P300/CBP-Associated Factor Activates Cardiac Fibroblasts by SMAD2 Acetylation

Various heart diseases cause cardiac remodeling, which in turn leads to ineffective contraction. Although it is an adaptive response to injury, cardiac fibrosis contributes to this remodeling, for which the reactivation of quiescent myofibroblasts is a key feature. In the present study, we investigated the role of the p300/CBP-associated factor (PCAF), a histone acetyltransferase, in the activation of cardiac fibroblasts. An intraperitoneal (i.p.) injection of a high dose (160 mg/kg) of isoproterenol (ISP) induced cardiac fibrosis and reduced the amount of the PCAF in cardiac fibroblasts in the mouse heart. However, the PCAF activity was significantly increased in cardiac fibroblasts, but not in cardiomyocytes, obtained from ISP-administered mice. An in vitro study using human cardiac fibroblast cells recapitulated the in vivo results; an treatment with transforming growth factor-β1 (TGF-β1) reduced the PCAF, whereas it activated the PCAF in the fibroblasts. PCAF siRNA attenuated the TGF-β1-induced increase in and translocation of fibrosis marker proteins. PCAF siRNA blocked TGF-β1-mediated gel contraction and cell migration. The PCAF directly interacted with and acetylated mothers against decapentaplegic homolog 2 (SMAD2). PCAF siRNA prevented TGF-β1-induced phosphorylation and the nuclear localization of SMAD2. These results suggest that the increase in PCAF activity during cardiac fibrosis may participate in SMAD2 acetylation and thereby in its activation.

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Up-Regulating Relaxin Expression by G-Quadruplex Interactive Ligand to Achieve Antifibrotic Action

Endocrinology ◽

10.1210/en.2012-1114 ◽

2012 ◽

Vol 153 (8) ◽

pp. 3692-3700 ◽

Cited By ~ 28

Author(s):

Hui-Ping Gu ◽

Sen Lin ◽

Ming Xu ◽

Hai-Yi Yu ◽

Xiao-Jun Du ◽

...

Keyword(s):

Rna Polymerase Ii ◽

Cardiac Fibrosis ◽

Cardiac Fibroblasts ◽

Heart Diseases ◽

Gene Promoter ◽

Binding Motif ◽

Endogenous Expression ◽

G Quadruplex

Myocardial fibrosis is a key pathological change in a variety of heart diseases contributing to the development of heart failure, arrhythmias, and sudden death. Recent studies have shown that relaxin prevents and reverses cardiac fibrosis. Endogenous expression of relaxin was elevated in the setting of heart disease; the extent of such up-regulation, however, is insufficient to exert compensatory actions, and the mechanism regulating relaxin expression is poorly defined. In the rat relaxin-1 (RLN1, Chr1) gene promoter region we found presence of repeated guanine (G)-rich sequences, which allowed formation and stabilization of G-quadruplexes with the addition of a G-quadruplex interactive ligand berberine. The G-rich sequences and the G-quadruplexes were localized adjacent to the binding motif of signal transducer and activator of transcription (STAT)3, which negatively regulates relaxin expression. Thus, we hypothesized that the formation and stabilization of G-quadruplexes by berberine could influence relaxin expression. We found that berberine-induced formation of G-quadruplexes did increase relaxin gene expression measured at mRNA and protein levels. Formation of G-quadruplexes significantly reduced STAT3 binding to the promoter of relaxin gene. This was associated with consequent increase in the binding of RNA polymerase II and STAT5a to relaxin gene promoter. In cardiac fibroblasts and rats treated with angiotensin II, berberine was found to suppress fibroblast activation, collagen synthesis, and extent of cardiac fibrosis through up-regulating relaxin. The antifibrotic action of berberine in vitro and in vivo was similar to that by exogenous relaxin. Our findings document a novel therapeutic strategy for fibrosis through up-regulating expression of endogenous relaxin.

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Celastrol-Induced Suppression of the MiR-21/ERK Signalling Pathway Attenuates Cardiac Fibrosis and Dysfunction

Cellular Physiology and Biochemistry ◽

10.1159/000445554 ◽

2016 ◽

Vol 38 (5) ◽

pp. 1928-1938 ◽

Cited By ~ 31

Author(s):

Mian Cheng ◽

Gang Wu ◽

Yue Song ◽

Lin Wang ◽

Ling Tu ◽

...

Keyword(s):

Myocardial Fibrosis ◽

Transforming Growth Factor Beta ◽

Cardiac Dysfunction ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Smooth Muscle Actin ◽

Signalling Pathway ◽

Tgf Β1

Backgroud: Myocardial fibrosis results in myocardial remodelling and dysfunction. Celastrol, a traditional oriental medicine, has been suggested to have cardioprotective effects. However, its underlying mechanism is unknown. This study investigated the ability of celastrol to prevent cardiac fibrosis and dysfunction and explored the underlying mechanisms. Methods: Animal and cell models of cardiac fibrosis were used in this study. Myocardial fibrosis was induced by transverse aortic constriction (TAC) in mice. Cardiac hypertrophy and fibrosis were evaluated based on histological and biochemical measurements. Cardiac function was evaluated by echocardiography. The levels of transforming growth factor beta 1 (TGF-β1), extracellular signal regulated kinases 1/2 (ERK1/2) signalling were measured using Western blotting, while the expression of miR-21was analyzed by real-time qRT-PCR in vitro and in vivo. In vitro studies, cultured cardiac fibroblasts (CFs) were treated with TGF-β1 and transfected with microRNA-21(miR21). Results: Celastrol treatment reduced the increased collagen deposition and down-regulated α-smooth muscle actin (α-SMA), atrial natriuretic peptide (ANP), brain natriuretic peptides (BNP), beta-myosin heavy chain (β-MHC), miR-21 and p-ERK/ERK. Cardiac dysfunction was significantly attenuated by celastrol treatment in the TAC mice model. Celastrol treatment reduced myocardial fibroblast viability and collagen content and down-regulated α-SMA in cultured CFs in vitro. Celastrol also inhibited the miR-21/ERK signalling pathway. Celastrol attenuated miR-21 up-regulation by TGF-β1 and decreased elevated p-ERK/ERK levels in CFs transfected with miR-21. Conclusion: MiR-21/ERK signalling could be a potential therapeutic pathway for the prevention of myocardial fibrosis. Celastrol ameliorates myocardial fibrosis and cardiac dysfunction, these probably related to miR-21/ERK signaling pathways in vitro and in vivo.

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Long Non-Coding RNA 554 Promotes Cardiac Fibrosis via TGF-β1 Pathway in Mice Following Myocardial Infarction

Frontiers in Pharmacology ◽

10.3389/fphar.2020.585680 ◽

2020 ◽

Vol 11 ◽

Author(s):

Bihui Luo ◽

Zhiyu He ◽

Shijun Huang ◽

Jinping Wang ◽

Dunzheng Han ◽

...

Keyword(s):

Myocardial Infarction ◽

Cardiac Fibrosis ◽

Cardiac Fibroblasts ◽

Non Coding Rna ◽

Novel Target ◽

And Function ◽

Tgf Β1

Rationale: Cardiac fibrosis is observed in nearly every form of myocardial disease. Long non-coding RNAs (lncRNAs) have been shown to play an important role in cardiac fibrosis, but the detailed molecular mechanism remains unknown.Object: We aimed at characterizing lncRNA 554 expression in murine cardiac fibroblasts (CFs) after myocardial infarction (MI) to identify CF-enriched lncRNA and investigate its function and contribution to cardiac fibrosis and function.Methods and Results: In this study, we identified lncRNA NONMMUT022554 (lncRNA 554) as a regulator of MI-induced cardiac fibrosis. We found that lncRNA 554 was significantly up-regulated in the mouse hearts following MI. Further study showed that lncRNA 554 was predominantly expressed in cardiac fibroblasts, indicating a potential role of lncRNA 554 in cardiac fibrosis. In vitro knockdown of lncRNA 554 by siRNA suppressed fibroblasts migration and expression of extracellular matrix (ECM); while overexpression of lncRNA 554 promoted expression of ECM genes. Consistently, lentivirus mediated in vivo knockdown of lncRNA 554 could inhibit cardiac fibrosis and improve cardiac function in mouse model of MI. More importantly, TGF-β1 inhibitor (TEW-7197) could reverse the pro-fibrotic function of lncRNA 554 in CFs. This suggests that the effects of lncRNA 554 on cardiac fibrosis is TGF-β1 dependent.Conclusion: Collectively, our study illustrated the role of lncRNA 554 in cardiac fibrosis, suggested that lncRNA 554 might be a novel target for cardiac fibrosis.

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Abstract 331: Mir-433 Controls Cardiac Fibrosis

Circulation Research ◽

10.1161/res.117.suppl_1.331 ◽

2015 ◽

Vol 117 (suppl_1) ◽

Author(s):

Lichan Tao ◽

Xiaoting Wu ◽

Ping Chen ◽

Shanshan Li ◽

Xiaomin Zhang ◽

...

Keyword(s):

Myocardial Infarction ◽

Cardiac Fibrosis ◽

Cardiac Fibroblasts ◽

Heart Diseases ◽

Mice Model ◽

Ki 67 ◽

End Stage ◽

Common Manifestation

Background: Cardiac fibrosis, a result of multiple injurious insults in heart, is a final common manifestation of chronic heart diseases and can lead to end-stage cardiac failure. MicroRNAs (miRNAs, miRs) participate in many essential biological processes and their dysfunction has been implicated in a variety of cardiovascular diseases including fibrosis. miR-433 has recently been implicated in renal fibrosis, however, its role in cardiac fibrosis is unclear. Methods and results: miR-433 was increased in heart samples from dilated cardiomyopathy patients as determined by qRT-PCRs. In addition, miR-433 was also consistently upregulated in mice model of cardiac fibrosis after myocardial infarction or heart failure. Additionally, miR-433 was found to be enriched in fibroblasts compared to cardiomyocytes. In neonatal cardiac fibroblasts, forced expression of miR-433 promoted cell proliferation as indicated by EdU and Ki-67 staining. Moreover, miR-433 overexpression promoted the transdifferentiation of fibroblasts into myofibroblasts as determined by qRT-PCR and western blot for α-SMA and collagen whether in the presence of TGF-β or not, indicating that miR-433 is sufficient to induce fibrosis. In addition, knockdown of miR-433 inhibited proliferation and the transdifferentiation into myofibroblasts, indicating that miR-433 is required for cardiac fibrosis. Interestingly, miR-433 did not affect the migration of cardiac fibroblast. Importantly, miR-433 antagomir could partially attenuate cardiac fibrosis induced by myocardial infarction in mice. Conclusion: both in vitro and in vivo. Inhibition of miR-433 represents a novel therapeutic strategy for cardiac fibrosis.

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Old Drug, New Trick: Tilorone, a Broad-Spectrum Antiviral Drug as a Potential Anti-Fibrotic Therapeutic for the Diseased Heart

Pharmaceuticals ◽

10.3390/ph14030263 ◽

2021 ◽

Vol 14 (3) ◽

pp. 263

Author(s):

Duncan Horlock ◽

David M. Kaye ◽

Catherine E. Winbanks ◽

Xiao-Ming Gao ◽

Helen Kiriazis ◽

...

Keyword(s):

Collagen Synthesis ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Systolic Function ◽

Cardiac Fibroblasts ◽

Antiviral Drug ◽

Mouse Heart ◽

Novel Therapy ◽

Human Cardiac Fibroblasts

Cardiac fibrosis is associated with most forms of cardiovascular disease. No reliable therapies targeting cardiac fibrosis are available, thus identifying novel drugs that can resolve or prevent fibrosis is needed. Tilorone, an antiviral agent, can prevent fibrosis in a mouse model of lung disease. We investigated the anti-fibrotic effects of tilorone in human cardiac fibroblasts in vitro by performing a radioisotopic assay for [3H]-proline incorporation as a proxy for collagen synthesis. Exploratory studies in human cardiac fibroblasts treated with tilorone (10 µM) showed a significant reduction in transforming growth factor-β induced collagen synthesis compared to untreated fibroblasts. To determine if this finding could be recapitulated in vivo, mice with established pathological remodelling due to four weeks of transverse aortic constriction (TAC) were administered tilorone (50 mg/kg, i.p) or saline every third day for eight weeks. Treatment with tilorone was associated with attenuation of fibrosis (assessed by Masson’s trichrome stain), a favourable cardiac gene expression profile and no further deterioration of cardiac systolic function determined by echocardiography compared to saline treated TAC mice. These data demonstrate that tilorone has anti-fibrotic actions in human cardiac fibroblasts and the adult mouse heart, and represents a potential novel therapy to treat fibrosis associated with heart failure.

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Activation of SIRT3 by resveratrol ameliorates cardiac fibrosis and improves cardiac function via the TGF-β/Smad3 pathway

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00454.2014 ◽

2015 ◽

Vol 308 (5) ◽

pp. H424-H434 ◽

Cited By ~ 78

Author(s):

Tongshuai Chen ◽

Jingyuan Li ◽

Junni Liu ◽

Na Li ◽

Shujian Wang ◽

...

Keyword(s):

Growth Factor ◽

Cardiac Function ◽

Calorie Restriction ◽

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Cardiac Fibroblasts ◽

Transforming Growth Factor Β ◽

Wild Type

Sirtuins [sirtuin (SIRT)1–SIRT7] mediate the longevity-promoting effects of calorie restriction in yeast, worms, flies, and mice. Additionally, SIRT3 is the only SIRT analog whose increased expression has been shown to be associated with longevity in humans. The polyphenol resveratrol (RSV) is the first compound discovered able to mimic calorie restriction by stimulating SIRTs. In the present study, we report that RSV activated SIRT3 in cardiac fibroblasts both in vivo and in vitro. Moreover, in wild-type mice, RSV prevented cardiac hypertrophy in response to hypertrophic stimuli. However, this protective effect was not observed in SIRT3 knockout mice. Additionally, the activation of SIRT3 by RSV ameliorated collagen deposition and improved cardiac function. In isolated cardiac fibroblasts, pretreatment with RSV suppressed fibroblast-to-myoblast transformation by inhibiting the transforming growth factor-β/Smad3 pathway. Therefore, these data indicate that the activation of SIRT3 by RSV could ameliorate cardiac fibrosis and improve cardiac function via the transforming growth factor-β/Smad3 pathway.

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Enhanced expression of fibrillin-1, a constituent of the myocardial extracellular matrix in fibrosis

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00151.2005 ◽

2005 ◽

Vol 289 (3) ◽

pp. H982-H991 ◽

Cited By ~ 51

Author(s):

Fatiha Bouzeghrane ◽

Dieter P. Reinhardt ◽

Tim L. Reudelhuber ◽

Gaétan Thibault

Keyword(s):

Cardiac Fibrosis ◽

Transforming Growth Factor ◽

Cardiac Fibroblasts ◽

Transforming Growth Factor Β1 ◽

Ang Ii ◽

Rat Hearts ◽

Enhanced Expression ◽

Fibrillin 1

Fibrillin-1 localization in the myocardium and the modulation of its expression in cardiac fibrosis were examined. In normal rat hearts, fibrillin-1 was abundant throughout the myocardium as thin fibers that crossed over the perimysium and around arteries. After cardiac fibrosis was induced in rats by either 14-day ANG II infusion or 21-day DOCA-salt treatment [a high endothelin-1 (ET-1) model], fibrillin-1 immunostaining was stronger in the interstitium (2.8-fold and 4.4-fold increases, respectively, in each model), extended between myocytes, and accumulated in microscopic scars and in the perivascular area of both ventricles. mRNA analysis confirmed its enhanced ventricular expression in both groups of rats (2.5-fold and 6.6-fold increments, respectively, in each model). In 1B normotensive and 2C hypertensive transgenic mice, two lines expressing an ANG II fusion protein in cardiac myocytes, strong fibrillin-1 immunoreactivity was observed in the interstitium and around arteries (3.7-fold and 7-fold increases, respectively). ANG II and transforming growth factor-β1 enhanced fibrillin-1 synthesis by cardiac fibroblasts. Some fibrillin-1 fragments interacted with RGD-dependent integrins, including α8β1-integrin, of cardiac fibroblasts but not necessarily through the RGD motif. Our findings illustrate that fibrillin-1 is an important constituent of the myocardium. In vitro and in vivo evidence suggests that ANG II can directly induce fibrillin-1 expression in cardiac fibroblasts. This protein can thus contribute to reactive and reparative processes.

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Abstract 410: Rhein Administration Prevents Cardiac Fibrosis by Interfering with the TGF-β Pathway

Circulation Research ◽

10.1161/res.121.suppl_1.410 ◽

2017 ◽

Vol 121 (suppl_1) ◽

Author(s):

David Barbosa ◽

Melanie Wehmöller ◽

Maximilian R Spinner ◽

Ulrich Rüther ◽

Margriet Ouwens

Keyword(s):

Cardiac Fibrosis ◽

Heart Function ◽

Cardiac Fibroblasts ◽

Heart Diseases ◽

Collagen Gel ◽

Pressure Overload ◽

Protein Levels ◽

Adverse Remodeling

Fibrosis, which occurs in various heart diseases like acute myocardial ischemia and pressure overload, is triggered by the differentiation of fibroblasts into myofibroblasts. Dysregulation of this reparative mechanism results in excessive collagen accumulation leading to cardiac stiffness and impaired heart function. The aim of this study was to determine whether the rhubarb anthraquinone Rhein, a drug already used as treatment for chondroarthritis, prevents the transdifferentiation of cardiac fibroblasts. We observed that Rhein pre-treatment ameliorates the cardiac function and reduces adverse remodeling after acute myocardial infarction in mice, in vivo . In primary human cardiac fibroblasts, Rhein incubation dose-dependently inhibited the TGF-β-mediated upregulation of α-SMA, the master marker for myofibrolasts, and prevented the contraction of fibroblast-populated collagen gel lattices upon TGF-β stimulation. Further, Rhein reduced TGFβ-R1 expression in primary human cardiac fibroblast, resulting in decreased SMAD2 phosphorylation and blunting of the fibrogenic response. Furthermore, Rhein stabilized protein levels of SMAD7, a key inhibitor of TGF-β signaling. Collectively, these data show for the first time that Rhein administration prevents cardiac fibrosis in vivo and in vitro by blunting the TGF-β signaling pathway, and identify Rhein as potential therapeutic treatment to prevent excessive fibrosis and adverse remodeling in cardiac pathologies.

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Regorafenib-Attenuated, Bleomycin-Induced Pulmonary Fibrosis by Inhibiting the TGF-β1 Signaling Pathway

International Journal of Molecular Sciences ◽

10.3390/ijms22041985 ◽

2021 ◽

Vol 22 (4) ◽

pp. 1985

Author(s):

Xiaohe Li ◽

Ling Ma ◽

Kai Huang ◽

Yuli Wei ◽

Shida Long ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Signaling Pathway ◽

Transforming Growth Factor ◽

Kinase Inhibitor ◽

In Vivo Experiments ◽

Age Related ◽

And Migration ◽

Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a fatal and age-related pulmonary disease. Nintedanib is a receptor tyrosine kinase inhibitor, and one of the only two listed drugs against IPF. Regorafenib is a novel, orally active, multi-kinase inhibitor that has similar targets to nintedanib and is applied to treat colorectal cancer and gastrointestinal stromal tumors in patients. In this study, we first identified that regorafenib could alleviate bleomycin-induced pulmonary fibrosis in mice. The in vivo experiments indicated that regorafenib suppresses collagen accumulation and myofibroblast activation. Further in vitro mechanism studies showed that regorafenib inhibits the activation and migration of myofibroblasts and extracellular matrix production, mainly through suppressing the transforming growth factor (TGF)-β1/Smad and non-Smad signaling pathways. In vitro studies have also indicated that regorafenib could augment autophagy in myofibroblasts by suppressing TGF-β1/mTOR (mechanistic target of rapamycin) signaling, and could promote apoptosis in myofibroblasts. In conclusion, regorafenib attenuates bleomycin-induced pulmonary fibrosis by suppressing the TGF-β1 signaling pathway.

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Abstract 277: Microrna-33 Promotes Cardiac Fibrosis by Maintaining Cellular Lipid Homeostasis

Circulation Research ◽

10.1161/res.119.suppl_1.277 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Masataka Nishiga ◽

Takahiro Horie ◽

Yasuhide Kuwabara ◽

Osamu Baba ◽

Tetsushi Nakao ◽

...

Keyword(s):

Cardiac Fibrosis ◽

Cardiac Fibroblasts ◽

Pressure Overload ◽

Cholesterol Content ◽

Atp Binding Cassette Transporter ◽

Primary Fibroblasts ◽

Proliferation In Vitro ◽

Altered Lipid

Background: A highly conserved microRNA, miR-33 is considered as a potential therapeutic target for atherosclerosis, because recent reports, including ours, indicated miR-33 has atherogenic effects by reducing HDL-C. However, the functions of miR-33 in heart failure remain to be elucidated. Methods and results: To clarify the functions of miR-33 involved in cardiac hypertrophy and fibrosis in vivo, we investigated the responses to pressure overload by transverse aortic constriction (TAC) in miR-33 deficient (KO) mice. When subjected to TAC, miR-33 expression level was significantly up-regulated in wild-type (WT) left ventricles, whereas miR-33 KO hearts displayed no less hypertrophic responses than WT hearts. However, interestingly, histological and gene expression analyses showed ameliorated cardiac fibrosis in miR-33 KO hearts compared to WT hearts. Furthermore, we generated cardiac fibroblast specific miR-33 deficient mice, which also showed ameliorated cardiac fibrosis when they were subjected to TAC. We also found that cardiac fibroblasts were mainly responsible for miR-33 expression in the heart, because its expression was about 4-folds higher in isolated primary cardiac fibroblasts than cardiomyocytes. Deficiency of miR-33 impaired cell proliferation in primary fibroblasts, which was considered due to altered lipid raft cholesterol content by up-regulated ATP-binding cassette transporter A1/G1. Conclusion: Deficiency of miR-33 impaired fibroblast proliferation in vitro, and ameliorated cardiac fibrosis induced by pressure overload in vivo.

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