wnts in endosomes.

scholarly journals Bone Morphogenetic Protein 9 Reduces Cardiac Fibrosis and Improves Cardiac Function in Heart Failure

Circulation ◽  
2018 ◽  
Vol 138 (5) ◽  
pp. 513-526 ◽  
Author(s):  
Kevin J. Morine ◽  
Xiaoying Qiao ◽  
Sam York ◽  
Peter S. Natov ◽  
Vikram Paruchuri ◽  
...  
Keyword(s):  
Heart Failure ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Human Subjects ◽  
Lv Function ◽  
Transverse Aortic Constriction ◽  
Aortic Constriction ◽  
Bone Morphogenetic Protein 9 ◽  
Human Cardiac Fibroblasts ◽  
Tgf Β1

Background: Heart failure is a growing cause of morbidity and mortality worldwide. Transforming growth factor beta (TGF-β1) promotes cardiac fibrosis, but also activates counterregulatory pathways that serve to regulate TGF-β1 activity in heart failure. Bone morphogenetic protein 9 (BMP9) is a member of the TGFβ family of cytokines and signals via the downstream effector protein Smad1. Endoglin is a TGFβ coreceptor that promotes TGF-β1 signaling via Smad3 and binds BMP9 with high affinity. We hypothesized that BMP9 limits cardiac fibrosis by activating Smad1 and attenuating Smad3, and, furthermore, that neutralizing endoglin activity promotes BMP9 activity. Methods: We examined BMP9 expression and signaling in human cardiac fibroblasts and human subjects with heart failure. We used the transverse aortic constriction–induced model of heart failure to evaluate the functional effect of BMP9 signaling on cardiac remodeling. Results: BMP9 expression is increased in the circulation and left ventricle (LV) of human subjects with heart failure and is expressed by cardiac fibroblasts. Next, we observed that BMP9 attenuates type I collagen synthesis in human cardiac fibroblasts using recombinant human BMP9 and a small interfering RNA approach. In BMP9 –/– mice subjected to transverse aortic constriction, loss of BMP9 activity promotes cardiac fibrosis, impairs LV function, and increases LV levels of phosphorylated Smad3 (pSmad3), not pSmad1. In contrast, treatment of wild-type mice subjected to transverse aortic constriction with recombinant BMP9 limits progression of cardiac fibrosis, improves LV function, enhances myocardial capillary density, and increases LV levels of pSmad1, not pSmad3 in comparison with vehicle-treated controls. Because endoglin binds BMP9 with high affinity, we explored the effect of reduced endoglin activity on BMP9 activity. Neutralizing endoglin activity in human cardiac fibroblasts or in wild-type mice subjected to transverse aortic constriction–induced heart failure limits collagen production, increases BMP9 protein levels, and increases levels of pSmad1, not pSmad3. Conclusions: Our results identify a novel functional role for BMP9 as an endogenous inhibitor of cardiac fibrosis attributable to LV pressure overload and further show that treatment with either recombinant BMP9 or disruption of endoglin activity promotes BMP9 activity and limits cardiac fibrosis in heart failure, thereby providing potentially novel therapeutic approaches for patients with heart failure.

Abstract 18942: Inhibition of β-arrestin 1 Prevents post-MI Maladaptive Ventricular Remodeling

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Md Abdur Razzaque ◽  
Xianyao Xu ◽  
Abbasali Badami ◽  
Peter Klepacz ◽  
Mei Han ◽  
...  
Keyword(s):  
Cardiac Fibrosis ◽  
Ventricular Remodeling ◽  
Cardiac Fibroblasts ◽  
Male Rats ◽  
Lv Function ◽  
Adenoviral Delivery ◽  
Potential Therapeutic Role ◽  
Targeted Inhibition ◽  
Human Cardiac Fibroblasts

Remote (non-infarct) territory fibrosis is a significant cause of post-myocardial infarction (MI) heart failure (HF). We have previously shown that increased activity of β-arrestin 1 in adult human cardiac fibroblasts (CF) isolated from failing hearts is an important mechanism of cardiac fibrosis. This study investigates the potential therapeutic role of β-arrestin 1 inhibition on CF biology in vivo. Adult male rats underwent LAD ligation to induce post-MI HF. β-arrestin 1 was inhibited by intra-coronary adenoviral-mediated delivery of a β-arrestin 1 inhibitor (Ad-Barr1ct) immediately following LAD ligation (n=11). Ad-Barr1ct contains a rat β-arrestin 1 C-terminal fragment (aa. 369-418). Control rats received a null adenovirus (n=10). Animals were studied prior to and up to 8 weeks (wks) post-MI and adenoviral delivery. There was a significant decline in LV function at 8 wks post-MI in Ad-null rats vs. pre-MI. Remote territory (non-infarct area) fibrosis increased by 8 wks post-MI consistent with adverse remodeling. Intra-coronary delivery of Ad-Barr1ct following LAD ligation significantly inhibited post-MI LV dysfunction vs. Ad-Null as measured by improved fractional shortening and ejection fraction. Ad-Barr1ct also decreased peri-infarct and remote territory fibrosis. Consistent with these findings, Ad-arr1ct resulted in decreased α-SMA, collagen I, collegen III and fibronectin expression in CF isolated 8 wks post-MI vs. Ad-Null providing evidence of decreased post-MI CF activation and myofibroblast transformation with Ad-Barr1ct. Targeted inhibition of β-arrestin 1 in the heart may represent a novel therapeutic approach to prevent pathological fibrosis and maladaptive remodeling post-MI.

scholarly journals Soluble St2 Induces Cardiac Fibroblast Activation and Collagen Synthesis via Neuropilin-1

Cells ◽  
2020 ◽  
Vol 9 (7) ◽  
pp. 1667 ◽  
Author(s):  
Lara Matilla ◽  
Vanessa Arrieta ◽  
Eva Jover ◽  
Amaia Garcia-Peña ◽  
Ernesto Martinez-Martinez ◽  
...  
Keyword(s):  
Collagen Synthesis ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Interleukin 1 ◽  
Cardiac Fibroblast ◽  
Pathogenic Role ◽  
Fibroblast Activation ◽  
Neuropilin 1 ◽  
Human Cardiac Fibroblasts

Circulating levels of soluble interleukin 1 receptor-like 1 (sST2) are increased in heart failure and associated with poor outcome, likely because of the activation of inflammation and fibrosis. We investigated the pathogenic role of sST2 as an inductor of cardiac fibroblasts activation and collagen synthesis. The effects of sST2 on human cardiac fibroblasts was assessed using proteomics and immunodetection approaches to evidence the upregulation of neuropilin-1 (NRP-1), a regulator of the profibrotic transforming growth factor (TGF)-β1. In parallel, sST2 increased fibroblast activation, collagen and fibrosis mediators. Pharmacological inhibition of nuclear factor-kappa B (NF-κB) restored NRP-1 levels and blocked profibrotic effects induced by sST2. In NRP-1 knockdown cells, sST2 failed to induce fibroblast activation and collagen synthesis. Exogenous NRP-1 enhanced cardiac fibroblast activation and collagen synthesis via NF-κB. In a pressure overload rat model, sST2 was elevated in association with cardiac fibrosis and was positively correlated with NRP-1 expression. Our study shows that sST2 induces human cardiac fibroblasts activation, as well as the synthesis of collagen and profibrotic molecules. These effects are mediated by NRP-1. The blockade of NF-κB restored NRP-1 expression, improving the profibrotic status induced by sST2. These results show a new pathogenic role for sST2 and its mediator, NRP-1, as cardiac fibroblast activators contributing to cardiac fibrosis.

scholarly journals TGF-β and WNT signaling pathways in cardiac fibrosis: non-coding RNAs come into focus

2020 ◽  
Vol 18 (1) ◽  
Author(s):  
Fatemeh Yousefi ◽  
Zahra Shabaninejad ◽  
Sina Vakili ◽  
Maryam Derakhshan ◽  
Ahmad Movahedpour ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathways ◽  
Cardiac Fibrosis ◽  
Non Coding Rnas

scholarly journals Activation of the Transcription Factor GLI1 by WNT Signaling Underlies the Role of SULFATASE 2 as a Regulator of Tissue Regeneration

2013 ◽  
Vol 288 (29) ◽  
pp. 21389-21398 ◽  
Author(s):  
Ikuo Nakamura ◽  
Maite G. Fernandez-Barrena ◽  
Maria C. Ortiz-Ruiz ◽  
Luciana L. Almada ◽  
Chunling Hu ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Wnt Signaling ◽  
Cyclin D1 ◽  
Signaling Pathways ◽  
Tissue Regeneration ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Hepatocyte Proliferation ◽  
Dependent Manner ◽  
Gli1 Expression

Tissue regeneration requires the activation of a set of specific growth signaling pathways. The identity of these cascades and their biological roles are known; however, the molecular mechanisms regulating the interplay between these pathways remain poorly understood. Here, we define a new role for SULFATASE 2 (SULF2) in regulating tissue regeneration and define the WNT-GLI1 axis as a novel downstream effector for this sulfatase in a liver model of tissue regeneration. SULF2 is a heparan sulfate 6-O-endosulfatase, which releases growth factors from extracellular storage sites turning active multiple signaling pathways. We demonstrate that SULF2-KO mice display delayed regeneration after partial hepatectomy (PH). Mechanistic analysis of the SULF2-KO phenotype showed a decrease in WNT signaling pathway activity in vivo. In isolated hepatocytes, SULF2 deficiency blocked WNT-induced β-CATENIN nuclear translocation, TCF activation, and proliferation. Furthermore, we identified the transcription factor GLI1 as a novel target of the SULF2-WNT cascade. WNT induces GLI1 expression in a SULF2- and β-CATENIN-dependent manner. GLI1-KO mice phenocopied the SULF2-KO, showing delayed regeneration and decreased hepatocyte proliferation. Moreover, we identified CYCLIN D1, a key mediator of cell growth during tissue regeneration, as a GLI1 transcriptional target. GLI1 binds to the cyclin d1 promoter and regulates its activity and expression. Finally, restoring GLI1 expression in the liver of SULF2-KO mice after PH rescues CYCLIN D1 expression and hepatocyte proliferation to wild-type levels. Thus, together these findings define a novel pathway in which SULF2 regulates tissue regeneration in part via the activation of a novel WNT-GLI1-CYCLIN D1 pathway.

scholarly journals Notum Protects Against Myocardial Infarction-Induced Heart Dysfunction By Alleviating Cardiac Fibrosis

2021 ◽  
Author(s):  
Wei Su ◽  
Ruonan Fang ◽  
Yue Li ◽  
Liangliang Li ◽  
Jing Zhang ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Wnt Signaling ◽  
Signaling Pathway ◽  
Cardiac Fibrosis ◽  
Cardiac Fibroblasts ◽  
Wnt Signaling Pathway ◽  
New Strategy ◽  
Feedback Inhibitor ◽  
Reparative Process ◽  
Organ Fibrosis

Abstract Cardiac fibrosis is a pathological reparative process that occurs subsequent to myocardial injury. It is associated with cardiac systolic and diastolic dysfunction and reduced cardiac compliance that eventually leads to heart failure. Delaying or inhibiting the progression of pathological myocardial fibrosis is of great significance for the treatment of many cardiovascular diseases. The Wnt signaling pathway is closely related to the occurrence of organ fibrosis, and Notum is a highly conserved secreted feedback inhibitor of Wnt signaling. It has been shown that Notum acts as a regulator in many organs, such as the aging intestinal epithelium, adult ventricular-subventricular zone neurogenesis, and mouse tooth root development. However, the role and mechanism of Notum on cardiac fibrosis are not well-understood. In this study, we found that Notum significantly increased survival rate and improved cardiac function following myocardial infarction in mice. More importantly, Notum inhibited the Wnt/β-catenin signaling pathway and senescence of cardiac fibroblasts, thereby decreasing the activation of cardiac fibroblasts, reducing the excessive deposition of extracellular matrix, and ultimately inhibiting the occurrence of cardiac fibrosis. Taken together, our findings demonstrated the anti-fibrotic effects of Notum on maladaptive cardiac fibrosis, and suggest that it may be a new strategy for the treatment of cardiac fibrosis.

264The long noncoding RNA H19 modulates cardiac remodeling after infarction

2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
O K Choong ◽  
C Y Chen ◽  
J H Lin ◽  
P J Lin ◽  
J H Zhang ◽  
...  
Keyword(s):  
Cardiac Remodeling ◽  
Noncoding Rna ◽  
Cardiac Fibrosis ◽  
Cardiac Development ◽  
Cardiac Fibroblasts ◽  
Noncoding Rnas ◽  
Interacting Protein ◽  
Functional Studies ◽  
Rna Purification

Abstract Noncoding RNAs account for 80% of human transcripts, but functional studies on noncoding RNAs are relatively few and limited. Long noncoding RNAs (lncRNAs) are known to have an important role in cardiac development, and lately, high-throughput RNA sequencing has been extensively utilized to profile and explore the transcriptome landscape of lncRNAs in failing hearts. These studies have revealed that lncRNAs are mostly dysregulated in failing hearts and their expression signature can discriminate failing hearts of different etiologies. H19 is abundantly expressed in failing human hearts and its polymorphism was shown to possess a significant correlation with the risk of coronary artery diseases. In our study using murine hearts, we discovered that H19 was significantly up regulated in the heart after ischemic injury, with predominant expression in cardiac fibroblasts. This finding piqued our interest to further investigate the function of H19 in the heart. We demonstrated that ectopic overexpression of H19 using the AAV approach led to severe cardiac fibrosis in mouse hearts following myocardial infarction. In light of this finding, we generated H19 knockout mice to further investigate the functionality of H19 and we found that cardiac fibrosis was attenuated in these mice. Altogether, these findings suggested that H19 is a fibrosis regulator during cardiac remodeling process after infarction. Due to the multiple regulatory roles of lncRNAs, we then took advantage of chromatin isolation by RNA purification (ChIRP) to identify the H19-interacting protein, YB-1. Surprisingly, mice with YB-1 knockdown displayed severe cardiac fibrosis even without injury. Furthermore, we demonstrated that YB-1 is a transcriptional suppressor of collagen 1A1. Knockout of H19 in YB-1 knockdown partially suppressed Col1a1 expression, which suggests a negative regulatory role of H19 on YB-1 towards the expression of Col1a1. Taking into account all of these findings, we concluded that H19 mediates collagen expression in fibroblasts through the inhibition of YB-1 activity during cardiac remodeling.

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