The novel thromboxane prostanoid receptor mediates CTGF production to drive human nasal fibroblast self-migration through NF-κB and PKCδ-CREB signaling pathways.

Chronic rhinosinusitis without nasal polyp (CRSsNP) is characterized by tissue repair/remodeling and the subepithelial stroma region in whose nasal mucosa has been reported by us to have thromboxane A2 (TXA2) prostanoid (TP) receptor and overexpress connective tissue growth factor (CTGF). Therefore, this study aimed to investigate the relationship between TP receptor activation and CTGF production/function in human CRSsNP nasal mucosa stromal fibroblasts. We found that TP agonists including U46619 and IBOP ([1S-[1α,2α(Z),3ß(1E,3 S*),4α]]-7-[3-[3-hydroxy-4-(4-iodophenoxy)-1-butenyl]-7-oxabicyclo[2.2.1]hept-2-yl]-5-heptenoic acid) could promote CTGF protein/messenger RNA expression and secretion. The pharmacological intervention and TP activation assay with U46619 identified the possible participation of PKCµ, PKCδ, nuclear factor-κB (NF-κB), and cyclic AMP response element-binding protein (CREB) phosphorylation/activation in the CTGF induction. Moreover, a phorbol ester-phorbol-12-myristate 13-acetate (PMA) exhibited a similar cellular signaling and CTGF production profile to that elicited by TP activation. However, further small interfering RNA interference analysis revealed that only NF-κB and PKCδ-CREB pathways were necessarily required for TP-mediated CTGF production, which could not be completely supported by those findings from PMA. Finally, in a functional assay, although CTGF did not affect fibroblast proliferation, TP-mediated CTGF could drive novel self-migration in fibroblasts both in the scratch/wound healing and transwell apparatus assays. Meanwhile, the overall staining for stress fibers and formation of the lamellipodia and filopodia-like structures was concomitantly increased in the treated migrating cells. Collectively, we provided here that novel TP mediates CTGF production and self-migration in human nasal fibroblasts through NF-κB and PKCδ-CREB signaling pathways. More importantly, we also demonstrated that thromboxane, TP receptor, CTGF, and stromal fibroblasts may act in concert in the tissue remodeling/repair process during CRSsNP development and progression.

The Presence of TGFß3 in Human Ovarian Intrafollicular Fluid and Its Involvement in Thromboxane Generation in Follicular Granulosa Cells through a Canonical TGFßRI, Smad2/3 Signaling Pathway and COX-2 Induction.

Ovarian follicular fluid (FF) has a direct impact on oocyte quality, playing key roles in fertilization, implantation, and early embryo development. In our recent study, we found FF thromboxane (TX) to be a novel factor inversely correlated with oocyte maturation and identified thrombin, transforming growth factor ß (TGFß), TNF-α, and follicular granulosa cells (GCs) as possible contributors to FF TX production. Therefore, this study sought to investigate the role of TGFß3 in regulating TX generation in human ovarian follicular GCs. TGFß3 was differentially and significantly present in the FF of large and small follicles obtained from IVF patients with average concentrations of 68.58 ± 12.38 and 112.55 ± 14.82 pg/mL, respectively, and its levels were correlated with oocyte maturity. In an in vitro study, TGFß3 induced TX generation/secretion and the converting enzyme-COX-2 protein/mRNA expression both in human HO23 and primary cultured ovarian follicular GCs. While TGFßRI and Smad2/3 signaling was mainly required for COX-2 induction, ERK1/2 appeared to regulate TX secretion. The participation of Smad2/3 and COX-2 in TGFß3-induced TX generation/secretion could be further supported by the observations that Smad2/3 phosphorylation and nuclear translocation and siRNA knockdown of COX-2 expression compromised TX secretion in GCs challenged with TGFß3. Taken together, the results presented here first demonstrated that FF TGFß3 levels differ significantly in IVF patients' large preovulatory and small mid-antral follicles and are positively associated with oocyte maturation. TGFß3 can provoke TX generation by induction of COX-2 mRNA/protein via a TGFßR-related canonical Smad2/3 signaling pathway, and TX secretion possibly by ERK1/2. These imply that TGFß3 is one of the inducers for yielding FF TX in vivo, which may play a role in folliculogenesis and oocyte maturation.

Eugenol Suppresses Platelet Activation and Mitigates Pulmonary Thromboembolism in Humans and Murine Models.

Platelets assume a pivotal role in the pathogenesis of cardiovascular diseases (CVDs), emphasizing their significance in disease progression. Consequently, addressing CVDs necessitates a targeted approach focused on mitigating platelet activation. Eugenol, predominantly derived from clove oil, is recognized for its antibacterial, anticancer, and anti-inflammatory properties, rendering it a valuable medicinal agent. This investigation delves into the intricate mechanisms through which eugenol influences human platelets. At a low concentration of 2 µM, eugenol demonstrates inhibition of collagen and arachidonic acid (AA)-induced platelet aggregation. Notably, thrombin and U46619 remain unaffected by eugenol. Its modulatory effects extend to ATP release, P-selectin expression, and intracellular calcium levels ([Ca2+]i). Eugenol significantly inhibits various signaling cascades, including phospholipase Cγ2 (PLCγ2)/protein kinase C (PKC), phosphoinositide 3-kinase/Akt/glycogen synthase kinase-3ß, mitogen-activated protein kinases, and cytosolic phospholipase A2 (cPLA2)/thromboxane A2 (TxA2) formation induced by collagen. Eugenol selectively inhibited cPLA2/TxA2 phosphorylation induced by AA, not affecting p38 MAPK. In ADP-treated mice, eugenol reduced occluded lung vessels by platelet thrombi without extending bleeding time. In conclusion, eugenol exerts a potent inhibitory effect on platelet activation, achieved through the inhibition of the PLCγ2-PKC and cPLA2-TxA2 cascade, consequently suppressing platelet aggregation. These findings underscore the potential therapeutic applications of eugenol in CVDs.

The role of thromboxane prostanoid receptor signaling in gastric ulcer healing.

The process of gastric ulcer healing includes cell migration, proliferation, angiogenesis and re-epithelialization. Platelets contain angiogenesis stimulating factors that induce angiogenesis. Thromboxane A2 (TXA2 ) not only induces platelet activity but also angiogenesis. This study investigated the role of TXA2 in gastric ulcer healing using TXA2 receptor knockout (TPKO) mice. Gastric ulcer healing was suppressed by treatment with the TXA2 synthase inhibitor OKY-046 and the TXA2 receptor antagonist S-1452 compared with vehicle-treated mice. TPKO showed delayed gastric ulcer healing compared with wild-type mice (WT). The number of microvessels and CD31 expression were lower in TPKO than in WT mice, and TPKO suppressed the expression of transforming growth factor beta (TGF-ß) and vascular endothelial growth factor A (VEGF-A) in areas around gastric ulcers. Immunofluorescence assays showed that TGF-ß and VEGF-A co-localized with platelets. Gastric ulcer healing was significantly reduced in WT mice transplanted with TPKO compared with WT bone marrow. These results suggested that TP signalling on platelets facilitates gastric ulcer healing through TGF-ß and VEGF-A.

Correlation of platelet-derived growth factor and thromboxane A2 expression with platelet parameters and coagulation indices in chronic altitude sickness patients.

NEW FINDINGS: What is the central question of this study? Is the expression of platelet-derived growth factor (PDGF) and thromboxane A2 (TXA2) elevated in chronic altitude patients, and are they related to thrombosis in chronic mountain sickness? What is the main finding and its importance? The expression of PDGF and TXA2 in both the bone marrow and the peripheral blood of patients with chronic mountain sickness is elevated, and they are considered to be correlated in the mechanism of thrombosis in the chronic mountain sickness. ABSTRACT: The purpose of this study was to evaluate the expression of platelet-derived growth factor (PDGF) and thromboxane A2 (TXA2) along with platelet parameters and coagulation indices in chronic mountain sickness (CMS) patients and healthy individuals on the Qinghai-Tibet Plateau. The levels of PDGF and TXA2 were examined in 22 CMS patients (age, 52.77 ± 9.92 years, haemoglobin, 219 ± 13 g/l) and 25 healthy individuals (age, 47.80 ± 9.78 years, haemoglobin, 146 ± 18 g/l), and the association between platelet parameters and coagulation indices was investigated. Mean platelet volume and fibrinogen degradation product were higher in the CMS compared to the control group (10.58 ± 0.83 vs. 8.92 ± 1.61, 7.50 ± 2.15 vs. 4.40 ± 2.51), platelet count and plateletcrit were lower in the CMS compared to the control group (0.13 (0.80, 0.16) vs. 0.23 (0.18, 0.24), 109 ± 46 vs. 204 ± 86). The levels of PDGF and TXA2 in the bone marrow and peripheral blood of CMS patients were higher (P < 0.01) in comparison to the control group. The two factors had no statistically significant relationship with platelet parameters or coagulation indices (P > 0.159). According to the current findings, platelets in CMS patients were activated, resulting in aberrant coagulation and PDGF and TXA2 expression, which could be due to physiological adjustments to the plateau's high altitude. To summarize, PDGF and TXA2 levels in CMS patients were not correlated with coagulation or platelet parameters, implying that the mechanism behind their increased expression warrants additional investigation.

Signalling pathway of U46619-induced vascular smooth muscle contraction in mouse coronary artery.

BACKGROUND: Thromboxane A2 (TXA2 ) participates in many pathophysiological processes of coronary artery disease. However, its mechanism of TXA2 -induced contraction in the coronary artery remains to be clarified. A multi myograph system was used to measure the isometric tension of the mouse coronary arteries and identify the effect and pathway of TXA2 analogues U46619. Confocal laser scanning microscopy was used to measure the intracellular calcium concentration ([Ca2+ ]i ) in mouse coronary artery smooth muscle cells. Results from the experiment had shown that contraction in coronary artery was generated by U46619 in a concentration-dependent manner, which was completely abolished by a specific TXA2 receptor blocker, GR32191. PI-PLC inhibitors U73122 and D609 and Rho-Kinase inhibitor Y-27632 can block the U46619 elicited coronary artery contraction in a dose-dependent manner. Then, the vasoconstriction response to U46619 was obviously inhibited by two pan-PKC inhibitors chelerythrine or GÓ§6983, and a selective PKCδ inhibitor rottlerin, but was not blocked by a selective PKCζ inhibitor PKC-PS or a selective PKCß inhibitor hispidin. Meanwhile, the PKC activator PDBu-induced vasoconstriction was significantly inhibited by 1 µmol/L nifedipine, then mostly inhibited by 100 µmol/L 2-APB and 10 µmol/L Y27632. We further found that the response to U46619 was inhibited, respectively, by three calcium channel blockers nifedipine, SKF96356 or 2-APB in a concentration-dependent manner. Although Store-operated Ca2+ (SOC) channels generated the increase of [Ca2+ ]i in mouse coronary artery smooth muscle cells, SOC channels did not contribute to the vasoconstriction in mouse coronary arteries. Caffeine-induced sarcoplasmic reticulum (SR) Ca2+ release could obviously induce coronal vasoconstriction. In addition, NPPB, a cell membrane Ca2+ activated C1- channel blocker, could obviously inhibit the U46619-induced vasoconstriction. The U46619-induced mouse coronary artery contraction was involved in the increase in [Ca2+ ]i mediated by Cav1.2, TRPC channels and SR release through the activation of G-protein-coupled TP receptors and the kinases signalling pathway in TP downstream proteins, while SOC channels did not participate in the vasoconstriction.

Molecular Pathomechanisms of Impaired Flow-Induced Constriction of Cerebral Arteries Following Traumatic Brain Injury: A Potential Impact on Cerebral Autoregulation.

(1) Background: Traumatic brain injury (TBI) frequently occurs worldwide, resulting in high morbidity and mortality. Here, we hypothesized that TBI impairs an autoregulatory mechanism, namely the flow-induced constriction of isolated rat middle cerebral arteries (MCAs). (2) Methods: TBI was induced in anaesthetized rats by weight drop model, and then MCAs were isolated and transferred into a pressure-flow chamber. The internal diameter was measured by a video-microscopy. (3) Results: In MCAs from intact rats, increases in flow and pressure + flow elicited constrictions (-26 ± 1.9 µm and -52 ± 2.8 µm, p < 0.05), which were significantly reduced after TBI or in the presence of thromboxane-prostanoid (TP receptor) antagonist SQ 29,548. Flow-induced constrictions were significantly reduced by HET0016, inhibitor of cytochrome P450 4A (CYP450 4A). Arachidonic acid, (AA, 10-7 M), and CYP-450 4A metabolite 20-hydroxyeicosatetraenoic acid (20-HETE) elicited constrictions of intact MCA (-26 ± 2.3% and -31 ± 3.6%), which were significantly reduced after TBI (to 11 ± 1.3% and -16 ±2.5%). The TP receptor agonist U46619 (10-7 M) elicited substantial constrictions of MCA from intact rats (-21 ± 3.3%), which were also significantly reduced, after TBI (to -16 ± 2.4%). (4) Conclusions: Flow-induced constrictor response of MCA is impaired by traumatic brain injury, likely due to the reduced ability of cytochrome P450 4A to convert arachidonic acid to constrictor prostaglandins and the mitigated sensitivity of thromboxane-prostanoid receptors.

The functional link between microsomal prostaglandin E synthase-1 (mPGES-1) and peroxisome proliferator-activated receptor γ (PPARγ) in the onset of inflammation.

Many years have elapsed since the discovery of anti-inflammatories as effective therapeutics for the treatment of inflammatory-related diseases, but we are still uncovering their various mechanisms of action. Recent biochemical and pharmacological studies have shown that in different tissues and cell types lipid mediators from thearachidonic acid cascade, play a crucial role in the initiation and resolution of inflammation by shifting from pro-inflammatory prostaglandin (PG)E2 to anti-inflammatory PGD2 and PGJ2. Considering that until now very little is known about the biological effects evoked by microsomal prostaglandin E synthase-1 (mPGES-1) and contextually by peroxisome proliferator-activated receptor γ (PPARγ) modulation (key enzymes involved in PGE2 and PGD2/PGJ2metabolism), in this opinion paper we sought to define the coordinate functional regulation between these two enzymes at the "crossroads of phlogistic pathway" involved in the induction and resolution of inflammation.

Mechanism of thromboxane receptor-induced vasoconstriction in human saphenous vein.

Saphenous vein (SV) is one of the most widely used graft material in patients undergoing coronary artery bypass graft surgery (CABG). Thromboxane A2 (TXA2) is implicated in graft failure by inducing vasoconstriction and platelet aggregation. The aim of this study is to investigate the mechanism involved in TXA2-induced vasoconstriction in human SV. The role of different inhibitors and blockers on U46619 (TXA2-mimetic)-induced vasoconstriction is investigated by using an isolated organ bath system. Relaxation responses to several mediators are evaluated in SV pre-contracted with U46619 and compared with those pre-contracted with phenylephrine. Our results demonstrate that U46619-induced contraction is completely blocked by myosin light chain kinase inhibitor ML-9 or TP receptor antagonist BAY u3405. Furthermore, U46619-induced contraction is partially inhibited by phospholipase C inhibitor U73122, protein kinase C inhibitor calphostin C, Rho-kinase inhibitor Y-27632, L-type calcium channel blocker nifedipine, store-operated channel inhibitor SKF96365 or removal of extracellular calcium. Relaxation responses to NO donor (sodium nitroprusside), guanylate cyclase (GC) stimulator (riociguat), phosphodiesterase (PDE) inhibitors (sildenafil, IBMX), adenylate cyclase (AC) activator (forskolin) and acetylcholine (ACh) are markedly reduced when U46619 is used as a pre-contraction agent. Our results demonstrate that influx of extracellular Ca2+ (through L-type calcium channels and store-operated calcium channels) and intracellular Ca2+ release together with Ca2+ sensitization (through Rho-kinase activation) are necessary components for TXA2-induced vasoconstriction in SV. Moreover, more pronounced decrease in vasorelaxation induced by several mediators (SNP, riociguat, sildenafil, IBMX, forskolin, and ACh) in the presence of U46619 when compared with phenylephrine suggests that there is a crosstalk between the TP receptor signaling pathway and PDE, AC, GC enzymes. We believe that the investigation of mechanism of the TXA2-induced vasoconstriction in SV will provide additional information for the prevention of SV graft failure.

Gonadal function protects against organ culture-induced vascular damage. Involvement of prostanoids.

Androgen deprivation induces vascular dysfunction in which altered release and action of prostanoids has been extensively studied. On the other hand, the vascular organ-culture system has been reported as a valid model for phenotypic changes that occur in several cardiovascular pathologies. Since there are no studies analyzing the impact of androgenic loss on vascular vulnerability during induced vascular damage, the objective of this study was to analyze the possible preventive role of male sex hormones on the organ culture-induced vascular damage in rat aorta. The link to possible changes in gross structure was also analyzed. For this purpose, fresh and 20 h-cultured aortic arterial segments from intact and orchidectomized rats were used to analyze: (i) the release and vasomotor effect of the thromboxane A2 (TXA2), prostaglandin (PG) E2, PGF2α and PGI2; (ii) the vasodilator response induced by acetylcholine (ACh) as well as the involvement of prostanoids, in particular TXA2, in the ACh-induced response; (iii) the effect of activation of thromboxane/prostaglandin (TP) receptors on the ACh-induced response; and (iv) the vascular structure. The results showed that organ culture: i) increased production of prostanoids; ii) increased prostanoids-induced vasomotor responses; iii) decreased ACh-induced relaxation after incubation with indomethacin, a blocker of cyclooxygenases; iv) increased the ACh-induced relaxation after incubation with the TXA2 synthase inhibitor, furegrelate, more in arteries from orchidectomized rats than in those of intact rats; v) diminished ACh-induced relaxation after U-46619 incubation only in arteries from orchidectomized rats; and vi) preserved the integrity of the different vascular layers. These results showed the protective role of male sex hormones against the induced vascular damage, since a decreased deleterious effect of prostanoids, in particular that of TXA2, was observed in arteries from rats with intact gonadal function.

Tetrahydroxystilbene glucoside-induced relaxation of the superior mesenteric artery via both endothelium-dependent and endothelium-independent mechanisms.

Tetrahydroxystilbene glucoside (TSG) is the main water-soluble component in Polygonum multiflorum Thunb, and it has many cardioprotective effects. Although TSG is able to relax blood vessels, its relaxation of rat superior mesenteric arteries and the underlying mechanism of this process are not clearly understood. The aim of the present study was to use in vivo and in vitro models to investigate the arterial relaxation effect of TSG on rat superior mesenteric arteries and the mechanisms involved. We found that TSG concentration-dependently relaxed the superior mesenteric artery with or without endothelium. The vasorelaxation induced by TSG is not related to the vasodilator derived factor NO but is rather by the inhibition of COX-2 activity and decreased TXA2. We also found that the vasorelaxation induced by TSG was attenuated by 4­AP. Moreover, TSG also inhibited the contraction induced by an increase in external calcium concentration in Ca2+-free medium plus KCl (60 mM). These results suggest that TSG induces relaxation in mesenteric arterial rings through an endothelium-dependent pathway that involves the inhibition of COX-2 activity and decreased in TXA2 and through an endothelium-independent pathway via opening of a voltage-dependent K+ channel, blockade of Ca2+ influx and release of intracellular Ca2+.

Lactoferrin Induces the Synthesis of Vitamin B6 and Protects HUVEC Functions by Activating PDXP and the PI3K/AKT/ERK1/2 Pathway.

As a nutritional active protein in foods, multiple studies of the biological activities of lactoferrin had been undertaken, including antioxidant, antiviral, anti-inflammatory, antitumor, antibiosis, and antiparasitic effects, while the mechanism related with its protection of cardiovascular system remained elusive. In the present work, the effect of lactoferrin on the viability of HUVECs (human umbilical vein endothelial cells) was detected to select the proper doses. Moreover, transcriptomics detection and data analysis were performed to screen out the special genes and the related pathways. Meanwhile, the regulation of lactoferrin in the functional factors thromboxane A2 (TXA2) and prostacyclin (PGI2) was detected. Then, the small interfering RNA (SiRNA) fragment of the selected gene pyridoxal phosphatase (PDXP) was transfected into HUVECs to validate its role in protecting HUVECs function. Results showed that lactoferrin inhibited the expression of TXA2 and activated expression of PGI2, as well as activated expression of PDXP, which significantly up-regulated the synthesis of vitamin B6 (VB6) and the phosphoinositide 3-kinase (PI3K)/ serine/threonine-protein kinase (AKT)/ extracellular regulated protein kinases (ERK) 1/2 pathway. For the first time, we revealed that lactoferrin could induce the synthesis of VB6 and protect HUVECs function through activating PDXP gene and the related pathway.

Positive influence of aqua exercise and burdock extract intake on fitness factors and vascular regulation substances in elderly.

Health issues in elderly individuals are often complex and tend to lead to chronic diseases; such issues can be due to a decline in fitness resulting from lack of physical activity. Aqua exercise and burdock are positive effects on cardiovascular disease and vascular health. This study investigated the changes due to aqua exercise and burdock extract intake in senior fitness, prostaglandin I2 (PGI2), and thromboxane A2 (TXA2) in elderly women. Forty elderly women (65-80 years) volunteered for this study. After baseline measurements, participants were randomized into control (n = 8), aqua exercise (n = 11), aqua exercise and burdock extract intake combination (n = 11), and burdock extract intake groups (n = 10). The variables of senior fitness tests, PGI2 and TXA2 were measured in all participants before and after the 12-week study. Blood collections were carried out at the beginning- and the end of aqua exercise training. Muscular strength, endurance, flexibility, and cardiorespiratory endurance of aqua exercise and burdock extract intake group at post-test significantly increased compared to pre-test (p<0.05). There were no significant differences in PGI2 and TXA2 between pre- and post-training programs. In conclusion, our findings indicated that the aqua exercise and burdock extract intake improves senior fitness factors in elderly Korean women. Also, the program participation led to a balance between PGI2 and TXA2. Additionally, burdock extract intake may be useful in vascular health by playing a secondary role in disease prevention and health promotion.

Thromboxane A2 Receptor Inhibition Suppresses Multiple Myeloma Cell Proliferation by Inducing p38/c-Jun N-terminal Kinase (JNK) Mitogen-activated Protein Kinase (MAPK)-mediated G2/M Progression Delay and Cell Apoptosis.

Multiple myeloma (MM) is a plasma cell malignancy without effective therapeutics. Thromboxane A2 (TxA2)/TxA2 receptor (T prostanoid receptor (TP)) modulates the progression of some carcinomas; however, its effects on MM cell proliferation remain unclear. In this study, we evaluated cyclooxygenase (COX) enzymes and downstream prostaglandin profiles in human myeloma cell lines RPMI-8226 and U-266 and analyzed the effects of COX-1/-2 inhibitors SC-560 and NS-398 on MM cell proliferation. Our observations implicate COX-2 as being involved in modulating cell proliferation. We further incubated MM cells with prostaglandin receptor antagonists or agonists and found that only the TP antagonist, SQ29548, suppressed MM cell proliferation. TP silencing and the TP agonist, U46619, further confirmed this finding. Moreover, SQ29548 and TP silencing promoted MM cell G2/M phase delay accompanied by reducing cyclin B1/cyclin-dependent kinase-1 (CDK1) mRNA and protein expression. Notably, cyclin B1 overexpression rescued MM cells from G2/M arrest. We also found that the TP agonist activated JNK and p38 MAPK phosphorylation, and inhibitors of JNK and p38 MAPK depressed U46619-induced proliferation and cyclin B1/CDK1 protein expression. In addition, SQ29548 and TP silencing led to the MM cell apoptotic rate increasing with improving caspase 3 activity. The knockdown of caspase 3 reversed the apoptotic rate. Taken together, our results suggest that TxA2/TP promotes MM cell proliferation by reducing cell delay at G2/M phase via elevating p38 MAPK/JNK-mediated cyclin B1/CDK1 expression and hindering cell apoptosis. The TP inhibitor has potential as a novel agent to target kinase cascades for MM therapy.

Sildenafil (Viagra®) Prevents Cox-1/ TXA2 Pathway-Mediated Vascular Hypercontractility in ApoE-/- Mice.

BACKGROUND/AIMS: The atherosclerotic apolipoprotein E-deficient (apoE-/-) mouse exhibits impaired vasodilation and enhanced vasoconstriction responsiveness. The objectives of this study were: a) to determine the relative contribution of cyclooxygenases (Cox-1 and Cox-2), thromboxane A2 (TXA2) and endothelin-1 (ET-1) to enhancing vascular hyperresponsiveness in this model of atherosclerosis and b) to investigate the beneficial effects of the phosphodiesterase 5 inhibitor sildenafil on this endothelial dysfunction. METHODS: Adult male apoE-/- mice were treated with sildenafil (40 mg/kg/day, for 3 weeks) and compared with non-treated ApoE-/- and wild-type mice. The beneficial effects of sildenafil on vascular contractile response to phenylephrine (PE) in aortic rings were evaluated before and after incubation with Cox-1 (SC-560) or Cox-2 (NS-398) inhibitors or the TP antagonist SQ-29548, and on contractile responsiveness to ET-1. RESULTS: ApoE-/- mice exhibited enhanced vasoconstriction to PE (Rmax ∼35%, p<0.01), which was prevented by treatment with sildenafil. The enhanced PE-induced contractions were abolished by both Cox-1 inhibition and TP antagonist, but were not modified by Cox-2 inhibition. Aortic rings from ApoE-/- mice also exhibited enhanced contractions to ET-1 (Rmax ∼30%, p<0.01), which were attenuated in sildenafil-treated ApoE-/- mice. In addition, we observed augmented levels of vascular proinflammatory cytokines in ApoE-/- mice, which were partially corrected by treatment with sildenafil (IL-6, IL-10/IL-6 ratio and MCP-1). CONCLUSION: The present data show that the Cox-1/TXA2 pathway prevails over the Cox-2 isoform in the mediation of vascular hypercontractility observed in apoE-/-mice. The results also show a beneficial effect of sildenafil on this endothelial dysfunction and on the proinflammatory cytokines in atherosclerotic animals, opening new perspectives for the treatment of other endothelium-related cardiovascular abnormalities.

Lipidomic profiling of bioactive lipids by mass spectrometry during microbial infections.

Bioactive lipid mediators play crucial roles in promoting the induction and resolution of inflammation. Eicosanoids and other related unsaturated fatty acids have long been known to induce inflammation. These signaling molecules can modulate the circulatory system and stimulate immune cell infiltration into the site of infection. Recently, DHA- and EPA-derived metabolites have been discovered to promote the resolution of inflammation, an active process. Not only do these molecules stop the further infiltration of immune cells, they prompt non-phlogistic phagocytosis of apoptotic neutrophils, stimulating the tissue to return to homeostasis. After the rapid release of lipid precursors from the plasma membrane upon stimulation, families of enzymes in a complex network metabolize them to produce a large array of lipid metabolites. With current advances in mass spectrometry, the entire lipidome can be accurately quantified to assess the immune response upon microbial infection. In this review, we discuss the various lipid metabolism pathways in the context of the immune response to microbial pathogens, as well as their complex network interactions. With the advancement of mass spectrometry, these approaches have also been used to characterize the lipid mediator response of macrophages and neutrophils upon immune stimulation in vitro. Lastly, we describe the recent efforts to apply systems biology approaches to dissect the role of lipid mediators during bacterial and viral infections in vivo.

Leukotriene Receptor Antagonists and Antiallergy Drugs.

As one of the candidates of the therapeutic strategy for asthma in addition to inhaled corticosteroids (ICS), leukotriene receptor antagonists (LTRAs) are known to be useful for long-term management of asthma patients complicated by allergic rhinitis (AR) or exercise-induced asthma (EIA). Currently available LTRAs are pranlukast hydrate, zafirlukast, and montelukast. These LTRAs have a bronchodilator action and inhibit airway inflammation, resulting in a significant improvement of asthma symptoms, respiratory function, inhalation frequency of as-needed inhaled ß2-agonist, airway inflammation, airway hyperresponsiveness, dosage of ICSs, asthma exacerbations, and patients' QOL. Although cys-LTs are deeply associated with the pathogenesis of asthma, LTRAs alone are less effective compared with ICS. However, the effects of LTRAs in combination with ICS are the same as those of LABAs in combination with ICS in steroid-naïve asthmatic patients. Concerning antiallergy drugs other than LTRAs, some mediator-release suppressants, H1 histamine receptor antagonists (H1RAs), thromboxane A2 (TXA2) inhibitors/antagonists, and Th2 cytokine inhibitor had been used mainly in Japan until the late 1990s. However, the use of these agents rapidly decreased after ICS/long acting beta agonist (LABA) combination was introduced and recommended for the management of asthma in the early 2000s. The effectiveness of other antiallergic agents on asthma management seems to be quite limited, and the safety of oral antiallergic agents has not been demonstrated in fetuses during pregnancy. Further effectiveness studies are needed to determine the true value of these orally administered agents in combination with ICS as an anti-asthma treatment.

Platelet testing in flow diversion: a review of the evidence.

OBJECTIVE Although the use of dual antiplatelet therapy with flow diversion is recommended and commonplace, the testing of platelet inhibition is more controversial. METHODS The authors reviewed the medical literature to establish and describe the physiology of platelet adhesion, the pharmacology of antiplatelet medications, and the mechanisms of the available platelet function tests. Additionally, they present a review of the pertinent neurointerventional and interventional cardiology literature. RESULTS Competing reports in the neurointerventional literature argue for and against the use of routine platelet function testing, with adjustments to the dosage or medications based on the results. The interventional cardiology literature has also wrestled with this dilemma after percutaneous coronary interventions, with conflicting reports of the benefits of platelet function testing. CONCLUSIONS Despite its prevalence, the benefits of platelet function testing prior to flow diversion are unproven. This practice will likely remain controversial until the level of evidence improves through more rigorous testing and reporting.

Effects of astaxanthin on blood coagulation, fibrinolysis and platelet aggregation in hyperlipidemic rats.

CONTEXT: Astaxanthin (ASTX) is a xanthophyll carotenoid that reduces hemostasis in hyperlipidemic organisms. Its antihemostatic mechanisms remain unclear. OBJECTIVE: The effects of ASTX on coagulation, the fibrinolytic system and platelet aggregation were investigated in hyperlipidemic rats. MATERIALS AND METHODS: Different doses of ASTX (5, 10 and 30 mg/kg/day, p.o.) were administered for four weeks to high-fat diet-induced hyperlipidemic rats. Serum lipid and lipoprotein levels were measured with an automatic biochemical analyzer. The prothrombin time (PT), activated partial thromboplastin time (APTT) and maximum platelet aggregation rate (MAR) were determined by a coagulation analyzer. The activities of the tissue-type plasminogen activator (t-PA), type-1 plasminogen activator inhibitor (PAI-1) and endothelial nitric oxide synthase (eNOS), as well as the levels of thromboxane B(2) [TXB(2)], 6-keto prostaglandin F(1α) [6-keto-PGF(1α)] and platelet granule membrane protein (GMP-140), were measured with enzyme-linked immunosorbent assay kits. Gene and protein expression levels were analyzed by reverse transcriptase polymerase chain reaction and Western blot, respectively. RESULTS: ASTX (30 mg/kg) treatment in hyperlipidemic rats reduced serum TG (0.58 ± 0.14 versus 1.12 ± 0.24 mmol/L), serum TC (1.77 ± 0.22 versus 2.24 ± 0.21 mmol/L), serum LDL-C (1.13 ± 0.32 versus 2.04 ± 0.48 mmol/L), serum MDA (69%), plasma MAR (55%), serum TXB2/6-keto-PGF1α (34%) and serum GMP-140 levels (25%), plasma PAI-1 activity (48%) and downregulated the mRNA (33%) and protein (23%) expression of aorta eNOS, the mRNA (79%) and protein (72%) expression levels of aorta PAI-1. However, ASTX (30 mg/kg/d) treatment increased serum SOD activity (2.1 fold), serum GPx activity (1.8 fold), plasma PT (1.3 fold), plasma APTT (1.7 fold), serum NO (1.4-fold), serum 6-keto-PGF1α (1.3 fold). CONCLUSIONS: ASTX reduced blood coagulation and platelet aggregation and promoted fibrinolytic activity in hyperlipidemic rats. These activities were closely correlated with ASTX, maintaining the balance of t-PA/PAI-1, NO/ROS and TXA2/PGI2 in vivo.

Antiplatelet and antithrombotic effects of cordycepin-enriched WIB-801CE from Cordyceps militaris ex vivo, in vivo, and in vitro.

BACKGROUND: A species of the fungal genus Cordyceps has been used as a complementary and alternative medicine of traditional Chinese medicine, and its major component cordycepin and cordycepin-enriched WIB-801CE are known to have antiplatelet effects in vitro. However, it is unknown whether they have also endogenous antiplatelet and antithrombotic effects. In this study, to resolve these doubts, we prepared cordycepin-enriched WIB-801CE, an ethanol extract from Cordyceps militaris-hypha, then evaluated its ex vivo, in vivo, and in vitro antiplatelet and antithrombotic effects. METHODS: Ex vivo effects of WIB-801CE on collagen- and ADP-induced platelet aggregation, serotonin release, thromboxane A2 (TXA2) production and its associated activities of enzymes [cyclooxygenase-1 (COX-1), TXA2 synthase (TXAS)], arachidonic acid (AA) release and its associated phosphorylation of phospholipase Cß3, phospholipase Cγ2 or cytosolic phospholipase A2, mitogen-activated protein kinase (MAPK) [p38 MAPK, extracellular signal-regulated kinase (ERK)], and blood coagulation time in rats were investigated. In vivo effects of WIB-801CE on collagen plus epinephrine-induced acute pulmonary thromboembolism, and tail bleeding time in mice were also inquired. In vitro effects of WIB-801CE on cytotoxicity, and fibrin clot retraction in human platelets, and nitric oxide (NO) production in RAW264.7 cells or free radical scavenging activity were studied. RESULTS: Cordycepin-enriched WIB-801CE inhibited ex vivo platelet aggregation, TXA2 production, AA release, TXAS activity, serotonin release, and p38 MAPK and ERK2 phosphorylation in collagen- and ADP-activated rat platelets without affecting blood coagulation. Furthermore, WIB-801CE manifested in vivo inhibitory effect on collagen plus epinephrine-induced pulmonary thromboembolism mice model. WIB-801CE inhibited in vitro NO production and fibrin clot retraction, but elevated free radical scavenging activity without affecting cytotoxicity against human platelets. CONCLUSION: WIB-801CE inhibited collagen- and ADP-induced platelet activation and its associated thrombus formation ex vivo and in vivo. These were resulted from down-regulation of TXA2 production and its related AA release and TXAS activity, and p38MAPK and ERK2 activation. These results suggest that WIB-801CE has therapeutic potential to treat platelet activation-mediated thrombotic diseases in vivo.