Zerumbone, a humulane sesquiterpene from Syringa pinnatifolia, attenuates cardiac fibrosis by inhibiting of the TGF-β1/Smad signaling pathway after myocardial infarction in mice.

Zerumbone (ZER) is a humulane sesquiterpene isolated from Syringa pinnatifolia Hemsl., a representative Mongolian herbal medicine that is used to treat cardiovascular diseases. Cardiac fibrosis is a common pathological process in cardiovascular disease that results from the excessive accumulation of extracellular matrix (ECM), and the transforming growth factor (TGF)-β/Smad pathway is a canonical signaling pathway that directly induces expressions of ECM-related genes. Currently, the cardioprotective effect and underlying mechanisms of ZER on the inhibition of cardiac fibrosis are not well known. To explore the cardioprotective properties and pharmacological mechanism of ZER against cardiac fibrosis via the TGF-β1/Smad signaling pathway. Myocardial infarction (MI) model was induced by ligation of the left anterior descending coronary artery in ICR mice. The mice were randomly divided into six groups: sham, model, low-dose ZER (ZER-L), medium-dose ZER (ZER-M), high-dose ZER (ZER-H) and fosinopril. Mice in each group were intragastrically administered treatments for 21 days, and cardiac function was evaluated by 2D echocardiography. The pathological structure of the heart was examined by hematoxylin and eosin (HE) and Masson staining. Content of collagen I and collagen III were assessed by immunofluorescence methods. The inhibitory effect of ZER on TGF-β1 protein expression was predicted by molecular docking technology. Reverse transcriptase polymerase chain reaction (RT-PCR) and western blotting were used to measure the levels of genes and proteins expressed in the TGF-β1/Smad signaling pathway and MMPs. TGF-β1-treated cardiac fibroblasts (CFs) of neonatal SD rats were adopted for in vitro studies. Cardiac ejection fraction (EF) and fractional shortening (FS) in the model group were markedly decreased compared with those in the sham group, indicating that the MI model was successfully established. ZER and fosinopril elevated EF and FS values, suggesting cardioprotective effects. Pathological staining and immunofluorescence analysis showed that the content of collagen I and collagen III increased in the cardiac tissue of mice in model group, while ZER treatment obviously reduced collagen levels. The molecular docking simulations predicted the hydrophobic interactions between ZER and TGF-β1. In addition, the expression of TGF-β1, p-Smad2/3 and MMPs in the ZER treatment group was significantly decreased compared with the model group. In vitro studies further confirmed that α-smooth muscle actin (α-SMA) and p-Smad2/3 increased markedly in cardiac fibroblasts after incubation with TGF-β1, and treatment with ZER suppressed the expression of α-SMA and TGF-β1 downstream proteins in cardiac fibroblasts. ZER rescues cardiac function by attenuating cardiac fibrosis, and the antifibrotic effect may be mediated by blocking the TGF-β1/Smad pathway.

Li J ，Ge F ，Wuken S ，Jiao S ，Chen P ，Huang M ，Gao X ，Liu J ，Tu P ，Chai X ，Huang L ... -  《-》

Qishen granule attenuates cardiac fibrosis by regulating TGF-β /Smad3 and GSK-3β pathway.

Cardiac fibrosis is a common pathological progress of cardiovascular disease resulting from the excessive accumulation of extracellular matrix (ECM). Transforming growth factor (TGF)-β/SMADs pathway is a canonical signaling pathway which directly induces expressions of ECM related genes. Qishen Granule (QSG), a traditional Chinese formula developed from Zhen-Wu Decoration for heart failure (HF), has been proven to have definite therapeutic effects on cardiac fibrosis. However, its underlying mechanisms remain unclear. To investigate the effects of QSG on TGF-β pathway and the downstream mediators including Smad3 and Glycogen synthase kinase (GSK)-3β. HF model was induced by ligation of left coronary artery on male Sprague-Dawley (SD) rats. Rat were randomly divided into four groups including sham group, model group, QSG group and Fosinopril control group. Rats in each group were treated for 28 days, and 2D echocardiography was adopted to evaluate the heart function. The degree of cardiac fibrosis was assessed by Hematoxylin-Eosin (HE), Masson's trichrome and Picrosirius red (PSR) staining. Contents of collagen Ⅰ and Ⅲ were assessed by immunohistochemical method. Expressions of genes and proteins in TGF-β/SMADs and PI3K-GSK-3 signaling pathways were detected by Real-time Fluorescence Quantitative PCR (RT-qPCR) and Western blot respectively. TGF-β1-treated cardiac fibroblasts of neonatal SD rats were adopted for in vitro studies. 28 days after the surgery, cardiac ejection fraction (EF) and fractional shortening (FS) values in the model group showed a remarkable decrease, indicating the induction of HF model. QSG and Fosinopril elevated the EF and FS values, demonstrating cardio-protective effects. Pathological staining and immunohistochemistry showed that the contents of collagen I and III dramatically increased in the cardiac tissue of the model group compared with the sham group while QSG treatment reduced collagen contents. Furthermore, expressions of TGF-β1, p-Smad3 and p-GSK-3β were significantly decreased in the QSG treatment group compared with the model group, suggesting that the QSG may attenuate cardiac fibrosis through regulating TGF-β/Smad3 pathway. In vitro study further showed that the productions of type Ⅰ and Ⅲ collagen and α-smooth muscle actin (α-SMA) of cardiac fibroblasts were significantly increased by incubation with TGF-β1. QSG could markedly reduce the secretion of collagen Ⅰ and Ⅲ and α-SMA expression. Protein expressions of p-Smad3, PI3K, p-Akt and p-GSK-3β were significantly up-regulated by stimulation of TGF-β1. Treatment with QSG could suppress the activity of Smad3 and PI3K-GSK-3β signaling pathway in cardiac fibroblasts. QSG improves cardiac function through inhibiting cardiac fibrosis. The anti-fibrotic effects are potentially mediated by the inhibition of the TGF-β/Smad3 pathway and the phosphorylation of GSK-3β.

Zeng Z ，Wang Q ，Yang X ，Ren Y ，Jiao S ，Zhu Q ，Guo D ，Xia K ，Wang Y ，Li C ，Wang W ... -  《-》

Linggui Zhugan Decoction () Inhibits Ventricular Remodeling after Acute Myocardial Infarction in Mice by Suppressing TGF-β(1)/Smad Signaling Pathway.

Wang L ，Shi H ，Huang JL ，Xu S ，Liu PP ... -  《-》

Zerumbone, a major sesquiterpene from Syringa pinnatifolia Hemsl., exerts the sedative effect by regulating GABAergic nervous system.

Zerumbone (ZER) is a humulane sesquiterpenoid isolated from Syringa pinnatifolia Hemsl. (SP), its content accounts for 64.7% of volatile oil and 0.86% of total ethanol extract (TEE), representing one of characteristic ingredient of SP. As a representative Mongolian medicine with anti-"Khii", anti-asthma, and clearing-heat effects, SP has been used for the treatment of cardiovascular diseases, upset, insomnia, and other symptoms. Previous results showed that TEE has sedative effect, but the pharmacological substances and its sedative mechanism remains unclear. This study aims to determine whether ZER, as one of major and characteristic sesquiterpenoids of SP, contributes to the sedative effect of SP and its underlying mechanism. Locomotor activity and threshold dose of pentobarbital sodium sleep experiments were used to evaluate the sedative effects in mice. ELISA assay was used to examine the level of GABA/Glu ratio in rats hippocampus, cortex and hypothalamus tissue. The binding ability of ZER with glutamic acid decarboxylase 67 (GAD67) and Gephyrin protein were predicted by molecular docking. Western blot and Immunohistochemistry assay were used to determine the expression of GABAergic nerve system related proteins (GAD67, Gephyrin) in rat's hypothalamus. ZER was co-administrated with flumazenil and bicuculline (GABAA antagonist) to determine whether it acts on GABAA receptor. Furthermore, MQAE assay was used to test the effect of ZER on the chloride ion concentration in cerebellar granule cells. Current data demonstrated that ZER dose-dependently (5-20 mg/kg) reduces the locomotor activity and sleep latency of mice, and extend sleeping time of mice. The results of ELISA showed that ZER increases the level of GABA/Glu in rats brain tissue, in particular in hypothalamus. Molecular docking results revealed that ZER has a strong affinity to GAD67 and Gephyrin protein. The Western blot and Immunohistochemistry data indicated that ZER up-regulates the expression of GAD67 and Gephyrin protein in rat's hypothalamus. Antagonism test results demonstrated that flumazenil and bicuculline reverse the effect of ZER on threshold dose of pentobarbital sodium sleep experiments. In addition, ZER also could dose-dependently (5-20 μM) increase the chloride ion concentration in cerebellar granule cell, suggesting that ZER induces the opening of chloride channel, exerts central inhibitory effect. ZER has a significant sedative effect in mice and rat, and the effect is associated with GABAergic nervous system. The present results suggest that ZER, as one of the major bioactive ingredients of SP, contributes to the sedative effect and provide substantial evidence for its traditional use of anti-"Khii" in clinic of Syringa pinnatifolia.

Wuken S ，Li J ，Gao X ，Jiao S ，Ma X ，Chen S ，Tu P ，Huang L ，Chai X ... -  《-》

Effect of alprostadil on myocardial fibrosis in rats with diabetes mellitus via TGF-β1/Smad signaling pathway.

Tu JH ，Xu Y ，Dai Y ，Dang L ... -  《-》