Ellagic acid protects dopamine neurons from rotenone-induced neurotoxicity via activation of Nrf2 signalling.

Parkinson's disease (PD) is the second most prevalent central nervous system (CNS) degenerative disease. Oxidative stress is one of key contributors to PD. Nuclear factor erythroid-2-related factor 2 (Nrf2) is considered to be a master regulator of many genes involved in anti-oxidant stress to attenuate cell death. Therefore, activation of Nrf2 signalling provides an effective avenue to treat PD. Ellagic acid (EA), a natural polyphenolic contained in fruits and nuts, possesses amounts of pharmacological activities, such as anti-oxidant stress and anti-inflammation. Recent studies have confirmed EA could be used as a neuroprotective agent in neurodegenerative diseases. Here, mice subcutaneous injection of rotenone (ROT)-induced DA neuronal damage was performed to investigate EA-mediated neuroprotection. In addition, adult Nrf2 knockout mice and different cell cultures including MN9D-enciched, MN9D-BV-2 and MN9D-C6 cell co-cultures were applied to explore the underlying mechanisms. Results demonstrated EA conferred neuroprotection against ROT-induced DA neurotoxicity. Activation of Nrf2 signalling was involved in EA-mediated DA neuroprotection, as evidenced by the following observations. First, EA activated Nrf2 signalling in ROT-induced DA neuronal damage. Second, EA generated neuroprotection with the presence of astroglia and silence of Nrf2 in astroglia abolished EA-mediated neuroprotection. Third, EA failed to produce DA neuroprotection in Nrf2 knockout mice. In conclusion, this study identified EA protected against DA neuronal loss via an Nrf2-dependent manner.

Inflammation-Related MicroRNAs Are Associated with Plaque Stability Calculated by IVUS in Coronary Heart Disease Patients.

OBJECTIVES: This study aimed to investigate the association between inflammation-related microRNAs (miR-21, 146a, 155) and the plaque stability in coronary artery disease patients. METHODS: The expression of miR-21, 146a, and 155 was measured by real-time PCR in 310 consecutive patients. The level of hs-CRP, IL-6, and IL-8 was measured by ELISA. The plaque stability of coronary stenotic lesions was evaluated with intravascular ultrasound (IVUS). RESULTS: (1) The levels of hs-CRP, IL-6, and IL-8 were significantly increased in the UAP and AMI groups compared with the CPS group (P < 0.01). (2) The expression of miR-21 and miR-146a in peripheral blood mononuclear cells (PBMCs) and plasma was significantly higher in CAD patients compared with non-CAD patients, whereas the miR-155 expression in PBMCs and plasma was significantly lower in patients with CAD. (3) The miR-21 expression in PBMCs was higher in UAP and AMI groups compared with CPS group. The miR-146a expression in PBMCs was higher in SAP, UAP, and AMI groups than in CPS group. Although the level of miR-155 in PBMCs was lower in SAP, UAP, and AMI groups than in CPS group. The expression patterns of miR-21, miR-146a, and miR-155 in plasma were consistent with those of PBMCs. (4) The expressions of miR-21 and miR-146a in PBMCs and plasma were significantly higher in the vulnerable plaque group than those in stable plaque group. While miR-155 in PBMCs and plasma was significantly lower in vulnerable plaque group compared with stable plaque group. (5) The levels of miR-21 and miR-146a in PBMCs and plasma were significantly higher in soft plaque group than in fibrous plaque group and calcified plaque group. However, miR-155 in PBMCs and plasma was significantly lower in soft plaque group. CONCLUSIONS: The expression of miR-21 and miR-146a are associated with the plaque stability in coronary stenotic lesions, whereas miR-155 expression is inversely associated with the plaque stability.

Ellagic acid supports neuron by regulating astroglia Nrf2.

Astroglia support neuron by providing substrates for neuronal metabolism, glutamate clearance, and antioxidative protection. Nuclear factor erythroid 2-related factor 2 (Nrf2) participates in the antioxidative defense response. Also, Nrf2 signaling is recognized to activate the neurotrophic pathway to replace/protect damaged organelles. Ellagic acid (EA), an extraction component of fruits and nuts, presents many pharmacological activities such as anti-inflammation, antioxidation, and neuroprotection. However, few studies have been focused on the neurotrophic properties of EA. Our study investigated whether EA could increase neuronal survival and the target cells. Thus, primary neuron-enriched cultures and primary astroglia-enriched cultures were applied to detect whether EA-elicited neurotrophic effects were mediated by astroglia Nrf2. This study indicated that EA promoted neuronal survival. Further, astroglia Nrf2 participate in EA-elicited neuronal survival with the following scenarios. First, EA elicited astroglia proliferation, glial cell line-derived neurotrophic factor (GDNF) release, and Nrf2 activation. Second, after silencing astroglia Nrf2, EA-induced astrogliosis, GDNF release, and neuronal survival disappeared. Thus, EA-mediated astroglia Nrf2 activation is important to enhance neurotrophic effects on neurons, which might provide new insights for neurodegenerative disease.

Dual specific phosphatase 12 ameliorates cardiac hypertrophy in response to pressure overload.

Pathological cardiac hypertrophy is an independent risk factor of heart failure. However, we still lack effective methods to reverse cardiac hypertrophy. DUSP12 is a member of the dual specific phosphatase (DUSP) family, which is characterized by its DUSP activity to dephosphorylate both tyrosine and serine/threonine residues on one substrate. Some DUSPs have been identified as being involved in the regulation of cardiac hypertrophy. However, the role of DUSP12 during pathological cardiac hypertrophy is still unclear. In the present study, we observed a significant decrease in DUSP12 expression in hypertrophic hearts and cardiomyocytes. Using a genetic loss-of-function murine model, we demonstrated that DUSP12 deficiency apparently aggravated pressure overload-induced cardiac hypertrophy and fibrosis as well as impaired cardiac function, whereas cardiac-specific overexpression of DUPS12 was capable of reversing this hypertrophic and fibrotic phenotype and improving contractile function. Furthermore, we demonstrated that JNK1/2 activity but neither ERK1/2 nor p38 activity was increased in the DUSP12 deficient group and decreased in the DUSP12 overexpression group both in vitro and in vivo under hypertrophic stress conditions. Pharmacological inhibition of JNK1/2 activity (SP600125) is capable of reversing the hypertrophic phenotype in DUSP12 knockout (KO) mice. DUSP12 protects against pathological cardiac hypertrophy and related pathologies. This regulatory role of DUSP12 is primarily through c-Jun N-terminal kinase (JNK) inhibition. DUSP12 could be a promising therapeutic target of pathological cardiac hypertrophy. DUSP12 is down-regulated in hypertrophic hearts. An absence of DUSP12 aggravated cardiac hypertrophy, whereas cardiomyocyte-specific DUSP12 overexpression can alleviate this hypertrophic phenotype with improved cardiac function. Further study demonstrated that DUSP12 inhibited JNK activity to attenuate pathological cardiac hypertrophy.

Smad Nuclear Interacting Protein 1 Acts as a Protective Regulator of Pressure Overload-Induced Pathological Cardiac Hypertrophy.

BACKGROUND: Smad nuclear interacting protein 1 (SNIP1) plays a critical role in cell proliferation, transformation of embryonic fibroblasts, and immune regulation. However, the role of SNIP1 in cardiac hypertrophy remains unclear. METHODS AND RESULTS: Here we examined the role of SNIP1 in pressure overload-induced cardiac hypertrophy and its mechanisms. Our results demonstrated that SNIP1 expression was downregulated in human dilated cardiomyopathic hearts, aortic banding-induced mice hearts, and angiotensin II-treated cardiomyocytes. Accordingly, SNIP1 deficiency significantly exacerbated aortic banding-induced cardiac hypertrophy, fibrosis, and contractile dysfunction, whereas cardiac-specific overexpression of SNIP1 markedly recovered pressure overload-induced cardiac hypertrophy and fibrosis. Besides that, SNIP1 protected neonatal rat cardiomyocytes against angiotensin II-induced hypertrophy in vitro. Moreover, we identified that SNIP1 suppressed nuclear factor-κB signaling during pathological cardiac hypertrophy, and inhibition of nuclear factor-κB signaling by a cardiac-specific conditional inhibitor of κBS32A/S36A transgene blocked these adverse effects of SNIP1 deficiency on hearts. CONCLUSIONS: Together, our findings demonstrated that SNIP1 had protective effects in pressure overload-induced pathological cardiac hypertrophy via inhibition of nuclear factor-κB signaling. Thus, SNIP1 may be a novel approach for the treatment of heart failure.

Growth Arrest-Specific 6 Exacerbates Pressure Overload-Induced Cardiac Hypertrophy.

Growth arrest-specific 6 (GAS6) is a member of the vitamin K-dependent protein family that is involved in the regulation of the cardiovascular system, including vascular remodeling, homeostasis, and atherosclerosis. However, there is still no study that systemically elucidates the role of GAS6 in cardiac hypertrophy. Here, we found that GAS6 was upregulated in human dilated cardiomyopathic hearts, hypertrophic murine hearts, and angiotensin II-treated cardiomyocytes. Next, we examined the influence of GAS6 expression in response to a cardiac stress by inducing chronic pressure overload with aortic banding in wild-type and GAS6-knockout mice or cardiac-specific GAS6 overexpressing mice. Under basal conditions, the GAS6-knockout mice had normal left ventricular structure and function but after aortic banding, the mice demonstrated less hypertrophy, fibrosis, and contractile dysfunction when compared with wild-type mice. Conversely, cardiac-specific overexpression of GAS6 exacerbated aortic banding-induced cardiac hypertrophy, fibrosis, and dysfunction. Furthermore, we demonstrated that GAS6 activated the mitogen-activated protein kinase kinase 1/2-extracellular signal-regulated kinase 1/2 pathway during pressure overload-induced cardiac hypertrophy, and the pharmacological mitogen-activated protein kinase kinase 1/2 inhibitor U0126 almost completely reversed GAS6 overexpression-induced cardiac hypertrophy and fibrosis, resulting in improved cardiac function. Collectively, our data support the notion that GAS6 impairs ventricular adaptation to chronic pressure overload by activating mitogen-activated protein kinase kinase 1/2-extracellular signal-regulated kinase 1/2 signaling. Our findings suggest that strategies to reduce GAS6 activity in cardiac tissue may be a novel approach to attenuate the development of congestive heart failure.

[Application of zebra fishes in studies on traditional Chinese medicines].

The zebra fish model, as an integral animal model, features small volume, high throughput, low cost, short cycle and reliable experimental results, thus has been widely used in medical studies. Traditional Chinese medicines (TCM) constitute a complex system, their active ingredients and action mechanisms are among study hotspots in during the development of modern TCMs. Along with the constant improvement of advanced technologies and methods, zebra fishes have been increasingly applied in studies on TCMs and shown advantages in active screening, and toxicity and metabolism studies. In this paper, TCM studies by using zebra fishes in recent years are summarized to provide new ideas and methods for basic studies on TCMs.

Four new bis-iridoids isolated from the traditional Tibetan herb Pterocephalus hookeri.

Pterocenoids A-E (1-4), which Pterocenoids A(1) is one novel dimer containing a pyridine monoterpene alkaloid; and Pterocenoids B-E (2-4) are rare arranged non-glycosidic bis-iridoids were isolated from Pterocephlus hookeri. The structures of the compounds were established by 1D and 2D NMR spectroscopy and mass spectrometry. All bis-iridoids isolated from P. hookeri were found to possess secoiridoid/iridoid subtype skeletons. Hence, bis-iridoids can be regarded as the chemotaxonomic markers of P. hookeri. The origins of the new bis-iridoids (1-4) were postulated and their activities of inhibition of the NF-κB pathway were assayed and compounds 1-3 showed moderate activity in inhibiting NF-κB.

MicroRNA-155 inhibits angiotensin II-induced vascular smooth muscle cell proliferation.

BACKGROUND: MicroRNA-155 (miR-155) is a multifunctional signal microRNA that participates in a variety of cardiovascular diseases and is involved in physiological and pathological processes in different cell types. OBJECTIVE: The objective of this article is to examine the effect of miR-155 on angiotensin II (Ang II)-induced primary mice vascular smooth muscle cell (VSMC) proliferation. METHODS: Primary cultured VSMCs from the aorta of C57/BL6 mice were incubated with Ang II and miR-155. Cells were counted using CCK-8 and EdU, and flow cytometric analysis of cell cycle progression was performed. Angiotensin II 1 type receptor (AT1R) gene and protein expression were measured by real-time polymerase chain reaction and Western blotting. RESULTS: 1) Ang II increased the viability of VSMCs in a dose- and time-dependent manner. 2) miR-155 opposed the Ang II-induced increase in VSMC viability. 3) miR-155 inhibited Ang II-induced proliferation of VSMCs. 4) miR-155 increased the number of VSMCs in the G1 phase compared to G2 and M cell cycle phases. 5) miR-155 decreased ATR1 gene and protein expression. CONCLUSION: miR-155 downregulation of Ang II-induced VSMC viability identifies it as an important regulator of cell proliferation.

microRNA-155 is inversely associated with severity of coronary stenotic lesions calculated by the Gensini score.

OBJECTIVES: This study aimed to investigate the association between microRNA-155 (miR-155) and the severity and extent of coronary stenotic lesions. PATIENTS AND METHODS: We measured the miR-155 expression by real-time PCR in 110 consecutive patients undergoing coronary angiography for suspected coronary artery disease. The severity and extent of coronary stenotic lesions were evaluated on the basis of coronary angiography findings by the Gensini score. RESULTS: The miR-155 expression was significantly lower in 56 patients with coronary heart disease than those in 54 controls (P<0.01). The level of miR-155 in peripheral blood mononuclear cells or plasma was lower in patients with unstable angina pectoris and acute myocardial infarction than in patients with chest pain syndrome, whereas no statistically significant differences were observed between patients with stable angina pectoris and chest pain syndrome. Spearman's correlation analysis showed that the expression of miR-155 in plasma correlated positively with the expression in peripheral blood mononuclear cells. The levels of miR-155 in the patients with diseased vessels of two and three or more were significantly lower than in those with diseased vessel of zero and one. The levels of miR-155 were not significantly different among groups with diseased vessels of zero and one. miR-155 were associated negatively with Gensini scores (r = -0.663, P<0.001). The miR-155 expression was correlated significantly to age (r = -0.227), hypertension (r = -0.440), total cholesterol (r = 0.239), high-density lipoprotein cholesterol (r = 0.280), low-density lipoprotein cholesterol (r = -0.315), tobacco use (r = -0.363), angiotensin-converting enzyme inhibitor (r = -0.250), statins (r = -0.368), and high-sensitivity C-reactive protein (r = -0.515). CONCLUSION: miR-155 expression is associated inversely with complicated proatherogenic metabolic risk factors, and the severity of coronary stenotic lesions calculated by Gensini scores.

miR-155 inhibits oxidized low-density lipoprotein-induced apoptosis of RAW264.7 cells.

Macrophage apoptosis is a prominent feature of advanced atherosclerotic plaques. Here, we examined the hypothesis that the apoptotic machinery is regulated by microRNA-155 (miR-155). Constitutive expression of miR-155 was detected in RAW264.7 cells, which was increased following stimulation with oxidized low-density lipoprotein (OxLDL) in a dose- and time-dependent manner. OxLDL-treated RAW264.7 cells showed a marked time- and dose-dependent increase in apoptosis, which was suppressed in the presence of mimics and increased with antagonists of miR-155. Bioinformatics analysis revealed Fas-associated death domain-containing protein (FADD) as a putative target of miR-155. Luciferase reporter assay and Western blot further disclosed that miR-155 inhibits FADD expression by directly targeting the 3'-UTR region. We propose that miR-155 attenuates the macrophage apoptosis, at least in part, through FADD regulation, since forced expression of FADD blocked the ability of miR-155 to inhibit apoptosis. Our results collectively suggest that miR-155 attenuates apoptosis of OxLDL-mediated RAW264.7 cells by targeting FADD, supporting a possible therapeutic role in atherosclerosis.

Functional blockage of EMMPRIN ameliorates atherosclerosis in apolipoprotein E-deficient mice.

BACKGROUND: Extracellular matrix metalloproteinase inducer (EMMPRIN), a 58-kDa cell surface glycoprotein, has been identified as a key receptor for transmitting cellular signals mediating metalloproteinase activities, as well as inflammation and oxidative stress. Clinical evidence has revealed that EMMPRIN is expressed in human atherosclerotic plaque; however, the relationship between EMMPRIN and atherosclerosis is unclear. To evaluate the functional role of EMMPRIN in atherosclerosis, we treated apolipoprotein E-deficient (ApoE(-/-)) mice with an EMMPRIN function-blocking antibody. METHODS AND RESULTS: EMMPRIN was found to be up-regulated in ApoE(-/-) mice fed a 12-week high-fat diet in contrast to 12 weeks of normal diet. Administration of a function-blocking EMMPRIN antibody (100 µg, twice per week for 4 weeks) to ApoE(-/-) mice, starting after 12 weeks of high-fat diet feeding caused attenuated and more stable atherosclerotic lesions, less reactive oxygen stress generation on plaque, as well as down-regulation of circulating interleukin-6 and monocyte chemotactic protein-1 in ApoE(-/-) mice. The benefit of EMMPRIN functional blockage was associated with reduced metalloproteinases proteolytic activity, which delayed the circulating monocyte transmigrating into atherosclerotic lesions. CONCLUSION: EMMPRIN antibody intervention ameliorated atherosclerosis in ApoE(-/-) mice by the down-regulation of metalloproteinase activity, suggesting that EMMPRIN may be a viable therapeutic target in atherosclerosis.

New isoprenylated flavonoids and cytotoxic constituents from Artocarpus tonkinensis.

Two new isoprenylated flavonoids, artotonins A and B (1 and 2), along with 13 known compounds (3-15), were isolated from the roots of Artocarpus tonkinensis A. Chev. ex Gagnep. The structures were elucidated by spectroscopic methods. Cyclocommunol (6), isocyclomulberrin (7), cudraflavone C (11), and morusin (13) exhibited cytotoxicity against hepatocellular carcinoma (SMMC-7721) and gastric carcinoma (BGC-823 and SGC-7901) cell lines.

[Effects of cycloartocarpin A and artocarpin extracted from Fructus Artocarpin Heterophylli on apoptosis of SMMC-7721 and SGC-7901 cells].

OBJECTIVE: To investigate the effects of cycloartocarpin A (ACR-2) and artocarpin (ACR-3), monomeric compounds isolated from Fructus Artocarpi Heterophylli, on apoptosis of SMMC-7721 and SGC-7901 cell lines. METHODS: SMMC-7721 and SGC-7901 cells were routinely cultured, and divided into experiment group and control group. The SMMC-7721 cells were treated with different concentrations of ACR-2 (3.46 x 10(-3), 13.82 x (-3), 55.30 x 10(-3) mmol/L) and ACR-3 (6.88 x 10(-3), 27.52 x 10(-3), 110.09 x 10(-3) mmol/L), and the SGC-7901 cells were also treated with different concentrations of ACR-2 (8.06 x 10(-3), 32.26 x 10(-3), 129.03 x 10(-3) mmol/L) and ACR-3 (2.87 x 10(-3), 11.47 x 10(-3), 45.87 x 10(-3) mmol/L), with PBS (DMSO<0.1%) as control treatment. Cell apoptosis was measured by double labeled staining with Hoechst33342/propidium iodide (PI) and TdT-mediated dUTP-biotin nick end labeling (TUNEL) and flow cytometry. RESULTS: ACR-2 and ACR-3 could induce apoptosis of SMMC-7721 and SGC-7901 cells. Some of SMMC-7721 and SGC-7901 cells demonstrated typical apoptosis after being treated with ACR-2 and ACR-3. Hoechst33342/PI staining showed that cells were fraught with overlapping nuclei and nuclear debris or lobule, and the nuclear appeared light blue. TUNEL showed that cells permeated with overlapping nuclei and nuclear debris or lobule, and the nuclear appeared brown. Less apoptotic cells were observed in negative control group, and the nuclear appeared light blue. The apoptosis rates of SMMC-7721 and SGC-7901 cells in the ACR-2 and ACR-3 treated groups were significant higher than those in the control group (P<0.05, P<0.01). CONCLUSION: ACR-2 and ACR-3 can induce apoptosis of SMMC-7721 and SGC-7901 cells.

[Effects of three compounds extracted from Tripterygium wilfordii Hook on angiogenesis in chick chorioallantoic membrane].

OBJECTIVE: To investigate the effects of three compounds extracted from Tripterygium wilfordii Hook (TW) on angiogenesis in the chick chorioallantoic membrane (CAM). METHODS: Fifty fresh Hongkong Mahua chicken eggs were divided into five groups: PBS-treated group, TW1-, TW2- and TW3-treated groups and Rg3-treated group. After disinfection, the eggs were incubated for six days in a constant temperature box with the temperature being controlled within 37.8 degrees C, then exposed CAM, laid the filter papers with specimen on the CAM, and the eggs were incubated for another two days. CAM was fixed with the mixture of methyl alcohol and acetone at room temperature for about 15 min, and then cutting the CAM, taking photos and observing the angiogenesis in the CAM. RESULTS: There were many CAM vessels in the PBS-treated group and the blood vessel net could be seen clearly. The number of CAM vessels in the TW1-, TW2- and TW3-treated groups (10 microg/egg) was much less than that in the PBS-treated group. Furthermore, the frame of the vessels was not clear, and the color was obscure. Inhibition rates of angiogenesis in the TW1-, TW2- and TW3-treated groups were 80%, 60% and 100% respectively, while the inhibition rate of angiogenesis in the Rg3-treated group (10 microg/egg) was only 10%. CONCLUSION: TW1, TW2 and TW3 can obviously restrain the angiogenesis in CAM and still need further study.

Cytotoxic and antifungal isoprenylated xanthones and flavonoids from Cudrania fruticosa.

A new isoprenylated xanthone, cudrafrutixanthone A, was isolated from the roots of Cudrania fruticosa, together with 18 known compounds. Their structures were elucidated by spectroscopic methods. Most isoprenylated xanthones exhibited cytotoxicity against HCT-116, SMMC-7721, SGC-7901, and BGC-823 cell lines with IC (50) values of 1 - 5 microg/mL. Toxyloxanthone C and wighteone showed antifungal activity against Candida albicans with MICs of 25 and 12.5 microg/mL, respectively.

Isoprenylated xanthones and flavonoids from Cudrania tricuspidata.

Further phytochemical investigation on the roots of Cudrania tricuspidata afforded a new isoprenylated xanthone, cudratricusxanthone I (1), two new isoprenylated flavanones, cudraflavanones C and D (2 and 3, resp.), and seven known compounds, 1,7-dihydroxy-3,6-dimethoxyxanthone (4), macluraxanthone C (5), cudraxanthones E, K, and L (6, 7, and 8, resp.), cudraflavanone A (9), and cudraflavone C (10). Their structures were identified by spectroscopic methods. Cudratricusxanthone H (12), macluraxanthone B (13), two xanthones previously isolated from this plant, and 5, showed significant inhibitory effects on four kinds of human digestive apparatus tumor cell lines (HCT-116, SMMC-7721, SGC-7901, and BGC-823) with IC50 values of 2.70-12.66 microM.

Cytotoxic isoprenylated xanthones from Cudrania tricuspidata.

Eight new isoprenylated xanthones, cudratricusxanthones A-H (1-8), were isolated from the roots of Cudrania tricuspidata, together with ten known compounds, cudraxanthones H (9) and M (10), xanthone V(1a) (11), toxyloxanthone C (12), macluraxanthone B (13), 1-hydroxy-3, 6, 7-trimethoxyxanthone (14), cycloartocarpesin (15), artocarpesin (16), cudraflavone B (17), and kaempferol (18). Their structures were characterized by spectroscopic methods. Xanthones 5, 7, 10, and 12 showed inhibitory effects on four kinds of human digestive apparatus tumor cell lines (HCT-116, SMMC-7721, SGC-7901, and BGC-823) with IC(50) values of 1.6-11.8 microg/mL. Xanthones 2, 4, and 11 displayed significant cytotoxicity against HCT-116, SMMC-7721, and SGC-7901 (IC(50)=1.3-9.8 microg/mL). Flavonoids 15-17 were almost inactive.