scholarly journals Jiedu Tongluo Decoction Attenuates Myocardial Fibrosis Through Inhibition of The TGF-β1/Smad2/3 Pathway

2021 ◽  
Author(s):  
Ju Hui ◽  
Le Li ◽  
Huan Zhang ◽  
Bin Wang ◽  
Jiaxin Shi ◽  
...  
Keyword(s):  
Chinese Medicine ◽  
Growth Factor ◽  
Myocardial Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Myocardial Tissue ◽  
Type Iii Collagen ◽  
Isoproterenol Hydrochloride ◽  
Tgf Β1 ◽  
Masson Staining

Abstract Background: Jiedu Tongluo (JDTL) decoction is a traditional Chinese medicine (TCM) formula containing three herbal ingredients that is widely used to treat myocardial fibrosis (MF). This study aimed to investigate the molecular mechanism of action of JDTL decoction for the treatment of MF. Methods: The chemical constituents of Traditional Chinese medicine were analyzed by HPLC, Forty Wistar rats were randomly divided into normal control group, model group, Jiedu Tongluo decoction low-dose and high-dose treatment groups, with 10 rats in each group. Rat myocardial fibrosis model was induced by subcutaneous injection of 5 mg•kg-1 isoproterenol hydrochloride. 5 and 50 g•(kg.d) -1 were given intragastric administration for 7 days. Hematoxylin-eosin (HE) and Masson staining were performed for histological evaluation of myocardial tissue, MTT method detects the activity of cardiac fibroblasts, The alkaline water method is used to determine the content of hydroxyproline in myocardial tissue and myocardial fibroblasts, Immunohistochemical method was used to detect the expression of myocardial tissue transforming growth factor (TGF) β1, p smad2/3, and type III collagen (Col III) expression, Immunofluorescence to detect the expression of α-SMA/Vimentin in myocardial tissue and myocardial fibroblasts; Western blot was used to detect the protein expression levels of TGFβ1 and psmad2/3 in cardiac fibroblasts.Results: In this study, six compounds of JDTL decoction were identified by high-performance liquid chromatography (HPLC). Hematoxylin and eosin (HE) and Masson staining showed that in the isoproterenol hydrochloride (ISO)-induced MF rat model, JDTL treatment protected the myocardial structure and inhibited type III collagen (COL3) expression(p<0.05). The immunohistochemistry (IHC) results also showed that JDTL treatment significantly reduced the expression of vimentin, alpha-smooth muscle actin (α-SMA), and transforming growth factor-beta 1 (TGF-β1) as well as the phosphorylation of Smad2/3 in the rat MF model(p<0.05). Rat cardiac fibroblasts (RCFs) were used for the following assays performed in vitro: hydroxyproline detection, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, wound-healing test, western blot, and double immunofluorescence staining; the results of these assays confirmed the inhibitory effects of JDTL decoction on the proliferation, migration, and transdifferentiation abilities of RCFs as well as the molecular mechanisms underlying these effects, including the inhibition of the TGF-β1/Smad2/3 pathway through downregulation of TGF-β1 expression and phosphorylation of Smad2/3 as well as inhibition of vimentin and α-SMA expression(p<0.05). Conclusion: JDTL decoction can prolong the process of MF through inhibition of the TGF-β1/Smad2/3 signaling pathway.

scholarly journals Featured Article: TGF-β1 dominates extracellular matrix rigidity for inducing differentiation of human cardiac fibroblasts to myofibroblasts

2018 ◽  
Vol 243 (7) ◽  
pp. 601-612 ◽  
Author(s):  
Nathan Cho ◽  
Shadi E Razipour ◽  
Megan L McCain
Keyword(s):  
Extracellular Matrix ◽  
Growth Factor ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Matrix Rigidity ◽  
Growth Factor Beta ◽  
Human Cardiac Fibroblasts ◽  
Tgf Β1

Cardiac fibroblasts and their activated derivatives, myofibroblasts, play a critical role in wound healing after myocardial injury and often contribute to long-term pathological outcomes, such as excessive fibrosis. Thus, defining the microenvironmental factors that regulate the phenotype of cardiac fibroblasts and myofibroblasts could lead to new therapeutic strategies. Both chemical and biomechanical cues have previously been shown to induce myofibroblast differentiation in many organs and species. For example, transforming growth factor beta 1, a cytokine secreted by neutrophils, and rigid extracellular matrix environments have both been shown to promote differentiation. However, the relative contributions of transforming growth factor beta 1 and extracellular matrix rigidity, two hallmark cues in many pathological myocardial microenvironments, to the phenotype of human cardiac fibroblasts are unclear. We hypothesized that transforming growth factor beta 1 and rigid extracellular matrix environments would potentially have a synergistic effect on the differentiation of human cardiac fibroblasts to myofibroblasts. To test this, we seeded primary human adult cardiac fibroblasts onto coverslips coated with polydimethylsiloxane of various elastic moduli, introduced transforming growth factor beta 1, and longitudinally quantified cell phenotype by measuring expression of α-smooth muscle actin, the most robust indicator of myofibroblasts. Our data indicate that, although extracellular matrix rigidity influenced differentiation after one day of transforming growth factor beta 1 treatment, ultimately transforming growth factor beta 1 superseded extracellular matrix rigidity as the primary regulator of myofibroblast differentiation. We also measured expression of POSTN, FAP, and FSP1, proposed secondary indicators of fibroblast/myofibroblast phenotypes. Although these genes partially trended with α-smooth muscle actin expression, they were relatively inconsistent. Finally, we demonstrated that activated myofibroblasts incompletely revert to a fibroblast phenotype after they are re-plated onto new surfaces without transforming growth factor beta 1, suggesting differentiation is partially reversible. Our results provide new insights into how microenvironmental cues affect human cardiac fibroblast differentiation in the context of myocardial pathology, which is important for identifying effective therapeutic targets and dictating supporting cell phenotypes for engineered human cardiac disease models. Impact statement Heart disease is the leading cause of death worldwide. Many forms of heart disease are associated with fibrosis, which increases extracellular matrix (ECM) rigidity and compromises cardiac output. Fibrotic tissue is synthesized primarily by myofibroblasts differentiated from fibroblasts. Thus, defining the cues that regulate myofibroblast differentiation is important for understanding the mechanisms of fibrosis. However, previous studies have focused on non-human cardiac fibroblasts and have not tested combinations of chemical and mechanical cues. We tested the effects of TGF-β1, a cytokine secreted by immune cells after injury, and ECM rigidity on the differentiation of human cardiac fibroblasts to myofibroblasts. Our results indicate that differentiation is initially influenced by ECM rigidity, but is ultimately superseded by TGF-β1. This suggests that targeting TGF-β signaling pathways in cardiac fibroblasts may have therapeutic potential for attenuating fibrosis, even in rigid microenvironments. Additionally, our approach can be leveraged to engineer more precise multi-cellular human cardiac tissue models.

Upregulation of α8β1-integrin in cardiac fibroblast by angiotensin II and transforming growth factor-β1

2001 ◽  
Vol 281 (5) ◽  
pp. C1457-C1467 ◽  
Author(s):  
Gaétan Thibault ◽  
Marie-Josée Lacombe ◽  
Lynn M. Schnapp ◽  
Alexandre Lacasse ◽  
Fatiha Bouzeghrane ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Growth Factor ◽  
Matrix Protein ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Transforming Growth Factor Β1 ◽  
Cardiac Fibroblast ◽  
Extracellular Matrix Protein ◽  
Ang Ii ◽  
Tgf Β1

Using a novel pharmacological tool with125I-echistatin to detect integrins on the cell, we have observed that cardiac fibroblasts harbor five different RGD-binding integrins: α8β1, α3β1, α5β1, αvβ1, and αvβ3. Stimulation of cardiac fibroblasts by angiotensin II (ANG II) or transforming growth factor-β1 (TGF-β1) resulted in an increase of protein and heightening by 50% of the receptor density of α8β1-integrin. The effect of ANG II was blocked by an AT1, but not an AT2, receptor antagonist, or by an anti-TGF-β1 antibody. ANG II and TGF-β1 increased fibronectin secretion, smooth muscle α-actin synthesis, and formation of actin stress fibers and enhanced attachment of fibroblasts to a fibronectin matrix. The α8- and β1-subunits were colocalized by immunocytochemistry with vinculin or β3-integrin at focal adhesion sites. These results indicate that α8β1-integrin is an abundant integrin on rat cardiac fibroblasts. Its positive modulation by ANG II and TGF-β1 in a myofibroblast-like phenotype suggests the involvement of α8β1-integrin in extracellular matrix protein deposition and cardiac fibroblast adhesion.

scholarly journals Ac-SDKP inhibits transforming growth factor-β1-induced differentiation of human cardiac fibroblasts into myofibroblasts

2010 ◽  
Vol 298 (5) ◽  
pp. H1357-H1364 ◽  
Author(s):  
Hongmei Peng ◽  
Oscar A. Carretero ◽  
Edward L. Peterson ◽  
Nour-Eddine Rhaleb
Keyword(s):  
Cell Proliferation ◽  
Smooth Muscle ◽  
Growth Factor ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Transforming Growth Factor Β1 ◽  
Collagen Production ◽  
Smad7 Expression ◽  
Human Cardiac Fibroblasts ◽  
Tgf Β1

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) inhibits collagen production and cell proliferation in cultured rat cardiac fibroblasts, but its effect on differentiation of fibroblasts into myofibroblasts is not known. High amounts of transforming growth factor-β1 (TGF-β1) have been found in fibrotic cardiac tissue. TGF-β1 converts fibroblasts into myofibroblasts, which produce more extracellular matrix proteins than fibroblasts. We hypothesized that 1) Ac-SDKP inhibits TGF-β1-induced differentiation of fibroblasts into myofibroblasts; and 2) this effect is mediated in part by blocking phosphorylation of small-mothers-against-decapentaplegic (Smad) 2 and extracellular signal-regulated kinase (ERK) 1/2. For this study, we used human fetal cardiac fibroblasts (HCFs), which do not spontaneously become myofibroblasts when cultured at low passages. We investigated the effect of Ac-SDKP on TGF-β1-induced HCF transformation into myofibroblasts, Smad2 and ERK1/2 phosphorylation, Smad7 expression, cell proliferation, and collagen production. We also investigated TGF-β1 production by HCFs stimulated with endothelin-1 (ET-1). As expected, HCFs treated with TGF-β1 transformed into myofibroblasts as indicated by increased expression of α-smooth muscle actin and a higher proportion of the embryonic isoform of smooth muscle myosin compared with untreated cells. TGF-β1 also increased Smad2 and ERK1/2 phosphorylation but did not affect Smad7 expression. In addition, TGF-β1 stimulated HCF proliferation as indicated by an increase in mitochondrial dehydrogenase activity and collagen production (hydroxyproline assay). Ac-SDKP significantly inhibited all of the effects of TGF-β1. It also inhibited ET-1-stimulated TGF-β1 production. We concluded that Ac-SDKP markedly suppresses differentiation of human cardiac fibroblasts into myofibroblasts, probably by inhibiting the TGF-β/Smad/ERK1/2 signaling pathway, and thus mediating its anti-fibrotic effects.

scholarly journals Platelet-derived growth factor-D promotes fibrogenesis of cardiac fibroblasts

2013 ◽  
Vol 304 (12) ◽  
pp. H1719-H1726 ◽  
Author(s):  
Tieqiang Zhao ◽  
Wenyuan Zhao ◽  
Yuanjian Chen ◽  
Victoria S. Li ◽  
Weixin Meng ◽  
...  
Keyword(s):  
Growth Factor ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Platelet Derived Growth Factor ◽  
Type I ◽  
Fibroblast Proliferation ◽  
Infarcted Heart ◽  
Collagen Secretion ◽  
Tgf Β1

Platelet-derived growth factor (PDGF)-D is a newly recognized member of the PDGF family with its role just now being understood. Our previous study shows that PDGF-D and its receptors (PDGFR-β) are significantly increased in the infarcted heart, where PDGFR-β is primarily expressed by fibroblasts, indicating the involvement of PDGF-D in the development of cardiac fibrosis. In continuing with these findings, the current study explored the molecular basis of PDGF-D on fibrogenesis. Rat cardiac fibroblasts were isolated and treated with PDGF-D (200 ng/ml medium). The potential regulation of PDGF-D on fibroblast growth, phenotype change, collagen turnover, and the transforming growth factor (TGF)-β pathway were explored. We found: 1) PDGF-D significantly elevated cardiac fibroblast proliferation, myofibroblast (myoFb) differentiation, and type I collagen secretion; 2) matrix metalloproteinase (MMP)-1, MMP-2, and MMP-9 protein levels were significantly elevated in PDGF-D-treated cells, which were coincident with increased expressions of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2; 3) PDGF-D significantly enhanced TGF-β1 synthesis, which was eliminated by TGF-β blockade with small-interfering RNA (siRNA); 4) the stimulatory role of PDGF-D on fibroblast proliferation and collagen synthesis was abolished by TGF-β blockade; and 5) TGF-β siRNA treatment significantly suppressed PDGF-D synthesis in fibroblasts. These observations indicate that PDGF-D promotes fibrogenesis through multiple mechanisms. Coelevations of TIMPs and MMPs counterbalance collagen degradation. The profibrogenic role of PDGF-D is mediated through activation of the TGF-β1 pathway. TGF-β1 exerts positive feedback on PDGF-D synthesis. These findings suggest the potential therapeutic effect of PDGFR blockade on interstitial fibrosis in the infarcted heart.

scholarly journals Resveratrol Inhibits the TGF-β1-Induced Proliferation of Cardiac Fibroblasts and Collagen Secretion by Downregulating miR-17 in Rat

2018 ◽  
Vol 2018 ◽  
pp. 1-10 ◽  
Author(s):  
Yao Zhang ◽  
Yuan Lu ◽  
Machuki Jeremiah Ong’achwa ◽  
Liqi Ge ◽  
Yun Qian ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Myocardial Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Inhibitory Effect ◽  
Neonatal Rat ◽  
Protein Levels ◽  
Resveratrol Treatment ◽  
Collagen Secretion ◽  
Tgf Β1

Myocardial fibrosis (MF) can cause heart remodeling and it is an independent risk factor for malignant arrhythmias, sudden cardiac death, and other malignant cardiovascular events. It is often characterized by myocardial interstitial collagen deposition and hyperproliferation of cardiac fibroblasts (CFs). The transforming growth factor-β1 (TGF-β1) is the most influential profibrogenic factor. Resveratrol (RSV) is an active polyphenol substance that inhibits myocardial fibrosis. The mechanism of RSV-mediated inhibition of the proliferation of CFs at the microRNA level is not fully understood. We used TGF-β1 to induce CFs proliferation to simulate the pathogenesis of myocardial fibrosis. Neonatal rat CFs were treated with TGF-β1 in the presence or absence of resveratrol. Cell proliferation was measured using the CCK-8 and EdU assay. Collagen secretion was measured using hydroxyproline kit. Further, qPCR analysis was performed to determine microRNA levels after TGF-β1 or resveratrol treatment. To identify the target gene for miR-17, miR-17 was overexpressed or silenced, and the mRNA and protein levels of Smad7 were assessed. The effects of miR-17 silencing or Smad7 overexpression on cell proliferation and collagen secretion were also examined. Resveratrol treatment significantly decreased the TGF-β1-induced CF proliferation and collagen secretion. Resveratrol also decreased the levels of miR-17, miR-34a, and miR-181a in TGF-β1-treated CFs. Overexpression of miR-17 decreased the Smad7 mRNA and protein levels while silencing miR-17 increased them. Additionally, silencing miR-17 or overexpressing Smad7 decreased the TGF-β1-induced CFs proliferation and collagen secretion. In conclusion, resveratrol inhibits TGF-β1-induced CFs proliferation and collagen secretion. This inhibitory effect of resveratrol is orchestrated by the downregulation of miR-17 and the regulation of Smad7.

scholarly journals Salvia miltiorrhizaandCarthamus tinctoriusExtract Prevents Cardiac Fibrosis and Dysfunction after Myocardial Infarction by Epigenetically Inhibiting Smad3 Expression

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Jing Yang ◽  
Bo Wang ◽  
Na Li ◽  
Qingqing Zhou ◽  
Wenhui Zhou ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Growth Factor ◽  
Cardiac Function ◽  
Myocardial Fibrosis ◽  
Histone Methylation ◽  
Cardiac Fibrosis ◽  
Transforming Growth Factor ◽  
Cardiac Fibroblasts ◽  
Salvia Miltiorrhiza ◽  
Left Ventricular

The incidence of cardiac dysfunction after myocardial infarction (MI) continues to increase despite advances in treatment. Excessive myocardial fibrosis plays a vital role in the development of adverse cardiac remodeling and deterioration of cardiac function. Understanding the molecular and cellular mechanism of the fibrosis process and developing effective therapeutics are of great importance.Salvia miltiorrhizaandCarthamus tinctoriusextract (SCE) is indicated for angina pectoris and other ischemic cardiovascular diseases in China. SCE has been shown to inhibit the platelet activation and aggregation, ameliorate ROS-induced myocardial necrosis by inhibiting mitochondrial permeability transition pore opening, and promote angiogenesis by upregulating the expression of vascular endothelial growth factor (VEGF). However, whether SCE has effect on cardiac fibrosis after MI is not fully clear. Here, a mouse model of MI was established to observe the effect of SCE upon survival, cardiac function, myocardial fibrosis, and inflammation. Quantitative PCR and western blot assays were used to determine the expression of genes related to transforming growth factor-β(TGF-β) cascade and inflammatory responsesin vivo. Additionally, the effects of SCE upon the collagen production, TGF-β/Smad3 (SMAD family member 3) signaling, and the levels of histone methylation in primary cardiac fibroblasts were detected. We found that SCE treatment significantly improved survival and left ventricular function in mice after MI. Inhibition of inflammation and fibrosis, as well as decreased expression of Smad3, was observed with SCE treatment. In TGF-β-stimulated cardiac fibroblasts, SCE significantly decreased the expression of collagen,α-smooth muscle actin (α-SMA), and Smad3. Furthermore, SCE treatment downregulated the levels of H3K4 trimethylation (H3K4me3) and H3K36 trimethylation (H3K36me3) at theSmad3promoter region of cardiac fibroblasts, leading to inhibition ofSmad3transcription. Our findings suggested that SCE prevents myocardial fibrosis and adverse remodeling after MI with a novel mechanism of suppressing histone methylation of theSmad3promoter and its transcription.

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