Quarter-Century Explorations of Bioactive Polyphenols: Diverse Health Benefits.

Polyphenols, members of phytochemical superfamily rich in vegetables and fruits, include flavonoids, non-flavonoids, and phenolic acids. Their biological effects includes classical antioxidation (e.g., radical-scavenging, metal chelating, NOX inhibition, attenuation on mitochondrial respiration, inhibition on xanthine oxidase, and upregulations on endogenous antioxidant enzymes), multiple regulations on cell signaling (e.g., AMPK activation, SirT1 activation, eNOS activation, FOXO activation, NFκB inactivation, PI3K/AkT inhibition, mTORC1 inhibition, PKC inhibition, MAPK inhibition, ERK inhibition, JAK/STAT inhibition, IKK/JNK inhibition, PDE inhibition, ß-catenin inactivation, downregulation on TLR expression, ACE inhibition, adiponectin elevation, attenuated ET-1 production, and K+ channel activation), and many other actions (e.g., inhibition on α-glucosidase, anticoagulation, γ-secretase inhibition, monoamine oxidase inhibition, LPL upregulation, ANGPTL4 suppression, upregulation on paraoxonase 1, PAI-1 downregulation, tPA upregulation, immunoregulation, epigenetic modulation, and altered gut microbiota). Such multi- targeting and functions exhibiting antioxidative stress and antiinflammation as major pillars along with many other antagonisms could not only afford healthy polyphenols suitable supplements for promoting health, but also advance them to therapeutic applications. This review aims to translate diverse polyphenolic biochemical actions to clinical applications in fighting against non-communicable diseases such as CVD, cancer, diabetes, obesity, neurodegeneration, inflammatory diseases (e.g., IBD, IBS, NAFLD, etc.), AMD, allergy, and autoimmunity as well as communicable infection (e.g., bacteria, fungal, and viral).

Antagonism by bioactive polyphenols against inflammation: a systematic view.

Through pattern recognition receptors, infections and tissue injuries drive innate immune cells to trigger inflammation with elevated cytokines, chemokines, growth factors, and other mediators. Inflammation resolves upon removal of pathogenic signals and the presence of pro-resolving conditions including combating adaptive immunity. Failure of resolution progresses into chronic inflammation, manifesting as detrimental disease development known as inflammatory diseases including cardiovascular diseases, diabetes, obesity, cancers, etc. Inflammation typically involves activations of many intracellular signaling pathways such as PI3K/AkT/mTORC1, PI3K/AkT/IKK(JNK), Ras/Raf/MEK/ERK, JAK/STAT, etc.; these pathways could in turn mediate the upregulations of proinflammatory transcription factors (e.g., NFκB, activator protein 1 (AP-1), HIF, signal transducer and activator of transcription (STAT), etc.). Furthermore, the resulting FOXO inactivation ensures inflammatory proceeding. This review provides a systematic view that polyphenols target multiple inflammatory components and reinforce anti-inflammatory mechanisms by antioxidant potentials, AMPK activation, PI3K/AkT inhibition, IKK/JNK inhibition, mTORC1 inhibition, JAK/STAT inhibition, TLR suppression, and ACE inhibition. As a result, polyphenols readily lead to NFκB, AP-1, HIF, and STAT inactivations with reduced proinflammatory mediator generation. In conclusion, polyphenols sustain resolution of inflammation and antagonize against proinflammation, which is readily consistent with diverse anti-inflammatory actions. The promoted, restored, and maintained tissue homeostasis beyond its anti-inflammatory effects also extends to diverse health benefits for disease preventions and interventions.

Tissue factor, blood coagulation, and beyond: an overview.

Emerging evidence shows a broad spectrum of biological functions of tissue factor (TF). TF classical role in initiating the extrinsic blood coagulation and its direct thrombotic action in close relation to cardiovascular risks have long been established. TF overexpression/hypercoagulability often observed in many clinical conditions certainly expands its role in proinflammation, diabetes, obesity, cardiovascular diseases, angiogenesis, tumor metastasis, wound repairs, embryonic development, cell adhesion/migration, innate immunity, infection, pregnancy loss, and many others. This paper broadly covers seminal observations to discuss TF pathogenic roles in relation to diverse disease development or manifestation. Biochemically, extracellular TF signaling interfaced through protease-activated receptors (PARs) elicits cellular activation and inflammatory responses. TF diverse biological roles are associated with either coagulation-dependent or noncoagulation-mediated actions. Apparently, TF hypercoagulability refuels a coagulation-inflammation-thrombosis circuit in "autocrine" or "paracrine" fashions, which triggers a wide spectrum of pathophysiology. Accordingly, TF suppression, anticoagulation, PAR blockade, or general anti-inflammation offers an array of therapeutical benefits for easing diverse pathological conditions.

Polycations selectively blocking tissue factor-dependent FVII activation: collective in vitro anticoagulation studies.

Tissue factor (TF) is an initiator of the extrinsic blood coagulation, which is often susceptible to upregulation by tissue injury, advanced glycation end-product, or diverse inflammation. TF hypercoagulability is accompanied by elevated generation of clotting factors (e.g., FVIIa, FXa, and thrombin) and fibrin production, all of which are proinflammatory. In this laboratory, our in vitro experimental results show that polycationic anticoagulants (compound 48/80, ruthenium red, polybrene, protamine, Buforin I, and cationic polyamino acids) intervene TF hypercoagulability at posttranslational level. Polycations preferentially suppress TF-dependent FVII activation with diminished FVIIa formation shown on Western blotting, resulting in non- or un-competitive inhibition on FVIIa amidolytic activity. In contrast, polycations have no effect on FVIIa catalysis, FXa activity, or thrombin activity per se. Polycations could present a new class of anticoagulants with such unique upstream downregulation of blood coagulation. In view of coagulation-dependent inflammation and the new paradigm of blood coagulation-inflammation-thrombosis circuit, the polycations as a new class of anticoagulants could effectively contribute to antiinflammation, antithrombosis, and cardioprotection. Further development of effective anticoagulants is of biopharmaceutical significance in broadly easing disease conditions.

Blood coagulation as an intrinsic pathway for proinflammation: a mini review.

Blood coagulation could be recognized as intrinsic inflammation. The coagulant mediators (FVIIa, FXa, thrombin (FIIa), FXIIa) and fibrin(ogen) activate cellular signaling, eliciting the production of cytokines, chemokines, growth factors, and other proinflammatory mediators. Hypercoagulability with elevated coagulant mediators would certainly trigger hyper-inflammatory state not to mention about the direct hypercoagulable actions on thrombosis, and platelet and complement activations, all of which contribute to inflammatory events. Furthermore, anticoagulant's anti-inflammatory effects readily reinforce the proposal that blood coagulation results in inflammation. The observations on protease activated receptor (PAR) activation and PAR antagonists modulating inflammation are also in line with the concept of coagulation-dependent inflammation.

Role of tissue factor in thrombosis. Coagulation-inflammation-thrombosis circuit.

Tissue factor (TF) plays a role in thrombogenesis. TF initiates blood coagulation resulting in the generation of protease coagulant mediators (FVIIa, FXa, and FIIa) and fibrin production. TF hypercoagulablility directly contributes to thrombus formation resulting from the major events of fibrin deposition and FIIa-induced platelet activation/aggregation. In addition, blood coagulation indirectly promotes thrombogenicity via the coagulation-inflammation cycle in which TF plays a diverging and converging role. As the consequence of coagulation-dependent inflammation in which protease-activated receptor (PAR) mediates the coagulant signaling to elicit cytokines, selectins, and growth factors, such inflammation facilitates thrombosis by platelet aggregation and leukocyte recruitment. As TF hypercoagulability concerned, anti-thrombotic strategies involve the prevention by anticoagulation and PAR antagonism. Anticoagulants block the direct and indirect thrombotic contributions, while PAR antagonists arrest coagulation-dependent inflammation. With respect to both thrombosis and inflammation being cardiovascular risk factors, such strategies offer diverse benefits to cardioprotection.

Tissue factor upregulation drives a thrombosis-inflammation circuit in relation to cardiovascular complications.

The extrinsic coagulation is recognized as an 'inducible' signalling cascade resulting from tissue factor (TF) upregulation by exposure to clotting zymogen FVII upon inflammation or tissue injury. Following the substantial initiation, an array of proteolytic activation generates mediating signals (active serine proteases: FVIIa, FXa and FIIa) that lead to hypercoagulation with fibrin overproduction manifesting thrombosis. In addition, TF upregulation plays a central role in driving a thrombosis-inflammation circuit. Coagulant mediators (FVIIa, FXa and FIIa) and endproduct (fibrin) are proinflammatory, eliciting tissue necrosis factor, interleukins, adhesion molecules and many other intracellular signals in different cell types. Such resulting inflammation could ensure 'fibrin' thrombosis via feedback upregulation of TF. Alternatively, the resulting inflammation triggers platelet/leukocyte/polymononuclear cell activation thus contributing to 'cellular' thrombosis. TF is very vulnerable to upregulation resulting in hypercoagulability and subsequent thrombosis and inflammation, either of which presents cardiovascular risks. The prevention and intervention of TF hypercoagulability are of importance in cardioprotection. Blockade of inflammation reception and its intracellular signalling prevents TF expression from upregulation. Natural (activated protein C, tissue factor pathway inhibitor, or antithrombin III) or pharmacological anticoagulants readily offset the extrinsic hypercoagulation mainly through FVIIa, FXa or FIIa inhibition. Therefore, anticoagulants turn off the thrombosis-inflammation circuit, offering not only antithrombotic but anti-inflammatory significance in the prevention of cardiovascular complications.

Tissue factor mediates inflammation.

The role of tissue factor (TF) in inflammation is mediated by blood coagulation. TF initiates the extrinsic blood coagulation that proceeds as an extracellular signaling cascade by a series of active serine proteases: FVIIa, FXa, and thrombin (FIIa) for fibrin clot production in the presence of phospholipids and Ca2+. TF upregulation resulting from its enhanced exposure to clotting factor FVII/FVIIa often manifests not only hypercoagulable but also inflammatory state. Coagulant mediators (FVIIa, FXa, and FIIa) are proinflammatory, which are largely transmitted by protease-activated receptors (PAR) to elicit inflammation including the expression of tissue necrosis factor, interleukins, adhesion molecules (MCP-1, ICAM-1, VCAM-1, selectins, etc.), and growth factors (VEGF, PDGF, bFGF, etc.). In addition, fibrin, and its fragments are also able to promote inflammation. In the event of TF hypercoagulability accompanied by the elevations in clotting signals including fibrin overproduction, the inflammatory consequence could be enormous. Antagonism to coagulation-dependent inflammation includes (1) TF downregulation, (2) anti-coagulation, and (3) PAR blockade. TF downregulation and anti-coagulation prevent and limit the proceeding of coagulation cascade in the generation of proinflammatory coagulant signals, while PAR antagonists block the transmission of such signals. These approaches are of significance in interrupting the coagulation-inflammation cycle in contribution to not only anti-inflammation but also anti-thrombosis for cardioprotection.

Prevention of colorectal cancer using COX-2 inhibitors: basic science and clinical applications.

Cyclooxygenase-2 (COX-2), an inducible prostaglandin G/H synthase, is overexpressed in pre-neoplastic tissues and several human cancers including colorectal cancer. Evidence linking COX-2 activity to carcinogenesis was derived from epidemiologic studies and animal models with defect adenomatous polyposis coli (APC) gene. PGE2 induced by COX-2 exerts several biological properties that may be advantageous for tumorigenesis: 1) Promoting angiogenesis (increased VEGF, bFGF, and PDGF production), 2) Anti-apoptosis mechanism (via increased bcl-2 and Akt activity), 3) Stimulating tumor metastasis (by increasing matrix metalloproteinases) and 4) Decreased immune surveillance (decreased cytokine production and NK activity). In addition, COX-2 reaction can cause DNA oxidation and induce mutations. Chemoprevention of colorectal cancer has attracted great attention in recent years. Epidemiologic data showed that chronic intake of traditional nonsteroidal anti-inflammatory drugs (NSAIDs) could reduce the incidence of colorectal cancer. Recent clinical trial studies showed that celecoxib, a selective COX-2 inhibitor, is equally effective in reducing colorectal adenomas in animal models and patients with familial adenomatous polyposis (FAP), yet with superior GI safety. Two COX-2 inhibitors (celecoxib and refocoxib) have been approved by FDA as adjuncts to usual care in FPA patients, and are currently being studied in patients with sporadic adenomas and other types of cancers. These studies are expected to generate evidence in favor of targeting COX-2 and its gene products as chemopreventive strategies, which may provide an alternative in current approach to reducing the morbidity and mortality of this disease.

Biochemical strategies to anticoagulation: a comparative overview.

Hypercoagulability is widely associated with sepsis, inflammation, diabetes, cancers, aging, and many pathological conditions, resulting in life-threatening disseminated intravascular coagulation (DIC), venous thrombosis, thromboembolism, cardiovascular complications, or even deadly multiple organ failure. Relieving coagulation dysfunction is not only a task for research scientists but also a challenge for physicians. The development of effective anticoagulants is under way with the basic understanding of the pathophysiology of hypercoagulable state. In this overview, various anticoagulants will be discussed according to the proposed inhibitory target-sites along the extrinsic pathway that is believed to play an integral role in homeostasis. Anticoagulants generally fall into two broad categories as natural or pharmacological ones. Antithrombin (AT), activated protein C (APC), and tissue factor pathway inhibitor (TFPI) mainly constitute the natural anticoagulant system apart from the recently reported physiological components such as lipoproteins, sphingosine, thrombomodulin (TM) or cellular Marcks protein. Pharmacological anticoagulants include warfarin, FVIIa inhibitors, FXa inhibitors, and thrombin inhibition by its direct inhibitors or heparins. In addition, a group of novel compounds inhibiting TF-dependent FVII activation result in anticoagulation; such upstream downregulation in the extrinsic pathway awaits further research to establish their in vivo benefits. The molecular genetic approaches such as developing soluble TF, FVII and thrombin mutants provide unique downregulation. Anticoagulation also extends its significance to anti-inflammation, making broad impacts on the improvement of human health.

Poly-L-histidine downregulates fibrinolysis.

The elevated level of histidine-rich glycoprotein was considered a risk factor of inherited thrombophilia. However, the mode of action remains largely unclear. In the current study, we employ poly-l-histidine (PLH) mimicking the histidine-rich region and determine whether PLH modulates urokinase (uPA)-dependent fibrinolysis. In an in vitro model, turbidity appearance and clearance monitored fibrin polymer formation and lysis, respectively. Fibrin polymer formed upon fibrinogen incubation with thrombin. In the presence of uPA or plasmin, fibrin polymer lysis took place in a dose-dependent manner as a function of time. We demonstrated that PLH significantly downregulated uPA-dependent fibrinolysis. PLH had no effect on plasminogen activation, as evidenced by no inhibitions on either uPA amidolytic activity or plasmin formation derived from its zymogen. Nor did PLH show any inhibition on plasmin amidolytic activity. PLH caused a profound delay of plasmin-dependent fibrinolysis upon pre-incubation of either plasmin or fibrinogen with PLH. The observations taken together suggest that the complex [plasmin-PLH-fibrin] formation significantly delayed plasmin-dependent fibrinolysis.

Anticoagulant potential of an antibody against factor VII.

BACKGROUND: Hypercoagulability often resulting from sepsis, trauma, and other conditions is widely associated with thrombotic and cardiovascular disorders. The development of effective and safe anticoagulation is in great demand to relieve complications and improve human health. OBJECTIVE: We study the anticoagulant potential of a polyclonal antibody to human FVII (anti-hFVII Ab). METHODS AND RESULTS: Preincubating FVII with anti-hFVII Ab, we showed the significantly blocked tissue factor (TF)-dependent FVII activation monitored by a two-stage chromogenic assay. Consistently, the antibody depressed TF/FVII-catalyzed FX activation was shown on Western blotting analysis. As a result, TF procoagulation derived from rabbit brain thromboplastin was prolonged significantly by the preincubation of human normal plasma with the antibody, which mimicked FVII-deficient plasma in a single-stage clotting assay. In contrast, the anti-hFVII Ab had no effect on either FVIIa amidolytic activity or TF/FVIIa binary complex. CONCLUSIONS: Anti-hFVII Ab readily blocked clot formation, which was mediated by the upstream downregulation of the extrinsic coagulation of inhibiting FVII activation. Further research warrants establishing its in vivo application as an anticoagulant.

Novel anticoagulant activity of polyamino acid offsets bacterial endotoxin-induced extrinsic hypercoagulation: downregulation of monocytic tissue factor-dependent FVII activation.

The extrinsic hypercoagulation often resulting from sepsis could contribute to disseminated intravascular coagulation and cardiovascular complications. The effective prevention and intervention remained largely complex and unclear. In a cell model of human leukemia THP-1 monocytes following bacterial endotoxin (LPS) exposure, we show the novel anticoagulant ability of polyamino acid (polyAA) to suppress the extrinsic hypercoagulation. LPS-induced monocytic tissue factor (mTF) procoagulation was readily offset by poly-L-lysine (PLK), poly-L-arginine (PLR), or poly-L-ornithine (POR) included in single-stage clotting assays. IC50 was estimated at 0.35, 0.30, or 0.58 microM for PLR, POR, or PLK, respectively, whereas, poly-L-asparatic acid (PLD) remained ineffective. In a separate approach, inclusion of cationic polyAA in human plasma significantly prolonged prothrombin time, confirming the depressed extrinsic coagulation. In chromogenic assays dissecting the extrinsic pathway, we further determined the inhibitory site(s). PLK, PLR, or POR significantly inhibited LPS-induced FVII activation, which was consistent with the diminished FVIIa formation shown on Western blotting analysis. In contrast, polyAA did not show any additional effect on either FVIIa/FXa amidolytic activities or mTF/FVIIa-catalyzed FX activation. Nor did polyAA show any effect on FVII activation directly catalyzed by FXa. Taken together, PLK, PLR, or POR preferentially inhibited mTF-dependent FVII activation, accounting for their novel anticoagulant activities. PolyAA might present the specific antagonists to arrest the extrinsic hypercoagulation following inflammation.

Novel anticoagulant polyethylenimine: inhibition of thrombin-catalyzed fibrin formation.

Hypercoagulability is often associated with a variety of disease states, leading to cardiovascular complications. Polyethylenimine (PEI) prolonged prothrombin time, demonstrating its anticoagulant potential. In vitro, PEI at low concentration (nM) significantly blocked thrombin-catalyzed fibrin formation, accounting for its mode of anticoagulation. The uncompetitive inhibition by PEI of fibrin formation was independent of the concentration of fibrinogen (FBG), thrombin, or NaCl. PEI showed no effect on thrombin amidolytic activity, suggesting that the blockade of thrombin interaction with FBG could account for the inhibition on fibrin formation. PEI drastically depressed rabbit brain thromboplastin procoagulation monitored by a single-stage clotting assay using human plasma. In a THP-1 monocytic hypercoagulation model, a 4-h exposure to bacterial endotoxin or Ca(2+) ionophore A23187, respectively, resulted in a 5- or 10-fold enhancement in monocytic tissue factor (mTF) procoagulation. mTF hypercoagulation was offset by PEI included in the assay mixture. PEI showed the potential to arrest mTF hypercoagulation with IC(50) around 1.2 nM. Using a chromogenic assay to dissect the extrinsic pathway, we further assessed whether PEI has any effect on other clotting factors. PEI was not an inhibitor for either FVIIa or FXa, having no effect on not only the amidolytic but also their corresponding functionally catalytic activities. Although PEI upregulated TF-dependent FVII activation under the low-salt condition, the effective downstream inhibition of fibrin formation readily abolished and overrode the upstream enhancement, demonstrating the overall anticoagulation. PEI could present a new class of anticoagulant.

Possible role of Marcks in the cellular modulation of monocytic tissue factor-initiated hypercoagulation.

The enhanced extrinsic tissue factor (TF)-initiated coagulation, often resulting from sepsis, could lead to disseminated intravascular coagulation presenting cardiovascular complications. Using model human leukaemia THP-1 monocytes, we studied monocytic TF (mTF) hypercoagulation and its regulation. After an 8 h exposure to bacterial endotoxin [lipopolysaccharide (LPS); 100 ng/ml], mTF activity was significantly upregulated as the result of the enhanced mTF synthesis. Thereafter, LPS induction declined, exhibiting a "quiescent-desensitizing' phenomenon. Such diminished LPS induction was,however,associated with sustained LPS-enhanced mTF synthesis, revealing the possible occurrence of a post-translational downregulation. It was noted that LPS desensitization was accompanied by the increased expression of myristoylated alanine-rich C kinase substrate (Marcks). In contrast, A23187 (20 micromol/l) or Quin-2AM (20 micromol/l) drastically activated mTF activity without detectable effect on mTF synthesis; both of which showed that sustained functional upregulation during 24 h culture did not enhance Marcks expression. These inverse correlations between mTF activity upregulation and Marcks expression suggested that Marcks could be inhibitory. Marcks phosphorylation site domain (151-175) (Marcks PSD) readily inhibited mTF-dependent FVII activation and diminished FVIIa formation in LPS-challenged cells. As a result, Marcks PSD offset LPS-induced mTF hypercoagulation upon inclusion in the single-stage clotting assays. The anticoagulant activity was confirmed by showing that Marcks PSD significantly blocked rabbit brain thromboplastin (rbTF) procoagulation and inhibited rbTF-dependent FVII activation as well as FVIIa formation. Our study suggests that Marcks expression plays a role in a novel cellular modulation to downregulate mTF hypercoagulation.