scholarly journals Investigation of cardiac fibroblasts using myocardial slices

2017 ◽  
Vol 114 (1) ◽  
pp. 77-89 ◽  
Author(s):  
Filippo Perbellini ◽  
Samuel A Watson ◽  
Martina Scigliano ◽  
Samha Alayoubi ◽  
Sebastian Tkach ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Growth Factor ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Transforming Growth Factor Β ◽  
Mechanical Stimuli ◽  
New Model ◽  
Phenotypic Changes

Abstract Aims Cardiac fibroblasts (CFs) are considered the principal regulators of cardiac fibrosis. Factors that influence CF activity are difficult to determine. When isolated and cultured in vitro, CFs undergo rapid phenotypic changes including increased expression of α-SMA. Here we describe a new model to study CFs and their response to pharmacological and mechanical stimuli using in vitro cultured mouse, dog and human myocardial slices. Methods and results Unloading of myocardial slices induced CF proliferation without α-SMA expression up to 7 days in culture. CFs migrating onto the culture plastic support or cultured on glass expressed αSMA within 3 days. The cells on the slice remained αSMA(−) despite transforming growth factor-β (20 ng/ml) or angiotensin II (200 µM) stimulation. When diastolic load was applied to myocardial slices using A-shaped stretchers, CF proliferation was significantly prevented at Days 3 and 7 (P < 0.001). Conclusions Myocardial slices allow the study of CFs in a multicellular environment and may be used to effectively study mechanisms of cardiac fibrosis and potential targets.

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

2015 ◽  
Vol 308 (5) ◽  
pp. H424-H434 ◽  
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.

Abstract P475: Microrna-129-5p Rescues Myocardial Fibrosis And Calcification By Targeting Asporin And Sox9 In Cardiac Fibroblasts

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Lejla Medzikovic ◽  
Laila Aryan ◽  
Gregoire Ruffenach ◽  
Min Li ◽  
Nicoletta Savalli ◽  
...  
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Systolic Dysfunction ◽  
Transforming Growth Factor Β ◽  
Calcium Deposition ◽  
Systemic Delivery ◽  
Myocardial Calcification

Myocardial fibrosis promotes heart failure (HF) progression by impairing myocardial compliance, but also may predispose to myocardial calcification, further impairing cardiac function. Transition of resident cardiac fibroblast (CF) to pro-fibrotic myofibroblasts (MF) and osteogenic cell fates (OF) are key events which are partially controlled by microRNAs (miRs). To discover novel miRs involved in myocardial fibrosis and calcification, we compared online-available microarray datasets of left ventricles (LV) from failing human and mouse hearts. Assessing differentially-expressed miRs known to regulate fibrosis and calcification genes revealed that miR-129-5p is significantly downregulated in HF LV. Bioinformatic target analysis revealed small leucin-rich proteoglycan Asporin (Aspn) and SRY-Box Transcription Factor 9 (Sox9) as two novel miR-129-5p targets upregulated in both mouse and human diseased LV. Thus far, nothing is known about miR-129-5p in cardiac fibrosis and calcification. Additionally, the role of Asporin in myocardial fibrosis and the roles of either Asporin or Sox9 in myocardial calcification remain undiscovered. We show that miR-129-5p is expressed in CF in mouse and human hearts and is downregulated in CF of both HF patients and Angiotensin II (AngII)-injured mice, while Asporin and Sox9 are upregulated in CF of HF LV. In vitro , AngII or transforming growth factor-β downregulated miR-129-5p expression in primary adult mouse CF. Overexpression of miR-129-5p in CF inhibited expression of MF and OF transition markers, reduced migration, collagen production and calcium deposition. We validated Asporin and Sox9 as direct targets of miR-129-5p. Accordingly, silencing of Asporin and Sox9 in CF attenuated molecular and functional characteristics of MF and OF transition. Strikingly, systemic delivery of miR-129-5p mimics in mice directly targets CF and is sufficient to rescue preexisting AngII-induced myocardial fibrosis, calcification, diastolic- and systolic dysfunction. In conclusion, miR-129-5p rescues myocardial fibrosis and calcification by attenuating MF and OF transition via inhibition of Asporin and Sox9 in CF and is a promising therapeutic target.

scholarly journals Retinoid X receptor agonists attenuates cardiomyopathy in streptozotocin-induced type 1 diabetes through LKB1-dependent anti-fibrosis effects

2020 ◽  
Vol 134 (6) ◽  
pp. 609-628 ◽  
Author(s):  
Dajun Chai ◽  
Xiaoyan Lin ◽  
Qiaowen Zheng ◽  
Changsheng Xu ◽  
Hong Xie ◽  
...  
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Transforming Growth Factor Β ◽  
Underlying Mechanism ◽  
Left Ventricular ◽  
Retinoid X Receptor ◽  
Sprague Dawley

Abstract Diabetic cardiac fibrosis increases ventricular stiffness and facilitates the occurrence of diastolic dysfunction. Retinoid X receptor (RXR) plays an important role in cardiac development and has been implicated in cardiovascular diseases. In the present study, we investigated the effects of RXR agonist treatment on streptozotocin (STZ)-induced diabetic cardiomyopathy (DCM) and the underlying mechanism. Sprague–Dawley (SD) rats induced by STZ injection were treated with either RXR agonist bexarotene (Bex) or vehicle alone. Echocardiography was performed to determine cardiac structure and function. Cardiac fibroblasts (CFs) were treated with high glucose (HG) with or without the indicated concentration of Bex or the RXR ligand 9-cis-retinoic acid (9-cis-RA). The protein abundance levels were measured along with collagen, body weight (BW), blood biochemical indexes and transforming growth factor-β (TGF-β) levels. The effects of RXRα down-regulation by RXRα small interfering RNA (siRNA) were examined. The results showed that bexarotene treatment resulted in amelioration of left ventricular dysfunction by inhibiting cardiomyocyte apoptosis and myocardial fibrosis. Immunoblot with heart tissue homogenates from diabetic rats revealed that bexarotene activated liver kinase B1 (LKB1) signaling and inhibited p70 ribosomal protein S6 kinase (p70S6K). The increased collagen levels in the heart tissues of DCM rats were reduced by bexarotene treatment. Treatment of CFs with HG resulted in significantly reduced LKB1 activity and increased p70S6K activity. RXRα mediated the antagonism of 9-cis-RA on HG-induced LKB1/p70S6K activation changes in vitro. Our findings suggest that RXR agonist ameliorates STZ-induced DCM by inhibiting myocardial fibrosis via modulation of the LKB1/p70S6K signaling pathway. RXR agonists may serve as novel therapeutic agents for the treatment of DCM.

scholarly journals Connective tissue growth factor expression after angiotensin II exposure is dependent on transforming growth factor-β signaling via the canonical Smad-dependent pathway in hypertensive induced myocardial fibrosis

2018 ◽  
Vol 19 (1) ◽  
pp. 147032031875935 ◽  
Author(s):  
Chloe Kok Sum Wong ◽  
Alec Falkenham ◽  
Tanya Myers ◽  
Jean-Francois Légaré
Keyword(s):  
Angiotensin Ii ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Transforming Growth Factor Β ◽  
Tissue Growth ◽  
Ang Ii ◽  
Qrt Pcr ◽  
Dependent Pathway

Introduction: Transforming growth factor-β (TGF-β) and connective tissue growth factor (CTGF) are often described as the initial pro-fibrotic mediators upregulated early in fibrosis models dependent on angiotensin II (Ang-II). In the present study, we explore the mechanistic link between TGF-β and CTGF expression by using a novel TGF-β trap. Materials and methods: NIH/3T3 fibroblasts were subjected to TGF-β with or without TGF-β trap or 1D11 antibody, CTGF or CTGF plus TGF-β for six or 24 hours, and then used for quantitative real-time polymerase chain reaction (qRT-PCR) or immunocytochemistry. Male C57BL/6 mice were infused with Ang-II and randomly assigned TGF-β trap for six or 24 hours. Hearts were harvested for histological analyses, qRT-PCR and western blotting. Results: Exogenous TGF-β-induced fibroblasts resulted in significant upregulation of CTGF, TGF-β and type I collagen transcript levels in vitro. Additionally, TGF-β promoted the differentiation of fibroblasts into α-SMA+ myofibroblasts. CTGF expression was reduced by the addition of TGF-β trap or neutralizing antibody, confirming that its expression is dependent on TGF-β signaling. In contrast, exogenous CTGF did not appear to have an effect on fibroblast production of pro-fibrotic transcripts or fibroblast differentiation. Ang-II infusion in vivo led to a significant increase in TGF-β and CTGF mRNA expression at six and 24 hours with corresponding changes in Smad2 phosphorylation (pSmad2), indicative of increased TGF-β signaling. Ang-II animals that received the TGF-β trap demonstrated reduced CTGF mRNA levels and pSmad2 at six hours, suggesting that early CTGF expression is dependent on TGF-β signaling. Conclusions: We demonstrated that CTGF expression is dependent on TGF-β signaling both in vitro and in vivo in a model of myocardial fibrosis. This also suggests that early myocardial CTGF mRNA expression (six hours) after Ang-II exposure is likely dependent on latent TGF-β activation via the canonical Smad-dependent pathway in resident cardiac cells.

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