Trilobatin attenuates cerebral ischaemia/reperfusion-induced blood-brain barrier dysfunction by targeting matrix metalloproteinase 9: The legend of a food additive.

BACKGROUND AND PURPOSE: Blood-brain barrier (BBB) breakdown is one of the crucial pathological changes of cerebral ischaemia-reperfusion (I/R) injury. Trilobatin (TLB), a naturally occurring food additive, exerts neuroprotective effects against cerebral I/R injury as demonstrated in our previous study. This study was designed to investigate the effect of TLB on BBB disruption after cerebral I/R injury. EXPERIMENTAL APPROACH: Rats with focal cerebral ischaemia caused by transient middle cerebral artery occlusion were studied along with brain microvascular endothelial cells and human astrocytes to mimic BBB injury caused by oxygen and glucose deprivation/reoxygenation (OGD/R). KEY RESULTS: The results showed that TLB effectively maintained BBB integrity and inhibited neuronal loss following cerebral I/R challenge. Furthermore, TLB increased tight junction proteins including ZO-1, Occludin and Claudin 5, and decreased the levels of apolipoprotein E (APOE) 4, cyclophilin A (CypA) and phosphorylated nuclear factor kappa B (NF-κB), thereby reducing proinflammatory cytokines. TLB also decreased the Bax/Bcl-2 ratio and cleaved-caspase 3 levels along with a reduced number of apoptotic neurons. Molecular docking and transcriptomics predicted MMP9 as a prominent gene evoked by TLB treatment. The protective effects of TLB on cerebral I/R-induced BBB breakdown was largely abolished by overexpression of MMP9, and the beneficial effects of TLB on OGD/R-induced loss of BBB integrity in human brain microvascular endothelial cells and astrocyte co-cultures was markedly reinforced by knockdown of MMP9. CONCLUSIONS AND IMPLICATIONS: Our findings reveal a novel property of TLB: preventing BBB disruption following cerebral I/R via targeting MMP9 and inhibiting APOE4/CypA/NF-κB axis.

Icariside II mitigates myocardial infarction by balancing mitochondrial dynamics and reducing oxidative stress through the activation of Nrf2/SIRT3 signaling pathway.

Nuclear factor erythroid 2-related factor 2 (Nrf2)/silent mating type information regulation 2 homolog 3 (SIRT3) signaling pathway plays a pivotal role in regulating mitochondrial dynamics and oxidative stress, which are considered to be the principal pathogenesis of myocardial infarction (MI). Our previous study proved that pretreatment with icariside II (ICS II), a major active ingredient of Herbal Epimedii, exerts cardioprotective effect on MI, however, whether post-treatment with ICS II can alleviate MI and its underlying mechanism are still uncertain. Therefore, the present study was designed to investigate the therapeutic effect and the possible mechanism of ICS II on MI both in vivo and in vitro. The results revealed that post-treatment with ICS II markedly ameliorated myocardial injury in MI-induced mice and mitigated oxygen and glucose deprivation (OGD)-elicited cardiomyocyte injury. Further researches showed that ICS II promoted mitochondrial fusion, and suppressed mitochondrial fission and oxidative stress, which were achieved by facilitating the nuclear translocation of Nrf2 and activation of SIRT3. In summary, our findings indicate that ICS II mitigates MI-induced mitochondrial dynamics disorder and oxidative stress via activating the Nrf2/SIRT3 signaling pathway.

Icariside II Exerts Anti-Type 2 Diabetic Effect by Targeting PPARα/γ: Involvement of ROS/NF-κB/IRS1 Signaling Pathway.

Type 2 diabetes mellitus (T2DM) is a multisystem and complex metabolic disorder which is associated with insulin resistance and impairments of pancreatic ß-cells. Previous studies have shown that icariside II (ICS II), one of the main active ingredients of Herba Epimedii, exerts potent anti-inflammatory and anti-oxidative properties. In this study, we investigated whether ICS II exerted anti-T2DM profile and further explored its possible underlying mechanism both in vivo and in vitro. db/db mice were administered ICS II (10, 20, 40 mg·kg-1) for 7 weeks. We found that ICS II dose-dependently attenuated hyperglycemia and dyslipidemia, as well as inhibited hepatic steatosis and islet architecture damage in db/db mice. Moreover, ICS II not only dramatically reduced inflammatory cytokines and oxidative stress, but also up-regulated PPARα/γ protein expressions, phosphorylation of Akt, GSK3ß and IR, meanwhile, down-regulated phosphorylation of NF-κB(p65) and IRS1 in db/db mice. In palmitic acid (PA)-treated HepG2 or MIN6 cells, ICS II (5-20 µM) concentration-dependently promoted the cell viability via mediating PPARα/γ/NF-κB signaling pathway. PPARα/γ knockout by CRISPR-Cas9 system partly abolished the protective effects of ICS II on HepG2 or MIN6 cells following PA insults. These findings reveal that ICS II effectively confer anti-T2DM property by targeting PPARα/γ through mediation of ROS/NF-κB/IRS1 signaling pathway.

Icariside II, a Naturally Occurring SIRT3 Agonist, Protects against Myocardial Infarction through the AMPK/PGC-1α/Apoptosis Signaling Pathway.

Myocardial infarction (MI) refers to the death of cardiomyocytes triggered by a lack of energy due to myocardial ischemia and hypoxia, and silent mating type information regulation 2 homolog 3 (SIRT3) plays an essential role in protecting against myocardial oxidative stress and apoptosis, which are deemed to be the principal causes of MI. Icariside II (ICS II), one of the main active ingredients of Herbal Epimedii, possesses extensive pharmacological activities. However, whether ICS II can protect against MI is still unknown. Therefore, this study was designed to investigate the effect and possible underlying mechanism of ICS II on MI both in vivo and in vitro. The results showed that pretreatment with ICS II not only dramatically mitigated MI-induced myocardial damage in mice but also alleviated H9c2 cardiomyocyte injury elicited by oxygen and glucose deprivation (OGD), which were achieved by suppressing mitochondrial oxidative stress and apoptosis. Furthermore, ICS II elevated the phosphorylation level of adenosine monophosphate-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC-1α) expression, thereby activating SIRT3. However, these protective effects of ICS II on MI injury were largely abolished in SIRT3-deficient mice, manifesting that ICS II-mediated cardioprotective effects are, at least partly, due to the presence of SIRT3. Most interestingly, ICS II directly bound with SIRT3, as reflected by molecular docking, which indicated that SIRT3 might be a promising therapeutic target for ICS II-elicited cardioprotection in MI. In conclusion, our findings illustrate that ICS II protects against MI-induced oxidative injury and apoptosis by targeting SIRT3 through regulating the AMPK/PGC-1α pathway.

Icariside II Restores Vascular Smooth Muscle Cell Contractile Phenotype by Enhancing the Focal Adhesion Signaling Pathway in the Rat Vascular Remodeling Model.

Vascular smooth muscle cell (VSMC) phenotypic transition represents the fundamental pathophysiological alteration in the vascular remodeling process during the initiation and progression of cardiovascular diseases. Recent studies have revealed that Icariside II (ICS-II), a flavonol glycoside derived from the traditional Chinese medicine Herba Epimedii, exhibited therapeutic effects in various cardiovascular diseases. However, the therapeutic efficacy and underlying mechanisms of ICS-II regarding VSMC phenotypic transition were unknown. In this study, we investigated the therapeutic effects of ICS-Ⅱ on vascular remodeling with a rat's balloon injury model in vivo. The label-free proteomic analysis was further implemented to identify the differentially expressed proteins (DEPs) after ICS-II intervention. Gene ontology and the pathway enrichment analysis were performed based on DEPs. Moreover, platelet-derived growth factor (PDGF-BB)-induced primary rat VSMC was implemented to verify the restoration effects of ICS-II on the VSMC contractile phenotype. Results showed that ICS-II could effectively attenuate the vascular remodeling process, promote SMA-α protein expression, and inhibit OPN expression in vivo. The proteomic analysis identified 145 differentially expressed proteins after ICS-II intervention. Further, the bioinformatics analysis indicated that the focal adhesion signaling pathway was enriched in the ICS-II group. In vitro studies showed that ICS-II suppressed VSMC proliferation and migration, and promoted VSMC contractile phenotype by modulating the focal adhesion signaling pathway. Taken together, our results suggest that ICS-II attenuates the vascular remodeling process and restores the VSMC contractile phenotype by promoting the focal adhesion pathway.

Icariside II Attenuates Vascular Remodeling Via Wnt7b/CCND1 Axis.

ABSTRACT: Angioplasty often fails due to the abnormal proliferation of vascular smooth muscle cells (VSMCs). Success rates of angioplasty may increase following the administration of an agent that effectively ameliorates aberrant vascular remodeling. Icariside II (ICS-II) is a natural flavonol glycoside extract from the Chinese herbal medicine Epimedii that possesses several medicinal qualities that are beneficial in humans. Nevertheless, the role of ICS-II in addressing aberrant vascular remodeling have yet to be clarified. The current investigation studies the molecular effects of ICS-Ⅱ on balloon-inflicted neointimal hyperplasia in rats in vivo and on platelet-derived growth factor-induced vascular proliferation in primary rat aortic smooth muscle cells (VSMCs) in vitro. ICS-II was found to be as effective as rapamycin, the positive control used in this study. ICS-II inhibited neointimal formation in injured rat carotid arteries and notably reduced the expression of Wnt7b. ICS-Ⅱ significantly counteracted platelet-derived growth factor-induced VSMCs proliferation. Cell cycle analysis showed that ICS-II triggered cell cycle arrest during the G1/S transition. Western blot analysis further indicated that this cell cycle arrest was likely through Wnt7b suppression that led to CCND1 inhibition. In conclusion, our findings demonstrate that ICS-II possesses significant antiproliferative qualities that counteracts aberrant vascular neointimal hyperplasia. This phenomenon most likely occurs due to the suppression of the Wnt7b/CCND1 axis.

Icarrin prevents cardiomyocyte apoptosis in spontaneously hypertensive rats by inhibiting endoplasmic reticulum stress pathways.

OBJECTIVES: This study aimed to explore whether icarrin (ICA) can protect cardiomyocytes from hypertension-induced damage by inhibiting endoplasmic reticulum stress (ERS). METHODS: Spontaneously hypertensive rats (SHRs) were orally administered water or ICA at 10, 20 and 40 mg/kg once daily for 12 weeks, and Wistar-Kyoto (WKY) rats were used as control. Changes in the growth and blood pressure of rats were assessed. Cardiac function was determined by ultrasound and the left ventricle mass was calculated. Myocardial tissue structure was assessed by haematoxylin and eosin staining, cardiomyocyte apoptosis was observed by TUNEL staining and the expression of ERS-related proteins was determined by western blotting. RESULTS: In the SHR group, blood pressure was significantly high, left ventricular function decreased and left ventricular mass index increased. Additionally, left ventricular cardiomyocyte hypertrophy, disordered myofilament arrangement and increased cardiomyocyte apoptosis were observed by histological staining. ERS-induced proteins associated with apoptosis, including GRP78, PERK, ATF-6, ATF-4, CHOP, DR5, Caspase 12, c-JUN and ASK-1 were found to be highly expressed. ICA treatment reduced blood pressure and regulated the expression of proteins induced by ERS. Cardiomyocyte apoptosis decreased and left ventricular function improved. CONCLUSIONS: ICA can inhibit ERS-induced apoptosis of cardiomyocytes and protect ventricular function in SHR.

Osthole improves pulmonary artery hypertension by inducing apoptosis in pulmonary artery smooth muscle cells.

OBJECTIVES: The objectives of this study were to explore the effect of Osthole (Ost) on apoptosis in pulmonary artery smooth muscle cells (PASMCs) and investigate the potential mechanism of this effect. METHODS: Rats were injected subcutaneously with monocrotaline (MCT) to establish a PAH model, and Ost were intragastrically administrated from day 1 to day 35. After 35 days administration, the mean pulmonary artery pressure and lung weight index were measured. HE and TUNEL staining were used to observe the morphology of pulmonary artery and the apoptosis of PASMCs. In addition, the apoptosis of PASMCs were detected by flow cytometry in cultured PASMCs. The proteins of Bax and Bcl-2, and the levels of p-ASK1 and cleaved caspase 3 were measured by Western blot. KEY FINDINGS: Ost decreased the mean pulmonary artery pressure and lung weight index in MCT-induced rats, and promoted apoptosis in PASMCs in MCT-induced rats and PDGF-BB stimulated PASMCs. Ost increased the ratio of Bax/Bcl-2 and the levels of p-ASK1, cleaved caspase 3 in MCT-induced rats and PDGF-BB stimulated PASMCs. CONCLUSION: Ost promoted apoptosis in PASMCs in vivo and in vitro, and the mechanism may be associated with upregulation of ASK1 and the Bax/Bcl-2-caspase 3 signalling pathway.

Sildenafil improves right ventricular remodelling in monocrotaline-induced rats by decreasing myocardial apoptosis and activating peroxisome proliferator-activated receptors.

OBJECTIVES: To assess the effect of sildenafil on monocrotaline-induced right ventricular (RV) remodeling and investigate the possible mechanism. METHODS: Rats were subcutaneously injected with monocrotaline to establish an RV remodeling model and then administered sildenafil (25 mg/kg) from days 1 to 28. After 28 days of administration, the RV systolic pressure and the RV hypertrophy index (RVHI) were measured. The morphology of the right ventricle was observed by H&E staining. The ultrastructure of the right ventricle was observed using a transmission electron microscope. The myocardial apoptosis of the right ventricle was evaluated by TUNEL staining. The protein expression of apoptosis-related proteins and PPARs were examined by western blotting. KEY FINDINGS: The results indicated that sildenafil decreased the RV systolic pressure and RVHI, and improved the microstructure and ultrastructure of the right ventricle in monocrotaline-induced rats. In addition, sildenafil suppressed myocardial apoptosis and promoted the protein expression of PPARs of the right ventricle in monocrotaline-induced rats. CONCLUSION: Sildenafil inhibits RV remodeling in monocrotaline-induced rats, which might be partially mediated by reducing myocardial apoptosis and activating PPARs.

Osthole Alleviates Neointimal Hyperplasia in Balloon-Induced Arterial Wall Injury by Suppressing Vascular Smooth Muscle Cell Proliferation and Downregulating Cyclin D1/CDK4 and Cyclin E1/CDK2 Expression.

Percutaneous coronary intervention (PCI) is the most widely used therapy for treating ischemic heart disease. However, intimal hyperplasia and restenosis usually occur within months after angioplasty. Modern pharmacological researchers have proven that osthole, the major active coumarin of Cnidium monnieri (L.) Cusson, exerts potent antiproliferative effects in lung cancer cells, the human laryngeal cancer cell line RK33 and TE671 medulloblastoma cells, and its mechanism of action is related to cell cycle arrest. The goal of the present study was to observe the effect of osthole on vascular smooth muscle cell (VSMC) proliferation using platelet-derived growth factor-BB (PDGF-BB)-stimulated VSMCs isolated from rats and vascular balloon injury as models to further elucidate the molecular mechanisms underlying this activity. We detected the relative number of VSMCs by the MTT assay and EdU staining and examined cell cycle progression by flow cytometry. To more deeply probe the mechanisms, the protein expression levels of PCNA, the cyclin D1/CDK4 complex and the cyclin E1/CDK2 complex in balloon-treated rat carotid arteries and the mRNA and protein expression levels of the cyclin D1/CDK4 and cyclin E1/CDK2 complexes in VSMCs were detected by real-time RT-PCR and western blotting. The data showed that osthole significantly inhibited the proliferation of VSMCs induced by PDGF-BB. Furthermore, osthole caused apparent VSMC cycle arrest early in G0/G1 phase and decreased the expression of cyclin D1/CDK4 and cyclin E1/CDK2. Our results demonstrate that osthole can significantly inhibit PDGF-BB-induced VSMC proliferation and that its regulatory effects on cell cycle progression and proliferation may be related to the downregulation of cyclin D1/CDK4 and cyclin E1/CDK2 expression as well as the prevention of cell cycle progression from G0/G1 phase to S phase. The abovementioned mechanism may be responsible for the alleviation of neointimal hyperplasia in balloon-induced arterial wall injury by osthole.

Osthole inhibits cell proliferation by regulating the TGF-ß1/Smad/p38 signaling pathways in pulmonary arterial smooth muscle cells.

Pulmonary artery smooth muscle cell (PASMC) proliferation contributes to pulmonary vascular remodeling, which ultimately leads to pulmonary arterial hypertension (PAH). Osthole has been previously shown to inhibit tumor cell growth. Our previous experiments demonstrated that osthole could prevent monocrotaline-induced PAH and pulmonary artery remodeling in rats and that its effects might be associated with inhibiting PASMC proliferation. However, the exact mechanism remains unclear. In this study, we observed the inhibitory effect of osthole on platelet-derived growth factor (PDGF)-BB-induced rat PASMC growth, cell cycle progression and proliferating cell nuclear antigen (PCNA) expression, as measured by CCK-8 assay, flow cytometric analysis and western blotting, respectively. We also detected the expression and activities of the cell cycle regulators cyclin D1/CDK4, cyclin E1/CDK2, p53, p27 and p21 and the TGF-ß1/Smad/p38 signaling pathways in rat PASMCs by western blotting. Our results show that osthole effectively suppressed PDGF-BB-stimulated proliferation, PCNA protein expression, and cell cycle progression in rat PASMCs in vitro. We further demonstrated that treatment with osthole significantly induced cell cycle arrest at the G0/G1 phase in PASMCs, which was supported by the finding that osthole significantly decreased cyclin D1/CDK4 and cyclin E1/CDK2 protein levels and increased p53, p27 and p21 protein levels. These effects may partly be attributed to the downregulation of TGF-ß1/Smad/p38 signaling pathway activation. Our findings suggest that osthole is a potential therapeutic candidate that warrants further investigation regarding its potential use for the treatment of PAH.

Icariside II improves myocardial fibrosis in spontaneously hypertensive rats by inhibiting collagen synthesis.

OBJECTIVES: We aimed to investigate the effects of icariside II (ICS II) on myocardial fibrosis in spontaneously hypertensive rats (SHRs) and to explore the possible mechanisms. METHODS: We used SHRs as animal models, and we administered ICS II (4, 8 or 16 mg/kg) orally by gavage for 12 consecutive weeks (Fu et al., Biomed Pharmacother 2018; 100: 64). The left ventricular morphology of the rats was observed using haematoxylin-eosin (HE) staining. The occurrence of myocardial interstitial fibrosis was detected by Masson's trichrome staining. The protein levels of alpha smooth muscle actin (α-SMA), Collagen I, III, matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9, respectively), tissue inhibitor of metalloproteinase 1 (TIMP-1), transforming growth factor-ß1 (TGF-ß1), phospho-Smad2 (p-Smad2), phospho-Smad3 (p-Smad3) and phospho-p38 (p-p38) were examined by Western blotting. KEY FINDINGS: The results suggested that ICS II improved myocardial interstitial and perivascular collagen deposition and decreased Collagen I/III and α-SMA expression. ICS II (8 and 16 mg/kg) downregulated the expression of MMP-2 and MMP9 and upregulated the expression of TIMP1. In addition, the protein levels of p-Smad2/3, TGF-ß1 and p-p38 were decreased by ICS II treatment. CONCLUSIONS: The results suggest that ICS II can inhibit the expression of Collagen I and Collagen III through the MMP/TIMP-1 and TGF-ß1/Smad2,3/p-p38 signalling pathways and that it has therapeutic effects on myocardial fibrosis.

Icariside II prevents hypertensive heart disease by alleviating endoplasmic reticulum stress via the PERK/ATF-4/CHOP signalling pathway in spontaneously hypertensive rats.

OBJECTIVES: Reducing endoplasmic reticulum stress (ERS)-induced cardiomyocyte apoptosis is a key strategy for preventing hypertensive heart disease. In our previous study, Icariside II can improve left ventricular remodelling in spontaneously hypertensive rats (SHRs). This study aims to determine whether Icariside II can exert its effect by inhibiting ERS-induced cardiomyocyte apoptosis via the PERK/ATF-4/CHOP signalling pathway. METHODS: Spontaneously hypertensive rats were randomly divided into model group and Icariside II groups. The rats in the Icariside II groups were intragastrically administrated with Icariside II 4, 8 and 16 mg/kg from 14 to 26 week-age, respectively. The left ventricular function was measured at the 18, 22 and 26 week-age by small animal ultrasound. At the end of the 26th week, cardiomyocyte apoptosis was analysed and the levels of GRP78, PERK, ATF-4 and CHOP gene and protein were detected. KEY FINDINGS: The function of left ventricular became declined with age in SHRs, but improved in Icariside II groups. Myocardial apoptosis was aggravated in SHRs, but alleviated in Icariside II groups. Icariside II could reduce the levels of GRP78, PERK, ATF-4, CHOP gene and protein that increased in SHRs. CONCLUSIONS: Icariside II prevents hypertensive heart disease by alleviating ERS-induced cardiomyocyte apoptosis, and its mechanism is related to the impediment of the PERK/ATF-4/CHOP signalling pathway.

Icariside II attenuates myocardial fibrosis by inhibiting nuclear factor-κB and the TGF-ß1/Smad2 signalling pathway in spontaneously hypertensive rats.

Studies have demonstrated that icariin plays important roles in preventing hypertension and improving myocardial hypertrophy, inflammatory and infiltration. Icariside (ICS II) is the main metabolite of icariin, which has anti-inflammatory and anti-oxidant activities and protects against ischaemic brain injury. Whether ICS II improves myocardial fibrosis in spontaneously hypertensive rats (SHRs) and the related mechanism remain unknown. Some studies have suggested that TGF-ß and the nuclear factor κB signalling pathway play a key role in the progression of myocardial fibrosis. Therefore, in the current study, we aimed to evaluate the effects of ICS II on induced myocardial fibrosis in SHRs and explore the mechanism underlying this activity. The SHRs were treated with ICS II (4, 8, and 16 mg/kg) via daily gavage for 12 weeks. Left ventricular function was detected using the Vevo2100 system, and the collagen area was measured by Masson staining. The results indicated that ICS II markedly improved left ventricular function and decreased the left ventricular myocardial collagen area compared with the SHR group. To further investigate the mechanism underlying this activity, we measured the protein expression of interleukin-1ß (IL-1ß), tumour necrosis factor-α (TNF-α), transforming growth factor-ß1 (TGF-ß1), Smad2, inhibitory κB (IκB), and nuclear factor κB (NF-κB) p65 by Western blot. The results showed that ICS II inhibited NF-κB p65 expression and the TGF-ß1/Smad2 signalling pathways. In conclusion, the present results suggest that ICS II suppresses myocardial fibrosis in SHRs, and this effect might be at least partially mediated through suppression of NF-kB signalling and the TGF-ß1/Smad2 signalling pathway.

IcarisideII improves left ventricular remodeling in spontaneously hypertensive rats by inhibiting the ASK1-JNK/p38 signaling pathway.

Inhibition or removal of excess reactive oxygen species can effectively protect cellular function or reduce cell death because oxidative stress is the main cause of cellular damage in many diseases. The flavonoid compound IcarisideII having a slight inhibitory effect on PDE5, is the main active components of epimedium in vivo and has a wide range of pharmacological effects on oxidation and apoptosis. However, whether IcarisideII has the same protective effect on ventricular remodeling in spontaneously hypertensive rats (SHR) is unknown. We found that compared with WKY rats, SHRs exhibited noticeable arterial hypertension. Additionally, echocardiography showed that the diameter of the left ventricle was enlarged, wall thickness was increased, and ejection fraction and short axis shortening rate were reduced. H&E staining demonstrated that SHR cells were disordered and noticeably hypertrophic. Masson trichrome staining revealed significant myocardial fibrosis in the myocardium. Tunel staining indicated that 4.39 times the percent of apoptotic cells were present in SHRs compared to WKY rats. In our study, intra-gastric administration of IcarisideII decreased blood pressure, promoted heart function recovery and improved ventricular remodeling in SHRs. Additionally, it reduced myocardial fibrosis, inhibited myocardial apoptosis, decreased the generation of reactive oxygen species and improved SOD activity. IcarisideII down-regulated the activation of the oxidative stress associated proteins ASK1, p38 and JNK; inhibited the expression of p53, Bax and cleaved-caspase3 in the mitochondrial apoptosis pathway; and up-regulated the expression of Bcl-2. In conclusion, this study indicates that IcarisideII can inhibit myocardial apoptosis and improve left ventricular remodeling in SHRs. It can be inferred that this mechanism may be related to the inhibition of the ASK1-JNK/p38 signaling pathway.

Osthole attenuates right ventricular remodeling via decreased myocardial apoptosis and inflammation in monocrotaline-induced rats.

Osthole (Ost) is a coumarin that exhibits wide pharmacological effects in the cardiovascular system. However, whether Ost can inhibit apoptosis and inflammation in right ventricle (RV) cardiomyocytes and prevent RV remodeling is not clear. This study was designed to investigate the effect of Ost on RV remodeling and the underlying mechanism. By applying a monocrotaline (MCT)-induced rat model, the effect of Ost on RV remodeling was investigated. Rats were given a single dose of MCT (50mg/kg) subcutaneously (s.c.) to establish the RV remodeling model, followed by treatment with 10 or 20mg/kg Ost via daily gavage for 28 days. The RV pressure was measured, and a histological analysis was performed. The results suggested that Ost remarkably decreased RV pressure and improved myocardial hypertrophy and mitochondrial swelling, vacuolization, and sarcoplasmic reticulum enlargement when compared with the model group. To further investigate the roles of apoptosis and inflammation in the effects of Ost on MCT-induced RV remodeling, apoptosis-related factors and inflammatory-associated factors were examined by western blot. Ost was found to inhibit myocardial apoptosis and inflammation in the RV. Overall, the present results indicate that Ost suppresses the RV remodeling process induced by MCT in rats, which may be at least partially mediated through the reduction of myocardial apoptosis and inflammation.

Osthole attenuates pulmonary arterial hypertension in monocrotaline­treated rats.

Pulmonary arterial hypertension (PAH) is an insidious and progressive disease that is triggered by various cardiopulmonary diseases. Inflammation has an important role in the progression of PAH. Osthole (Ost) is a coumarin that has clear anti­inflammatory properties. The present study aimed to investigate the effects of Ost on PAH, and to explore the mechanism underlying this effect. Using the monocrotaline (MCT)­induced PAH rat model, the effects of Ost on PAH were investigated. Rats were subcutaneously administered a single dose of MCT (50 mg/kg) to establish the PAH model, followed by daily treatment with Ost (10 or 20 mg/kg) by gavage for 28 days. The mean pulmonary arterial pressure (mPAP) was measured and histological analysis was performed. The results demonstrated that Ost significantly decreased mPAP, and reduced thickening of the pulmonary artery, compared with in rats in the MCT group. To further determine whether the effects of Ost on MCT­induced PAH were associated with inflammatory responses, the nuclear factor­κB (NF­κB) p65 signaling pathway was investigated by western blot analysis. The results demonstrated that Ost increased inhibition of the NF­κB p65 signaling pathway. In conclusion, the results of the present study demonstrate that Ost may suppress the progression of MCT­induced PAH in rats, which may be, at least partially, mediated through modulation of the NF­κB p65 signaling pathway.

Osthole inhibits intimal hyperplasia by regulating the NF-κB and TGF-ß1/Smad2 signalling pathways in the rat carotid artery after balloon injury.

Osthole (7-methoxy-8-isopentenoxy-coumarin), a compound extracted from Cnidiummonnieri (L.) Cusson seeds, has been found to exhibit potent therapeutic effects in cancer due to its ability to inhibit inflammation and cell proliferation. However, its effects on arterial wall hypertrophy-related diseases remain unclear. Therefore, in this study, we aimed to investigate the effects of Osthole on intimal hyperplasia in a rat model of carotid artery balloon injury. We established the balloon-induced carotid artery injury rat model in male Sprague-Dawley rats, after which we administered Osthole (20mg/kg/day or 40mg/kg/day) or volume-matched normal saline orally by gavage for 14 consecutive days. Intimal hyperplasia and the degree of vascular smooth muscle cell proliferation were then evaluated by histopathological examination of the changes in the carotid artery, as well as by examination of proliferating cell nuclear antigen (PCNA) expression. Tumour necrosis factor-ɑ (TNF-α), interleukin-1ß (IL-1ß), transforming growth factor-beta (TGF-ß1) and PCNA mRNA expression levels were examined by real-time RT-PCR, while nuclear factor-κB (NF-κB (p65)), IκB-α, TGF-ß1 and phospho-Smad2 (p-Smad2) protein expression levels were analysed by immunohistochemistry or western blot analysis. We found that Osthole significantly attenuated neointimal thickness and decreased the elevations in PCNA protein expression induced by balloon injury. Moreover, Osthole down-regulated the pro-inflammatory factors TNF-α and IL-1ß and NF-κB (p65), whose expression had been upregulated after balloon injury. Moreover, IκB-α protein expression levels increased following Osthole treatment. In addition, the elevations in TGF-ß1 and p-Smad2 protein expression induced by balloon injury were both significantly attenuated by Osthole administration. We concluded that Osthole significantly inhibited neointimal hyperplasia in balloon-induced rat carotid artery injury and that the mechanism by which this occurs may involve NF-κB, IL-1ß and TNF-ɑ down-regulation, which alleviates the inflammatory response, and TGF-ß1/Smad2 signalling pathway inhibition.

Icariin prevents hypertension-induced cardiomyocyte apoptosis through the mitochondrial apoptotic pathway.

A prior study demonstrated that icariin (ICA) could repress angiotensin II-induced apoptosis in H9c2 cells. The activation of mitochondrial apoptotic pathways may play a crucial role in this phenomenon. In this study, we explored the potential protective roles of ICA in apoptosis in cardiomyocytes, cardiac remodelling, and the underlying mechanisms with regard to the mitochondrial apoptotic pathway in rats with spontaneous hypertension. The oral administration of ICA (20 and 40mg/kg/d) inhibited cardiomyocyte apoptosis and ameliorated left heart ventricle remodelling and abnormal mitochondria. ICA also decreased the blood pressure of model rats. ICA treatment increased the expression of Bcl-2 and decreased the expression of p53, Bax, Bok and cleaved caspase 3 in model rats, which suggests the potential mechanism underlying this effect. In summary, ICA inhibits the apoptosis of cardiomyocytes and ameliorates cardiac remodelling. The potential mechanism may relate to the inhibition of the mitochondrial apoptotic pathway.