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Cardiac fibrosis is characterized by the over-proliferation, over-transdifferentiation and over-deposition of extracellular matrix (ECM) of cardiac fibroblasts (CFs). Cardiac sympathetic activation is one of the leading causes of myocardial fibrosis. Meanwhile, cardiac fibrosis is often together with cardiac inflammation, which accelerates fibrosis by mediating inflammatory cytokines secretion. Recently, the Janus kinase/signal transducer and activator of transcription (JAK/STAT3) signaling pathway has been confirmed by its vital role during the progression of cardiac fibrosis. Thus, JAK/STAT3 signaling pathway is thought to be a potential therapeutic target for cardiac fibrosis. Baricitinib (BR), a novel JAK1/2 inhibitor, has been reported excellent effects of anti-fibrosis in multiple fibrotic diseases. However, little is known about whether and how BR ameliorates cardiac fibrosis caused by chronic sympathetic activation. Isoproterenol (ISO), a β-Adrenergic receptor (β-AR) nonselective agonist, was used to modulate chronic sympathetic activation in mice. As expected, our results proved that BR ameliorated ISO-induced cardiac dysfunction. Meanwhile, BR attenuated ISO-induced cardiac fibrosis and cardiac inflammation in mice. Moreover, BR also inhibited ISO-induced cardiac fibroblasts activation and macrophages pro-inflammatory secretion. As for mechanism studies, BR reduced ISO-induced cardiac fibroblasts by JAK2/STAT3 and PI3K/Akt signaling, while reduced ISO-induced macrophages pro-inflammatory secretion by JAK1/STAT3 and NF-κB signaling. In summary, BR alleviates cardiac fibrosis and inflammation caused by chronic sympathetic activation. The underlying mechanism involves BR-mediated suppression of JAK1/2/STAT3, PI3K/Akt and NF-κB signaling., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Aims: Heart failure (HF) is one of the most devastating consequences of cardiovascular diseases. Regardless of etiology, cardiac fibrosis is present and promotes the loss of heart function in HF patients. Cardiac resident fibroblasts, in response to a host of pro-fibrogenic stimuli, trans-differentiate into myofibroblasts to mediate cardiac fibrosis, the underlying mechanism of which remains incompletely understood., Methods: Fibroblast-myofibroblast transition was induced in vitro by exposure to transforming growth factor (TGF-β). Cardiac fibrosis was induced in mice by either transverse aortic constriction (TAC) or by chronic infusion with angiotensin II (Ang II)., Results: Through bioinformatic screening, we identified Kruppel-like factor 6 (KLF6) as a transcription factor preferentially up-regulated in cardiac fibroblasts from individuals with non-ischemic cardiomyopathy (NICM) compared to the healthy donors. Further analysis showed that nuclear factor kappa B (NF-κB) bound to the KLF6 promoter and mediated KLF6 trans-activation by pro-fibrogenic stimuli. KLF6 knockdown attenuated whereas KLF6 over-expression enhanced TGF-β induced fibroblast-myofibroblast transition in vitro. More importantly, myofibroblast-specific KLF6 depletion ameliorated cardiac fibrosis and rescued heart function in mice subjected to the TAC procedure or chronic Ang II infusion., Significance: In conclusion, our data support a role for KLF6 in cardiac fibrosis., Competing Interests: Declaration of competing interest None., (Copyright © 2024. Published by Elsevier Inc.)

This study investigates how dynamic fluctuations in matrix stiffness affect the behavior of cardiac fibroblasts (CFs) within a three-dimensional (3D) hydrogel environment. Using hybrid hydrogels with tunable stiffness, we created an in vitro model to mimic the varying stiffness of the cardiac microenvironment. By manipulating hydrogel stiffness, we examined CF responses, particularly the expression of α-smooth muscle actin (α-SMA), a marker of myofibroblast differentiation. Our findings reveal that increased matrix stiffness promotes the differentiation of CFs into myofibroblasts, while matrix softening reverses this process. Additionally, we identified the role of focal adhesions and integrin β1 in mediating stiffness-induced phenotypic switching. This study provides significant insights into the mechanobiology of cardiac fibrosis and suggests that modulating matrix stiffness could be a potential therapeutic strategy for treating cardiovascular diseases., (© 2024. The Author(s).)

Background: Cardiac fibrosis after myocardial infarction (MI) has been considered an important part of cardiac pathological remodeling. Immune cells, especially macrophages, are thought to be involved in the process of fibrosis and constitute a niche with fibroblasts to promote fibrosis. However, the diversity and variability of fibroblasts and macrophages make it difficult to accurately depict interconnections., Methods: We collected and reanalyzed scRNA-seq and snRNA-seq datasets from 12 different studies. Differentiation trajectories of these subpopulations after MI injury were analyzed by using scVelo, PAGA and Slingshot. We used CellphoneDB and NicheNet to infer fibroblast-macrophage interactions. Tissue immunofluorescence staining and in vitro experiments were used to validate our findings., Results: We discovered two subsets of ECM-producing fibroblasts, reparative cardiac fibroblasts (RCFs) and matrifibrocytes, which appeared at different times after MI and exhibited different transcriptional profiles. We also observed that CTHRC1 + fibroblasts represent an activated fibroblast in chronic disease states. We identified a macrophage subset expressing the genes signature of SAMs conserved in both human and mouse hearts. Meanwhile, the SPP1 hi macrophages were predominantly found in the early stages after MI, and cell communication analysis indicated that SPP1 hi macrophage-RCFs interactions are mainly involved in collagen deposition and scar formation., Conclusions: Overall, this study comprehensively analyzed the dynamics of fibroblast and macrophage subsets after MI and identified specific subsets of fibroblasts and macrophages involved in scar formation and collagen deposition., (© 2024. The Author(s).)

The response of cardiac fibroblast proliferation to detrimental stimuli is one of the main pathological factors causing heart remodeling. Reactive oxygen species (ROS) mediate the proliferation of cardiac fibroblasts. However, the exact molecular mechanism remains unclear. In vivo, we examined the oxidative modification of miRNAs with miRNA immunoprecipitation with O 8 G in animal models of cardiac fibrosis induced by Ang II injection or ischemia‒reperfusion injury. Furthermore, in vitro, we constructed oxidation-modified miR-30c and investigated its effects on the proliferation of cardiac fibroblasts. Additionally, luciferase reporter assays were used to identify the target of oxidized miR-30c. We found that miR-30c oxidation was modified by Ang II and PDGF treatment and mediated by excess ROS. We demonstrated that oxidative modification of G to O 8 G occurred at positions 4 and 5 of the 5' end of miR-30c (4,5-oxo-miR-30c), and this modification promoted cardiac fibroblast proliferation. Furthermore, CDKN2C is a negative regulator of cardiac fibroblast proliferation. 4,5-oxo-miR-30c misrecognizes CDKN2C mRNA, resulting in a reduction in protein expression. Oxidized miR-30c promotes cardiac fibroblast proliferation by mismatch mRNA of CDKN2C., (© 2024. The Author(s).)

Hypertension induces cardiac fibrotic remodelling characterised by the phenotypic switching of cardiac fibroblasts (CFs) and collagen deposition. We tested the hypothesis that Wnt1-inducible signalling pathway protein-1 (WISP-1) promotes CFs' phenotypic switch, type I collagen synthesis, and in vivo fibrotic remodelling. The treatment of human CFs (HCFs, n = 16) with WISP-1 (500 ng/mL) induced a phenotypic switch (α-smooth muscle actin-positive) and type I procollagen cleavage to an intermediate form of collagen (pC-collagen) in conditioned media after 24h, facilitating collagen maturation. WISP-1-induced collagen processing was mediated by Akt phosphorylation via integrin β1, and disintegrin and metalloproteinase with thrombospondin motifs 2 (ADAMTS-2). WISP-1 wild-type (WISP-1 +/+ ) mice and WISP-1 knockout (WISP-1 -/- ) mice (n = 5-7) were subcutaneously infused with angiotensin II (AngII, 1000 ng/kg/min) for 28 days. Immunohistochemistry revealed the deletion of WISP-1 attenuated type I collagen deposition in the coronary artery perivascular area compared to WISP-1 +/+ mice after a 28-day AngII infusion, and therefore, the deletion of WISP-1 attenuated AngII-induced cardiac fibrosis in vivo. Collectively, our findings demonstrated WISP-1 is a critical mediator in cardiac fibrotic remodelling, by promoting CFs' activation via the integrin β1-Akt signalling pathway, and induced collagen processing and maturation via ADAMTS-2. Thereby, the modulation of WISP-1 levels could provide potential therapeutic targets in clinical treatment.

Excessive extracellular matrix (ECM) deposition is a defining feature of cardiac fibrosis. Most notably, it is characterized by a significant change in the concentration and volume fraction of collagen I, a disproportionate deposition of collagen subtypes, and a disturbed ECM network arrangement, which directly affect the systolic and diastolic functions of the heart. Immune cells that reside within or infiltrate the myocardium, including macrophages, play important roles in fibroblast activation and consequent ECM remodeling. Through both direct and indirect connections to fibroblasts, monocyte-derived macrophages and resident cardiac macrophages play complex, bidirectional, regulatory roles in cardiac fibrosis. In this review, we discuss emerging interactions between fibroblasts and macrophages in physiology and pathologic conditions, providing insights for future research aimed at targeting macrophages to combat cardiac fibrosis.

Aims: Transforming growth factor β (TGF-β) signalling is one of the critical pathways in fibroblast activation, and several drugs targeting the TGF-β/Smad signalling pathway in heart failure with cardiac fibrosis are being tested in clinical trials. Some caveolins and cavins, which are components of caveolae on the plasma membrane, are known for their association with the regulation of TGF-β signalling. Cavin-2 is particularly abundant in fibroblasts; however, the detailed association between Cavin-2 and cardiac fibrosis is still unclear. We tried to clarify the involvement and role of Cavin-2 in fibroblasts and cardiac fibrosis., Methods and Results: To clarify the role of Cavin-2 in cardiac fibrosis, we performed transverse aortic constriction (TAC) operations on four types of mice: wild-type (WT), Cavin-2 null (Cavin-2 KO), Cavin-2 flox/flox , and activated fibroblast-specific Cavin-2 conditional knockout (Postn-Cre/Cavin-2 flox/flox , Cavin-2 cKO) mice. We collected mouse embryonic fibroblasts (MEFs) from WT and Cavin-2 KO mice and investigated the effect of Cavin-2 in fibroblast trans-differentiation into myofibroblasts and associated TGF-β signalling. Four weeks after TAC, cardiac fibrotic areas in both the Cavin-2 KO and the Cavin-2 cKO mice were significantly decreased compared with each control group (WT 8.04 ± 1.58% vs. Cavin-2 KO 0.40 ± 0.03%, P < 0.01; Cavin-2 flox/flox , 7.19 ± 0.50% vs. Cavin-2 cKO 0.88 ± 0.44%, P < 0.01). Fibrosis-associated mRNA expression (Col1a1, Ctgf, and Col3) was significantly attenuated in the Cavin-2 KO mice after TAC. α1 type I collagen deposition and non-vascular αSMA-positive cells (WT 43.5 ± 2.4% vs. Cavin-2 KO 25.4 ± 3.2%, P < 0.01) were reduced in the heart of the Cavin-2 cKO mice after TAC operation. The levels of αSMA protein (0.36-fold, P < 0.05) and fibrosis-associated mRNA expression (Col1a1, 0.69-fold, P < 0.01; Ctgf, 0.27-fold, P < 0.01; Col3, 0.60-fold, P < 0.01) were decreased in the Cavin-2 KO MEFs compared with the WT MEFs. On the other hand, αSMA protein levels were higher in the Cavin-2 overexpressed MEFs compared with the control MEFs (2.40-fold, P < 0.01). TGF-β1-induced Smad2 phosphorylation was attenuated in the Cavin-2 KO MEFs compared with WT MEFs (0.60-fold, P < 0.01). Heat shock protein 90 protein levels were significantly reduced in the Cavin-2 KO MEFs compared with the WT MEFs (0.69-fold, P < 0.01)., Conclusions: Cavin-2 loss suppressed fibroblast trans-differentiation into myofibroblasts through the TGF-β/Smad signalling. The loss of Cavin-2 in cardiac fibroblasts suppresses cardiac fibrosis and may maintain cardiac function., (© 2023 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

Abstract Background Cardiac fibrosis after myocardial infarction (MI) has been considered an important part of cardiac pathological remodeling. Immune cells, especially macrophages, are thought to be involved in the process of fibrosis and constitute a niche with fibroblasts to promote fibrosis. However, the diversity and variability of fibroblasts and macrophages make it difficult to accurately depict interconnections. Methods We collected and reanalyzed scRNA-seq and snRNA-seq datasets from 12 different studies. Differentiation trajectories of these subpopulations after MI injury were analyzed by using scVelo, PAGA and Slingshot. We used CellphoneDB and NicheNet to infer fibroblast-macrophage interactions. Tissue immunofluorescence staining and in vitro experiments were used to validate our findings. Results We discovered two subsets of ECM-producing fibroblasts, reparative cardiac fibroblasts (RCFs) and matrifibrocytes, which appeared at different times after MI and exhibited different transcriptional profiles. We also observed that CTHRC1+ fibroblasts represent an activated fibroblast in chronic disease states. We identified a macrophage subset expressing the genes signature of SAMs conserved in both human and mouse hearts. Meanwhile, the SPP1hi macrophages were predominantly found in the early stages after MI, and cell communication analysis indicated that SPP1hi macrophage-RCFs interactions are mainly involved in collagen deposition and scar formation. Conclusions Overall, this study comprehensively analyzed the dynamics of fibroblast and macrophage subsets after MI and identified specific subsets of fibroblasts and macrophages involved in scar formation and collagen deposition.

Objective(s): Slit guidance ligand 3 (SLIT3) has been identified as a potential therapeutic regulator against fibroblast activity and fibrillary collagen production in an autocrine manner. However, this research aims to investigate the potential role of SLIT3 in cardiac fibrosis and fibroblast differentiation and its underlying mechanism. Materials and Methods: C57BL/6 mice (male, 8-10 weeks, n=47) were subcutaneously infused with Ang II (2.0 mg/kg/day) for 4 weeks. One to two-day-old Sprague-Dawley (SD) rats were anesthetized by intraperitoneal injection of 1% pentobarbital sodium (60 mg/kg) and ketamine (50 mg/kg) and the cardiac fibroblast was isolated aseptically. The mRNA and protein expression were analyzed using RT-qPCR and Western blotting. Results: The SLIT3 expression level was increased in Ang II-induced mice models and cardiac fibroblasts. SLIT3 significantly increased migrated cells and α-smooth muscle actin (α-SMA) expression in cardiac fibroblasts. Ang II-induced increases in mRNA expression of collagen I (COL1A1), and collagen III (COL3A1) was attenuated by SLIT3 inhibition. SLIT3 knockdown attenuated the Ang II-induced increase in mRNA expression of ACTA2 (α-SMA), Fibronectin, and CTGF. SLIT3 suppression potentially reduced DHE expression and decreased malondialdehyde (MDA) content, and the superoxide dismutase (SOD) and catalase (CAT) levels were significantly increased in cardiac fibroblasts. Additionally, SLIT3 inhibition markedly decreased RhoA and ROCK1 protein expression, whereas ROCK inhibitor Y-27632 (10 μM) markedly attenuated the migration of cardiac fibroblasts stimulated by Ang II and SLIT3. Conclusion: The results speculate that SLIT3 could significantly regulate cardiac fibrosis and fibroblast differentiation via the RhoA/ROCK1 signaling pathway. [ABSTRACT FROM AUTHOR]

Myocardial infarction is one of the leading causes of mortality globally. Currently, the pleiotropic inflammatory cytokine interleukin-6 (IL-6) is considered to be intimately related to the severity of myocardial injury during myocardial infarction. Interventions targeting IL-6 are a promising therapeutic option for myocardial infarction, but the underlying molecular mechanisms are not well understood. Here, we report the novel role of IL-6 in regulating adverse cardiac remodeling mediated by fibroblasts in a mouse model of myocardial infarction. It was found that the elevated expression of IL-6 in myocardium and cardiac fibroblasts was observed after myocardial infarction. Further, fibroblast-specific knockdown of Il6 significantly attenuated cardiac fibrosis and adverse cardiac remodeling and preserved cardiac function induced by myocardial infarction. Mechanistically, the role of Il6 contributing to cardiac fibrosis depends on signal transduction and activation of transcription (STAT)3 signaling activation. Additionally, Stat3 binds to the Il11 promoter region and contributes to the increased expression of Il11, which exacerbates cardiac fibrosis. In conclusion, these results suggest a novel role for IL-6 derived from fibroblasts in mediating Stat3 activation and substantially augmented Il11 expression in promoting cardiac fibrosis, highlighting its potential as a therapeutic target for cardiac fibrosis. [ABSTRACT FROM AUTHOR]

In heart disease patients, myocyte loss or malfunction invariably leads to fibrosis, involving the activation and accumulation of cardiac fibroblasts that deposit large amounts of extracellular matrix. Apart from the vital replacement fibrosis that follows myocardial infarction, ensuring structural integrity of the heart, cardiac fibrosis is largely considered to be maladaptive. Much work has focused on signaling pathways driving the fibrotic response, including TGF-ß signaling and biomechanical strain. However, currently there are very limited options for reducing cardiac fibrosis, with most patients suffering from chronic fibrosis. The adult heart has very limited regenerative capacity. However, cardiac regeneration has been reported in humans perinatally, and reproduced experimentally in neonatal mice. Furthermore, model organisms such as the zebrafish are able to fully regenerate their hearts following massive cardiac damage into adulthood. Increasing evidence points to a transient immunofibrotic response as being key for cardiac regeneration to occur. The mechanisms at play in this context are changing our views on fibrosis, and could be leveraged to promote beneficial remodeling in heart failure patients. This review summarizes our current knowledge of fibroblast properties associated with the healthy, failing or regenerating heart. Furthermore, we explore how cardiac fibroblast activity could be targeted to assist future therapeutic approaches. [ABSTRACT FROM AUTHOR]

Abstract Aims Transforming growth factor β (TGF‐β) signalling is one of the critical pathways in fibroblast activation, and several drugs targeting the TGF‐β/Smad signalling pathway in heart failure with cardiac fibrosis are being tested in clinical trials. Some caveolins and cavins, which are components of caveolae on the plasma membrane, are known for their association with the regulation of TGF‐β signalling. Cavin‐2 is particularly abundant in fibroblasts; however, the detailed association between Cavin‐2 and cardiac fibrosis is still unclear. We tried to clarify the involvement and role of Cavin‐2 in fibroblasts and cardiac fibrosis. Methods and results To clarify the role of Cavin‐2 in cardiac fibrosis, we performed transverse aortic constriction (TAC) operations on four types of mice: wild‐type (WT), Cavin‐2 null (Cavin‐2 KO), Cavin‐2flox/flox, and activated fibroblast‐specific Cavin‐2 conditional knockout (Postn‐Cre/Cavin‐2flox/flox, Cavin‐2 cKO) mice. We collected mouse embryonic fibroblasts (MEFs) from WT and Cavin‐2 KO mice and investigated the effect of Cavin‐2 in fibroblast trans‐differentiation into myofibroblasts and associated TGF‐β signalling. Four weeks after TAC, cardiac fibrotic areas in both the Cavin‐2 KO and the Cavin‐2 cKO mice were significantly decreased compared with each control group (WT 8.04 ± 1.58% vs. Cavin‐2 KO 0.40 ± 0.03%, P

Altered cardiac innate immunity is highly associated with the progression of cardiac disease states and heart failure. S100A8/A9 is an important component of damage-associated molecular patterns (DAMPs) that is critically involved in the pathogenesis of heart failure, thus considered a promising target for pharmacological intervention. In the current study, initially, we validated the role of S100A8/A9 in contributing to cardiac injury and heart failure via the overactivation of the β-adrenergic pathway and tested the potential use of paquinimod as a pharmacological intervention of S100A8/A9 activation in preventing cardiac dysfunction, collagen deposition, inflammation, and immune cell infiltration in β-adrenergic overactivation–mediated heart failure. This finding was further confirmed by the cardiomyocyte-specific silencing of S100A9 via the use of the adeno-associated virus (AAV) 9-mediated short hairpin RNA (shRNA) gene silencing system. Most importantly, in the assessment of the underlying cellular mechanism by which activated S100A8/A9 cause aggravated progression of cardiac fibrosis and heart failure, we discovered that the activated S100A8/A9 can promote fibroblast-macrophage interaction, independent of inflammation, which is likely a key mechanism leading to the enhanced collagen production. Our results revealed that targeting S100A9 provides dual beneficial effects, which is not only a strategy to counteract cardiac inflammation but also preclude cardiac fibroblast-macrophage interactions. The findings of this study also indicate that targeting S100A9 could be a promising strategy for addressing cardiac fibrosis, potentially leading to future drug development. [ABSTRACT FROM AUTHOR]

Cardiac fibrosis, the hallmark of cardiovascular disease, is characterized by excessive deposition of extracellular matrix in the heart. Emerging evidence indicates that cardiac fibroblasts (CFs) play pivotal roles in driving cardiac fibrosis. However, due to incomplete insights into CFs, there are limited effective approaches to prevent or reverse cardiac fibrosis currently. Palmatine, a protoberberine alkaloid extracted from traditional Chinese botanical remedies, possesses diverse biological effects. This study investigated the potential therapeutic value and mechanism of palmatine against cardiac fibrosis. Adult male C57BL/6 mice were treated with vehicle, isoproterenol (ISO), or ISO plus palmatine for one week. After echocardiography assessment, mice hearts were collected for histopathology, real-time polymerase chain reaction, and Western blot analyses. Primary rat CFs were utilized in vitro. Compared to control, ISO-treated mice exhibited cardiac hypertrophy and structural abnormalities; however, treatment with palmatine ameliorated these effects of ISO. Moreover, palmatine treatment mitigated ISO-induced cardiac fibrosis. Network pharmacology and molecular docking analysis showed that palmatine strongly binds the regulators of cardiac fibrosis including signal transducer and activator of transcription 3 (STAT3) and mammalian target of rapamycin. Furthermore, palmatine reduced the elevated fibrotic factor expressions and overactivated STAT3 induced by ISO, Transformed growth factor β1 (TGF-β1), or interleukin-6 both in vivo and in vitro. Additionally, blocking STAT3 suppressed the TGF-β1-induced CF activation. Collectively, these data demonstrated that palmatine attenuated cardiac fibrosis partly by inhibiting fibroblast activation through the STAT3 pathway. This provides an experimental basis for the clinical treatment of cardiac fibrosis with palmatine. Palmatine ameliorates abnormal heart structure and fibrosis induced by ISO. STAT3 participates in the activation of cardiac fibroblasts. Palmatine inhibits the activation of cardiac fibroblast via the STAT3 signaling pathway. [ABSTRACT FROM AUTHOR]