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1.
Clin Biochem ; 74: 54-59, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31669512

RESUMO

BACKGROUND: In order to manage risks of bleeding and thrombosis after some surgical procedures, platelet function is often measured repeatedly over days or weeks using laboratory tests of platelet function. To interpret test results in the perioperative period, it is necessary to understand analytical, biological and between-person variation. METHODS: We collected three separate blood specimens from 16 healthy volunteers on the first study day, and one additional specimen from each volunteer 1, 2, and 3 months later. Arachidonic acid-induced and adenosine diphosphate (ADP)-induced platelet function were measured in duplicate by whole blood impedance aggregometry using Multiplate (ASPI/ADP tests) and VerifyNow (Aspirin Reaction Units [ARU] and P2Y12 Reaction Units [PRU]). The analytical variation (CVA), within-subject variation (CVI), between-subject variation (CVG), index of individuality (II), and reference change values (RCV) were calculated. RESULTS: VerifyNow ARU demonstrated the smallest short-term and long-term variability (CVA, CVI, and CVG ~1%), resulting in short- and long-term RCV values <5%. II was also higher (1.92) for VerifyNow ARU than other platelet function tests. Multiplate ASPI and ADP tests had the highest RCV both short-(19.0% and 25.2%, respectively) and long-term (32.1% and 39.6%, respectively) due to increased CVA (>5%) and CVI (3.9-13.1%). VerifyNow PRU had a lower RCV than Multiplate ADP; but was the only test with II <0.6. CONCLUSIONS: VerifyNow ARU results can be interpreted relative to a fixed cut-off or population-based reference interval; or relative to small changes in an individual's previous values. VerifyNow PRU and Multiplate ASPI and ADP tests should only be interpreted based upon relative change; and can only distinguish relatively large (>23%) changes over several weeks.


Assuntos
Variação Biológica da População/fisiologia , Testes de Função Plaquetária , Difosfato de Adenosina/farmacologia , Ácido Araquidônico/farmacologia , Aspirina/farmacologia , Feminino , Seguimentos , Hemorragia/prevenção & controle , Humanos , Masculino , Distribuição Normal , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação de Plaquetas/farmacologia , Valores de Referência , Trombose/prevenção & controle
2.
Cells ; 8(7)2019 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-31323976

RESUMO

It has long been known that the conditionally essential polyunsaturated arachidonic acid (AA) regulates cerebral blood flow (CBF) through its metabolites prostaglandin E2 and epoxyeicosatrienoic acid, which act on vascular smooth muscle cells and pericytes to vasorelax cerebral microvessels. However, AA may also elicit endothelial nitric oxide (NO) release through an increase in intracellular Ca2+ concentration ([Ca2+]i). Herein, we adopted Ca2+ and NO imaging, combined with immunoblotting, to assess whether AA induces intracellular Ca2+ signals and NO release in the human brain microvascular endothelial cell line hCMEC/D3. AA caused a dose-dependent increase in [Ca2+]i that was mimicked by the not-metabolizable analogue, eicosatetraynoic acid. The Ca2+ response to AA was patterned by endoplasmic reticulum Ca2+ release through type 3 inositol-1,4,5-trisphosphate receptors, lysosomal Ca2+ mobilization through two-pore channels 1 and 2 (TPC1-2), and extracellular Ca2+ influx through transient receptor potential vanilloid 4 (TRPV4). In addition, AA-evoked Ca2+ signals resulted in robust NO release, but this signal was considerably delayed as compared to the accompanying Ca2+ wave and was essentially mediated by TPC1-2 and TRPV4. Overall, these data provide the first evidence that AA elicits Ca2+-dependent NO release from a human cerebrovascular endothelial cell line, but they seemingly rule out the possibility that this NO signal could acutely modulate neurovascular coupling.


Assuntos
Ácido Araquidônico/farmacologia , Sinalização do Cálcio , Cálcio/metabolismo , Células Endoteliais/efeitos dos fármacos , Óxido Nítrico/metabolismo , Encéfalo/irrigação sanguínea , Canais de Cálcio/metabolismo , Células Cultivadas , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Humanos , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Microcirculação , Canais de Cátion TRPV/metabolismo
3.
Biomed Pharmacother ; 116: 108836, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31004838

RESUMO

BACKGROUND: An impaired trophoblast invasion ability contributes to the development of pre-eclampsia (PE), and can be induced by the altered expression of various microRNAs (miRs). MiR-141 and CXCL12ß (C-X-C motif chemokine ligand 12) signaling regulate trophoblast invasion and vascularization capabilities during PE pathogenesis; however, their interactions and underlying mechanisms of action remain unclear. We investigated how miR-141 modulates trophoblast invasion, with a focus on its interaction with CXCL12ß signaling. METHODS: A PE model was established by using HTR-8/SVneo cells, which were first cultured with 2% O2 for 48 h, and then with 5% O2. The expression of miR-141 in human villous trophoblast HTR-8/SVneo cells was modulated with mimics or an inhibitor, and analyzed by quantitative RT-PCR. CXCL12ß levels were determined by ELISA. Cell apoptosis was determined by flow cytometry, and the invasion and vascularization capabilities of trophoblasts were evaluated by Transwell and tube formation assays, respectively. Binding of miR-141 with CXCL12ß mRNA was verified by the dual luciferase assay. Protein levels were estimated by western blotting. RESULTS: MiR-141 expression was significantly induced by hypoxia in HTR-8/SVneo cells. MiR-141 was found to promote apoptosis and inhibit the invasion and vascularization abilities of HTR-8/SVneo cells under conditions of hypoxia. MiR-141 could directly bind with the 3'UTR region of CXCL12ß mRNA and inhibit its translation. In addition, we proved that miR-141 could inhibit the invasion and vascularization abilities, and promote the apoptosis of HTR-8/SVneo cells by targeting CXCL12ß under hypoxic conditions. Furthermore, we demonstrated that arachidonic acid could reverse the invasion and apoptosis abilities of HTR-8/SVneo cells mediated by CXCL12ß during hypoxia. In terms of mechanism, MiR-141 could downregulate MMP2, p62, and LC3B expression, and upregulate ROCK1 and RhoA expression in HTR-8/SVneo cells by targeting the CXCL12ß gene during hypoxia. The effects of CXCL12ßon HTR-8/SVneo cells could be reversed by arachidonic acid (ARA). CONCLUSION: Induction of miR-141 by hypoxia promotes apoptosis, and inhibits the invasion and vascularization capabilities of HTR-8/SVneo cells by suppressing the CXCL12ß and CXCR2/4 signaling pathways.


Assuntos
Apoptose , Quimiocina CXCL12/metabolismo , MicroRNAs/metabolismo , Neovascularização Fisiológica , Receptores CXCR4/metabolismo , Receptores de Interleucina-8B/metabolismo , Transdução de Sinais , Trofoblastos/patologia , Apoptose/efeitos dos fármacos , Ácido Araquidônico/farmacologia , Sequência de Bases , Hipóxia Celular/efeitos dos fármacos , Hipóxia Celular/genética , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , MicroRNAs/genética , Modelos Biológicos , Neovascularização Fisiológica/efeitos dos fármacos , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo
4.
Molecules ; 24(8)2019 Apr 13.
Artigo em Inglês | MEDLINE | ID: mdl-31013947

RESUMO

Many Premna species have been used in traditional medicine to treat hypertension and cardiac insufficiency, and as a tonic for cardiac-related problems. Some have been reported to possess cardiovascular protective activity through several possible mechanisms, but not Premna foetida. In the present study, the methanol extract of P. foetida leaves (PFM) and its isolated compounds were evaluated for their ability to inhibit copper-mediated human low-density lipoprotein (LDL) oxidation and arachidonic acid (AA)- and adenosine diphosphate (ADP)-induced platelet aggregation. Six flavonoids, three triterpenoids, vanillic acid and stigmasterol were successfully isolated from PFM. Of the isolated compounds, quercetin was the most active against LDL oxidation (IC50 4.25 µM). The flavonols were more active than the flavones against LDL oxidation, suggesting that hydroxyl group at C-3 and the catechol moiety at B-ring may play important roles in protecting LDL from oxidation. Most tested flavonoids showed stronger inhibition towards AA-induced than the ADP-induced platelet aggregation with apigenin exhibiting the strongest effect (IC50 52.3 and 127.4 µM, respectively) while quercetin and kaempferol showed moderate activity. The results suggested that flavonoids, especially quercetin, apigenin and kaempferol were among the major constituents of P. foetida responsible for anti-LDL oxidation and anti-platelet aggregation.


Assuntos
Plaquetas/metabolismo , Flavonoides , Lamiaceae/química , Lipoproteínas LDL/metabolismo , Extratos Vegetais , Folhas de Planta/química , Inibidores da Agregação de Plaquetas , Agregação Plaquetária/efeitos dos fármacos , Difosfato de Adenosina/farmacologia , Ácido Araquidônico/farmacologia , Plaquetas/citologia , Flavonoides/química , Flavonoides/farmacologia , Humanos , Lipoproteínas LDL/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Inibidores da Agregação de Plaquetas/química , Inibidores da Agregação de Plaquetas/farmacologia
5.
Vet Clin Pathol ; 48(1): 11-18, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30924545

RESUMO

BACKGROUND: Dogs with Babesia rossi infection display a normocoagulable thromboelastogram, despite being markedly thrombocytopenic, which is purportedly due to large-scale platelet activation. Thromboelastographic platelet mapping (TEG-PM) evaluates individual contributions of thrombin, fibrinogen, and platelets to clot formation, and may elucidate some of the pathomechanisms of thrombocytopenia-associated hemostatic alterations. OBJECTIVE: This study investigated potential differences in TEG-PM variables in dogs with complicated B rossi infection compared with healthy controls, and whether these variables correlated with platelet activation indices. METHODS: The maximum amplitude (MA) following thrombin generation (MAThrombin ) was determined using kaolin-activated TEG. The TEG-PM variables included MA following the addition of platelet agonists arachidonic acid (MAAA ) and adenosine diphosphate (MAADP ), and MA due to fibrin alone (MAFibrin ). In addition, platelet indices and fibrinogen concentrations were determined. RESULTS: Thirteen dogs with complicated B rossi infection and five healthy controls were included. The median MAFibrin and fibrinogen concentrations were significantly higher (P < 0.01 for both) and median platelet count was significantly lower (P < 0.01) in the babesiosis group vs the control group. No significant differences were found for MAThrombin and MAAA/ADP . maximum amplitude due to fibrin alone was positively correlated with fibrinogen concentration (r = 0.735), mean platelet volume (r = 0.517), and mean platelet mass (r = 0.498), and negatively correlated with hematocrit (r = -0.685), platelet count (r = -0.476), and plateletcrit (r = -0.479) (P < 0.05 for all). CONCLUSIONS: This study suggests that the presence of hyperfibrinogenemia offsets the severe thrombocytopenia associated with B rossi to result in normal thromboelastograms and lack of overt clinical bleeding.


Assuntos
Babesia , Babesiose/sangue , Plaquetas/fisiologia , Doenças do Cão/sangue , Tromboelastografia/veterinária , Animais , Apirase/farmacologia , Ácido Araquidônico/farmacologia , Plaquetas/química , Plaquetas/efeitos dos fármacos , Estudos de Casos e Controles , Doenças do Cão/parasitologia , Cães , Feminino , Fibrinogênio/análise , Masculino
6.
Artigo em Inglês | MEDLINE | ID: mdl-30826550

RESUMO

Overdevelopment of adipose tissue in cultured fish is one of the biggest issues plaguing current aquaculture industry, leading to unhealthy status of fishes and production losses. Diet supplemented with 0.30% arachidonic acid (ARA) has been found to reduce adipogenesis and inflammation in grass carp, but the potential mechanism is not comprehensively understood. To fully reveal the effects of dietary ARA on the mRNA profiles of adipose tissue, transcriptome techniques were applied in this study. A 10-weeks feeding experiment was performed using two isonitrogenous and isoenergetic purified diets, namely ARA-free (control) and 0.30% ARA (ARA group). Results showed increased ARA content and decreased intraperitoneal fat index and adipocyte size in the adipose tissue of fish fed ARA (P < 0.05). A total of 611 and 973 genes of the adipose tissue were significantly up-regulated and down-regulated, respectively, in fish fed ARA (P < 0.05). Dietary ARA upregulated LOX pathway but downregulated CYP450 pathway annotated genes expression. A total of 65 cell development annotated genes including 30 adipocyte proliferation, 21 adipocyte differentiation, and 14 cell apoptosis annotated genes were down-regulated in the ARA group. In addition, 19 lipid catabolism annotated genes were increased. The mRNA expression levels of 5 chemokines, 10 cytokines, 26 cytokine and chemokine receptors, 15 cell adhesion, 6 oxidative stress, and 6 angiogenesis annotated genes were all down-regulated in fish fed ARA. Finally, dietary ARA also decreased the expression of transcripts annotated with glucose transportation, glycolysis and gluconeogenesis. Overall, our results demonstrate that dietary ARA has a fat reducing role, and tends to retard adipocyte development and attenuate chronic inflammation based on these adipose transcript expression results in grass carp.


Assuntos
Ração Animal , Aquicultura , Ácido Araquidônico/farmacologia , Carpas/fisiologia , Alimento Funcional , Adipogenia/efeitos dos fármacos , Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/fisiologia , Ração Animal/análise , Animais , Carpas/genética , Alimento Funcional/análise , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/genética , Inflamação/prevenção & controle , Inflamação/veterinária , Lipólise/efeitos dos fármacos
7.
Food Chem ; 288: 256-261, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-30902290

RESUMO

The effect of elicitation with arachidonic and jasmonic acids on the production of phenolic compounds as well as the antioxidant and anti-inflammatory properties of phenolic extracts of wheatgrass was evaluated. The qualitative and quantitative analysis of phenolic compounds carried out with the UPLC-MS technique indicated that luteolin and apigenin derivatives were the dominant flavonoids, while ferulic acid derivatives and syringic acid were the main components in the phenolic acid fraction in the wheatgrass. No qualitative changes in the examined phenolic compounds were observed in the case of the control and elicited plants, while there was an increase in the content of some compounds. The antioxidant activity increased in the elicited samples (with the exception of reducing power) and this elevation was partially correlated with the increase in the polyphenol content in the studied plants. Elicitation with 0.01 µM arachidonic acid also caused improvement of potential anti-inflammatory properties of the wheatgrass.


Assuntos
Anti-Inflamatórios/química , Antioxidantes/química , Ácido Araquidônico/farmacologia , Ciclopentanos/farmacologia , Oxilipinas/farmacologia , Fenóis/química , Triticum/efeitos dos fármacos , Anti-Inflamatórios/metabolismo , Flavonoides/análise , Lipoxigenase/química , Lipoxigenase/metabolismo , Fenóis/análise , Fenóis/metabolismo , Extratos Vegetais/química , Polifenóis/análise , Sementes/química , Sementes/efeitos dos fármacos , Sementes/crescimento & desenvolvimento , Triticum/química , Triticum/crescimento & desenvolvimento
8.
Int J Mol Sci ; 20(4)2019 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-30813326

RESUMO

OBJECTIVES: Obesity is a complex disease that has a strong association with diet and lifestyle. Dietary factors can influence the expression of key genes connected to insulin resistance, lipid metabolism, and adipose tissue composition. In this study, our objective was to determine gene expression and fatty acid (FA) profiles in visceral adipose tissue (VAT) from lean and morbidly obese individuals. We also aimed to study the agonist effect of dietary factors on glucose metabolism. DESIGN AND METHODS: Lean and low and high insulin resistance morbidly obese subjects (LIR-MO and HIR-MO) were included in this study. The gene expression of liver X receptor type alpha (LXR-α) and glucose transporter type 4 (GLUT4) and the FA profiles in VAT were determined. Additionally, the in vivo and in vitro agonist effects of oleic acid (OA), linoleic acid (LA), and arachidonic acid (AA) by peroxisome proliferator-activated receptor type gamma 2 (PPAR-γ2) on the activity of GLUT4 were studied. RESULTS: Our results showed a dysregulation of GLUT4 and LXR-α in VAT of morbidly obese subjects. In addition, a specific FA profile for morbidly obese individuals was found. Finally, AA was an PPAR-γ2 agonist that activates the expression of GLUT4. CONCLUSIONS: Our study suggests a dysregulation of LXR-α and GLUT4 expression in VAT of morbidly obese individuals. FA profiles in VAT could elucidate their possible role in lipolysis and adipogenesis. Finally, AA binds to PPAR-γ2 to activate the expression of GLUT4 in the HepG2 cell line, showing an alternative insulin-independent activation of GLUT4.


Assuntos
Ácido Araquidônico/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Transportador de Glucose Tipo 4/genética , PPAR gama/metabolismo , Adulto , DNA/metabolismo , Regulação para Baixo/genética , Feminino , Transportador de Glucose Tipo 4/metabolismo , Células Hep G2 , Humanos , Resistência à Insulina , Gordura Intra-Abdominal/metabolismo , Ligantes , Receptores X do Fígado/genética , Receptores X do Fígado/metabolismo , Masculino , Obesidade Mórbida/genética , PPAR gama/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Magreza/genética
9.
Domest Anim Endocrinol ; 67: 28-36, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30677541

RESUMO

Many studies have shown positive effects of prostaglandins (PGs) on various steps of skeletal muscle formation such as myoblast proliferation and myotube hypertrophy. In animals, PGs are synthesized through the action of the rate-limiting enzymes cyclooxygenase (COX) -1 and COX-2 from arachidonic acid (AA), a conditionally essential fatty acid. As a step toward exploring the possibility of using dietary supplementation of AA to improve skeletal muscle growth in cattle, which are major meat-producing animals, we determined the effects of AA and its major PG derivatives PGE2, PGF2α, and PGI2 on proliferation, differentiation, and fusion of primary bovine myoblasts in vitro. In the proliferation experiment, myoblasts were cultured in a growth medium to which was added 10 µM AA, 1 µM PGE2, 1 µM PGF2α, 1 µM PGI2, or vehicle control for 24 h, and the proliferating cells were identified by 5-ethynyl-2'-deoxyuridine (EdU) labeling. This experiment revealed that AA, PGE2, PGF2α, and PGI2 each increased the number of proliferating cells by 13%, 24%, 16%, and 16%, respectively, compared to the control (n = 7, P < 0.05). In the differentiation and fusion test, myoblasts were induced to differentiate and fuse into myotubes in the presence of the aforementioned treatments for 0, 24, 48, and 72 h. Based on quantitative reverse transcription PCR analyses of mRNAs of myoblast differentiation and fusion markers (myogenin; myosin heavy chain 3; creatine kinase, muscle; myomaker) at 0, 24, and 48 h of differentiation, AA, PGE2, and PGF2α promoted myoblast differentiation (n = 6, P < 0.05). Based on Giemsa staining and counting the number of myotubes at 72 h of differentiation, PGE2 enhanced the number of formed myotubes by 14% (P < 0.05) compared to the control. Treating the myoblasts with AA and either the COX-1 and COX-2 common inhibitor indomethacin or the COX-2-specific inhibitor NS-398 reversed the stimulatory effect of AA on myoblast proliferation (n = 4, P < 0.05). Overall, this study demonstrates that exogenous AA stimulates bovine myoblast proliferation and differentiation in culture. The results of this study suggest that AA stimulates myoblast proliferation through its metabolites PGE2, PGF2α, or PGI2, and that AA stimulates myoblast differentiation through PGE2.


Assuntos
Ácido Araquidônico/farmacologia , Bovinos/fisiologia , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Mioblastos/citologia , Prostaglandinas/farmacologia , Animais , Fusão Celular , Células Cultivadas , Meios de Cultura , Dinoprosta/farmacologia , Dinoprostona/farmacologia , Epoprostenol/farmacologia , Masculino , Fibras Musculares Esqueléticas/citologia , Músculo Esquelético/crescimento & desenvolvimento , Mioblastos/efeitos dos fármacos
10.
Cell Chem Biol ; 26(3): 420-432.e9, 2019 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-30686757

RESUMO

The initiation and execution of cell death can be regulated by various lipids. How the levels of environmental (exogenous) lipids impact cell death sensitivity is not well understood. We find that exogenous monounsaturated fatty acids (MUFAs) potently inhibit the non-apoptotic, iron-dependent, oxidative cell death process of ferroptosis. This protective effect is associated with the suppression of lipid reactive oxygen species (ROS) accumulation at the plasma membrane and decreased levels of phospholipids containing oxidizable polyunsaturated fatty acids. Treatment with exogenous MUFAs reduces the sensitivity of plasma membrane lipids to oxidation over several hours. This effect requires MUFA activation by acyl-coenzyme A synthetase long-chain family member 3 (ACSL3) and is independent of lipid droplet formation. Exogenous MUFAs also protect cells from apoptotic lipotoxicity caused by the accumulation of saturated fatty acids, but in an ACSL3-independent manner. Our work demonstrates that ACSL3-dependent MUFA activation promotes a ferroptosis-resistant cell state.


Assuntos
Ácidos Graxos Monoinsaturados/farmacologia , Lipídeos/química , Animais , Ácido Araquidônico/química , Ácido Araquidônico/metabolismo , Ácido Araquidônico/farmacologia , Linhagem Celular , Membrana Celular/química , Membrana Celular/metabolismo , Coenzima A Ligases/metabolismo , Ácidos Graxos Monoinsaturados/química , Ácidos Graxos Monoinsaturados/metabolismo , Gotículas Lipídicas/química , Gotículas Lipídicas/metabolismo , Camundongos , Oxirredução , /metabolismo , Espécies Reativas de Oxigênio/química , Espécies Reativas de Oxigênio/metabolismo
11.
Channels (Austin) ; 13(1): 36-47, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-30661462

RESUMO

The current knowledge of electrogenesis in mesenchymal stromal cells (MSCs) remains scarce. Earlier, we demonstrated that in MSCs from the human adipose tissue, transduction of certain agonists involved the phosphoinositide cascade. Its pivotal effector PLC generates DAG that can regulate ion channels directly or via its derivatives, including arachidonic acid (AA). Here we showed that AA strongly hyperpolarized MSCs by stimulating instantly activating, outwardly rectifying TEA-insensitive K+ channels. Among AA-regulated K+ channels, K2P channels from the TREK subfamily appeared to be an appropriate target. The expression of K2P channels in MSCs was verified by RT-PCR, which revealed TWIK-1, TREK-1, and TASK-5 transcripts. The TREK-1 inhibitor spadin antagonized the electrogenic action of AA, which was simulated by the channel activator BL 1249. This functional evidence suggested that TREK-1 channels mediated AA-dependent hyperpolarization of MSCs. Being mostly silent at rest, TREK-1 negligibly contributed to the "background" K+ current. The dramatic stimulation of TREK-1 channels by AA indicates their involvement in AA-dependent signaling in MSCs.


Assuntos
Tecido Adiposo/efeitos dos fármacos , Tecido Adiposo/metabolismo , Ácido Araquidônico/farmacologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Canais de Potássio de Domínios Poros em Tandem/agonistas , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Tecido Adiposo/citologia , Adulto , Células Cultivadas , Relação Dose-Resposta a Droga , Humanos , Masculino , Células-Tronco Mesenquimais/metabolismo , Pessoa de Meia-Idade , Peptídeos/farmacologia , Relação Estrutura-Atividade
12.
Am J Respir Cell Mol Biol ; 60(5): 554-568, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30648905

RESUMO

Obesity is an important risk factor for severe asthma exacerbations, which are mainly caused by respiratory infections. Dietary fatty acids, which are increased systemically in obese patients and are further increased after high-fat meals, affect the innate immune system and may contribute to dysfunctional immune responses to respiratory infection. In this study we investigated the effects of dietary fatty acids on immune responses to respiratory infection in pulmonary fibroblasts and a bronchial epithelial cell line (BEAS-2B). Cells were challenged with BSA-conjugated fatty acids (ω-6 polyunsaturated fatty acids [PUFAs], ω-3 PUFAs, or saturated fatty acids [SFAs]) +/- the viral mimic polyinosinic:polycytidylic acid (poly[I:C]) or bacterial compound lipoteichoic acid (LTA), and release of proinflammatory cytokines was measured. In both cell types, challenge with arachidonic acid (AA) (ω-6 PUFA) and poly(I:C) or LTA led to substantially greater IL-6 and CXCL8 release than either challenge alone, demonstrating synergy. In epithelial cells, palmitic acid (SFA) combined with poly(I:C) also led to greater IL-6 release. The underlying signaling pathways of AA and poly(I:C)- or LTA-induced cytokine release were examined using specific signaling inhibitors and IB. Cytokine production in pulmonary fibroblasts was prostaglandin dependent, and synergistic upregulation occurred via p38 mitogen-activated protein kinase signaling, whereas cytokine production in bronchial epithelial cell lines was mainly mediated through JNK and p38 mitogen-activated protein kinase signaling. We confirmed these findings using rhinovirus infection, demonstrating that AA enhances rhinovirus-induced cytokine release. This study suggests that during respiratory infection, increased levels of dietary ω-6 PUFAs and SFAs may lead to more severe airway inflammation and may contribute to and/or increase the severity of asthma exacerbations.


Assuntos
Ácido Araquidônico/farmacologia , Células Epiteliais/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Ácido Palmítico/farmacologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Adulto , Idoso , Linhagem Celular , Ácidos Docosa-Hexaenoicos/farmacologia , Ácido Eicosapentaenoico/farmacologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Fibroblastos/imunologia , Fibroblastos/patologia , Regulação da Expressão Gênica , Células HeLa , Humanos , Interleucina-6/genética , Interleucina-6/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Lipopolissacarídeos/farmacologia , Pulmão/imunologia , Pulmão/patologia , MAP Quinase Quinase 4/genética , MAP Quinase Quinase 4/imunologia , Masculino , Pessoa de Meia-Idade , Poli I-C/farmacologia , Cultura Primária de Células , Rhinovirus/efeitos dos fármacos , Rhinovirus/crescimento & desenvolvimento , Transdução de Sinais/imunologia , Ácidos Teicoicos/farmacologia , Ácido alfa-Linoleico/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética
13.
Nutrition ; 57: 52-58, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30153579

RESUMO

OBJECTIVES: Defects in the insulin-signaling pathway have been implicated in the pathogenesis of impaired glucose uptake, insulin resistance, and type 2 diabetes. However, the specific defects that precipitate these abnormalities are yet to be fully elucidated. After binding to insulin, the plasma membrane-embedded insulin receptor transmembrane protein initiates a cascade of phosphorylation that leads to the activation of protein kinase B (AKT) and subsequently to the initiation of some metabolic actions of insulin. The activities of this receptor, insulin binding, and tyrosine kinase activation is dependent on its plasma lipid environment. Published data on the influence of omega-3 and -6 polyunsaturated fatty acids on insulin response are scarce. Moreover, the findings of the published investigations, most of which used omega-3 and -6, polyunsaturated fatty-acid blends, have been inconclusive. Hence, further, well thought out research is needed. The aim of the current study was to elucidate the effect of treatments with linoleic acid (LNA), arachidonic acid (ARA), alpha-linolenic acid (ALA), docoshexaenoic acid (DHA), and eicosapentaenoic acid (EPA) on cell membrane composition and consequently on the insulin-signaling pathway and specifically AKT phosphorylation. METHODS: Human colon adenocarcinoma (HT29) and liver hepatocellular (HepG2) cells were treated with or without 40 µM of LNA, ARA, ALA, EPA, or DHA for 48 h, the fatty-acid composition of phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdEtn) from the treated cells by capillary gas liquid chromatograph. Cells were incubated for 30 min with or without human insulin (50 ng/mL), and the phosphorylation of AKT was assessed with the use of Western blotting. RESULTS: The fatty acids were incorporated in the PtdCho and PtdEtn of both cell lines, but the level of incorporation was higher in HT29. Phosphorylation of AKT increased when HT29 was treated with LNA (P < 0.05) and ARA (P < 0.01) but not with ALA, EPA, or DHA. A similar but non-significant increase in AKT phosphorylation was observed in LNA- and ARA- treated HepG2 cells. CONCLUSIONS: The finding of this investigation demonstrates that plasma membrane lipid bilayer enrichment with LNA or ARA treatment enhances insulin action by AKT activation.


Assuntos
Ácido Araquidônico/farmacologia , Insulina/metabolismo , Ácido Linoleico/farmacologia , Proteínas Proto-Oncogênicas c-akt/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Células Cultivadas , Células HT29 , Células Hep G2 , Humanos , Técnicas In Vitro , Fosforilação
14.
Dev Psychobiol ; 61(1): 5-16, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30311214

RESUMO

The present study sought to determine whether supplementation of long-chain polyunsaturated fatty acids (LCPUFA) during the first year of life influenced brain function, structure, and metabolism at 9 years of age. Newborns were randomly assigned to consume formula containing either no LCPUFA (control) or formula with 0.64% of total fatty acids as arachidonic acid (ARA; 20:4n6) and variable amounts of docosahexaenoic acid (DHA; 22:6n3) (0.32%, 0.64%, or 0.96% of total fatty acids) from birth to 12 months. At age 9 years (±0.6), 42 children enrolled in a follow-up multimodal magnetic resonance imaging (MRI) study including functional (fMRI, Flanker task), resting state (rsMRI), anatomic, and proton magnetic resonance spectroscopy (1 H MRS). fMRI analysis using the Flanker task found that trials requiring greater inhibition elicited greater brain activation in LCPUFA-supplemented children in anterior cingulate cortex (ACC) and parietal regions. rsMRI analysis showed that children in the 0.64% group exhibited greater connectivity between prefrontal and parietal regions compared to all other groups. In addition, voxel-based analysis (VBM) revealed that the 0.32% and 0.64% groups had greater white matter volume in ACC and parietal regions compared to controls and the 0.96% group. Finally, 1 H MRS data analysis identified that N-acetylaspartate (NAA) and myo-inositol (mI) were higher in LCPUFA groups compared to the control group. LCPUFA supplementation during infancy has lasting effects on brain structure, function, and neurochemical concentrations in regions associated with attention (parietal) and inhibition (ACC), as well as neurochemicals associated with neuronal integrity (NAA) and brain cell signaling (mI).


Assuntos
Ácido Araquidônico/farmacologia , Atenção/fisiologia , Encéfalo/anatomia & histologia , Encéfalo/fisiologia , Desenvolvimento Infantil/fisiologia , Suplementos Nutricionais , Ácidos Docosa-Hexaenoicos/farmacologia , Fórmulas Infantis , Fenômenos Fisiológicos da Nutrição do Lactente , Imagem por Ressonância Magnética/métodos , Ácido Araquidônico/administração & dosagem , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Criança , Ácidos Docosa-Hexaenoicos/administração & dosagem , Feminino , Seguimentos , Neuroimagem Funcional , Humanos , Lactente , Recém-Nascido , Masculino , Imagem Multimodal , Espectroscopia de Prótons por Ressonância Magnética
15.
Ann Hematol ; 98(3): 581-588, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30446804

RESUMO

The thrombopoietin receptor agonist romiplostim is used for the long-term treatment of chronic immune thrombocytopenia (ITP). ITP patients have an increased thrombotic risk, which could be exacerbated if romiplostim increased platelet hyperreactivity or caused spontaneous platelet aggregation. To investigate this possibility, this study examined platelet function in romiplostim-treated ITP patients and healthy subjects. Light transmission platelet aggregometry utilizing arachidonic acid, collagen, epinephrine, ristocetin, ADP, and saline (to assess spontaneous aggregation) was performed for each subject. In addition, the ADP AC50 (ADP concentration that induced half-maximal aggregation) was determined for each patient as a sensitive measurement of altered platelet reactivity. Fifteen ITP patients and 7 healthy subjects entered the study. All ITP patients had active disease and were receiving weekly romiplostim as the sole ITP-directed therapy. Platelet aggregation in response to the strong agonists arachidonic acid, collagen, and ristocetin was not significantly different between ITP patients and healthy subjects (P = 0.2442, P = 0.0548, and P = 0.0879, respectively). Platelet aggregation in response to weak agonists was significantly reduced in ITP patients compared with that in healthy subjects: median (range) aggregation to ADP, 45% (15-84%) versus 89% (70-95%) (P = 0.0010), and epinephrine, 21% (1.6-90%) versus 88% (79-94%) (P = 0.0085). The median AC50 of ADP was threefold higher in ITP patients versus that in healthy subjects (6.3 µM vs 2.1 µM) (P = 0.0049). Significant spontaneous aggregation was not observed in any patient. Platelets from romiplostim-treated ITP patients do not show evidence for spontaneous aggregation or hyperreactivity, but instead have a modestly reduced aggregation response to ADP and epinephrine.


Assuntos
Transtornos Plaquetários/induzido quimicamente , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Receptores Fc/uso terapêutico , Proteínas Recombinantes de Fusão/uso terapêutico , Trombopoetina/uso terapêutico , Difosfato de Adenosina/farmacologia , Adulto , Idoso , Ácido Araquidônico/farmacologia , Colágeno/farmacologia , Epinefrina/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Agregação Plaquetária/efeitos dos fármacos , Púrpura Trombocitopênica Idiopática/sangue , Receptores de Trombopoetina/agonistas , Proteínas Recombinantes de Fusão/efeitos adversos , Proteínas Recombinantes de Fusão/farmacologia , Ristocetina/farmacologia , Trombofilia/induzido quimicamente , Trombopoetina/efeitos adversos , Trombopoetina/farmacologia , Adulto Jovem
16.
Platelets ; 30(8): 1001-1007, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30580677

RESUMO

The use of arachidonic acid (AA) to stimulate platelets is considered as a specific approach to study aspirin treatment efficacy. However, very high concentrations of AA are used, and it has been previously reported that AA can induce cell lysis in other settings. Several clinical studies have reported decreased responses to AA in whole blood tests in the presence of clopidogrel. Our aim was to investigate whether unspecific effects contribute to AA-induced aggregation and platelet activation in light transmission aggregometry (LTA) in platelet-rich plasma (PRP), and in assays using whole blood, multiple electrode aggregometry (MEA, Multiplate®), and flow cytometry. We report that cell lysis, especially of red blood cells, does occur at concentrations of AA used in the clinical tests and that ADP is very important for the AA-induced platelet activation responses. In flow cytometry, very limited platelet activation was detected before reaching AA concentrations in the millimolar range, where cell lysis also occurred, making it problematic to develop a reliable flow cytometry assay using AA as reagent. We conclude that cell lysis and ADP release contribute to AA-induced platelet responses, most markedly in whole blood assays. This finding could potentially explain some differences between studies comparing methods using whole blood and PRP and also how clopidogrel treatment could influence AA-induced aggregation results in previously published studies. Our findings highlight some issues with AA as reagent for platelet activation, which also have an impact on how platelet activation assays using AA should be interpreted.


Assuntos
Ácido Araquidônico/uso terapêutico , Células Sanguíneas/metabolismo , Ativação Plaquetária/fisiologia , Testes de Função Plaquetária/métodos , Ácido Araquidônico/farmacologia , Feminino , Humanos , Masculino
17.
Int J Mol Sci ; 19(12)2018 Dec 04.
Artigo em Inglês | MEDLINE | ID: mdl-30518038

RESUMO

Arachidonic and docosahexaenoic acids (ARA and DHA) are important during pregnancy. However, the effects of dietary supplementation on fetal growth and oxidative stress are inconclusive. We aimed to assess the effect of high ARA and DHA diet during rat gestation on: (1) ARA and DHA availability in plasma and placenta, (2) fetal growth, and (3) placental oxidative stress, analyzing the influence of sex. Experimental diet (ED) was prepared by substituting soybean oil in the control diet (CD) by a fungi/algae-based oil containing ARA and DHA (2:1). Rats were fed with CD or ED during gestation; plasma, placenta, and fetuses were obtained at gestational day 20. DHA, ARA, and their precursors were analyzed in maternal plasma and placenta by gas chromatography/mass spectrophotometry. Fetuses and placentas were weighed, the proportion of fetuses with intrauterine growth restriction (IUGR) determined, and placental lipid and protein oxidation analyzed. ED fetuses exhibited lower body weight compared to CD, being >40% IUGR; fetal weight negatively correlated with maternal plasma ARA, but not DHA. Only ED female placenta exhibited higher lipid and protein oxidation compared to its CD counterparts; lipid peroxidation is negatively associated with fetal weight. In conclusion, high ARA during gestation associates with IUGR, through placental oxidative stress, with females being more susceptible.


Assuntos
Ácido Araquidônico/farmacologia , Suplementos Nutricionais , Ácidos Docosa-Hexaenoicos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Placenta/patologia , Animais , Ácido Araquidônico/sangue , Dieta , Ácidos Docosa-Hexaenoicos/sangue , Feminino , Desenvolvimento Fetal/efeitos dos fármacos , Peso Fetal/efeitos dos fármacos , Feto/anatomia & histologia , Feto/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Tamanho do Órgão/efeitos dos fármacos , Oxirredução , Placenta/efeitos dos fármacos , Gravidez , Resultado da Gravidez , Ratos
18.
Respir Res ; 19(1): 211, 2018 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-30390648

RESUMO

BACKGROUND: The obesity paradox in COPD describes protective effects of obesity on lung pathology and inflammation. However, the underlying relationships between obesity, diet and disease outcomes in COPD are not fully understood. In this study we measured the response to dietary fatty acids upon markers of inflammation and remodelling in human lung cells from people with and without COPD. METHODS: Pulmonary fibroblasts were challenged with ω-3 polyunsaturated fatty acids (PUFAs), ω-6 PUFAs, saturated fatty acids (SFAs) or the obesity-associated cytokine TNFα. After 48-72 h release of the pro-inflammatory cytokines interleukin (IL)-6 and CXCL8 was measured using ELISA and mRNA expression and deposition of the extracellular matrix (ECM) proteins fibronectin, type I collagen, tenascin and perlecan were measured using qPCR or ECM ELISA, respectively. RESULTS: Challenge with the ω-6 PUFA arachidonic acid (AA), but not ω-3 PUFAs or SFAs, resulted in increased IL-6 and CXCL8 release from fibroblasts, however IL-6 and CXCL8 release was reduced in COPD (n = 19) compared to non-COPD (n = 36). AA-induced cytokine release was partially mediated by downstream mediators of cyclooxygenase (COX)-2 in both COPD and non-COPD. In comparison, TNFα-induced IL-6 and CXCL8 release was similar in COPD and non-COPD, indicating a specific interaction of AA in COPD. In patients with or without COPD, regression analysis revealed no relationship between BMI and cytokine release. In addition, AA, but not SFAs or ω-3 PUFAs reduced the basal deposition of fibronectin, type I collagen, tenascin and perlecan into the ECM in COPD fibroblasts. In non-COPD fibroblasts, AA-challenge decreased basal deposition of type I collagen and perlecan, but not fibronectin and tenascin. CONCLUSIONS: This study shows that AA has disease-specific effects on inflammation and ECM protein deposition. The impaired response to AA in COPD might in part explain why obesity appears to have less detrimental effects in COPD, compared to other lung diseases.


Assuntos
Ácido Araquidônico/efeitos adversos , Proteínas da Matriz Extracelular/antagonistas & inibidores , Proteínas da Matriz Extracelular/biossíntese , Ácidos Graxos Ômega-6/efeitos adversos , Mediadores da Inflamação/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Adulto , Idoso , Ácido Araquidônico/farmacologia , Células Cultivadas , Proteínas da Matriz Extracelular/genética , Ácidos Graxos Ômega-6/farmacologia , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Expressão Gênica , Humanos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Doença Pulmonar Obstrutiva Crônica/patologia
19.
Toxicol Appl Pharmacol ; 360: 141-149, 2018 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-30290169

RESUMO

Salinomycin, a monocarboxylic ionophore in Streptomyces albus, has been studied as an anti-cancer agent. However, we wondered whether salinomycin has another effect such as an anti-oxidant and hepatic protectant, because some chemical drugs treating human diseases were sometimes related with their toxic effects. Therefore, this study was conducted to examine the effects of salinomycin against oxidative stress and mitochondrial impairment in vivo and in vitro as well as the cellular mechanisms of action. In hepatocyte, salinomycin inhibited arachidonic acid (AA) + iron-induced apoptosis, mitochondrial dysfunction and ROS production. As a molecular mechanism, salinomycin induced autophagy through AMP-activated protein kinase (AMPK) activation, as assessed by the accumulation of acidic vesicle organelles, p62 and LC3-II. Moreover, these protective effects were blocked by AMPK inhibition, which indicates the importance of AMPK in the process of salinomycin's effects. In mice, oral administration of salinomycin protected against carbon tetrachloride (CCl4)-induced oxidative stress and liver injury, and also activated AMPK as well as autophagy-related proteins in the liver. Collectively, salinomycin had the ability to protect hepatocytes against AA+iron-induced reactive oxygen species production and mitochondrial dysfunction, as well as CCl4-induced liver injury. Although this beneficial effect was demonstrated under severe oxidative stress, this study showed that salinomycin protected the liver against the oxidative stress and liver damage through AMPK and autophagy, and suggest that salinomycin has a possibility to treat a broad range of diseases.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Autofagia/efeitos dos fármacos , Hepatopatias/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Piranos/farmacologia , Animais , Apoptose/efeitos dos fármacos , Ácido Araquidônico/farmacologia , Tetracloreto de Carbono/farmacologia , Linhagem Celular Tumoral , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Hepatopatias/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Fosforilação/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
20.
Vet Clin Pathol ; 47(4): 556-559, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30325547

RESUMO

BACKGROUND: Whole blood impedance platelet aggregometry (Multiplate-) can be performed with different agonists to evaluate platelet function. Although the manufacturer recommends disposal of stored reagents after 1 month at -20°C or 24 hours at 4°C, reagent integrity after reconstitution under different storage conditions is unknown. If reagent integrity is stable for longer periods, assay costs could decrease dramatically. OBJECTIVES: This study aimed to determine the stability of reconstituted arachidonic acid (AA) and adenosine diphosphate (ADP) platelet agonists stored at -20°C and -80°C for up to 6 months. METHODS: Aliquots of reconstituted AA and ADP were stored at -20°C and -80°C each month for a total of 6 months. Six healthy staff-owned dogs were enrolled in the study. A physical examination, CBC, diagnostic panel, urinalysis, and baseline platelet aggregometry assessment was performed on all of the dogs. Platelet aggregometry was performed using fresh and stored aliquots of AA and ADP reagents on the same day. The area under the curve (AUC) was recorded from each platelet aggregometry analysis. Repeated measures (RM) analysis (one-way ANOVA) was performed and subsequent time points (1, 2, 3, 4, 5, and 6 months) were compared with fresh AUC results. RESULTS: All dogs were clinically healthy, and all diagnostic tests were normal. There were no differences in AUC obtained from fresh samples at any time point or either temperature for AA or ADP. CONCLUSIONS: Whole blood impedance platelet aggregometry reagents, AA and ADP, were stable for up to 6 months when stored at -20°C or -80°C, obviating the need to discard viable reagents, and decreasing assay costs.


Assuntos
Apirase/farmacologia , Ácido Araquidônico/farmacologia , Coleta de Amostras Sanguíneas/veterinária , Agregação Plaquetária , Testes de Função Plaquetária/veterinária , Animais , Plaquetas/efeitos dos fármacos , Coleta de Amostras Sanguíneas/métodos , Cães/sangue , Impedância Elétrica , Agregação Plaquetária/efeitos dos fármacos
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