scholarly journals Mechanisms of Fibroblast Activation and Myocardial Fibrosis: Lessons Learned from FB-Specific Conditional Mouse Models

Cells ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 2412
Author(s):  
Prachi Umbarkar ◽  
Suma Ejantkar ◽  
Sultan Tousif ◽  
Hind Lal
Keyword(s):  
Myocardial Fibrosis ◽  
Mouse Models ◽  
Cardiac Fibrosis ◽  
Tissue Injury ◽  
Lessons Learned ◽  
Cardiac Diseases ◽  
Hippo Signaling ◽  
Fibroblast Activation ◽  
Extracellular Matrix Components

Heart failure (HF) is a leading cause of morbidity and mortality across the world. Cardiac fibrosis is associated with HF progression. Fibrosis is characterized by the excessive accumulation of extracellular matrix components. This is a physiological response to tissue injury. However, uncontrolled fibrosis leads to adverse cardiac remodeling and contributes significantly to cardiac dysfunction. Fibroblasts (FBs) are the primary drivers of myocardial fibrosis. However, until recently, FBs were thought to play a secondary role in cardiac pathophysiology. This review article will present the evolving story of fibroblast biology and fibrosis in cardiac diseases, emphasizing their recent shift from a supporting to a leading role in our understanding of the pathogenesis of cardiac diseases. Indeed, this story only became possible because of the emergence of FB-specific mouse models. This study includes an update on the advancements in the generation of FB-specific mouse models. Regarding the underlying mechanisms of myocardial fibrosis, we will focus on the pathways that have been validated using FB-specific, in vivo mouse models. These pathways include the TGF-β/SMAD3, p38 MAPK, Wnt/β-Catenin, G-protein-coupled receptor kinase (GRK), and Hippo signaling. A better understanding of the mechanisms underlying fibroblast activation and fibrosis may provide a novel therapeutic target for the management of adverse fibrotic remodeling in the diseased heart.

scholarly journals Oncofetal Protein CRIPTO Is Involved in Wound Healing and Fibrogenesis in the Regenerating Liver and Is Associated with the Initial Stages of Cardiac Fibrosis

Cells ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 3325
Author(s):  
Sofia Karkampouna ◽  
Danny van der Helm ◽  
Mario Scarpa ◽  
Bart van Hoek ◽  
Hein W. Verspaget ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Mouse Models ◽  
Cardiac Fibrosis ◽  
Ex Vivo ◽  
Pressure Overload ◽  
Collagen Type I ◽  
Type I ◽  
Liver Fibrogenesis ◽  
End Stage

Oncofetal protein, CRIPTO, is silenced during homeostatic postnatal life and often re-expressed in different neoplastic processes, such as hepatocellular carcinoma. Given the reactivation of CRIPTO in pathological conditions reported in various adult tissues, the aim of this study was to explore whether CRIPTO is expressed during liver fibrogenesis and whether this is related to the disease severity and pathogenesis of fibrogenesis. Furthermore, we aimed to identify the impact of CRIPTO expression on fibrogenesis in organs with high versus low regenerative capacity, represented by murine liver fibrogenesis and adult murine heart fibrogenesis. Circulating CRIPTO levels were measured in plasma samples of patients with cirrhosis registered at the waitlist for liver transplantation (LT) and 1 year after LT. The expression of CRIPTO and fibrotic markers (αSMA, collagen type I) was determined in human liver tissues of patients with cirrhosis (on a basis of viral hepatitis or alcoholic disease), in cardiac tissue samples of patients with end-stage heart failure, and in mice with experimental liver and heart fibrosis using immuno-histochemical stainings and qPCR. Mouse models with experimental chronic liver fibrosis, induced with multiple shots of carbon tetrachloride (CCl4) and acute liver fibrosis (one shot of CCl4), were evaluated for CRIPTO expression and fibrotic markers. CRIPTO was overexpressed in vivo (Adenoviral delivery) or functionally sequestered by ALK4Fc ligand trap in the acute liver fibrosis mouse model. Murine heart tissues were evaluated for CRIPTO and fibrotic markers in three models of heart injury following myocardial infarction, pressure overload, and ex vivo induced fibrosis. Patients with end-stage liver cirrhosis showed elevated CRIPTO levels in plasma, which decreased 1 year after LT. Cripto expression was observed in fibrotic tissues of patients with end-stage liver cirrhosis and in patients with heart failure. The expression of CRIPTO in the liver was found specifically in the hepatocytes and was positively correlated with the Model for End-stage Liver Disease (MELD) score for end-stage liver disease. CRIPTO expression in the samples of cardiac fibrosis was limited and mostly observed in the interstitial cells. In the chronic and acute mouse models of liver fibrosis, CRIPTO-positive cells were observed in damaged liver areas around the central vein, which preceded the expression of αSMA-positive stellate cells, i.e., mediators of fibrosis. In the chronic mouse models, the fibrosis and CRIPTO expression were still present after 11 weeks, whereas in the acute model the liver regenerated and the fibrosis and CRIPTO expression resolved. In vivo overexpression of CRIPTO in this model led to an increase in fibrotic markers, while blockage of CRIPTO secreted function inhibited the extent of fibrotic areas and marker expression (αSMA, Collagen type I and III) and induced higher proliferation of residual healthy hepatocytes. CRIPTO expression was also upregulated in several mouse models of cardiac fibrosis. During myocardial infarction CRIPTO is upregulated initially in cardiac interstitial cells, followed by expression in αSMA-positive myofibroblasts throughout the infarct area. After the scar formation, CRIPTO expression decreased concomitantly with the αSMA expression. Temporal expression of CRIPTO in αSMA-positive myofibroblasts was also observed surrounding the coronary arteries in the pressure overload model of cardiac fibrosis. Furthermore, CRIPTO expression was upregulated in interstitial myofibroblasts in hearts cultured in an ex vivo model for cardiac fibrosis. Our results are indicative for a functional role of CRIPTO in the induction of fibrogenesis as well as a potential target in the antifibrotic treatments and stimulation of tissue regeneration.

scholarly journals Androgen Receptor (AR) Pathophysiological Roles in Androgen Related Diseases in Skin, Metabolism Syndrome, Bone/Muscle and Neuron/Immune Systems: Lessons Learned from Mice Lacking AR in Specific Cells

2013 ◽  
Vol 11 (1) ◽  
pp. nrs.11001 ◽  
Author(s):  
Chawnshang Chang ◽  
Shuyuan Yeh ◽  
Soo Ok Lee ◽  
Ta-min Chang
Keyword(s):  
Androgen Receptor ◽  
Mouse Models ◽  
Bone Mineralization ◽  
Muscle Growth ◽  
Lessons Learned ◽  
Specific Cell ◽  
Cell Type ◽  
Vast Number ◽  
Tissue Specific

The androgen receptor (AR) is expressed ubiquitously and plays a variety of roles in a vast number of physiological and pathophysiological processes. Recent studies of AR knockout (ARKO) mouse models, particularly the cell type- or tissue-specific ARKO models, have uncovered many AR cell type- or tissue-specific pathophysiological roles in mice, which otherwise would not be delineated from conventional castration and androgen insensitivity syndrome studies. Thus, the AR in various specific cell types plays pivotal roles in production and maturation of immune cells, bone mineralization, and muscle growth. In metabolism, the ARs in brain, particularly in the hypothalamus, and the liver appear to participate in regulation of insulin sensitivity and glucose homeostasis. The AR also plays key roles in cutaneous wound healing and cardiovascular diseases, including atherosclerosis and abdominal aortic aneurysm. This article will discuss the results obtained from the total, cell type-, or tissue-specific ARKO models. The understanding of AR cell type- or tissue-specific physiological and pathophysiological roles using these in vivo mouse models will provide useful information in uncovering AR roles in humans and eventually help us to develop better therapies via targeting the AR or its downstream signaling molecules to combat androgens/AR-related diseases.

Curcumin Management of Myocardial Fibrosis and its Mechanisms of Action: A Review

2019 ◽  
Vol 47 (08) ◽  
pp. 1675-1710 ◽  
Author(s):  
Yonghui Yu ◽  
Jinghui Sun ◽  
Ru Wang ◽  
Jiangang Liu ◽  
Peili Wang ◽  
...  
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Cardiac Injury ◽  
Volume Overload ◽  
Curcumin Analogs ◽  
Natural Polyphenol ◽  
Anti Oxidant ◽  
Nlrp3 Inflammasomes

Myocardial fibrosis is implicated as a leading risk factor for heart failure, arrhythmia, and sudden death after cardiac injury, as the excessive interstitial extracellular matrix impedes heart contraction and electrical conduction. Complicated mechanisms involving oxidative stress, pro-inflammatory cytokines, chemokine families, NLRP3 inflammasomes, growth factors, and non-coding RNAs participate in cardiac fibrogenesis and make it difficult to designate specific and effective therapies. Oriental herbs have been popular for thousands of years in the health care of Asian residents, due to their multi-targeted, multi-faceted approaches and their multi-functional effects in fighting difficult and complicated diseases, including cardiovascular disorders such as myocardial fibrosis. Curcumin, a natural polyphenol and yellow pigment obtained from the spice turmeric, was found to have strong anti-oxidant and anti-inflammatory properties. Increasing evidence has shown that curcumin can be used to prevent and treat myocardial fibrosis, when the myocardium suffers pathological pro-fibrotic changes in vivo and in vitro. The present review focuses on recent studies elucidating the mechanisms of curcumin in treating different pathologic conditions, including ischemia, hypoxia/reoxygenation, pressure or volume overload, and hyperglycemia or high-fat-induced cardiac fibrosis. Novel analogs such as C66, B2BrBC, Y20, and J17 have been designed to maximize the therapeutic potentials of curcumin. These optimized curcumin analogs with improved bioavailability and pharmacokinetic profiles need to be clinically verified before curcumin could be recommended for the treatment of myocardial fibrosis.

scholarly journals Mir-21 Promotes Cardiac Fibrosis After Myocardial Infarction Via Targeting Smad7

2017 ◽  
Vol 42 (6) ◽  
pp. 2207-2219 ◽  
Author(s):  
Jinxia Yuan ◽  
Hongtao Chen ◽  
Dawei Ge ◽  
Yu Xu ◽  
Haihua Xu ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Critical Role ◽  
Luciferase Reporter Assay ◽  
In Vivo Study ◽  
Luciferase Reporter ◽  
Reporter Assay ◽  
Tgf Β1

Background/Aims: Cardiac fibrosis after myocardial infarction (MI) has been identified as an important factor in the deterioration of heart function. Previous studies have demonstrated that miR-21 plays an important role in various pathophysiological processes in the heart. However, the role of miR-21 in fibrosis regulation after MI remains unclear. Methods: To induce cardiac infarction, the left anterior descending coronary artery was permanently ligated of mice. First, we explored the expression of miR-21 in the infarcted zone in mice model of MI via RT-qPCR. Next, we examined the effects of TGF-β1 on miR-21 expression in cardiac fibroblasts (CFs). Then, CFs were infected with miR-21 mimics or miR-21 inhibitors to investigate the effects of miR-21 on the process of CFs activation in vitro. Further, bioinformatics analysis and luciferase reporter assay were performed to identify and validate the target gene of miR-21. At last, in-vivo study was done to confirm MiR-21 regulated myocardial fibrosis after MI in mice. Results: MiR-21 was up-regulated in the infarcted zone after MI in vivo. TGF-β1 treatment increased miR-21 expression in CFs. Overexpression of miR-21 promoted the effects of TGF-β1-induced activation of CFs, evidenced by increased expression of Col-1, α-SMA and F-actin, whereas inhibition of miR-21 attenuated the process of fibrosis. Bioinformatics, Western blot analysis and luciferase reporter assay demonstrated that Smad7 is a direct target of miR-21. In addition, in-vivo study revealed that MiR-21 regulated myocardial fibrosis after MI in mice. Conclusion: These findings suggested that miR-21 has a critical role in CF activation and cardiac fibrosis after MI through via TGF-β/Smad7 signaling pathway. Thus, miR-21 promises to be a potential therapy in treatment of cardiac fibrosis after MI.

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

2016 ◽  
Vol 38 (5) ◽  
pp. 1928-1938 ◽  
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.

scholarly journals Intermittent hypoxia mediated by TSP1 dependent on STAT3 induces cardiac fibroblast activation and cardiac fibrosis

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Qiankun Bao ◽  
Bangying Zhang ◽  
Ya Suo ◽  
Chen Liu ◽  
Qian Yang ◽  
...  
Keyword(s):  
Intermittent Hypoxia ◽  
Cardiac Fibrosis ◽  
Synergistic Effects ◽  
Cardiac Fibroblasts ◽  
Cardiac Fibroblast ◽  
Ang Ii ◽  
Fibroblast Activation ◽  
Obstructive Sleep

Intermittent hypoxia (IH) is the predominant pathophysiological disturbance in obstructive sleep apnea (OSA), known to be independently associated with cardiovascular diseases. However, the effect of IH on cardiac fibrosis and molecular events involved in this process are unclear. Here, we tested IH in angiotensin II (Ang II)-induced cardiac fibrosis and signaling linked to fibroblast activation. IH triggered cardiac fibrosis and aggravated Ang II-induced cardiac dysfunction in mice. Plasma thrombospondin-1 (TSP1) content was upregulated in both IH-exposed mice and OSA patients. Moreover, both in vivo and in vitro results showed IH-induced cardiac fibroblast activation and increased TSP1 expression in cardiac fibroblasts. Mechanistically, phosphorylation of STAT3 at Tyr705 mediated the IH-induced TSP1 expression and fibroblast activation. Finally, STAT3 inhibitor S3I-201 or AAV9 carrying a periostin promoter driving the expression of shRNA targeting Stat3 significantly attenuated the synergistic effects of IH and Ang II on cardiac fibrosis in mice. This work suggests a potential therapeutic strategy for OSA-related fibrotic heart disease.

scholarly journals BMP7 Ameliorated Viral Myocardial Fibrosis by Inhibiting Endothelial-to-Mesenchymal Transition

2020 ◽  
Author(s):  
Yeqing Xie ◽  
Jianquan Liao ◽  
Yong Yu ◽  
Ruizhen Chen ◽  
Xiaoqiang Ding
Keyword(s):  
Myocardial Fibrosis ◽  
Cardiac Fibrosis ◽  
Viral Myocarditis ◽  
Myocardial Damage ◽  
Mrna Levels ◽  
Mesenchymal Transition ◽  
Protein Levels ◽  
Cvb3 Infection ◽  
Endothelial To Mesenchymal Transition

Abstract Background: Endothelial-to-mesenchymal transition (Endo-MT) is associated with myocardial fibrosis in dilated cardiomyopathy (DCM). Endothelial-to-mesenchymal transition (Endo-MT) is induced by coxsackievirus B3 (CVB3) in cardiac microvascular endothelial cells (CMVECs). Bone morphogenetic protein 7 (BMP7) significantly inhibits Endo-MT and the progression of cardiac fibrosis. The study was aimed to investigate the effect and the underlying mechanism of BMP7 on Endo-MT in myocardial fibrosis induce by CVB3 infection in vivo. Methods: BALB/c mice were intraperitoneally injected by CVB3 to induce viral myocarditis (VMC). Mice were treated with BMP7 after CVB3 infection. Subsequently, all groups of mice were determined by echocardiography, histopathologic and molecular detection. Results: We found that the ratio of BMP7/TGF-β1 in mRNA levels was decreased obviously at different time points after CVB3 injection. BMP7 facilitated the recovery of cardiac function after CVB3 infection via inhibition of myocardial damage, collagen deposition. Double immunofluorescence staining indicated that Endo-MT was implicated in CVB3-induced myocardial fibrosis, which was attenuated by BMP7. The protein levels of pSmad3 and Smad4 were significantly upregulated in VMC group, as well as Wnt/β-catenin and the transcription factor snail. BMP7 treatment reversed the changes of these protein levels. Moreover, CO-IP demonstrated the crosstalk between β-catenin and Smad3 in VMC mice, which was downregulated by BMP7 treatment. Conclusions: These results indicated that BMP7 obviously ameliorated myocardial fibrosis in CVB3-infected mice via Endo-MT, which was involved in the TGF-β/Smad and Wnt/β-catenin pathway. β-Catenin/Smad3 interaction may be associated with Endo-MT in the development of viral myocardial fibrosis.

scholarly journals Adiponectin Modified BMSCs Alleviate Heart Fibrosis via Inhibition TGF-beta1/Smad in Diabetic Rats

2021 ◽  
Vol 9 ◽  
Author(s):  
Ke Meng ◽  
Huabo Cai ◽  
Simin Cai ◽  
Yucai Hong ◽  
Xiaoming Zhang
Keyword(s):  
Myocardial Fibrosis ◽  
High Glucose ◽  
Cardiac Fibrosis ◽  
Therapeutic Potential ◽  
Heart Diseases ◽  
Diabetic Rats ◽  
Diabetic Rat ◽  
H9c2 Cells

Background: Accumulating evidence suggested that bone marrow mesenchymal stem cells (BMSCs) have therapeutic potential for diabetes and heart diseases. However, the effects of BMSC on reducing myocardial fibrosis need to be optimized. This study aimed to investigate the mechanism of adiponectin (APN) modified BMSCs on myocardial fibrosis in diabetic model in vivo and in vitro.Methods: The high-fat diet combined with streptozotocin (STZ) injection were used to induced diabetic rat model. H9c2 cells were cultured under a high glucose medium as in vitro model. The BMSCs were modified by APN plasmid or APN small interfering RNA (siRNA), then transplanted to the diabetic rats by a single tail-vein injection, or co-cultured with H9c2 cells.Results: We demonstrated that diabetic rats showed typical diabetic symptoms, such as decreased cardiac function, accumulation of pathological lesions and collagen expression. However, these impairments were significantly prevented by the APN modified BMSCs treatment while no effects on APN siRNA modified BMSCs treated diabetic rats. Moreover, we confirmed that APN modified BMSCs could attenuate the expression of TGF-beta1/smad to suppress the myocardial fibrosis in the diabetic rats and high glucose induced H9c2 cells.Conclusion: The present results for the first time showed that APN modified BMSCs exerted protection on cardiac fibrosis via inhibiting TGF-beta1/smad signal pathway in diabetic rats. Our findings suggested that APN modified BMSCs might be a novel and optimal therapy for the diabetic cardiomyopathy in future.

scholarly journals Interleukin-9 in Immunopathology of Trypanosoma cruzi Experimental Infection

2021 ◽  
Vol 11 ◽  
Author(s):  
Nadjania Saraiva de Lira Silva ◽  
Cristina Mary Orikaza ◽  
Fabiana Rodrigues de Santana ◽  
Luana Aguiar dos Santos ◽  
Bruno Ramos Salu ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Chagas Disease ◽  
Experimental Infection ◽  
Cardiac Fibrosis ◽  
Tissue Injury ◽  
Chronic Phase ◽  
Protective Role

Chagas’ disease is a parasitosis caused by Trypanosoma cruzi, which affects approximately 8 million people worldwide. The balance between pro- and anti-inflammatory cytokines produced during immunological responses contributes to disease prognosis and progression. Parasite tissue persistence can induce chronic inflammatory stimuli, which can cause long-term tissue injury and fibrosis. Chronic Chagas’ patients exhibit increased levels of interleukin (IL)-9, an important cytokine in the regulation of inflammatory and fibrogenic processes. Data on the role of IL-9 in other pathologies are sometimes contradictory, and few studies have explored this cytokine’s influence in Chagas’ disease pathology. Hence, the aim of this study was to evaluate the role of IL-9 in the progression of T. cruzi infection in vivo and in vitro. In vitro infection demonstrated that IL-9 reduced the number of infected cells and decreased the multiplication of intracellular amastigotes in both C2C12 myoblasts and bone marrow-derived macrophages. In myoblasts, the increased production of nitric oxide (NO) was essential for reduced parasite multiplication, whereas macrophage responses resulted in increased IL-6 and reduced TGF-β levels, indicating that parasite growth restriction mechanisms induced by IL-9 were cell-type specific. Experimental infection of BALB/c mice with T. cruzi trypomastigotes of the Y strain implicated a major role of IL-9 during the chronic phase, as increased Th9 and Tc9 cells were detected among splenocytes; higher levels of IL-9 in these cell populations and increased cardiac IL-9 levels were detected compared to those of uninfected mice. Moreover, rIL9 treatment decreased serum IL-12, IL-6, and IL-10 levels and cardiac TNF-α levels, possibly attempting to control the inflammatory response. IL-9 neutralization increased cardiac fibrosis, synthesis of collagens I and III, and mastocyte recruitment in BALB/c heart tissue during the chronic phase. In conclusion, our data showed that IL-9 reduced the invasion and multiplication of T. cruzi in vitro, in both myoblasts and macrophages, favoring disease control through cell-specific mechanisms. In vivo, IL-9 was elevated during experimental chronic infection in BALB/c mice, and this cytokine played a protective role in the immunopathological response during this phase by controlling cardiac fibrosis and proinflammatory cytokine production.

scholarly journals Oncofetal protein CRIPTO regulates wound healing and fibrogenesis in regenerating liver and is associated with the initial stages of cardiac fibrosis

2021 ◽  
Author(s):  
Sofia Karkampouna ◽  
Danny van der Helm ◽  
Bart van Hoek ◽  
Hein W Verspaget ◽  
Marie Jose TH Goumans ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Mouse Models ◽  
Cardiac Fibrosis ◽  
Ex Vivo ◽  
Pressure Overload ◽  
Liver Fibrogenesis ◽  
One Year ◽  
End Stage

Background: Oncofetal protein, Cripto, is silenced during postnatal life and often re-expressed in different neoplastic processes. In the present study we investigated the potential role of Cripto in hepatic and cardiac fibrosis. In this study, the aim was to explore whether Cripto is expressed during liver fibrogenesis and whether this is related to the disease severity and pathogenesis of fibrogenesis. Furthermore, we aimed to identify the impact of Cripto expression on fibrogenesis in organs with high versus low regenerative capacity, represented by murine liver fibrogenesis and adult murine heart fibrogenesis. Methods: Circulating CRIPTO levels were measured in plasma samples of patients with cirrhosis registered at the waitlist for liver transplantation (LT) and one year after LT. The expression of Cripto and fibrotic markers (aSMA, collagen I) were determined in human liver tissues of patients with cirrhosis (on a basis of viral hepatitis or alcoholic disease), in cardiac tissue samples of patients with end-stage heart failure and of mice with experimental liver and heart fibrosis using immuno-histochemical stainings and qPCR. Mouse models with experimental chronic liver fibrosis, induced with multiple shots of carbon tetrachloride (CCl4) and acute liver fibrosis (one shot of CCl4) were evaluated for Cripto expression and fibrotic markers. Cripto was overexpressed in vivo (Adenoviral delivery) or functionally sequestered by ALK4Fc ligand trap in the acute liver fibrosis mouse model. Murine heart tissues were evaluated for Cripto and fibrotic markers, in three models of heart injury; following myocardial infarction, pressure overload and ex vivo induced fibrosis. Results: Patients with end-stage liver cirrhosis showed elevated Cripto levels in plasma, which had decreased one year after LT. Cripto expression was observed in fibrotic tissues of patients with end-stage liver cirrhosis and in patients with heart failure. The expression of Cripto in the liver was found specifically in the hepatocytes and was positively correlated with the Model for End-stage Liver Disease (MELD) score for end-stage liver disease. Cripto expression in the samples of cardiac fibrosis was limited and mostly observed in the interstitial cells. In the chronic and acute mouse models of liver fibrosis, Cripto-positve cells were observed in damaged liver areas around the central vein, which preceded the expression of aSMA-positive stellate cells, i.e. mediators of fibrosis. Whereas in the chronic mouse models the fibrosis and Cripto expression was still present after 11 weeks, in the acute model the liver regenerated and the fibrosis and Cripto expression resolved. In vivo overexpression of Cripto in this model, led to an increase in fibrotic markers while blockage of Cripto secreted function inhibited the extend of fibrotic areas and marker expression (αSMA, Collagen type I and III) and induced higher proliferation of residual healthy hepatocytes. Cripto expression was also upregulated in several mouse models of cardiac fibrosis. During myocardial infarction Cripto is upregulated initially in cardiac interstitial cells, followed by expression in αSMA-positive myofibroblasts throughout the infarct area. After the scar formation, Cripto expression decreased concomitantly with the aSMA expression. Temporal expression of Cripto in αSMA-positive myofibroblasts was also observed surrounding the coronary arteries in the pressure overload model of cardiac fibrosis. Furthermore, Cripto expression was upregulated in interstitial myofibroblasts in hearts cultured in an ex vivo model for cardiac fibrosis. Conclusion: Our results are indicative for a functional role of Cripto in induction of fibrogenesis and potential applications in antifibrotic treatments and stimulation of tissue regeneration.

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