6-keto Prostaglandin F1α Research Articles

A Simple and Highly Sensitive Quantitation of Eicosanoids in Biological Samples Using Nano-flow Liquid Chromatography/Mass Spectrometry.

A simple sample preparation method for eicosanoid was developed by the combination of deproteinization and nanoLC-ESI-MS/MS. Eicosanoids are a group of bioactive lipid mediators, present in trace amounts in the body. Therefore, an analytical method for eicosanoids requires superior sensitivity. The method described in this report, which takes advantage of the highly sensitive power of nanoLC-ESI-MS/MS, enabled a simplification of the sample-preparation process. Eicosanoid extraction was performed just by homogenization in methanol with subsequent phospholipid removal, and then the liquid phase was directly subjected to nanoLC-ESI-MS/MS analysis without a condensation process. The quantitation range achieved 0.01 - 100 ng/mL for thromboxane B2, and 0.05 - 100 ng/mL for prostaglandin E2, prostaglandin D2, prostaglandin F2, leukotriene B4, 6-keto prostaglandin F1α and 11-dehydro thromboxane B2. Rat brain sample analyses demonstrated the feasibility of the quantification of those seven eicosanoids from biological samples.

Ethanol extract of Schisandrae chinensis fructus ameliorates the extent of experimentally induced atherosclerosis in rats by increasing antioxidant capacity and improving endothelial dysfunction

Context: Schisandrae chinensis fructus, the dried ripe fruit of Schisandra chinensis (Turcz.) Baill. (Magnoliaceae) has been used for thousands of years as a traditional Chinese herb, which can attenuate and prevent the development of cardiovascular events.Objective: To evaluate the effects of the ethanol extracts from Schisandrae chinensis fructus fruit (EESC) on experimental atherosclerosis (AS) in rats.Materials and methods: Treatment with EESC (0.35, 0.7, 1.4 g/kg/d, i.g.) and simvastatin (4 mg/kg/d, i.g.) on AS rats for 3 weeks. Sprague–Dawley rats on normal chow and under water treatment were used as control. The content of schisandrin, schisandrin A and schisandrin B in EESC was detected by HPLC. Aortic pathology changes, serum biochemical indices and nuclear factor E2-related factor 2 (Nrf-2) and heame oxygenase-1 (HO-1) expressions were measured.Results: Schisandrin, schisandrin A and schisandrin B contents were 291.8, 81.46 and 279.1 mg/g of dry weight, respectively. EESC significantly reduced the aortic plaque area (76.5, 90.5 and 73.9% reduction), regulated the levels of serum lipid (p < 0.05), enhanced the antioxidant enzyme activities (p < 0.01), reduced the malondialdehyde levels (72.5, 69.3, 67.3%), and up-regulated the Nrf-2 and HO-1 expression (p < 0.05). Furthermore, EESC reduced the levels of oxidized-LDL and endothelin-1 and thromboxane B2 but increased that of 6-keto prostaglandin F1α (p < 0.05). Acute toxicity was calculated on mice to be LD50 > 20 g/kg.Conclusions: EESC positively affects the treatment of AS in vivo and the findings will provide a reliable theoretical basis for developing novel therapeutics.

Thrombocytic activity in patients with arterial hypertension associated with metabolic syndrome

This work was aimed to elucidate the level of in vitro and in vivo aggregation activity of platelets and the functional significance of individual mechanisms of its regulation in patients with grade III arterial hypertension and metabolic syndrome. The study included 29 adult patients (15 men and 14 women) and 25 clinically healthy subjects of similar age. Biochemical, hematological and statistical methods were used. Marked dyslipidemia was associated with active lipid peroxidation. Plasma thromboxane B2 level was increased by 84,8% while 6-ketoprostaglandin F1α level was decreased by 17,9% and the total amount of NO metabolites by 28,7%. The degree of platelet aggregation and their aggregation with collagen 25,0 and 27,5% lower than the respective control values while the respective indices of their aggregation with ristomycin were 25,7 and 46,4% higher. The degree of platelet aggregation and their aggregation with ADP inducer were 25,7 and 58,4% higher than in control while the platelet-discocyte levels were reduced to 48,6 ± 0,4%. The sum of active platelet forms reached 51,4 ± 0,12% vs 17,9 ± 0,09% in control was, the number of small and large aggregates 18,6 ± 0,08 and 5,4 ± 0,04 per 1000 free platelets respectively vs 2,9 ± 0,06 и 0,2 ± 0,06 in control. Excess platelet activity in the patients was due to their enhanced adhesive and aggregative potential and reduced ability to disaggregate. The most important causs of thrombocytopathy was AH, negative changes in plasma lipid composition, and enhanced lipid peroxidation.

Reduced platelet aggregation in women after intercourse: a possible role for the cyclooxygenase pathway.

We hypothesise that molecules in the cyclooxygenase pathway affect platelet activity when seminal fluid (SF) is present. We considered the influence of SF on platelet aggregation in women, and believe that the prostanoids in SF signalling are significant. Thirty-one female subjects were studied, 20 of whom were sexually active. Male partners were given either aspirin or indomethacin to inhibit cyclooxygenase. The 6-keto prostaglandin F1α (6-keto PGF1α) and prostaglandin E metabolite (PGE-M) in SF were measured by competitive assay. Platelets and prostanoids were evaluated in women, periodically, before and after intercourse. The platelets were tested with adenosine diphosphate (ADP) and arachidonic acid (AA). To block the interaction between the uterus and SF, some couples used condoms. We found that the 6-keto prostaglandin F1α in urine at 2hours post-intercourse (1418.75pg/mL, Std 688.39) was greater than pre-intercourse (772.68pg/mL, Std 116.54). Post-intercourse, a transient decrease in platelet aggregation was observed in women whose partners did not use condoms. Averages for platelet aggregation were 20.16% with ADP, and more significantly, 37.79% with AA after 2hours. In contrast, couples using condoms showed no changes, averaging 64.02% with ADP and 72.06% with AA. Women whose partners were taking aspirin or indomethacin also showed no changes. SF from men taking aspirin or indomethacin led to no reduction in platelet aggregometry in their partners. These results indicate that in cases of exposure to SF, the transient change in women's platelet activity could be related to the cyclooxygenase pathway.

Relaxin treatment reduces angiotensin II-induced vasoconstriction in pregnancy and protects against endothelial dysfunction†.

The peptide relaxin has gained considerable attention as a new vasoactive drug, largely through its beneficial therapeutic effects in cardiovascular disease. In this study, we tested the hypothesis that relaxin treatment alleviates systemic vascular dysfunction characteristic of hypertensive diseases of pregnancy. We investigated vascular effects and mechanisms of relaxin action in (i) pregnant relaxin-deficient (Rln-/-) mice with enhanced responses to angiotensin II (AngII) and (ii) arteries pre-incubated ex vivo in trophoblast conditioned media (TCM) to induce endothelial dysfunction. Pregnant Rln-/- mice received 0.5 μg/h recombinant human H2 relaxin (rhRLX: n = 5) or placebo (20 nM sodium acetate; n = 7) subcutaneously via osmotic minipumps for 5 days prior to gestational day 17.5. This treatment protocol significantly reduced AngII-mediated contraction of mesenteric arteries and increased plasma 6-keto prostaglandin F1α. These vascular effects were endothelium independent and likely involve smooth muscle-derived vasodilator prostanoids. In the second study, mesenteric arteries were incubated ex vivo for 24 h at 37°C in TCM, which contained high levels of soluble Flt-1 (>20 ng/ml) and soluble Eng (>1 ng/ml). TCM incubation caused significant reduction in endothelium-dependent relaxation and increased sensitivity to AngII. Co-incubation of arteries with rhRLX for 24 h (n = 6-16/treatment) prevented endothelial dysfunction but had no effect on AngII-mediated contraction. In conclusion, relaxin treatment prevents and/or reverses vascular dysfunction in mesenteric arteries, but acts through different vascular pathways depending on duration of relaxin treatment and type of vascular dysfunction. Overall, our data suggest that relaxin is a potential therapeutic to alleviate maternal systemic vascular dysfunction associated with hypertensive diseases in pregnant women.

Effects of Shuxuetong injection applied in acute ischemic stroke

ObjectiveTo study the effects of Shuxuetong injection in adjuvant treatment of ischemic stroke on the degree of nerve injury, lipid metabolism and blood coagulation function. MethodsPatients with ischemic stroke admitted in our hospital during the period from May 2012 to May 2015 were selected for retrospective analysis. They were divided into the control group receiving regular treatment and the Shuxuetong group receiving adjuvant treatment with Shuxuetong injection. One and the three months after treatment, serum was collected and nerve injury molecules, indexes of lipid metabolism and blood coagulation function were measured. ResultsOne month after treatment, the contents of neuron-specific enolase, S100 calcium binding protein B, total cholesterol, triglyceride, low-density lipoprotein, oxidized low-density lipoprotein, thromboxane B2, fibrinogen and D-dimer in the serum of patients from Shuxuetong group were significantly lower than those of control group. The contents of high-density lipoprotein and 6-keto prostaglandin F1α were significantly higher than those of control group. Three months after treatment, the contents of neuron-specific enolase, S100 calcium binding protein B, total cholesterol, triglyceride, low-density lipoprotein, oxidized low-density lipoprotein, thromboxane B2, fibrinogen and D-dimer in the serum of patients from Shuxuetong group were significantly lower than those of control group. The contents of high-density lipoprotein and 6-keto prostaglandin F1α were significantly higher than those of control group. ConclusionsAdjuvant treatment with Shuxuetong injection can reduce the injury of nerve function of patients with ischemic stroke and improve blood lipid metabolism and blood coagulation function, which is an effective drug for the treatment of ischemic stroke.

Mechanisms Underlying Enhanced Noradrenaline-Induced Femoral Arterial Contractions of Spontaneously Hypertensive Rats: Involvement of Endothelium-Derived Factors and Cyclooxygenase-Derived Prostanoids.

We investigated the relationship between noradrenaline (NAd)-induced contractions, endothelial function, and hypertension in femoral arteries isolated from spontaneously hypertensive rats (SHR). In the femoral arteries of SHR, vs. age-matched control Wistar Kyoto (WKY) rats, contractions induced by NAd were increased. These effects were enhanced by endothelial denudation, which abolished the differences between the two groups. NAd-induced contractions were enhanced by nitric oxide (NO) synthase inhibition, and further increased by the blockade of endothelium-derived hyperpolarizing factor (EDHF). Conversely, NAd-induced contractions were inhibited by cyclooxygenase (COX) inhibition. In addition, in SHR arteries, acetylcholine-induced relaxation was reduced, and components of endothelium-derived factors were altered, such as increased COX-derived vasoconstrictor prostanoids, reduced EDHF, and preserved NO-mediated relaxation. In the femoral arteries of SHR, the production of prostanoids [6-keto prostaglandin (PG)F1α (a metabolite of prostacyclin (PGI2), PGE2, and PGF2α] and COX-2 protein were increased compared with that in WKY rats. By contrast, contractions induced by beraprost (a stable PGI2 analogue), PGE2, and U46619 (thromboxane/prostanoid receptor agonist) were similar between the SHR and WKY groups. Thus, NAd-induced femoral arterial contractions are augmented in SHR resulting from endothelial dysfunction and increased COX-derived vasoconstrictor prostanoid levels.

Cyclooxygenase 1 (COX1) expression in Type 2 diabetes mellitus: A preliminary study from north India

Background and purposeThe 6th edition of International Diabetes Federation, 2014 shows an estimate of 387 million people with Type 2 diabetes mellitus (T2DM) worldwide, expected to rise to 592 million by 2035. T2DM is a metabolic disorder, one of the reasons being oxidative stress due to impairment in antioxidant enzymes. It leads to several complications such as micro and macrovascular diseases. Cyclooxygenase1 (COX1) enzyme is the rate limiting factor for the arachidonic pathway leading to vascular wall contraction with angiotensin II occurring in heart diseases resulting from T2DM. COX1 determines 6-Keto Prostaglandin F1α (6-k-PGF1α) level, plays a major role in vasodilation and restricts macrophage platelet aggregation. The aim of the present study was to compare the COX1 expression and level of reactive oxygen species (ROS) in T2DM patients and controls at different time periods in human macrophages in order to find a biomarker or drug target. Subjects and methodsThe study subjects consisted of 100 individuals, 50 each from T2DM patients and healthy sex/age matched controls. Cell proliferation by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and ROS measurement by 2′,7′-dichlorofluorescein diacetate (DCFDA) staining were performed at different time periods (24, 48, 72h). COX1 mRNA expression was checked by relative quantification method after real-time polymerase chain reaction (RT-PCR). ResultsThe MTT assay showed that cell viability was significantly higher at 48h (P<0.05). ROS production was found to be lowest at 24h by DCFDA staining. ROS levels were raised in T2DM patients as compared to controls. The quantitative RT-PCR analysis showed that the COX1 expression was higher in T2DM patients as compared to healthy controls although not significant (P>0.05). ConclusionAlthough COX1 is known to be a “housekeeping” gene, our study showed that its expression can be correlated with the disease condition and be used as a marker. However, further studies are required in more number of samples from other ethnic populations to confirm the findings.

Celecoxib coupled to dextran via a glutamic acid linker yields a polymeric prodrug suitable for colonic delivery.

Celecoxib, a selective cyclooxygenase-2 inhibitor, is potentially useful for the treatment of colonic diseases such as colorectal cancer and colitis. However, the cardiovascular toxicity of celecoxib limits its routine use in the clinic. Generally, colon-specific delivery of a drug both increases the therapeutic availability in the large intestine and decreases the systemic absorption of the drug, most likely resulting in enhanced therapeutic effects against colonic diseases such as colitis and reduced systemic side effects. To develop a colon-specific prodrug of celecoxib that could reduce its cardiovascular toxicity and improve its therapeutic activity, dextran–glutamic acid–celecoxib conjugate (glutam-1-yl celecoxib-dextran ester [G1CD]) was prepared and evaluated. While stable in pH 1.2 and 6.8 buffer solutions and small-intestinal contents, G1CD efficiently released celecoxib in cecal contents. Oral administration of G1CD to rats delivered a larger amount of celecoxib to the large intestine than free celecoxib. G1CD prevented the systemic absorption of celecoxib and did not decrease the serum level of 6-ketoprostaglandin F1α, an inverse indicator of cardiovascular toxicity of celecoxib. Collectively, G1CD may be a polymeric colon-specific celecoxib prodrug with therapeutic and toxicological advantages.

Analysis of fatty acid samples by hydrophilic interaction liquid chromatography and charged aerosol detector

HILIC/CAD techniques were used in analysis of samples containing fatty acids. Amine base column appeared to be the more retentive stationary phase compared to zwitterionic and BEH silica. The retention decreased with pH mobile phases changing from 3 to 5. Acetonitrile and acetone organic modifier were compared. Acetone gave higher eluotropic strength and better peak symmetry whereas acetonitrile led to higher efficiency. The retention decreased when ammonium acetate concentration increased from 5 to 20mM. The use of sub-2μm column did not show flat Van Deemter curves at high flow rates.A rapid separation of PGI2 and its main degradation product, 6-keto prostaglandin F1α was obtained in 1.6min with a Hypersil GOLD, 50mm×2.1mm, 1.9μm with; acetonitrile/acetate ammonium pH 5 at 20mM (85/15; v/v at 0.7ml/min).

Thyroid hormone affects both endothelial and vascular smooth muscle cells in rat arteries

Hypothyroidism impairs endothelium-dependent dilatations, while hyperthyroidism augments the production of endothelial nitric oxide. Thus, experiments were designed to determine if thyroid hormone causes endothelium-dependent responses, or alleviates diabetic endothelial dysfunction. Isometric tension was measured in rings with or without endothelium of arteries from normal and diabetic Sprague-Dawley rats. Release of 6-keto prostaglandin F1α and thromboxane B2 were measured by enzyme linked immunosorbent assay and protein levels [endothelial nitric oxide synthase (eNOS), cyclooxygenases (COX)] by immunoblotting. Triiodothyronine (T3) caused concentration-dependent (3×10−6–3×10−5M) relaxations in mesenteric (pEC50, 4.96±0.19) and femoral (pEC50, 4.57±0.35) arteries without endothelium. In femoral arteries of rats with diabetes, 5-methylamino-2-[[(2S,3R,5R,8S,9S)-3,5,9-trimethyl-2-(1-oxo-(1H-pyrrol-2- -yl)propan-2-yl)-1,7-dioxaspiro-(5,5)undecan-8-yl]methyl]benzooxazole-4-carboxylic acid (A23187, 3×10−7 to 10−6M) caused partly endothelium-dependent contractions. After chronic T3-treatment with (10μg/kg/day; four weeks), the contractions to A23187 of preparations with and without endothelium were comparable, the thromboxane B2-release was reduced (by 38.1±9.2%). The pEC50 of 9, 11-dideoxy-11α, 9α-epoxymethanoprostaglandin F2α (U46619, TP-receptor agonist) was increased in T3-treated diabetic rats compared with controls (8.53±0.06 vs 7.94±0.09). The protein expression of eNOS increased (by 228%) but that of COX-1 decreased (by 35%) after chronic T3 treatment. In human umbilical vein endothelial cells incubated for one week with T3 (10−10–10−7M) in the presence but not in the absence of interleukin-1β (1ng/ml), the expression of eNOS was increased compared to control. In conclusion, thyroid hormone acutely relaxes mesenteric and femoral vascular smooth muscle, but given chronically reduces the release of endothelium-derived vasoconstrictor prostanoids while enhancing the responsiveness of TP receptors of vascular smooth muscle.

Effects of danshensu on platelet aggregation and thrombosis: in vivo arteriovenous shunt and venous thrombosis models in rats.

Danshensu, a type of dihydroxyphenyl lactic acid, is one of the most abundant active phenolic acids in the dried root of Salvia miltiorrhizae (Lamiaceae)—widely used traditional Chinese medicine. The effects of danshensu on platelet aggregation and thrombus formation in rats were examined using various methods. It was found that danshensu significantly reduced thrombus weight in 2 experimental thrombosis models; dose-dependent inhibition of adenosine diphosphate (ADP) and arachidonic acid (AA)-induced platelet aggregation occurred in normal and blood stasis-induced rats; Danshensu also significantly mitigated blood viscosity, plasma viscosity and hematocrit levels. Moreover, danshensu significantly inhibited venous thrombosis-induced expression of cyclooxygenases-2 (COX-2) rather than cyclooxygenases-1(COX-1) in the venous walls, down regulated thromboxane B2 (TXB2) and up regulated 6-keto prostaglandin F1α (6-keto-PGF1α), normalizing the TXB2/6-keto-PGF1α ratio. In addition, danshensu did not induce gastric lesions and even had protective effects on aspirin-induced ulcer formation at doses as high as 60 mg/kg. These findings suggest that the antithrombotic and antiplatelet aggregation effects of danshensu are attributed to its highly selective inhibition of COX-2 and ability to normalize the thromboxane A2(TXA2)/prostacyclin(PGI2) balance. These findings suggest that danshensu have great prospects in antithrombotic and antiplatelet therapy.

Nano-LC-MS/MS for the quantitation of prostanoids in immune cells.

Prostanoids, derivatives of arachidonic acid, are involved in inflammation and immune reactions. To understand the role of prostanoids produced by diverse immune cells, a highly sensitive quantitation method for prostaglandin E2 (PGE2), prostaglandin D2 (PGD2), 6-keto prostaglandin F1α (6-keto PGF1α), prostaglandin F2α (PGF2α), and thromboxane B2 (TXB2) by means of nano-liquid chromatography-tandem mass spectrometry has been developed. It was validated according to the guidelines of the Food and Drug Administration (FDA) in terms of linearity, precision, accuracy, recovery, stability, and lower limit of quantitation (LLOQ). The LLOQ were 25pg/mL in the injected solution (75fg on column (o.c.)) for PGE2 and PGD2 and 37.5pg/mL (112.5fg on column) for 6-keto PGF1α, PGF2α, and TXB2, respectively. It was successfully applied to murine mast cells isolated from paws after zymosan injection and to CD4(+) and CD8(+) T lymphocytes from blood of sensitized versus non-sensitized mice in context of a delayed type hypersensitivity model. About 5,000 (T cells) to 40,000 (mast cells) cells were sufficient for quantitation. In the mast cells, the production of PGE2 increased at a significantly higher extent than the synthesis of the other prostanoids. The T lymphocytes did not show any difference in prostanoid production, no matterwhether they were obtained from sensitized mice or non-sensitized mice.

Abstract 19092: Cyclooxygenase 2 Mediates Increased Endothelium-Dependent Vascular Contraction in Hyperhomocysteinemia and Hyperlipidemia

Objectives: We recently reported that elevation of plasma homocysteine level_hyperhomocysteinemia (HHcy) accelerates atherosclerosis and vascular inflammation in hyperlipidemia (HLP). In this study, we examined the role of cyclooxygenase 2 (COX2)[[Unable to Display Character: —]]an inducible isoform of COX in responsible to inflammation, in HHcy/HPL-induced endothelial dysfunction (ED). Methods: Endothelial function was assessed in the thoracic aorta of a novel severe HHcy (plasma Hcy=221 μM) mouse model: human cystathionine β-synthase (CBS) transgenic, mouse CBS and apolipoprotine E gene deficient mouse fed with a high fat diet for 8 weeks (HHcy/HLP Cbs-/-¬ mice). Results: Endothelium-dependent relaxation to acetylcholine (ACh) was significantly impaired in HHcy/HLP Cbs-/- mice compared to HHcy/HLP Cbs+/+ mice. In the presence of eNOS inhibitor N(G)-nitro-L-Arg-methyl ester (L-NAME), ACh induced endothelium-dependent contraction in HHcy/HLP Cbs-/- but not in HHcy/HLP Cbs+/+ mice. L-NAME plus COX1/2 inhibitor indomethacin (INDO) completely blocked the HHcy/HLP-induced endothelium-dependent contraction. Moreover, INDO, COX2 inhibitor nimesulide, and thromboxane A2/prostaglandin H2 receptor antagonist SQ29548, but not COX1 inhibitor SC560, improved vascular relaxation to ACh in HHcy/HLP Cbs-/-¬ mice, indicating that COX2 plays a major role. Preincubation the aorta with antioxidants Tempol, polyethylene glycol superoxide dismutase and apocynin also improved endothelial function in HHcy/HLP Cbs-/- mice. Expression of COX2 and 3-nitrotyrosin (3-NT), but not COX1, was markedly increased in the endothelium of aorta of HHcy/HLP Cbs-/- mice. ICAM-1, VCAM-1, and TNFα were markedly increased in the aorta of HHcy/HLP Cbs-/- mice. Moreover, COX2 and 3-NT, but not COX1, was significantly increased in cultured human aortic endothelial cells treated with a combination of DL-Hcy (500 μM) and oxidized-LDL (200 μg/ml) for 48 hrs. Furthermore, DL-Hcy/oxidized-LDL-induced COX2 was prevented by PEG-SOD. Finally, HHcy markedly accelerated urinary 2,3-dinor TXB2, TXB2, 8-isoprostan and 6-keto prostaglandin F1α levels in HPL. Conclusions: COX2 mediates HHcy-induced endothelium-dependent vascular contraction in HPL via oxidative stress-dependent pathway.

Aspirin augments carotid-cardiac baroreflex sensitivity during muscle mechanoreflex and metaboreflex activation in humans

Muscle mechanoreflex activation decreases the sensitivity of carotid baroreflex (CBR)-heart rate (HR) control during local metabolite accumulation in humans. However, the contribution of thromboxane A2 (TXA2) toward this response is unknown. Therefore, the effect of inhibiting TXA2 production via low-dose aspirin on CBR-HR sensitivity during muscle mechanoreflex and metaboreflex activation in humans was examined. Twelve young subjects performed two trials during two visits, preceded by 7 days' low-dose aspirin (81 mg) or placebo. One trial involved 3-min passive calf stretch (mechanoreflex) during 7.5-min limb circulatory occlusion (CO). In another trial, CO was preceded by 1.5 min of 70% maximal voluntary contraction isometric calf exercise to accumulate metabolites during CO and stretch (mechanoreflex and metaboreflex). HR (ECG) and mean arterial pressure (Finometer) were recorded. CBR function was assessed using rapid neck pressures ranging from +40 to -80 mmHg. Aspirin significantly decreased baseline thromboxane B2 production by 84 ± 4% (P < 0.05) but did not affect 6-keto prostaglandin F1α. Following aspirin, stretch with metabolite accumulation significantly augmented maximal gain (GMAX) and operating point gain (GOP) of CBR-HR (GMAX; -0.71 ± 0.14 vs. -0.37 ± 0.08 and GOP; -0.69 ± 0.13 vs. -0.35 ± 0.12 beats·min(-1)·mmHg(-1) for aspirin and placebo, respectively; P < 0.05). CBR-HR function curves were reset similarly with aspirin and placebo during stretch with metabolite accumulation. In conclusion, these findings suggest that low-dose aspirin augments CBR-HR sensitivity during concurrent muscle mechanoreflex and metaboreflex activation in humans. This increased sensitivity appears linked to reduced TXA2 production, which likely plays a role in metabolite sensitization of muscle mechanoreceptors.

GW24-e3751 Oxidative stress-dependent cyclooxygenase 2-derived vasoconstrictive prostaglandins contribute to hyperhomocysteinemia-induced endothelial dysfunction in hyperlipidemia

ObjectivesWe recently reported that elevation of plasma homocysteine level—hyperhomocysteinemia (HHcy) accelerates atherosclerosis and vascular inflammation in hyperlipidemia. Prostanoids are local mediators of inflammation and are synthesised from arachidonic acid by...

Acidosis prevents and alkalosis augments endothelium-dependent contractions in mouse arteries

Changes in pH modulate the responsiveness of vascular smooth muscle cells to vasoconstrictor stimuli, but their effect on endothelium-dependent responses is unknown. Therefore, the influence of moderate changes in pH on responses to endothelium-dependent and -independent agonists was determined in aortae and carotid arteries of 15- to 26-week-old male C57BL/6N mice. Isolated rings were suspended in Halpern-Mulvany myographs for isometric tension recording. The preparations were exposed to either acidic (pH 7), control (pH 7.4) or alkaline (pH 7.8) modified Krebs-Ringer bicarbonate buffer solutions and their contractions and relaxations compared. Endothelium-dependent relaxations to acetylcholine (in the presence of meclofenamate or of the thromboxane-prostanoid (TP) receptor antagonist S18886) were comparable at the three pH values tested in contracted aortic or carotid arterial rings. Endothelium-dependent contractions of quiescent carotid arteries were reduced in acidosis and potentiated in alkalosis compared to control; these effects were reversible. The carotid arteries produced equal amounts of 6-keto prostaglandin F1α and thromboxane B2 at the different pH values tested. Contractions to the full TP receptor agonist U46619 were similar in the three milieus, but after inducing partial TP receptor blockade (with low concentrations of the TP receptor antagonist S18886) they were depressed in acidosis compared to alkalosis. Prostacyclin as a partial TP receptor activator also induced weaker contractions at low than at high pH, whereas its vasodilator effect was not affected. These findings demonstrate that changes in pH modulate endothelium-dependent contractions in mouse arteries primarily by altering the sensitivity of TP receptors of vascular smooth muscle to endothelium-derived contracting factors.

Effects of COX-2 inhibitor on ventilator-induced lung injury in rats

Mechanical ventilation especially with large tidal volume has been demonstrated to activate inflammatory response inducing lung injury, which could be attenuated by cyclooxygenase (COX)-2 inhibitors. As the main small integral membrane proteins that selectively conduct water molecules' transportation, aquaporin (AQP)-1 downregulation significantly related to lung edema and inflammation. This study aims to investigate the role of AQP1 in ventilator-induced lung injury in rats and evaluates the effects of COX-2 inhibition. Forty rats were allocated into four groups, where rats in Groups LD (low volume+DMSO) and LN (low volume+NS-398) were given intravenously 2ml DMSO and 8mg/kg NS-398 (a specific COX-2 inhibitor, dissolved in 2ml DMSO) before 4-hour lower tidal volume ventilation (8ml/kg), respectively, while DMSO and NS-398 were administrated in the same manner before 4-hour injurious ventilation (40ml/kg) in Groups HD (high volume+DMSO) and HN (high volume+NS-398). The arachidonic acid metabolites (6-keto prostaglandin F1α, thromboxane B2), inflammatory cytokines (tumor necrosis factor-α, interleukin-1β, 6, 8) and total protein levels in bronchoalveolar lavage (BAL) fluid and COX-2 mRNA and AQP1 protein expression in lung tissue were detected; water content and lung morphology were also evaluated. Compared to Groups LD and LN, the rats in Groups HD and HN suffered obvious lung morphological changes with higher wet-to-dry weight ratio and lung injury score, and the levels of arachidonic acid metabolites, inflammatory cytokines and total protein in BAL fluid were increased, the expression of COX-2 mRNA was significantly upregulated and AQP1 protein was downregulated in lung tissue (p<0.05). The changes in BAL fluid and the severity of lung injury were attenuated, and AQP1 expression was upregulated in Group HN as compared to HD (p<0.05). Ventilation with large tidal volume causes inflammatory mediator production and AQP1 downregulation, which could be attenuated by COX-2 inhibition.

Effect of Cyclooxygenase-2 Blockade on Renal Hypertrophy Development during Early Diabetes Mellitus

Diabetes mellitus is the leading cause of diabetic nephropathy; the early phase of diabetes is associated with kidney growth and hyperfiltration; several factors modulate these changes, among them, prostaglandins and angiotensin II. Previous studies have shown that cyclooxygenase-2 is implicated in experimental models of diabetes. The aim of this work was to study the effect of celecoxib treatment on renal hypertrophy development in early diabetes mellitus. In our rats with early streptozotocin-induced diabetes there was renal hypertrophy, and increased renal expression of cyclooxygenase-2, AT1 receptor, and transforming growth factor-β1. Treatment with the selective cyclooxygenase-2 inhibitor celecoxib reduced the urinary excretion of prostaglandins such as prostaglandin E2, 6-keto prostaglandin F1α, and thromboxane B2. Kidney hypertrophy was reversed by the treatment, and the renal expression of cyclooxygenase-2, AT1 receptor, and transforming growth factor-β1 decreased. The renoprotective effects of celecoxib were independent of the changes in plasma glucose levels. These results confirm that cyclooxygenase-2 inhibition in rats with streptozotocin-induced diabetes decrease renal hypertrophy; this effect in turn, may be mediated by reduction of the expression of AT1 receptors and transforming growth factor-b1 in the kidney.

Effects of Yerba Mate tea (Ilex paraguariensis) on vascular endothelial function and liver lipoprotein receptor gene expression in hyperlipidemic rats

Yerba Mate tea (Mate), an infusion made from the leaves of the tree Ilex paraguariensis, is a widely consumed beverage in South America. This study was performed to investigate the effect of Mate tea on vascular endothelial dysfunction and liver lipoprotein receptor gene expression in hyperlipidemic rats, with the aim of gaining insight into its known lipid-lowering protective mechanisms. Sixty male Sprague–Dawley rats were randomly divided into five groups: a normal control group (NC), a high-fat diet group (HC), and three Mate tea-treated groups. In the NC group, rats were fed with standard diet while in the other groups the rats were fed a high-fat diet for 8weeks. In the Mate tea-treated groups, the rats were fed a high-fat diet supplemented with low, moderate or high concentrations of aqueous Mate tea extract for the final 4weeks. Compared to the HC group, aqueous Mate tea extract significantly reduced endothelin (ET) and thromboxane B2 (TXB2) levels and increased nitric oxide (NO) and 6-keto prostaglandin F1α (6-keto-PGF1α) levels in the blood, reduced the pathological damage of vascular endothelial cells, decreased intercellular adhesion molecule-1 (ICAM-1) protein expression in the thoracic aorta, and upregulated mRNA expression of hepatic low density lipoprotein receptor (LDLR) and scavenger receptor B1 (SR-B1). These findings indicate that Mate tea administration might have a regulatory effect on blood fat and endothelial function in hyperlipidemia rats. The mechanism may involve protecting vascular endothelial cell function and upregulating the expression of LDLR and SR-B1 genes, thereby inhibiting the occurrence of atherosclerosis.