Novelty of Sphingolipids in the Central Nervous System Physiology and Disease: Focusing on the Sphingolipid Hypothesis of Neuroinflammation and Neurodegeneration.

For decades, lipids were confined to the field of structural biology and energetics as they were considered only structural constituents of cellular membranes and efficient sources of energy production. However, with advances in our understanding in lipidomics and improvements in the technological approaches, astounding discoveries have been made in exploring the role of lipids as signaling molecules, termed bioactive lipids. Among these bioactive lipids, sphingolipids have emerged as distinctive mediators of various cellular processes, ranging from cell growth and proliferation to cellular apoptosis, executing immune responses to regulating inflammation. Recent studies have made it clear that sphingolipids, their metabolic intermediates (ceramide, sphingosine-1-phosphate, and N-acetyl sphingosine), and enzyme systems (cyclooxygenases, sphingosine kinases, and sphingomyelinase) harbor diverse yet interconnected signaling pathways in the central nervous system (CNS), orchestrate CNS physiological processes, and participate in a plethora of neuroinflammatory and neurodegenerative disorders. Considering the unequivocal importance of sphingolipids in CNS, we review the recent discoveries detailing the major enzymes involved in sphingolipid metabolism (particularly sphingosine kinase 1), novel metabolic intermediates (N-acetyl sphingosine), and their complex interactions in CNS physiology, disruption of their functionality in neurodegenerative disorders, and therapeutic strategies targeting sphingolipids for improved drug approaches.

Targeting cyclooxygenase enzyme for the adjuvant COVID-19 therapy.

Despite vigorous efforts, the COVID-19 pandemic continues to take a toll on the global health. The contemporary therapeutic regime focused on the viral spike proteins, viral 3CL protease enzyme, immunomodulation, inhibition of viral replication, and providing a symptomatic relief encouraged the repurposing of drugs to meet the urgency of treatment. Similarly, the representative drugs that proved beneficial to alleviate SARS-CoV-1, MERS-CoV, HIV, ZIKV, H1N1, and malarial infection in the past presented a sturdy candidature for ameliorating the COVID-19 therapeutic doctrine. However, most of the deliberations for developing effective pharmaceuticals proved inconsequential, thereby encouraging the identification of new pathways, and novel pharmaceuticals for capping the COVID-19 infection. The COVID-19 contagion encompasses a burst release of the cytokines that increase the severity of the infection mainly due to heightened immunopathogenicity. The pro-inflammatory metabolites, COX-2, cPLA2, and 5-LOX enzymes involved in their generation, and the substrates that instigate the origination of the innate inflammatory response therefore play an important role in intensifying and worsening of the tissue morbidity related to the coronavirus infection. The deployment of representative drugs for inhibiting these overexpressed immunogenic pathways in the tissues invaded by coronaviruses has been a matter of debate since the inception of the pandemic. The effectiveness of NSAIDs such as Aspirin, Indomethacin, Diclofenac, and Celecoxib in COVID-19 coagulopathy, discouraging the SARS viral replication, the inflammasome deactivation, and synergistic inhibition of H5N1 viral infection with representative antiviral drugs respectively, have provided a silver lining in adjuvant COVID-19 therapy. Since the anti-inflammatory NSAIDs and COXIBs mainly function by reversing the COX-2 overexpression to modulate the overproduction of pro-inflammatory cytokines and chemokines, these drugs present a robust treatment option for COVID-19 infection. This commentary succinctly highlights the various claims that support the status of immunomodulatory NSAIDs, and COXIBs in the adjuvant COVID-19 therapy.

Long-term nitric oxide synthase inhibition prevents 17ß-estradiol-induced suppression of cyclooxygenase-dependent contractions and enhancement of endothelium-dependent hyperpolarization-like relaxation in mesenteric arteries of ovariectomized rats.

Endothelial dysfunction is associated with a reduced bioavailability of nitric oxide (NO). In this study, the effects of 17ß-estradiol supplement on endothelial function were examined in ovariectomized (OVX) rats following long-term inhibition of NO synthases with L-NAME. Female Sprague Dawley rats were ovariectomized at 12 weeks old. They were supplemented with 17ß-estradiol (25 µg/kg/day, intramuscularly) or its vehicle (olive oil) until they were killed. At 18 weeks old, they were administered daily with NO synthase inhibitor L-NAME (60 mg/kg, by gavage) or its vehicle (distilled water) for 6 weeks. Rats were then anesthetized for blood pressure measurement and for isolation of mesenteric arteries and aortae for isometric tension measurement. Long-term L-NAME-treatment, without or with 17ß-estradiol supplement, resulted in reduced plasma nitrite/nitrate level without causing an increase in blood pressure in OVX rats. Acute inhibition of cyclooxygenase (COX) with indomethacin improved relaxations of mesenteric arteries to the calcium ionophore A23187 in OVX rats, and in those with long-term L-NAME-treatment without or with 17ß-estradiol supplement, but not in those with female hormone supplement only. 17ß-estradiol supplement or long-term L-NAME-treatment resulted in a greater endothelium-dependent hyperpolarization-like relaxation in mesenteric arteries. In the quiescent aorta, 17ß-estradiol supplement or long-term L-NAME-treatment unmasked the COX-dependent components of A23187-induced contractions, but prevented that of the smooth muscle contractions to U46619 in OVX rats. In summary, long-term 17ß-estradiol-supplement results in differential effects in different blood vessel types, and its beneficial vascular effects are masked under the conditions with NO synthase inhibition.

Roles of Toll-like receptor 2/4, monoacylglycerol lipase, and cyclooxygenase in social defeat stress-induced prostaglandin E2 synthesis in the brain and their behavioral relevance.

Inflammation in the brain and periphery has been associated with stress-related pathology of mental illness. We have shown that prostaglandin (PG) E2, an arachidonic acid-derived lipid mediator, and innate immune receptors Toll-like receptor (TLR) 2/4 are crucial for repeated stress-induced behavioral changes in rodents. However, how the stress induces PGE2 synthesis in the brain and whether TLR2/4 are involved in the PGE2 synthesis remain unknown. Using mice lacking TLR2 and TLR4 in combination, here we show that social defeat stress (SDS) induced the PGE2 synthesis in subcortical, but not cortical, tissues in a TLR2/4-dependent manner. It is known that PGE2 in the brain is mainly derived by monoacylglycerol lipase (MAGL)-mediated conversion of endocannabinoid 2-arachidonoylglycerol to free-arachidonic acid, a substrate for cyclooxygenase (COX) for PGE2 synthesis. We found that TLR2/4 deletion reduced the mRNA expression of MAGL and COX1 in subcortical tissues after repeated SDS. Perturbation of MAGL and COX1 as well as COX2 abolished SDS-induced PGE2 synthesis in subcortical tissues. Furthermore, systemic administration of JZL184, an MAGL inhibitor, abolished repeated SDS-induced social avoidance. These results suggest that SDS induces PGE2 synthesis in subcortical regions of the brain via the MAGL-COX pathway in a TLR2/4-dependent manner, thereby leading to social avoidance.

Inhibidores de ciclooxigenasa para impedir la vitreorretinopatía proliferativa: ¿aptos para uso clínico? / Cyclooxygenases inhibitors for preventing the proliferative vitreoretinopathy: Ready for clinical use?

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Periapical bone response to bacterial lipopolysaccharide is shifted upon cyclooxygenase blockage.

OBJECTIVES: Infection, inflammation and bone resorption are closely related events in apical periodontitis development. Therefore, we sought to investigate the role of cyclooxygenase (COX) in osteoclastogenesis and bone metabolism signaling in periapical bone tissue after bacterial lipopolysaccharide (LPS) inoculation into root canals. METHODOLOGY: Seventy two C57BL/6 mice had the root canals of the first molars inoculated with a solution containing LPS from E. coli (1.0 mg/mL) and received selective (celecoxib) or non-selective (indomethacin) COX-2 inhibitor. After 7, 14, 21 and 28 days the animals were euthanized and the tissues removed for total RNA extraction. Evaluation of gene expression was performed by qRT-PCR. Statistical analysis was performed using analysis of variance (ANOVA) followed by post-tests (α=0.05). RESULTS: LPS induced expression of mRNA for COX-2 (Ptgs2) and PGE2 receptors (Ptger1, Ptger3 and Ptger4), indicating that cyclooxygenase is involved in periapical response to LPS. A signaling that favours bone resorption was observed because Tnfsf11 (RANKL), Vegfa, Ctsk, Mmp9, Cd36, Icam, Vcam1, Nfkb1 and Sox9 were upregulated in response to LPS. Indomethacin and celecoxib differentially modulated expression of osteoclastogenic and other bone metabolism genes: celecoxib downregulated Igf1r, Ctsk, Mmp9, Cd36, Icam1, Nfkb1, Smad3, Sox9, Csf3, Vcam1 and Itga3 whereas indomethacin inhibited Tgfbr1, Igf1r, Ctsk, Mmp9, Sox9, Cd36 and Icam1. CONCLUSIONS: We demonstrated that gene expression for COX-2 and PGE2 receptors was upregulated after LPS inoculation into the root canals. Additionally, early administration of indomethacin and celecoxib (NSAIDs) inhibited osteoclastogenic signaling. The relevance of the cyclooxygenase pathway in apical periodontitis was shown by a wide modulation in the expression of genes involved in both bone catabolism and anabolism.

The contribution of beta-2 adrenergic, muscarinic and histamine (H1) receptors, calcium and potassium channels and cyclooxygenase pathway in the relaxant effect of Allium cepa L. on the tracheal smooth muscle.

ETHNOPHARMACOLOGICAL RELEVANCE: There are report regarding therapeutic effects for Allium cepa L. (A. cepa) in Iranian traditional medicine and the plant has showed anti-inflammatory, anti-allergic, anti-hyperglycemic, antioxidant, anti-cancer, anti-hypertension, anti-hypercholesterolemia and anti-asthmatic activities in previous studies. AIM OF THE STUDY: In this study, the contribution of ß2 adrenergic, muscarinic and histamine (H1) receptors, calcium and potassium channels, and cyclooxygenase pathway in the relaxant effect of A. cepa extract on tracheal smooth muscle (TSM) was assessed. MATERIALS AND METHODS: TSM was contracted by KCl (60 mM) or methacholine (10 µM) for 5 min and cumulative concentrations of A. cepa extract (2, 4, 8, 16, 32 and 64 mg/ml) were added to organ bath every 5 min. Theophylline (0.2, 0.4, 0.6 and 0.8 mM) as positive control, and saline (1 ml) as negative control were also examined in non-incubated tissues. The relaxant effect of A. cepa extract was examined on non-incubated and incubated TSM with propranolol, chlorpheniramine, diltiazem, atropine, glibenclamide and indomethacin. RESULTS: A. cepa showed concentration-dependent relaxant effects on non-incubated TSM contracted by KCl (60 mM) or methacholine (10 µM), (P < 0.01 to p < 0.001). There was no significant difference in the relaxant effects of A. cepa between non-incubated and incubated tissues with glibenclamide, atropine, chlorpheniramine and indomethacin. The plant extract showed significant lower relaxant effects in incubated TSM with propranolol and diltiazem compared to non-incubated tissues. EC50 values of A. cepa extract in incubated TSM with propranolol and diltiazem were significantly lower than non-incubated tissues (p < 0.001 and p < 0.05, respectively). The relaxant effects of different concentrations of the extract of A. cepa were not significantly different with those of theophylline. The concentrations of A. cepa extract and theophylline were significant correlated with their relaxant effects (p < 0.05 to p < 0.001). In incubated TSM with propranolol and diltiazem, concentration ratio minus one (CR-1) values was positive (2.65 ±â€¯0.63 and 1.28 ±â€¯0.43 respectively). CONCLUSION: The A. cepa extract showed relatively potent relaxant effect on TSM which was comparable to the effect of theophylline. The results showed that ß2-adrenergic stimulatory and/or calcium channel blockade are the possible mechanisms for the relaxant effects of the plant.

Pharmacological strategies for targeting platelet activation in asthma.

The activation of platelets during host defence and inflammatory disorders has become increasingly documented. Clinical studies of patients with asthma reveal heightened platelet activation and accumulation into lung tissue. Accompanying studies in animal models of allergic lung inflammation, using protocols of experimentally induced thrombocytopenia proclaim an important role for platelets during the leukocyte recruitment cascade, tissue integrity, and lung function. The functions of platelets during these inflammatory events are clearly distinct to platelet functions during haemostasis and clot formation, and have led to the concept that a dichotomy (or polytomy, depending on what else platelets do) in platelet activation exists. The platelet, therefore, presents us with novel opportunities for modulating these inflammatory responses. This review discusses the rationale and effectiveness of current anti-platelet drugs in their use to supress inflammation with regard to asthma, and the need to consider novel possibilities for pharmacological modulation of platelet function associated with inflammation that are pharmacologically distinct to current anti-platelet therapies.

Enzymatically oxidized phospholipids assume center stage as essential regulators of innate immunity and cell death.

Enzymatically oxidized phospholipids (eoxPLs) are formed through regulated processes by which eicosanoids or prostaglandins are attached to phospholipids (PLs) in immune cells. These eoxPLs comprise structurally diverse families of biomolecules with potent bioactivities, and they have important immunoregulatory roles in both health and disease. The formation of oxPLs through enzymatic pathways and their signaling capabilities are emerging concepts. This paradigm is changing our understanding of eicosanoid, prostaglandin, and PL biology in health and disease. eoxPLs have roles in cellular events such as ferroptosis, apoptosis, and blood clotting and diseases such as arthritis, diabetes, and cardiovascular disease. They are increasingly recognized as endogenous bioactive mediators and potential targets for drug development. This review will describe recent evidence that places eoxPLs and their biosynthetic pathways center stage in immunoregulation.

Low-dose anti-inflammatory combinatorial therapy reduced cancer stem cell formation in patient-derived preclinical models for tumour relapse prevention.

BACKGROUND: Emergence of drug-resistant cancer phenotypes is a challenge for anti-cancer therapy. Cancer stem cells are identified as one of the ways by which chemoresistance develops. METHOD: We investigated the anti-inflammatory combinatorial treatment (DA) of doxorubicin and aspirin using a preclinical microfluidic model on cancer cell lines and patient-derived circulating tumour cell clusters. The model had been previously demonstrated to predict patient overall prognosis. RESULTS: We demonstrated that low-dose aspirin with a sub-optimal dose of doxorubicin for 72 h could generate higher killing efficacy and enhanced apoptosis. Seven days of DA treatment significantly reduced the proportion of cancer stem cells and colony-forming ability. DA treatment delayed the inhibition of interleukin-6 secretion, which is mediated by both COX-dependent and independent pathways. The response of patients varied due to clinical heterogeneity, with 62.5% and 64.7% of samples demonstrating higher killing efficacy or reduction in cancer stem cell (CSC) proportions after DA treatment, respectively. These results highlight the importance of using patient-derived models for drug discovery. CONCLUSIONS: This preclinical proof of concept seeks to reduce the onset of CSCs generated post treatment by stressful stimuli. Our study will promote a better understanding of anti-inflammatory treatments for cancer and reduce the risk of relapse in patients.

Periapical bone response to bacterial lipopolysaccharide is shifted upon cyclooxygenase blockage

Abstract Objectives: Infection, inflammation and bone resorption are closely related events in apical periodontitis development. Therefore, we sought to investigate the role of cyclooxygenase (COX) in osteoclastogenesis and bone metabolism signaling in periapical bone tissue after bacterial lipopolysaccharide (LPS) inoculation into root canals. Methodology: Seventy two C57BL/6 mice had the root canals of the first molars inoculated with a solution containing LPS from E. coli (1.0 mg/mL) and received selective (celecoxib) or non-selective (indomethacin) COX-2 inhibitor. After 7, 14, 21 and 28 days the animals were euthanized and the tissues removed for total RNA extraction. Evaluation of gene expression was performed by qRT-PCR. Statistical analysis was performed using analysis of variance (ANOVA) followed by post-tests (α=0.05). Results: LPS induced expression of mRNA for COX-2 (Ptgs2) and PGE2 receptors (Ptger1, Ptger3 and Ptger4), indicating that cyclooxygenase is involved in periapical response to LPS. A signaling that favours bone resorption was observed because Tnfsf11 (RANKL), Vegfa, Ctsk, Mmp9, Cd36, Icam, Vcam1, Nfkb1 and Sox9 were upregulated in response to LPS. Indomethacin and celecoxib differentially modulated expression of osteoclastogenic and other bone metabolism genes: celecoxib downregulated Igf1r, Ctsk, Mmp9, Cd36, Icam1, Nfkb1, Smad3, Sox9, Csf3, Vcam1 and Itga3 whereas indomethacin inhibited Tgfbr1, Igf1r, Ctsk, Mmp9, Sox9, Cd36 and Icam1. Conclusions: We demonstrated that gene expression for COX-2 and PGE2 receptors was upregulated after LPS inoculation into the root canals. Additionally, early administration of indomethacin and celecoxib (NSAIDs) inhibited osteoclastogenic signaling. The relevance of the cyclooxygenase pathway in apical periodontitis was shown by a wide modulation in the expression of genes involved in both bone catabolism and anabolism.

Prostacyclins in Cardiac Surgery: Coming of Age.

Prostacyclin (prostaglandin I2 [PGI2]) is an eicosanoid lipid mediator produced by the endothelial cells. It plays pivotal roles in vascular homeostasis by virtue of its potent vasodilatory and antithrombotic effects. Stable pharmacological analogues of PGI2 are used for treatment of pulmonary hypertension and right ventricular failure. PGI2 dose dependently inhibits platelet activation induced by adenosine-5'-diphosphate, arachidonic acid, collagen, and low-dose thrombin. This property has led to its use as an alternative to direct thrombin inhibitors in patients with type II heparin-induced thrombocytopenia (HIT) undergoing cardiac surgery. The aims of this review are the following: (1) to review the pharmacology of PGI2 and its derivatives, (2) to present the evidence for their use in pulmonary hypertension and right heart failure, and (3) to discuss their utility in the management of HIT in cardiac surgery.

Direct Involvement of Arachidonic Acid in the Development of Ear Edema via TRPV3.

Arachidonic acid (AA) plays a pivotal role in the development of edema via its oxidized metabolites derived from cyclooxygenase (COX) and lipoxygenase (LOX), and is recently recognized as an activator of TRPV3. However, it is not clear whether AA plays some TRPV3-mediated pathological roles in the development of edema. Pharmacological and histological studies using ICRTRPV3+/+ and ICRTRPV3-/- mice indicated that higher ear edema responses to topical application of AA were observed in ICRTRPV3+/+ mice compared with ICRTRPV3-/- mice. However, there was no difference in the ear edema response to 12-O-tetradecanoylphorbol 13-acetate, skin histology, and skin barrier function between these mouse strains. Furthermore, oxidized fatty acids from the lesional site were analyzed to elucidate the TRPV3-mediated pathological roles of AA, and the results revealed that there were no differences in the level of COX or LOX metabolites derived from AA between both mouse strains. We concluded that AA plays a role in the development of TRPV3-mediated ear edema and that this result may contribute to better understanding of the pathophysiological mechanisms involved in the development of a certain type of edema.

Replicative senescence promotes prothrombotic responses in endothelial cells: Role of NADPH oxidase- and cyclooxygenase-derived oxidative stress.

Endothelial senescence has been suggested to promote endothelial dysfunction in age-related vascular disorders. This study evaluated the prothrombotic properties of senescent endothelial cells (ECs) and the underlying mechanism. Serial passaging from passage (P)1 to P4 (replicative senescence) of porcine coronary artery ECs, or treatment of P1 ECs with the endothelial nitric oxide synthase (eNOS) inhibitor L-NAME (premature senescence) induced acquisition of markers of senescence including increased senescence-associated-ß-galactosidase (SA-ß-gal) activity and p53, p21, p16 expression. Approximately 55% of P3 cells were senescent with a high level oxidative stress, and decreased eNOS-derived nitric oxide (NO) formation associated with increased expression of NADPH oxidase subunits (gp91phox, p47phox), cyclooxygenase (COX)-2 but not COX-1, and a decreased eNOS expression leading to a reduced ability of ECs to inhibit platelet aggregation. P3 cells also presented increased expression and activity of tissue factor (TF), a key initiator of the coagulation cascade. Treatment of senesecent cells with a NADPH oxidase inhibitor (VAS-2870) or by a COX inhibitor (indomethacin) reduced oxidative stress, decreased TF activity and expression, and reduced the expression of gp91phox, p47phox and COX-2 and restored the ability of ECs to inhibit effectively platelet aggregation. Thus, replicative endothelial senescence promotes a prothrombotic response involving the down-regulation of the protective NO pathway and the upregulation of the NADPH oxidase- and COXs-dependent oxidative stress pathway promoting TF expression and activity.

Distinct Signature of Oxylipid Mediators of Inflammation during Infection and Asymptomatic Colonization by E. coli in the Urinary Bladder.

Urinary tract infection (UTI) is an extremely common infectious disease. Uropathogenic Escherichia coli (UPEC) is the predominant etiological agent of UTI. Asymptomatic bacteriuric E. coli (ABEC) strains successfully colonize the urinary tract resulting in asymptomatic bacteriuria (ABU) and do not induce symptoms associated with UTI. Oxylipids are key signaling molecules involved in inflammation. Based on the distinct clinical outcomes of E. coli colonization, we hypothesized that UPEC triggers the production of predominantly proinflammatory oxylipids and ABEC leads to production of primarily anti-inflammatory or proresolving oxylipids in the urinary tract. We performed quantitative detection of 39 oxylipid mediators with proinflammatory, anti-inflammatory, and proresolving properties, during UTI and ABU caused by genetically distinct E. coli strains in the murine urinary bladder. Our results reveal that infection with UPEC causes an increased accumulation of proinflammatory oxylipids as early as 6 h postinoculation, compared to controls. To the contrary, ABEC colonization leads to decreased accumulation of proinflammatory oxylipids at the early time point compared to UPEC infection but does not affect the level of proresolving oxylipids. This report represents the first comprehensive investigation on the oxylipidome during benign ABEC colonization observed in ABU and acute inflammation triggered by UPEC leading to UTI.

[Gene regulation of prostaglandin synthase and prostate diseases].

Prostaglandin synthase (PGS) can catalyze the production of various types of prostaglandins and regulate the expression levels of related substances. The regulation mechanisms of the PGS gene are closely related with the occurrence and development of prostate diseases. However, few studies are reported on the regulation mechanisms of PGS in prostatic diseases, such as benign prostatic hyperplasia (BPH) and prostate cancer (PCa), or on the relationship between PGS gene regulation and prostate diseases. This review aims to analyze their correlation and provide some ideas for the prevention and control of BPH and PCa by intervention of the prostaglandin synthase regulatory pathway.

Vascular Endothelium-Dependent and Independent Actions of Oleanolic Acid and Its Synthetic Oleanane Derivatives as Possible Mechanisms for Hypotensive Effects.

PURPOSE: Plant-derived oleanolic acid (OA) and its related synthetic derivatives (Br-OA and Me-OA) possess antihypertensive effects in experimental animals. The present study investigated possible underlying mechanisms in rat isolated single ventricular myocytes and in vascular smooth muscles superfused at 37°C. METHODS: Cell shortening was assessed at 1 Hz using a video-based edge-detection system and the L-type Ca2+ current (ICaL) was measured using the whole-cell patch-clamp technique in single ventricular myocytes. Isometric tension was measured using force transducer in isolated aortic rings and in mesenteric arteries. Vascular effects were measured in endothelium-intact and denuded vessels in the presence of various enzyme or channel inhibitors. RESULTS: OA and its derivatives increased cell shortening in cardiomyocytes isolated from normotensive rats but had no effect in those isolated from hypertensive animals. These triterpenes also caused relaxation in aortic rings and in mesenteric arteries pre-contracted with either phenylephrine or KCl-enriched solution. The relaxation was only partially inhibited by endothelium denudation, and also partly inhibited by the cyclooxygenase (COX) inhibitor indomethacin, with no additional inhibitory effect of the NO synthase inhibitor, N-ω-Nitro-L-arginine. A combination of both ATP-dependent channel inhibition by glibenclaminde and voltage-dependent K+ channel inhibition by 4-aminopyridine was necessary to fully inhibit the relaxation. CONCLUSION: These data indicate that the effects of OA and its derivatives are mediated via both endothelium-dependent and independent mechanisms suggesting the involvement of COX in the endothelium-dependent effects and of vascular muscle K+ channels in the endothelium-independent effects. Finally, our results support the view that the antihypertensive action of OA and its derivatives is due to a decrease of vascular resistance with no negative inotropic effect on the heart.

ß-Adrenergic-mediated vasodilation in young men and women: cyclooxygenase restrains nitric oxide synthase.

We tested the hypothesis that women exhibit greater vasodilator responses to ß-adrenoceptor stimulation compared with men. We further hypothesized women exhibit a greater contribution of nitric oxide synthase and cyclooxygenase to ß-adrenergic-mediated vasodilation compared with men. Forearm blood flow (Doppler ultrasound) was measured in young men (n = 29, 26 ± 1 yr) and women (n = 33, 25 ± 1 yr) during intra-arterial infusion of isoproterenol (ß-adrenergic agonist). In subset of subjects, isoproterenol responses were examined before and after local inhibition of nitric oxide synthase [N(G)-monomethyl-l-arginine (l-NMMA); 6 male/10 female] and/or cyclooxygenase (ketorolac; 5 male/5 female). Vascular conductance (blood flow ÷ mean arterial pressure) was calculated to assess vasodilation. Vascular conductance increased with isoproterenol infusion (P < 0.01), and this effect was not different between men and women (P = 0.41). l-NMMA infusion had no effect on isoproterenol-mediated dilation in men (P > 0.99) or women (P = 0.21). In contrast, ketorolac infusion markedly increased isoproterenol-mediated responses in both men (P < 0.01) and women (P = 0.04) and this rise was lost with subsequent l-NMMA infusion (men, P < 0.01; women, P < 0.05). ß-Adrenergic vasodilation is not different between men and women and sex differences in the independent contribution of nitric oxide synthase and cyclooxygenase to ß-mediated vasodilation are not present. However, these data are the first to demonstrate ß-adrenoceptor activation of cyclooxygenase suppresses nitric oxide synthase signaling in human forearm microcirculation and may have important implications for neurovascular control in both health and disease.

Lipopolysaccharide-induced blood-brain barrier disruption: roles of cyclooxygenase, oxidative stress, neuroinflammation, and elements of the neurovascular unit.

BACKGROUND: Disruption of the blood-brain barrier (BBB) occurs in many diseases and is often mediated by inflammatory and neuroimmune mechanisms. Inflammation is well established as a cause of BBB disruption, but many mechanistic questions remain. METHODS: We used lipopolysaccharide (LPS) to induce inflammation and BBB disruption in mice. BBB disruption was measured using (14)C-sucrose and radioactively labeled albumin. Brain cytokine responses were measured using multiplex technology and dependence on cyclooxygenase (COX) and oxidative stress determined by treatments with indomethacin and N-acetylcysteine. Astrocyte and microglia/macrophage responses were measured using brain immunohistochemistry. In vitro studies used Transwell cultures of primary brain endothelial cells co- or tri-cultured with astrocytes and pericytes to measure effects of LPS on transendothelial electrical resistance (TEER), cellular distribution of tight junction proteins, and permeability to (14)C-sucrose and radioactive albumin. RESULTS: In comparison to LPS-induced weight loss, the BBB was relatively resistant to LPS-induced disruption. Disruption occurred only with the highest dose of LPS and was most evident in the frontal cortex, thalamus, pons-medulla, and cerebellum with no disruption in the hypothalamus. The in vitro and in vivo patterns of LPS-induced disruption as measured with (14)C-sucrose, radioactive albumin, and TEER suggested involvement of both paracellular and transcytotic pathways. Disruption as measured with albumin and (14)C-sucrose, but not TEER, was blocked by indomethacin. N-acetylcysteine did not affect disruption. In vivo, the measures of neuroinflammation induced by LPS were mainly not reversed by indomethacin. In vitro, the effects on LPS and indomethacin were not altered when brain endothelial cells (BECs) were cultured with astrocytes or pericytes. CONCLUSIONS: The BBB is relatively resistant to LPS-induced disruption with some brain regions more vulnerable than others. LPS-induced disruption appears is to be dependent on COX but not on oxidative stress. Based on in vivo and in vitro measures of neuroinflammation, it appears that astrocytes, microglia/macrophages, and pericytes play little role in the LPS-mediated disruption of the BBB.