Erratum: Activation of Nrf2 Attenuates Pulmonary Vascular Remodeling via Inhibiting Endothelial-to-Mesenchymal Transition: an Insight from a Plant Polyphenol: Erratum.

[This corrects the article DOI: 10.7150/ijbs.20316.].

Baicalein attenuates rotenone-induced SH-SY5Y cell apoptosis through binding to SUR1 and activating ATP-sensitive potassium channels.

Dopaminergic neurons in the substantia nigra (SN) expressing SUR1/Kir6.2 type ATP-sensitive potassium channels (K-ATP) are more vulnerable to rotenone or metabolic stress, which may be an important reason for the selective degeneration of neurons in Parkinson's disease (PD). Baicalein has shown neuroprotective effects in PD animal models. In this study, we investigated the effect of baicalein on K-ATP channels and the underlying mechanisms in rotenone-induced apoptosis of SH-SY5Y cells. K-ATP currents were recorded from SH-SY5Y cells using whole-cell voltage-clamp recording. Drugs dissolved in the external solution at the final concentration were directly pipetted onto the cells. We showed that rotenone and baicalein opened K-ATP channels and increased the current amplitudes with EC50 values of 0.438 µM and 6.159 µM, respectively. K-ATP channel blockers glibenclamide (50 µM) or 5-hydroxydecanoate (5-HD, 250 µM) attenuated the protective effects of baicalein in reducing reactive oxygen species (ROS) content and increasing mitochondrial membrane potential and ATP levels in rotenone-injured SH-SY5Y cells, suggesting that baicalein protected against the apoptosis of SH-SY5Y cells by regulating the effect of rotenone on opening K-ATP channels. Administration of baicalein (150, 300 mg·kg-1·d-1, i.g.) significantly inhibited rotenone-induced overexpression of SUR1 in SN and striatum of rats. We conducted surface plasmon resonance assay and molecular docking, and found that baicalein had a higher affinity with SUR1 protein (KD = 10.39 µM) than glibenclamide (KD = 24.32 µM), thus reducing the sensitivity of K-ATP channels to rotenone. Knockdown of SUR1 subunit reduced rotenone-induced apoptosis and damage of SH-SY5Y cells, confirming that SUR1 was an important target for slowing dopaminergic neuronal degeneration in PD. Taken together, we demonstrate for the first time that baicalein attenuates rotenone-induced SH-SY5Y cell apoptosis through binding to SUR1 and activating K-ATP channels.

Puerarin Inhibits NLRP3-Caspase-1-GSDMD-Mediated Pyroptosis via P2X7 Receptor in Cardiomyocytes and Macrophages.

Diabetic cardiomyopathy (DCM) is a critical complication of long-term chronic diabetes mellitus, and it is characterized by myocardial fibrosis and myocardial hypertrophy. Previous studies have shown that the pyroptosis pathway was significantly activated in DCM and may be related to the P2X7 receptor. However, the role of the P2X7 receptor in the development of DCM with pyroptosis is still unclear. In this study, we aimed to explore the mechanism of puerarin and whether the P2X7 receptor can be used as a new target for puerarin in the treatment of DCM. We adopted systematic pharmacology and bioinformatic approaches to identify the potential targets of puerarin for treating DCM. Additionally, we employed D-glucose-induced H9C2 rat cardiomyocytes and lipopolysaccharide-treated RAW264.7 mouse mononuclear macrophages as the in vitro model on DCM research, which is close to the pathological conditions. The mRNA expression of cytokines in H9C2 cells and RAW264.7 macrophages was detected. The protein expressions of NLRP3, N-GSDMD, cleaved-caspase-1, and the P2X7 receptor were investigated with Western blot analysis. Furthermore, molecular docking of puerarin and the P2X7 receptor was conducted based on CDOCKER. A total of 348 puerarin targets and 4556 diabetic cardiomyopathy targets were detected, of which 218 were cross targets. We demonstrated that puerarin is effective in enhancing cardiomyocyte viability and improving mitochondrial function. In addition, puerarin is efficacious in blocking NLRP3-Caspase-1-GSDMD-mediated pyroptosis in H9C2 cells and RAW264.7 cells, alleviating cellular inflammation. On the other hand, similar experimental results were obtained by intervention with the P2X7 receptor antagonist A740003, suggesting that the protective effects of puerarin are related to the P2X7 receptor. The molecular docking results indicated key binding activity between the P2X7 receptor and puerarin. These findings indicate that puerarin effectively regulated the pyroptosis signaling pathway during DCM, and this regulation was associated with the P2X7 receptor.

Aesculin suppresses the NLRP3 inflammasome-mediated pyroptosis via the Akt/GSK3ß/NF-κB pathway to mitigate myocardial ischemia/reperfusion injury.

BACKGROUND: Aesculin (AES), an effective component of Cortex fraxini, is a hydroxycoumarin glucoside that has diverse biological properties. The nucleotide-binding domain leucine-rich repeat-containing receptor, pyrin domain-containing 3 (NLRP3) inflammasome has been heavily interwoven with the development of myocardial ischemia/reperfusion injury (MIRI). Nevertheless, it remains unclear whether AES makes a difference to the changes of the NLRP3 inflammasome in MIRI. PURPOSE: We used rats that were subjected to MIRI and neonatal rat cardiomyocytes (NRCMs) that underwent oxygen-glucose deprivation/restoration (OGD/R) process to investigate what impacts AES exerts on MIRI and the NLRP3 inflammasome activation. METHODS: The establishment of MIRI model in rats was conducted using the left anterior descending coronary artery ligation for 0.5 h ischemia and then untying the knot for 4 h of reperfusion. After reperfusion, AES were administered intraperitoneally using 10 and 30 mg/kg doses. We evaluated the development of reperfusion ventricular arrhythmias, hemodynamic changes, infarct size, and the biomarkers in myocardial injury. The inflammatory mediators and pyroptosis were also assessed. AES at the concentrations of 1, 3, and 10 µM were imposed on the NRCMs immediately before the restoration process. We also determined the cell viability and cell death in the NRCMs exposed to OGD/R insult. Furthermore, we also analyzed the levels of proteins that affect the NLRP3 inflammasome activation, pyroptosis, and the AKT serine/threonine kinase (Akt)/glycogen synthase kinase 3 beta (GSK3ß)/nuclear factor-kappa B (NF-κB) signaling pathway via western blotting. RESULTS: We found that AES notably attenuated reperfusion arrhythmias and myocardia damage, improved the hemodynamic function, and ameliorated the inflammatory response and pyroptosis of cardiomyocytes in rats and NRCMs. Additionally, AES reduced the NLRP3 inflammasome activation in rats and NRCMs. AES also enhanced the phosphorylation of Akt and GSK3ß, while suppressing the phosphorylation of NF-κB. Moreover, the allosteric Akt inhibitor, MK-2206, abolished the AES-mediated cardioprotection and the NLRP3 inflammasome suppression. CONCLUSIONS: These findings indicate that AES effectively protected cardiomyocytes against MIRI by suppressing the NLRP3 inflammasome-mediated pyroptosis, which may relate to the upregulated Akt activation and disruption of the GSK3ß/NF-κB pathway.

Systematic analysis of molecular mechanism of resveratrol for treating pulmonary hypertension based on network pharmacology technology.

Resveratrol is a polyphenolic antioxidant derived from plant products such as grapes. Previous studies explored the effects of resveratrol on pulmonary hypertension (PH). However, systematic research on the exact mechanism of action of resveratrol is still lacking; in particular, our knowledge on the molecule-gene interaction is limited. In this study, systematic pharmacology and bioinformatic approaches were employed to identify the potential targets of resveratrol for treating PH. Furthermore, core genes were identified by constructing a protein-protein interaction network and by conducting topology analyses. The results showed that the effect of resveratrol may be closely associated with targets such as AKT serine/threonine kinase 1 (AKT1), mitogen-activated protein kinase 3 (MAPK3), Sirtuin-1 (SIRT1) and proto-oncogene tyrosine-protein kinase Src (SRC), as well as biological processes such as cell proliferation, inflammatory response, and redox balance. The present study systematically elucidates the mechanisms by which resveratrol alleviates PH and provides a new perspective on drug research for this disease.

Puerarin attenuates the endothelial-mesenchymal transition induced by oxidative stress in human coronary artery endothelial cells through PI3K/AKT pathway.

Endothelial-mesenchymal transition (EndMT) is a process in which endothelial cells lose their specific morphology/markers and undergo a dramatic remodeling of the cytoskeleton. It has been implicated in the progression of cardiovascular diseases such as cardiac fibrosis and cardiac dysfunction. Recent study indicated that puerarin could inhibit EndMT against cardiac fibrosis. However, the precise role of puerarin in EndMT and the underlying molecular mechanisms remain unclear. EndMT was induced by H2O2 (150 µM) in human coronary artery endothelial cells (HCAECs). HCAECs were exposed to H2O2 for six days with or without puerarin pretreated 2 h. The protein changes of EndMT markers (CD31, VE-cadherin, FSP1 and α-SMA) in HCAECs were detected. The levels of phosphoinositide-3-kinase (PI3K) and protein kinase B (AKT) proteins were analyzed by Western Blot. Wound healing and transwell assay were carried out to examine cell chemotaxis. Puerarin mitigated H2O2-induced EndMT as indicated by alleviating the reduced expression of CD31 and VE-cadherin and inhibiting the upregulation of α-SMA and FSP1. Furthermore, the mechanisms study showed that puerarin activated the PI3K/AKT pathway by inhibiting reactive oxygen species and further attenuated EndMT. On the other hand, PI3K inhibitor LY294002 reversed this effect imposed by puerarin. Puerarin alleviated the migration of mesenchymal-like cells through reducing MMPs protein expression. These results implicated that puerarin exhibited cytoprotective effects against H2O2-induced EndMT in HCAECs through alleviating oxidative stress, activating the PI3K/AKT pathway and limiting cell migration.

Total flavonoids from Anchusa italica Retz. Improve cardiac function and attenuate cardiac remodeling post myocardial infarction in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The plant Anchusa italica Retz. (Anchusa azurea Mill.) has been traditionally used in Uygur medicine for the treatment of cardiovascular and cerebrovascular diseases in China. Our previous study showed that total flavonoids from Anchusa italica Retz. (TFAI) exhibited potent cardioprotection in acute ischemia/reperfusion injured rats. AIM OF THE STUDY: This study was undertaken to investigate the effects of TFAI on chronic myocardial infarction (MI) in mice and the underlying mechanism. MATERIALS AND METHODS: Total flavonoids were extracted from the whole herb of Anchusa italica Retz. and were characterized using HPLC-MS analysis. The left anterior descending branch of the coronary artery was ligated to simulate MI injury in mice. After surgery, mice were orally fed with TFAI at the doses of 10, 30 and 50 mg/kg body weight/day for a total of four weeks. Cardiac function and infarct size were measured, and inflammatory mediators were detected. Hematoxylin and eosin (H&E) staining and Masson's trichrome staining were performed on heart sections. The apoptotic factors, such as Bax, Bcl-2 and cleaved caspase 3, as well as the key proteins in the PI3K/Akt/mTOR signaling pathway were examined by Western blot. RESULTS: The content of total flavonoids in TFAI was 56.2%. Four weeks following the MI surgery, TFAI enhanced the survival rate in post-MI mice. TFAI treatment at the doses of 30 and 50 mg/kg remarkably reduced infarct size and improved cardiac function as indicated by elevated EF and FS. Assay of the inflammatory factors showed that sera levels of TNF-α, IL-1ß and IL-6 were markedly decreased by TFAI treatment compared to the MI group. H&E staining and Masson's trichrome staining demonstrated that TFAI suppressed myocyte hypertrophy and cardiac fibrosis as indicated by the decreased cross-section area and collagen volume. Western blot analysis showed that cleaved caspase 3 and Bax/Bcl-2 were significantly downregulated following TFAI treatment. Furthermore, TFAI treatment significantly suppressed the activation of the PI3K/Akt/mTOR signaling pathway. CONCLUSIONS: Our data suggest that TFAI exerts a potent protective effect against chronic MI injury, and its beneficial effects on cardiac function and cardiac remodeling might be attributable, at least in part, to anti-inflammation and inhibition of the PI3K/Akt/mTOR signaling pathway.

Formononetin ameliorates myocardial ischemia/reperfusion injury in rats by suppressing the ROS-TXNIP-NLRP3 pathway.

Formononetin (FN), a methoxy isoflavone abundant in many plants and herbs, has been evidently proven to possess multiple medicinal properties. Our study aimed to clarify the impact of FN on myocardial ischemia/reperfusion (I/R) injury (MIRI) and the involved mechanism. A rat model of MIRI was produced by ligation and loosening of the left anterior descending (LAD) branch of the coronary artery. Rats received 10 and 30 mg/kg of FN when the reperfusion started. At 24 h after surgery, cardiac function, infarct size, and sera levels of the cardiac markers and inflammatory mediators were measured. To mimic the inflammasome activation in cardiomyocytes, neonatal rat cardiomyocytes (NRCMs) were cultured and treated with lipopolysaccharide (LPS) plus nigericin. Cell death and reactive oxygen species (ROS) were determined. Myocardial expression and activation of the nucleotide-binding domain and leucine-rich repeat-containing protein 3 (NLRP3) inflammasome in rats were examined by western blotting. The level of thioredoxin interacting protein (TXNIP)-NLRP3 interaction was assessed. FN notably attenuated cardiac dysfunction, infarct size, release of cardiac markers, and elevation of TNF-α, IL-1ß, and IL-6. FN alleviated LPS plus nigericin-induced injury and ROS increase in NRCMs. Western blotting revealed that FN suppressed the activation of NLRP3 inflammasome and TXNIP-NLRP3 interaction in rats. These findings indicate that FN ameliorated MIRI in rats and inhibited the activation of the NLRP3 inflammasome, at least partially, attributable to suppression of the ROS-TXNIP-NLRP3 pathway.

Chemical and pharmacological research on the polyphenol acids isolated from Danshen: A review of salvianolic acids.

Danshen, the dried root of Salvia miltiorrhiza Bge, is a common medicinal herb in Traditional Chinese Medicine, which has been used for the treatment of a number of diseases for thousands of years. More than 2000 years ago, the Chinese early pharmacy monograph "Shennong Materia Medica" recorded that Danshen could be used for the treatment of gastrointestinal diseases, cardiovascular diseases, certain gynecological diseases, etc. Since then, Danshen has been widely used clinically in many different prescriptions for many different diseases, especially for the treatment of cardiovascular diseases. Nowadays, many pharmacological studies about the water-soluble components from Danshen have been reported, especially salvianolic acids. It turned out that salvianolic acids showed strong anti-lipid peroxidation and anti-thrombic activities, and among them, SalAA and SalAB were the most potent. This review focused on the achievements in research of salvianolic acids regarding their bioactivities and pharmacological effects. These studies not only shed light on the water-soluble active components of Danshen and their mechanisms at the molecular level, but also provided theoretical information for the development of new medicines from Danshen for the treatment of cardiovascular and cerebrovascular diseases, inflammatory diseases, metabolic diseases, etc.

Glucose consumption assay discovers coptisine with beneficial effect on diabetic mice.

Many drugs with anti-diabetic effects regulate glucose consumption in peripheral tissues. Via cellular glucose consumption assays, we identified that coptisine, a main effective constituent from the plant Coptis chinensis, enhanced hepatic and skeletal muscle glucose consumption. We further explored its effects on glucose metabolism in diabetic animals to elucidate its mechanism of action. Our results showed that coptisine did not show cytotoxicity. Intragastric administration of coptisine for ten days in normal ICR mice markedly decreased fasting blood-glucose levels without significant effects on body weight. In alloxan-induced type 1 diabetic mice, intragastric administration of coptisine for 28 days decreased fasting and non-fasting blood-glucose levels as well. In type 2 diabetic KKAy mice, intragastric administration of coptisine for nine weeks improved glucose tolerance. It decreased fasting/non-fasting blood-glucose and fructosamine levels. Coptisine decreased low-density lipoprotein and total cholesterol levels, however, had no significant effect on triglyceride levels. Coptisine increased AMPK phosphorylation while decreasing Akt phosphorylation in HepG2 hepatic cells and C2C12 myotubes. Coptisine also reduced mitochondrial respiration in isolated and cellular mitochondria, suggesting that coptisine lowered cellular energy levels. In particularly, coptisine administration (10-6 M) decreased the mitochondrial oxygen consumption rate (OCR) with a greater extracellular acidification rate (ECAR), resulting in an oxidative-to-glycolysis phosphorylation shifted for cellular energy generation. Our results demonstrate that coptisine acts as an enhancer of peripheral glucose consumption could improve glucose metabolism in diabetic animals. Coptisine may serve as a novel anti-diabetic agent and warrant further evaluation.

Cardioprotective Effects of Puerarin-V on Isoproterenol-Induced Myocardial Infarction Mice Is Associated with Regulation of PPAR-Υ/NF-κB Pathway.

Puerarin is a well-known traditional Chinese medicine which has been used for the treatment of cardiovascular diseases. Recently, a new advantageous crystal form of puerarin, puerarin-V, has been developed. However, the cardioprotective effects of puerarin-V on myocardial infarction (MI) heart failure are still unclear. In this research, we aim to evaluate the cardioprotective effects of puerarin-V on the isoproterenol (ISO)-induced MI mice and elucidate the underlying mechanisms. To induce MI in C57BL/6 mice, ISO was administered at 40 mg/kg subcutaneously every 12 h for three times in total. The mice were randomly divided into nine groups: (1) control; (2) ISO; (3) ISO + puerarin injection; (4⁻9) ISO + puerarin-V at different doses and timings. After treatment, cardiac function was evaluated by electrocardiogram (ECG), biochemical and histochemical analysis. In vitro inflammatory responses and apoptosis were evaluated in human coronary artery endothelial cells (HCAECs) challenged by lipopolysaccharide (LPS). LPS-induced PPAR-Υ/NF-κB and subsequently activation of cytokines were assessed by the western blot and real-time polymerase chain reaction (PCR). Administration of puerarin-V significantly inhibits the typical ST segment depression compared with that in MI mice. Further, puerarin-V treatment significantly improves ventricular wall infarction, decreases the incidence of mortality, and inhibits the levels of myocardial injury markers. Moreover, puerarin-V treatment reduces the inflammatory milieu in the heart of MI mice, thereby blocking the upregulation of proinflammatory cytokines (TNF-α, IL-1ß and IL-6). The beneficial effects of puerarin-V might be associated with the normalization in gene expression of PPAR-Υ and PPAR-Υ/NF-κB /ΙκB-α/ΙΚΚα/ß phosphorylation. In the in vitro experiment, treatment with puerarin-V (0.3, 1 and 3 µM) significantly reduces cell death and suppresses the inflammation cytokines expression. Likewise, puerarin-V exhibits similar mechanisms. The cardioprotective effects of puerarin-V treatment on MI mice in the pre + post-ISO group seem to be more prominent compared to those in the post-ISO group. Puerarin-V exerts cardioprotective effects against ISO-induced MI in mice, which may be related to the activation of PPAR-γ and the inhibition of NF-κB signaling in vivo and in vitro. Taken together, our research provides a new therapeutic option for the treatment of MI in clinic.

Activation of Nrf2 Attenuates Pulmonary Vascular Remodeling via Inhibiting Endothelial-to-Mesenchymal Transition: an Insight from a Plant Polyphenol.

The endothelial-to-mesenchymal transition (EndMT) has been demonstrated to be involved in pulmonary vascular remodeling. It is partly attributed to oxidative and inflammatory stresses in endothelial cells. In current study, we conducted a series of experiments to clarify the effect of salvianolic acid A (SAA), a kind of polyphenol compound, in the process of EndMT in human pulmonary arterial endothelial cells and in vivo therapeutic efficacy on vascular remodeling in monocrotaline (MCT)-induced EndMT. EndMT was induced by TGFß1 in human pulmonary arterial endothelial cells (HPAECs). SAA significantly attenuated EndMT, simultaneously inhibited cell migration and reactive oxygen species (ROS) formation. In MCT-induced pulmonary arterial hypertension (PAH) model, SAA improved vascular function, decreased TGFß1 level and inhibited inflammation. Mechanistically, SAA stimulated Nrf2 translocation and subsequent heme oxygenase-1 (HO-1) up-regulation. The effect of SAA on EndMT in vitro was abolished by ZnPP, a HO-1 inhibitor. In conclusion, this study indicates a deleterious impact of oxidative stress on EndMT. Polyphenol antioxidant treatment may provide an adjunctive action to alleviate pulmonary vascular remodeling via inhibiting EndMT.

Salvianolic Acid A, a Component of Salvia miltiorrhiza, Attenuates Endothelial-Mesenchymal Transition of HPAECs Induced by Hypoxia.

Salvianolic acid A (SAA), a polyphenols acid, is a bioactive ingredient from a traditional Chinese medicine called Dan shen (Salvia Miltiorrhiza Bunge). According to previous studies, it was shown to have various effects such as anti-oxidative stress, antidiabetic complications and antipulmonary hypertension. This study aimed to investigate the effect of SAA on pulmonary arterial endothelial-mesenchymal transition (EndoMT) induced by hypoxia and the underlying mechanisms. Primary cultured human pulmonary arterial endothelial cells (HPAECs) were exposed to 1% O2 for 48[Formula: see text]h with or without SAA treatment. SAA treatment improved the morphology of HPAECs and inhibited the cytoskeleton remodeling. A total of 3[Formula: see text][Formula: see text]M SAA reduced migration distances from 262.2[Formula: see text][Formula: see text]m to 198.4[Formula: see text][Formula: see text]m at 24[Formula: see text]h and 344.8[Formula: see text][Formula: see text]m to 109.3[Formula: see text][Formula: see text]m at 48[Formula: see text]h. It was observed that the production of ROS in cells was significantly reduced by the treatment of 3[Formula: see text][Formula: see text]M SAA. Meanwhile, SAA alleviated the loss of CD31 and slightly inhibited the expression of [Formula: see text]-SMA. The mechanisms study shows that SAA treatment increased the phosphorylation levels of Smad1/5, but inhibited that of Smad2/3. Furthermore, SAA attenuated the phosphorylation levels of ERK and Cofilin, which were enhanced by hypoxia. Based on these results, our study indicated that SAA treatment can protect HPAECs from endoMT induced by hypoxia, which may perform via the inhibition on ROS production and further through the downstream effectors of BMPRs or TGF[Formula: see text]R including Smads, ERK and ROCK/cofilin pathways.

The Protective Effect of DL0805 Derivatives on Pulmonary Artery Cells and the Underlying Mechanisms Study.

BACKGROUND: Pulmonary hypertension (PH) is a severe disease characterized by a progressive increase in pulmonary vascular resistance, initially due to abnormal pulmonary vasoconstriction in response to endothelial and smooth muscle cells injury. The discovery of new chemical entities having a protective effect on pulmonary artery cells could be meaningful for the treatment of PH. METHODS: We evaluated the protective effect of DL0805 derivatives (DL0805-1 and DL0805-2) on pulmonary artery vascular cells, including human pulmonary artery endothelial cells (HPAECs) and human pulmonary artery smooth muscle cells (HPASMCs). DL0805 derivatives are novel ROCKs (Rho-associated coiled-coil forming protein serine/threonine kinases) inhibitors. Treatment of HPAECs with DL0805-2 (10 µM) cultured under a hypoxic environment could significantly reduce the proliferation of cells. Meanwhile, the compounds inhibited the production of reactive oxygen species (ROS) in HPAECs at every dose tested. RESULTS: A Western Blot experiment showed that the protective effect of DL0805 derivatives might result from the down-regulation of RhoA (Ras homolog gene family, member A) expression and the inhibition of ROCKs activity. In addition, the compounds inhibited the proliferation of HPASMCs induced by fetal bovine serum (FBS) or platelet derived growth factor BB (PDGF-BB), and suppressed the F-actin remodeling induced by endothelin. CONCLUSION: The preliminary results from an immunofluorescence assay showed that DL0805 derivatives inhibited the activity of ROCKs in HPASMCs. The above mentioned results indicated that DL0805 derivatives have a protective effect on pulmonary artery cells, and the underlying mechanisms might be the result of inhibition of RhoA/ROCK signaling pathway.

Isolation and bioactivity of diterpenoids from the roots of Salvia grandifolia.

A phytochemical investigation of root extracts of Salvia grandifolia led to isolation of six previously unreported diterpenoids, grandifolias A-F, along with eight known compounds. The structures of grandifolias A-F were primarily established by extensive 1D and 2D NMR spectroscopic analyses, as well as HRESIMS data. Their absolute configurations were assigned by their calculated and experimental electronic circular dichroism spectra or by X-ray diffraction analysis. All of the diterpenoids were evaluated for their vasorelaxant effects. Grandifolia B and isograndifoliol both exhibited dose-dependent vasorelaxant effects on rat aortic rings, preconstricted by KCl or norepinephrine, with EC50 values of 36.36-74.51µg/mL.

Syntheses and structure-activity relationships in cytotoxicities of 13-substituted quaternary coptisine derivatives.

Twenty five 13-substituted quaternary coptisine derivatives were synthesized to test their cytotoxicities against several cancer cell-lines and on intestinal epithelial cell-6 (IEC-6) in vitro to evaluate structure-activity relationship (SAR). Introduction of the alkyl groups into the C-13 position of quaternary coptisine (1) led to significant increase of the cytotoxic activity, while the substitution of arylmethyl groups and others at the same position showed no effect on improving cytotoxicities against the same cancer cell-lines. The cytotoxicities of quaternary 13-alkylcoptisines was significantly reinforced as the length of the aliphatic chain increased, with quaternary 13-n-undecylcoptisine (4l) showing 7, 23, 12, and 9 times, respectively, more active than quaternary coptisine (1) against HCT, A549, Bel7402, and C33A, and being 4, 11, 2, and 3 times, respectively, more active than the positive control, fluorouracil (5-FU), against the same cell-lines, by IC50 values. In comparison to quaternary 13-n-undecylcoptisine (4l) and the above references, quaternary 13-n-dodecylcoptisine (4m) almost showed the same cytotoxicities. In contrast with the n-alkyl chains, the arylmethyl substituents at C-13 displayed low cytotoxicity, except for naphthyl rings or phenyl rings with CF3 or methyl substituents. However, their low cytotoxicity could make them useful as drug candidates for other diseases (bowel, etc).

Inhibitory effects of Brazilin on the vascular smooth muscle cell proliferation and migration induced by PDGF-BB.

Abnormal vascular smooth muscle cell (VSMC) proliferation and migration contribute to the pathogenesis of vascular diseases including atherosclerosis and restenosis. Brazilin isolated from the heartwood of Caesalpinia sappan L. has been reported to exhibit various biological activities, such as anti-platelet aggregation, anti-inflammation, vasorelaxation and pro-apoptosis. However, the functional effects of Brazilin on VSMCs remain unexplored. The present study investigated the potential effects of Brazilin on platelet-derived growth factor (PDGF)-BB induced VSMC proliferation and migration as well as the underlying mechanism of action. VSMC proliferation and migration were measured by Crystal Violet Staining, wound-healing and Boyden chamber assays, respectively. Cell cycle was analyzed by flow cytometry. Enzymatic action of matrix metalloproteinase-9 (MMP-9) was carried out by gelatin zymography. Expression of adhesion molecules, cell cycle regulatory proteins, the phosphorylated levels of PDGF receptor ß (PDGF-Rß), Src, extracellular signal regulated kinase (ERK) and Akt were tested by immunoblotting. The present study demonstrated that pretreatment with Brazilin dose-dependently inhibited PDGF-BB stimulated VSMC proliferation and migration, which were associated with a cell-cycle arrest at G0/G1 phase, a reduction in the adhesion molecule expression and MMP-9 activation in VSMCs. Furthermore, the increase in PDGF-Rß, Src, ERK1/2 and Akt phosphorylation induced by PDGF-BB were suppressed by Brazilin. These findings indicate that Brazilin inhibits PDGF-BB induced VSMC proliferation and migration, and the inhibitory effects of Brazilin may be associated with the blockade of PDGF-Rß - ERK1/2 and Akt signaling pathways. In conclusion, the present study implicates that Brazilin may be useful as an anti-proliferative agent for the treatment of vascular diseases.

Coptisine protects rat heart against myocardial ischemia/reperfusion injury by suppressing myocardial apoptosis and inflammation.

OBJECTIVE: Protecting the heart from myocardial ischemia and reperfusion (I/R) damage is the focus of intense research. Coptisine is an isoquinoline alkaloid isolated from Coptidis Rhizoma. The present study investigated the potential effect of coptisine on myocardial I/R damage in rats and the underlying mechanisms. METHODS AND RESULTS: Electrocardiogram examination showed that the administration of coptisine 10 min before ischemia significantly decreased I/R-induced arrhythmia after 30 min ischemia followed by 3 h reperfusion. The release of cardiac markers was also limited. Echocardiography was performed before ischemia and 24 h post-I/R, separately. The M-mode records showed that the reductions of ejection fraction (EF) and fractional shortening (FS) were attenuated in coptisine-treated rats compared with the I/R rats. Similar results were obtained with Evans Blue/triphenyl tetrazolium chloride (TTC) staining, in which coptisine notably reduced infarct size. Moreover, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay demonstrated coptisine suppressed myocardial apoptosis, which may be related to the upregulation of Bcl-2 protein and inhibition of caspase-3 activation. Coptisine treatment also attenuated the proinflammatory cytokines including interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α in heart tissue. Additionally, Western blot and immunohistochemical analysis showed that coptisine markedly reduced Rho, Rho-kinase 1 (ROCK1), and ROCK2 expression and attenuated the phosphorylation of myosin phosphatase targeting subunit-1, a downstream target of ROCK. CONCLUSIONS: Coptisine exerts pronounced cardioprotection in rats subjected to myocardial I/R likely through suppressing myocardial apoptosis and inflammation by inhibiting the Rho/ROCK pathway.

[Progress in study of pharmacological effect of Cortex Fraxini].

Cortex Fraxini is a commonly used traditional Chinese medicine. It has been indicated that Cortex Fraxini possess various pharmacological effects, including anti-pathogenic microorganism, anti-inflammatory, analgesic, anti-cancer, anti-oxidative stress, neuroprotective and vascular protect effects. Cortex Fraxini has been therapeutically used in the treatment of inflammations, bacillary dysentery, relieving fever, relieving cough and asthma. This paper reviewed recent progress in the studies on pharmacological effects and clinical applications of Cortex Fraxini.

Inhibitory effects of tetrandrine on the serum- and platelet-derived growth factor-BB-induced proliferation of rat aortic smooth muscle cells through inhibition of cell cycle progression, DNA synthesis, ERK1/2 activation and c-fos expression.

Tetrandrine (TET) is a well known naturally occurred nonspecific Ca(2+) channel blocker. It has long been used for the treatment of arrhythmia, hypertension, and occlusive cardiovascular disorders. The objective of the present study was to investigate the effect of TET on the proliferation of primary cultured rat aortic smooth muscle cells (RASMCs). TET significantly inhibited both 10% fetal bovine serum (FBS) and 50 ng/ml platelet-derived growth factor (PDGF)-BB-induced proliferation, [(3) H] ]thymidine incorporation into DNA, and p42/p44 mitogen-activated protein kinase (ERK1/2) phosphorylation at the concentration of 1.0 and 5.0 microM. Flow cytometry analysis of DNA content in synchronized cells revealed blocking of the FBS-inducible cell cycle progression by TET. In accordance with these findings, TET 5 microM caused a 48% decrease in the early elevation of c-fos expression induced after 10% FBS addition. Furthermore, in contrast to its distinguishable higher potency of Ca(2+) antagonistic activity, verapamil showed lower potent antiproliferative activities than TET. These results suggest that TET can exert antiproliferative effects against mitogenic stimuli for RASMCs in vitro by a mechanism that involves the MAPK pathway, altering cell cycle progression, and the inhibitory action cannot be limited to its Ca(2+) modulation.