Phellinus merrillii extracts induce apoptosis of vascular smooth muscle cells via intrinsic and extrinsic pathways.

Restenosis frequently occurs after balloon angioplasty. Percutaneous coronary intervention (PCI)-induced artery damage is a significant part of triggering restenosis of the vascular smooth muscles (VSMC). This study aimed to study how ethanol extract of Phellinus merrillii (EPM) affected balloon injury-induced overgrowth of VSMC, indicating neointima formation. Firstly, our results demonstrated that EPM notably decreased VSMC viability. A fragmentation assay and Annexin V/Propidium Iodide apoptosis assay showed that higher doses of EPM significantly induced the apoptosis of VSMC after 24 h of exposure. Total protein extracted from VSMC treated with EPM in various time and concentration periods was then conducted in Western blotting analysis. Our data demonstrated that EPM substantially elevated the p53, p21, Fas, Bax, p-p38, and active caspase-3 protein expressions. The results indicated that EPM induces VSMC apoptosis via intrinsic and extrinsic pathways. Also, our results demonstrated that EPM effectively attenuated the balloon injury-induced neointima formation. In conclusion, the information offers a mechanism of EPM in inducing the VSMC apoptosis, thus as a potential interference for restenosis.

The Wound-Healing Activity of PEDOT-PSS in Animals.

This study evaluated the wound-healing activity of a polymer, Poly(3,4-ethylenedioxythiophene):poly-(styrene sulfonate) (PEDOT: PSS), and determined its mechanism based on angiogenic activity in a full-thickness excision wound model in Spraque Dawley (SD) rats. Administering PEDOT: PSS (1.6) 1.5 ppm at a dose of 50 mg/kg/day significantly improved wound healing in the SD rats on the eleventh day after the incision was created. PEDOT: PSS-treated animals presented no anti-inflammatory skin effects; however, there was an increase in angiogenic behavior. VEGF was found to be significantly elevated in the PEDOT: PSS-treated groups seven days post-incision. However, only a higher concentration of PEDOT: PSS increased TGF-ß1 expression within the same time frame. Our results showed that PEDOT: PSS enhances wound healing activity, mainly in terms of its angiogenic effects. In this paper, we describe the highly conductive macromolecular material PEDOT: PSS, which demonstrated accelerated wound-healing activity in the animal incision model. The results will further provide information regarding the application of PEDOT: PSS as a dressing for medical use.

Carbonic Anhydrase VIII (CAVIII) Gene Mediated Colorectal Cancer Growth and Angiogenesis through Mediated miRNA 16-5p.

Carbonic anhydrase VIII (CAVIII) is a member of the CA family, while CA8 is the oncogene. Here we observed increased expression of CAVIII with high expression in colorectal cancer tissues. CAVIII is also expressed in more aggressive types of human colorectal cancer cells. Upregulated CAVIII expression in SW480 cell lines increased vascular endothelial growth factor (VEGF) and reduced miRNA16-5p. Conversely, knockdown of the CAVIII results in VEGF decline by up-regulated miRNA16-5p. Moreover, the collection of different grades of CAVIII expression CRC cells supernatant co-culture with endothelial progenitor cells (EPCs) promotes the ability of tube formation in soft agar and migration in the Transwell experiment, indicating that CAVIII might facilitate cancer-cell-released VEGF via the inhibition of miRNA16-5p signaling. Furthermore, in the xenograft tumor angiogenesis model, knockdown of CAVIII significantly reduced tumor growth and tumor-associated angiogenesis. Taken together, our results prove that the CAVIII/miR-16-5p signaling pathway might function as a metastasis suppressor in CRC. Targeting CAVIII/miR-16-5p may provide a strategy for blocking its metastasis.

Curcuminoids Inhibit Angiogenic Behaviors of Human Umbilical Vein Endothelial Cells via Endoglin/Smad1 Signaling.

BACKGROUND: Angiogenesis is primarily attributed to the excessive proliferation and migration of endothelial cells. Targeting the vascular endothelial growth factor (VEGF) is therefore significant in anti-angiogenic therapy. Although these treatments have not reached clinical expectations, the upregulation of alternative angiogenic pathways (endoglin/Smad1) may play a critical role in drug (VEGF-neutralizing agents) resistance. Enhanced endoglin expression following a VEGF-neutralizing therapy (semaxanib®) was noted in patients. Treatment with an endoglin-targeting antibody augmented VEGF expression in human umbilical vein endothelial cells (HUVECs). Therefore, approaches that inhibit both the androgen and VEGF pathways enhance the HUVECs cytotoxicity and reverse semaxanib resistance. The purpose of this study was to find natural-occurring compounds that inhibited the endoglin-targeting pathway. METHODS: Curcuminoids targeting endoglin were recognized from two thousand compounds in the Traditional Chinese Medicine Database@Taiwan (TCM Database@Taiwan) using Discovery Studio 4.5. RESULTS: Our results, obtained using cytotoxicity, migration/invasion, and flow cytometry assays, showed that curcumin (Cur) and demethoxycurcumin (DMC) reduced angiogenesis. In addition, Cur and DMC downregulated endoglin/pSmad1 phosphorylation. CONCLUSIONS: The study first showed that Cur and DMC demonstrated antiangiogenic activity via the inhibition of endoglin/Smad1 signaling. Synergistic effects of curcuminoids (i.e., curcumin and DMC) and semaxanib on HUVECs were found. This might be attributed to endoglin/pSmad1 downregulation in HUVECs. Combination treatment with curcuminoids and a semaxanib is therefore expected to reverse semaxanib resistance.

Honokiol, an active compound of Magnolia officinalis, is involved in restoring normal baroreflex sensitivity in hypercholesterolemic rabbits.

This study investigated the effects of methanol extract Magnolia officinalis (MEMO) on baroreceptor reflex sensitivity (BRS) in the hypercholesterolemic rabbits and the involved molecular mechanisms. Male New Zealand white rabbits were randomly divided into Control (normal diet), Cholesterol (0.5% w/w cholesterol diet), and Magnolia groups (0.5% w/w cholesterol diet plus 1% w/w MEMO). The animals were treated with the designated diet for 4 or 8 weeks. BRS in the control of heart rate was assessed by linear regression method. After 8 weeks of treatments, plasma total cholesterol (TC) was significantly elevated in the Cholesterol/Magnolia groups. The arterial blood pressure (aBP) was increased in the Cholesterol and Magnolia groups. The depression of BRS observed in the Cholesterol group was significantly ameliorated in the Magnolia group. After L-NAME (Nω-nitro-Larginine methyl ester, 20 mg/kg, iv), the BRS of the Cholesterol group was significantly improved. Results from our in vitro study further indicated that honokiol, the principle component of MEMO, would protect human umbilical vein endothelial cells (HUVECs) from H2O2-induced damages and inhibit H2O2-induced vascular smooth muscles cells (VSMCs) proliferation, which was evident by the decreased expression of pFAK, and p-Erk1/2. The results of the present study suggested that the improvement of BRS by MEMO in the hypercholesterolemic rabbits might be mediated by the antioxidant property of MEMO as indicated by the results from the L-NAME and in vitro honokiol studies.

Pipoxolan Exhibits Antitumor Activity Toward Oral Squamous Cell Carcinoma Through Reactive Oxygen Species-mediated Apoptosis.

Pipoxolan is frequently prescribed as a smooth muscle relaxant. Pipoxolan has also been shown to have anticancer activity. Our study investigated whether pipoxolan induced apoptosis in oral squamous cell carcinoma (OSCC). Cell cytotoxicity was evaluated by the MTT assay. Cell apoptosis and cell-cycle distribution were measured by annexin V/propidium iodide (PI) double staining and flow cytometry, respectively. Apoptotic-related proteins were assessed by western blotting. Reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP) were measured with fluorescent probes. Following exposure of TW206 OSCC cells to pipoxolan, a time-dependently decrease in MMP and an increase in ROS were observed. However, these effects were significantly abrogated by the free radical scavenger N-acetyl-L-cysteine. Since high levels of ROS were produced early in the treatment, intracellular ROS seemed to play a key role in pipoxolan-induced apoptosis. In HSC-3 OSCC cells, our results demonstrated that pipoxolan treatment caused a time-dependent increase of protein expression of active caspase-3 and -9, cytosolic cytochrome c, cleavage of poly (ADP-ribose) polymerase, and B-cell lymphoma 2 (BCL2)-like protein 4 (BAX). However, expression of BCL2 itself was reduced. Clearly, such an increase in BAX/BCL2 ratio would be associated with apoptosis. In addition, pipoxolan markedly suppressed the protein expression of phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) and phosphorylation of protein kinase B (AKT). These data suggest that pipoxolan acts against HSC-3 in vitro via intrinsic apoptotic signaling pathways, and inhibition of PI3K/AKT signaling.

Toxicological effects of NCKU-21, a phenanthrene derivative, on cell growth and migration of A549 and CL1-5 human lung adenocarcinoma cells.

BACKGROUND: Chemotherapy insensitivity continues to pose significant challenges for treating non-small cell lung cancer (NSCLC). The purposes of this study were to investigate whether 3,6-dimethoxy-1,4,5,8-phenanthrenetetraone (NCKU-21) has potential activity to induce effective toxicological effects in different ethnic NSCLC cell lines, A549 and CL1-5 cells, and to examine its anticancer mechanisms. METHODS: Mitochondrial metabolic activity and the cell-cycle distribution were analyzed using an MTT assay and flow cytometry in NCKU-21-treated cells. NCKU-21-induced cell apoptosis was verified by Annexin V-FITC/propidium iodide (PI) double-staining and measurement of caspase-3 activity. Western blotting and wound-healing assays were applied to respectively evaluate regulation of signaling pathways and cell migration by NCKU-21. Molecular interactions between target proteins and NCKU-21 were predicted and performed by molecular docking. A colorimetric screening assay kit was used to evaluate potential regulation of matrix metalloproteinase-9 (MMP-9) activity by NCKU-21. RESULTS: Results indicated that NCKU-21 markedly induced cytotoxic effects that reduced cell viability via cell apoptosis in tested NSCLC cells. Activation of AMP-activated protein kinase (AMPK) and p53 protein expression also increased in both NSCLC cell lines stimulated with NCKU-21. However, repression of PI3K-AKT activation by NCKU-21 was found in CL1-5 cells but not in A549 cells. In addition, increases in phosphatidylserine externalization and caspase-3 activity also confirmed the apoptotic effect of NCKU-21 in both NSCLC cell lines. Moreover, cell migration and translational levels of the gelatinases, MMP-2 and MMP-9, were obviously reduced in both NSCLC cell lines after incubation with NCKU-21. Experimental data obtained from molecular docking suggested that NCKU-21 can bind to the catalytic pocket of MMP-9. However, the in vitro enzyme activity assay indicated that NCKU-21 has the potential to increase MMP-9 activity. CONCLUSIONS: Our results suggest that NCKU-21 can effectively reduce cell migration and induce apoptosis in A549 and CL1-5 cells, the toxicological effects of which may be partly modulated through PI3K-AKT inhibition, AMPK activation, an increase in the p53 protein, and gelatinase inhibition.

Dual Inhibition of Key Proliferation Signaling Pathways in Triple-Negative Breast Cancer Cells by a Novel Derivative of Taiwanin A.

The treatment of breast cancer cells obtained by blocking the aberrant activation of the proliferation signaling pathways PI3K/Akt/mTOR and MEK/ERK has received considerable attention in recent years. Previous studies showed that Taiwanin A inhibited the proliferation of several types of cancer cells. In this study, we report that 3,4-bis-3,4,5-trimethoxybenzylidene-dihydrofuran (BTMB), a novel derivative of Taiwanin A, significantly inhibited the proliferation of triple-negative breast cancer (TNBC) cells both in vitro and in vivo The results show that BTMB inhibited the proliferation of human TNBC cells by the induction of cell-cycle arrest and apoptosis in a dose-dependent fashion. BTMB inhibited the expression of ß-catenin, cdc2 and the cell-cycle regulatory proteins, cyclin A, cyclin D1, and cyclin E. The mechanism of action was associated with the suppression of cell survival signaling through inactivation of the Akt and ERK1/2 signaling pathways. Moreover, BTMB induced cell apoptosis through an increase in the expression of BAX, cleaved caspase-3, and cleaved PARP. Moreover, BTMB inhibited TNBC cell colony formation and sensitized TNBC cells to cisplatin, a chemotherapeutic drug. In a TNBC mouse xenograft model, BTMB significantly inhibited the growth of mammary carcinomas through decreased expression of cyclin D1. BTMB was shown to significantly suppress the growth of mammary carcinoma and therefore to have potential as an anticancer therapeutic agent. Mol Cancer Ther; 16(3); 480-93. ©2016 AACR.

CCL3 promotes angiogenesis by dysregulation of miR-374b/ VEGF-A axis in human osteosarcoma cells.

Osteosarcoma is the most frequent bone tumor, characterized by a high metastatic potential. However, the crosstalk between chemokine (C-C motif) ligand 3 (CCL3), which facilitates tumor progression and metastasis. Vascular endothelial growth factor-A (VEGF-A), an angiogenesis inducer and a highly specific mitogen for endothelial cells, has not been well explored in human osteosarcoma. Here we demonstrate the correlation of CCL3 and VEGF-A expressions, quantified by immunohistochemistry, with the tumor stage of human osteosarcoma tissues. Furthermore, CCL3 promotes VEGF-A expression in human osteosarcoma cells that subsequently induces human endothelial progenitor cell (EPC) migration and tube formation. Phosphorylation of JNK, ERK, and p38 was found after CCL3 stimulation. In addition, JNK, ERK, and p38 inhibitors also abolished CCL3-induced VEGF-A expression and angiogenesis. We noted that CCL3 reduces the expression of miR-374b and miR-374b mimic by reversing CCL3-promoted VEGF-A expression and angiogenesis in vitro and in vivo. This study shows that CCL3 promotes VEGF-A expression and angiogenesis in human osteosarcoma cells by down-regulating miR-374b expression via JNK, ERK, and p38 signaling pathways. Thus, CCL3 may be a new molecular therapeutic target in osteosarcoma angiogenesis and metastasis.

The characteristics of thrombin in osteoarthritic pathogenesis and treatment.

Osteoarthritis (OA) is a mechanical abnormality associated with degradation of joints. It is characterized by chronic, progressive degeneration of articular cartilage, abnormalities of bone, and synovial change. The most common symptom of OA is local inflammation resulting from exogenous stress or endogenous abnormal cytokines. Additionally, OA is associated with local and/or systemic activation of coagulation and anticoagulation pathways. Thrombin plays an important role in the stimulation of fibrin deposition and the proinflammatory processes in OA. Thrombin mediates hemostatic and inflammatory responses and guides the immune response to tissue damage. Thrombin activates intracellular signaling pathways by interacting with transmembrane domain G protein coupled receptors (GPCRs), known as protease-activated receptors (PARs). In pathogenic mechanisms, PARs have been implicated in the development of acute and chronic inflammatory responses in OA. Therefore, discovery of thrombin signaling pathways would help us to understand the mechanism of OA pathogenesis and lead us to develop therapeutic drugs in the future.

Thalamic synaptic transmission of sensory information modulated by synergistic interaction of adenosine and serotonin.

The thalamic synapses relay peripheral sensory information to the cortex, and constitute an important part of the thalamocortical network that generates oscillatory activities responsible for different vigilance (sleep and wakefulness) states. However, the modulation of thalamic synaptic transmission by potential sleep regulators, especially by combination of regulators in physiological scenarios, is not fully characterized. We found that somnogen adenosine itself acts similar to wake-promoting serotonin, both decreasing synaptic strength as well as short-term depression, at the retinothalamic synapse. We then combined the two modulators considering the coexistence of them in the hypnagogic (sleep-onset) state. Adenosine plus serotonin results in robust synergistic inhibition of synaptic strength and dramatic transformation of short-term synaptic depression to facilitation. These synaptic effects are not achievable with a single modulator, and are consistent with a high signal-to-noise ratio but a low level of signal transmission through the thalamus appropriate for slow-wave sleep. This study for the first time demonstrates that the sleep-regulatory modulators may work differently when present in combination than present singly in terms of shaping information flow in the thalamocortical network. The major synaptic characters such as the strength and short-term plasticity can be profoundly altered by combination of modulators based on physiological considerations.

Deep sea water modulates blood pressure and exhibits hypolipidemic effects via the AMPK-ACC pathway: an in vivo study.

Deep sea water (DSW), originally pumped from the Pacific Rim off the coast of Hualien County (Taiwan), and its mineral constituents, were concentrated by a low-temperature vacuum evaporation system to produce a hardness of approximately 400,000 mg/L of seawater mineral concentrate. The primary composition of this seawater mineral concentrate was ionic magnesium (Mg²âº), which was approximately 96,000 mg/L. Referring to the human recommended daily allowance (RDA) of magnesium, we diluted the mineral concentrate to three different dosages: 0.1 × DSW (equivalent to 3.75 mg Mg²âº/kg DSW); 1 × DSW (equivalent to 37.5 mg Mg²âº/kg DSW); and 2 × DSW (equivalent to 75 mg Mg²âº/kg DSW). Additionally, a magnesium chloride treatment was conducted for comparison with the DSW supplement. The study indicated that 0.1 × DSW, 1 × DSW and 2 × DSW decreased the systolic and diastolic pressures in spontaneous hypertensive rats in an eight-week experiment. DSW has been shown to reduce serum lipids and prevent atherogenesis in a hypercholesterolemic rabbit model. Our results demonstrated that 1 × DSW and 2 × DSW significantly suppressed the serum cholesterol levels, reduced the lipid accumulation in liver tissues, and limited aortic fatty streaks. These findings indicated that the antiatherogenic effects of DSW are associated with 5'-adenosine monophosphate-activated protein kinase (AMPK) stimulation and the consequent inhibition of phosphorylation of acetyl-CoA carboxylase (ACC) in atherosclerotic rabbits. We hypothesize that DSW could potentially be used as drinking water because it modulates blood pressure, reduces lipids, and prevents atherogenesis.

Ethanol extracts of fruiting bodies of Antrodia cinnamomea exhibit anti-migration action in human adenocarcinoma CL1-0 cells through the MAPK and PI3K/AKT signaling pathways.

Cancer metastasis is a primary cause of cancer death. Antrodia cinnamomea (A. cinnamomea), a medicinal mushroom in Taiwan, has been shown antioxidant and anticancer activities. In this study, we first observed that ethanol extract of fruiting bodies of A. cinnamomea (EEAC) exerted a concentration-dependent inhibitory effect on migration and motility of CL1-0 cells in the absence of cytotoxicity. The results of a gelatin zymography assay showed that A. cinnamomea suppressed the activity of matrix metalloproteinase (MMP)-2 and MMP-9 in a concentration-dependent manner. Western blot results demonstrated that treatment with A. cinnamomea decreased the expression of MMP-9 and MMP-2; while the expression of the endogenous inhibitors of these proteins, i.e., tissue inhibitors of MMP (TIMP-1 and TIMP-2) increased. Two major compounds from EEAC codycepin and zhankuic acid A alone and together inhibited MMP-9 and MMP-2 expressions. Further investigation revealed that A. cinnamomea suppressed the phosphorylation of p38, and JNK1/2. A. cinnamomea also suppressed the expressions of PI3K and phosphorylation of AKT. This is the first report confirming the anti-migration activity of this potentially beneficial mushroom against human lung adenocarcinoma CL1-0.

Ethanol extracts of fruiting bodies of Antrodia cinnamomea suppress CL1-5 human lung adenocarcinoma cells migration by inhibiting matrix metalloproteinase-2/9 through ERK, JNK, p38, and PI3K/Akt signaling pathways.

Metastatic cancer attributes to a major cause of cancer death. In this pioneer study, we aimed to investigate how Antrodia cinnamomea (A. cinnamomea), indigenous to Taiwan, affects migration ability of highly metastatic human adenocarcinoma lung cancer cells CL1-5. Our result demonstrated that noncytotoxic ethanol extract of fruiting bodies of A. cinnamomea (EEAC) exhibited a dose-dependent inhibitory effect on motility and migration of the highly metastatic CL1-5 cells. Results of a gelatin zymography assay illustrated that A. cinnamomea repressed the activities of matrix metalloproteinase- (MMP-) 2 and 9 in a dose-dependent manner. A. cinnamomea administration decreased MMP-9 and MMP-2 protein expressions from Western blotting assay, whereas the expression of the tissue inhibitors of MMP (TIMP-1 and TIMP-2) increased. Additional study disclosed that A. cinnamomea suppressed FAK, ERK1/2, p38, AKT, and JNK1/2 phosphorylation, and also PI3K and Rac-1 were found decreased. Further, treatment of CL1-5 cells with inhibitors specific for PI3K (LY294002), ERK1/2 (PD98059), JNK (SP600125), and p38 MAPK (SB203580) decreased the expression of MMP-2 and MMP-9. Taken together, EEAC induced FAK phosphorylation and exhibited its antimigration activities via the PI3K/AKT and MAPK signalings in CL1-5 cells. This is the pioneer study verifying the antimigration activity of A. cinnamomea against human lung adenocarcinoma CL1-5 cancer cells [corrected].

Design and synthesis of iridium bis(carbene) complexes for efficient blue electrophosphorescence.

Five iridium bis(carbene) complexes, [Ir(pmi)(2)(pypz)] (1), [Ir(mpmi)(2)(pypz)] (2), [Ir(fpmi)(2)(pypz)] (3), [Ir(fpmi)(2)(pyim)] (4), and [Ir(fpmi)(2)(tfpypz)] (5) (pmi=1-phenyl-3-methylimdazolin-2-ylidene-C,C(2'); fpmi=1-(4-fluorophenyl)-3-methylimdazolin-2-ylidene-C,C(2'); mpmi=1-(4-methyl-phenyl)-3-methylimdazolin-2-ylidene-C,C(2'); pypz=2-(1H-pyrazol-5-yl)pyridinato; pyim=2-(1H-imidazol-2-yl)pyridinato; and tfpypz=2-(3-(trifluoromethyl)-1H-pyrazol-5-yl)pyridinato), were synthesized and their structures were characterized by NMR spectroscopy, mass spectroscopy and X-ray diffraction. These complexes showed phosphorescent emission with the emission maxima between 453 and 490 nm. Various spectrophotometric measurements, cyclic voltammetric studies, and density functional theory (DFT) calculations show that, unlike most of the phosphorescent cyclometalated iridium complexes, the lowest unoccupied molecular orbital (LUMO) energy and the emissive state of these iridium complexes are mainly controlled by the N,N'-heteroaromatic (N^N) ligand. Despite the fact that the LUMO levels of these complexes are mainly on the N^N ligands, the efficiencies of the electroluminescent (EL) devices are very high. For example, the EL devices using [Ir(mpmi)(2)(pypz)], [Ir(fpmi)(2)(pypz)], and [Ir(fpmi)(2)(tfpypz)] as the dopant emitters exhibited light- to deep-blue electrophosphorescence with external quantum efficiencies of 15.2, 14.1, and 7.6% and Commission Internationale d'Énclairage (x,y) coordinates (CIE(x,y)) of (0.14, 0.27), (0.14, 0.18) and (0.14, 0.10), respectively.

Synthesis and photo- and electroluminescence properties of 3,6-disubstituted phenanthrenes: alternative host material for blue fluorophores.

Three deep-blue fluorescent 9,10-bis(4-tert-butylphenyl)phenanthrenes with diphenyl, -naphthyl, and -pyrenyl moieties at C3 and C6 positions were synthesized and used as the host for doped blue fluorescent devices; one of these devices reveals excellent external quantum efficiency of 7.7% and current efficiency of 9.8 cd A(-1) with low efficiency roll-off, deep-blue color coordinates (0.14, 0.14) and long operational lifetime.

Trilinolein inhibits proliferation of human non-small cell lung carcinoma A549 through the modulation of PI3K/Akt pathway.

Trilinolein has been identified as one of the active constituents isolated from Panax notoginseng used widely in traditional Chinese medicine. Protective actions of Panax notoginseng against cerebral ischemia, beneficial effects on the cardiovascular system, and hemostatic, antioxidant, hypolipidemic, hepatoprotective, renoprotective and estrogen-like activities have been illustrated. In the present study, the effects of trilinolein on the growth of non-small cell lung carcinoma A549 were investigated. It was found that the exposure of A549 cells to trilinolein resulted in the growth inhibition and the induction of apoptosis in a dose- and time- dependent manner. Trilinolein treatment induced the upregulation of pro-apoptotic Bax, downregulation of anti-apoptotic Bcl-2 expression, which was associated with the proteolytic activation of caspases and the concomitant degradation of poly(ADP-ribose) polymerase (PARP) protein. Intracellular reactive oxygen species seem to play a role in the trilinolein-induced apoptosis, since ROS were produced early in the trilinolein treatment. Moreover, the activity of PI3K/Akt was downregulated in trilinolein-treated cells. Our results demonstrated that the most important regulators of trilinolein-induced apoptosis are Bcl-2 family and caspase-3, which are associated with cytochrome c release and dephosphorylation on the Akt signaling pathway.

Molecular mechanism of green microalgae, Dunaliella salina, involved in attenuating balloon injury-induced neointimal formation.

The pathological mechanism of restenosis is primarily attributed to excessive proliferation of vascular smooth muscle cells (VSMC). The preventive effects of ethanol extract of Dunaliella salina (EDS) on balloon injury-induced neointimal formation were investigated. To explore its molecular mechanism in regulating cell proliferation, we first showed that EDS markedly reduced the human aortic smooth muscle cell proliferation via the inhibition of 5'-bromo-2'-deoxyuridine (BrdU) incorporation at 40 and 80 microg/ml. This was further supported by the G0/G1-phase arrest using a flow cytometric analysis. In an in vivo study, EDS at 40 and 80 microg/ml was previously administered to the Sprague-Dawley rats and found that the thickness of neointima, and the ratio of neointima:media were also reduced. EDS inhibited VSMC proliferation in a dose-dependent manner following stimulation of VSMC cultures with 15 % fetal bovine serum (FBS). Suppressed by EDS were 15 % FBS-stimulated intracellular Raf, phosphorylated extracellular signal-regulated kinases (p-Erk) involved in cell-cycle arrest and proliferating cell nuclear antigen. Phosphorylated focal adhesion kinase (p-FAK) was also suppressed by EDS. Also active caspase-9, caspase-3 and cleaved poly(ADP-ribose) polymerase (PARP) protein expression levels were increased by administration with EDS; the apoptotic pathway may play an important role in the regulatory effects of EDS on cell growth. These observations provide a mechanism of EDS in attenuating cell proliferation, thus as a potential intervention for restenosis.

Pipoxolan inhibits proliferation of HL-60 human leukaemia cancer cells by arresting the cell cycle at the G0/G1 phase.

1. The aim of the present study was to investigate the molecular mechanisms by which pipoxolan exerts its inhibitory effects and apoptotic activity in human leukaemia HL-60 cells. 2. The effects of pipoxolan on the proliferation of HL-60 cells and on the distribution of cells within different phases of the cell cycle were investigated indirectly using a Trypan blue assay and a flow cytometer, respectively. The effects of pipoxolan on the apoptosis of HL-60 cells was investigated using DNA fragmentation and flow cytometer. The expression of factors affecting the cell cycle and apoptosis, including p53, p21, Bax, Bcl2, cytochrome c, caspase 3 and caspase 9, was examined by western blotting. 3. At 6.25 microg/mL, pipoxolan significantly induced apoptosis in human leukaemia HL-60 cells after 24 h exposure. In addition, HL-60 cells were arrested in the G(0)/G(1) phase via the induction of p53/p21 by pipoxolan. Apoptosis was associated with an increased Bax/Bcl-2 ratio, cytochrome c release, cleavage of procaspases-9 and -3 and hydrolysis of poly(ADP-ribose) polymerase. Intracellular reactive oxygen species (ROS) seem to play a key role in the pipoxolan-induced apoptosis, because high levels of ROS were produced early in the drug treatment. Apoptosis was significantly abrogated by the free radical scavenger N-acetylcysteine (NAC).

Analgesic and anti-inflammatory activities of a water extract of Trachelospermum jasminoides (Apocynaceae).

AIMS OF THE STUDY: This study investigated the analgesic and anti-inflammatory effects of a water extract of Trachelospermum jasminoides (WET) in ICR mice. MATERIALS AND METHODS: In HPLC analysis, the fingerprint chromatogram of WET was established. Acetic acid-induced writhing response and formalin-induced pain were examined the analgesics effects of WET. WET on lambda-Carrageenan(carr)-induced paw edema was performed. We investigate the anti-inflammatory mechanism of WET via studies of the activities of glutathione peroxidase (GPx), glutathione reductase (GRx) in the liver and the levels of malondialdehyde (MDA) and nitrite oxide (NO) in the edema paw. Serum NO and TNF-alpha were also measured. RESULTS: The fingerprint chromatogram of WET was established through HPLC analysis, and implies that WET contains the active ingredient gallic acid, chlorgenic acid, caffeic acid, taxifolin, isoquercitrin and quercetin. WET significantly inhibited the numbers of acetic acid-induced writhing responses and the formalin-induced pain in the late phase. In the anti-inflammatory test, WET inhibited the development of paw edema induced by carr. WET decreased the paw edema at the third, fourth and fifth hour after carr administration, and increased the activities of SOD, GPx and GRx in the liver tissue and decreased the MDA level in the edema paw at the third hour after carr injection. WET decreased the level of NO in edematous paw tissue and in serum level, and diminished the level of serum TNF-alpha at the fifth hour after carr injection. CONCLUSIONS: These results demonstrated that WET is an effective anti-inflammatory agent in carr-induced inflammation. WET probably exerts anti-inflammatory effects by suppressing TNF-alpha and NO. The anti-inflammatory mechanism of WET might be related to the decrease in the level of MDA in the edema paw via increasing the activities of SOD, GPx and GRx in the liver.