Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 29
Filtrar
1.
Circ Res ; 134(8): 954-969, 2024 Apr 12.
Artigo em Inglês | MEDLINE | ID: mdl-38501247

RESUMO

BACKGROUND: Acute ischemic stroke triggers endothelial activation that disrupts vascular integrity and increases hemorrhagic transformation leading to worsened stroke outcomes. rt-PA (recombinant tissue-type plasminogen activator) is an effective treatment; however, its use is limited due to a restricted time window and hemorrhagic transformation risk, which in part may involve activation of MMPs (matrix metalloproteinases) mediated through LOX-1 (lectin-like oxLDL [oxidized low-density lipoprotein] receptor 1). This study's overall aim was to evaluate the therapeutic potential of novel MMP-9 (matrix metalloproteinase 9) ± LOX-1 inhibitors in combination with rt-PA to improve stroke outcomes. METHODS: A rat thromboembolic stroke model was utilized to investigate the impact of rt-PA delivered 4 hours poststroke onset as well as selective MMP-9 (JNJ0966) ±LOX-1 (BI-0115) inhibitors given before rt-PA administration. Infarct size, perfusion, and hemorrhagic transformation were evaluated by 9.4-T magnetic resonance imaging, vascular and parenchymal MMP-9 activity via zymography, and neurological function was assessed using sensorimotor function testing. Human brain microvascular endothelial cells were exposed to hypoxia plus glucose deprivation/reperfusion (hypoxia plus glucose deprivation 3 hours/R 24 hours) and treated with ±tPA and ±MMP-9 ±LOX-1 inhibitors. Barrier function was assessed via transendothelial electrical resistance, MMP-9 activity was determined with zymography, and LOX-1 and barrier gene expression/levels were measured using qRT-PCR (quantitative reverse transcription PCR) and Western blot. RESULTS: Stroke and subsequent rt-PA treatment increased edema, hemorrhage, MMP-9 activity, LOX-1 expression, and worsened neurological outcomes. LOX-1 inhibition improved neurological function, reduced edema, and improved endothelial barrier integrity. Elevated MMP-9 activity correlated with increased edema, infarct volume, and decreased neurological function. MMP-9 inhibition reduced MMP-9 activity and LOX-1 expression. In human brain microvascular endothelial cells, LOX-1/MMP-9 inhibition differentially attenuated MMP-9 levels, inflammation, and activation following hypoxia plus glucose deprivation/R. CONCLUSIONS: Our findings indicate that LOX-1 inhibition and ± MMP-9 inhibition attenuate negative aspects of ischemic stroke with rt-PA therapy, thus resulting in improved neurological function. While no synergistic effect was observed with simultaneous LOX-1 and MMP-9 inhibition, a distinct interaction is evident.


Assuntos
Isquemia Encefálica , AVC Isquêmico , Acidente Vascular Cerebral , Ratos , Humanos , Animais , Ativador de Plasminogênio Tecidual , Metaloproteinase 9 da Matriz/metabolismo , AVC Isquêmico/tratamento farmacológico , Células Endoteliais/metabolismo , Ratos Sprague-Dawley , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/patologia , Hemorragia , Edema/tratamento farmacológico , Edema/patologia , Glucose/farmacologia , Infarto/tratamento farmacológico , Hipóxia
2.
Am J Physiol Cell Physiol ; 325(4): C951-C971, 2023 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-37642239

RESUMO

Endothelial integrity is critical in mitigating a vicious cascade of secondary injuries following acute ischemic stroke (AIS). Matrix metalloproteinase-9 (MMP-9), a contributor to endothelial integrity loss, is elevated during stroke and is associated with worsened stroke outcome. We investigated the FDA-approved selective sphingosine-1-phosphate receptor 1 (S1PR1) ligand, ozanimod, on the regulation/activity of MMP-9 as well as endothelial barrier components [platelet endothelial cell adhesion molecule 1 (PECAM-1), claudin-5, and zonula occludens 1 (ZO-1)] in human brain microvascular endothelial cells (HBMECs) following hypoxia plus glucose deprivation (HGD). We previously reported that S1PR1 activation improves HBMEC integrity; however, mechanisms underlying S1PR1 involvement in endothelial cell barrier integrity have not been clearly elucidated. We hypothesized that ozanimod would attenuate an HGD-induced increase in MMP-9 activity that would concomitantly attenuate the loss of integral barrier components. Male HBMECs were treated with ozanimod or vehicle and exposed to 3 h of normoxia (21% O2) or HGD (1% O2). Immunoblotting, zymography, qRT-PCR, and immunocytochemical labeling techniques assessed processes related to MMP-9 and barrier markers. We observed that HGD acutely increased MMP-9 activity and reduced claudin-5 and PECAM-1 levels, and ozanimod attenuated these responses. In situ analysis, via PROSPER, suggested that attenuation of MMP-9 activity may be a primary factor in maintaining these integral barrier proteins. We also observed that HGD increased intracellular mechanisms associated with augmented MMP-9 activation; however, ozanimod had no effect on these select factors. Thus, we conclude that ozanimod has the potential to attenuate HGD-mediated decreases in HBMEC integrity in part by decreasing MMP-9 activity as well as preserving barrier properties.NEW & NOTEWORTHY We have identified a potential novel mechanism by which ozanimod, a selective sphingosine-1-phosphate receptor 1 (S1PR1) agonist, attenuates hypoxia plus glucose deprivation (HGD)-induced matrix metalloproteinase-9 (MMP-9) activity and disruptions in integral human brain endothelial cell barrier proteins. Our results suggest that ischemic-like injury elicits increased MMP-9 activity and alterations of barrier integrity proteins in human brain microvascular endothelial cells (HBMECs) and that ozanimod via S1PR1 attenuates these HGD-induced responses, adding to its therapeutic potential in cerebrovascular protection during the acute phase of ischemic stroke.


Assuntos
AVC Isquêmico , Acidente Vascular Cerebral , Humanos , Masculino , Barreira Hematoencefálica/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Receptores de Esfingosina-1-Fosfato/metabolismo , Células Endoteliais/metabolismo , Claudina-5/metabolismo , AVC Isquêmico/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/metabolismo , Isquemia/metabolismo , Hipóxia/metabolismo , Glucose/metabolismo
3.
J Neurotrauma ; 39(19-20): 1429-1441, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35593008

RESUMO

Severe traumatic brain injury (TBI) results in cognitive dysfunction in part due to vascular perturbations. In contrast, the long-term vasculo-cognitive pathophysiology of mild TBI (mTBI) remains unknown. We evaluated mTBI effects on chronic cognitive and cerebrovascular function and assessed their interrelationships. Sprague-Dawley rats received midline fluid percussion injury (n = 20) or sham (n = 21). Cognitive function was assessed (3- and 6-month novel object recognition [NOR], novel object location [NOL], and temporal order object recognition [TOR]). Six-month cerebral blood flow (CBF) and cerebral blood volume (CBV) using contrast magnetic resonance imaging (MRI) and ex vivo circle of Willis artery endothelial and smooth muscle-dependent function were measured. mTBI rats showed significantly impaired NOR, with similar trends (non-significant) in NOL/TOR. Regional CBF and CBV were similar in sham and mTBI. NOR correlated with CBF in lateral hippocampus, medial hippocampus, and primary somatosensory barrel cortex, whereas it inversely correlated with arterial smooth muscle-dependent dilation. Six-month baseline endothelial and smooth muscle-dependent arterial function were similar among mTBI and sham, but post-angiotensin 2 stimulation, mTBI showed no change in smooth muscle-dependent dilation from baseline response, unlike the reduction in sham. mTBI led to chronic cognitive dysfunction and altered angiotensin 2-stimulated smooth muscle-dependent vasoreactivity. The findings of persistent pathophysiological consequences of mTBI in this animal model add to the broader understanding of chronic pathophysiological sequelae in human mild TBI.


Assuntos
Concussão Encefálica , Circulação Cerebrovascular , Cognição , Animais , Humanos , Ratos , Angiotensinas , Concussão Encefálica/complicações , Concussão Encefálica/patologia , Ratos Sprague-Dawley
4.
Am J Physiol Cell Physiol ; 320(6): C1055-C1073, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33788630

RESUMO

Vascular smooth muscle (VSM) cell phenotypic expression and autophagic state are dynamic responses to stress. Vascular pathologies, such as hypoxemia and ischemic injury, induce a synthetic VSM phenotype and autophagic flux resulting in a loss of vascular integrity and VSM cell death respectfully. Both clinical pilot and experimental stroke studies demonstrate that sphingosine-1-phosphate receptor (S1PR) modulation improves stroke outcome; however, specific mechanisms associated with a beneficial outcome at the level of the cerebrovasculature have not been clearly elucidated. We hypothesized that ozanimod, a selective S1PR type 1 ligand, will attenuate VSM synthetic phenotypic expression and autophagic flux in primary human brain VSM cells following acute hypoxia plus glucose deprivation (HGD; in vitro ischemic-like injury) exposure. Cells were treated with ozanimod and exposed to normoxia or HGD. Crystal violet staining, standard immunoblotting, and immunocytochemical labeling techniques assessed cellular morphology, vacuolization, phenotype, and autophagic state. We observed that HGD temporally decreased VSM cell viability and concomitantly increased vacuolization, both of which ozanimod reversed. HGD induced a simultaneous elevation and reduction in levels of pro- and antiautophagic proteins respectfully, and ozanimod attenuated this response. Protein levels of VSM phenotypic biomarkers, smoothelin and SM22, were decreased following HGD. Furthermore, we observed an HGD-induced epithelioid and synthetic morphological appearance accompanied by disorganized cytoskeletal filaments, which was rescued by ozanimod. Thus, we conclude that ozanimod, a selective S1PR1 ligand, protects against acute HGD-induced phenotypic switching and promotes cell survival, in part, by attenuating HGD-induced autophagic flux thus improving vascular patency in response to acute ischemia-like injury.


Assuntos
Autofagia/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Glucose/metabolismo , Hipóxia/tratamento farmacológico , Indanos/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Oxidiazóis/farmacologia , Receptores de Esfingosina-1-Fosfato/metabolismo , Encéfalo/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Hipóxia/metabolismo , Ligantes , Masculino , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Fenótipo , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/metabolismo
5.
Neurosci Lett ; 735: 135160, 2020 09 14.
Artigo em Inglês | MEDLINE | ID: mdl-32561451

RESUMO

Endothelial sphingosine-1-phosphate receptors are emerging as relevant therapeutic targets during acute ischemic stroke (AIS). Physiologically, the cerebrovascular endothelium plays a vital role in maintaining barrier integrity and cerebrovascular homeostasis. During a cerebral ischemic event, products from parenchymal cell death are released and trigger vascular endothelial dysfunction and vascular inflammation leading to barrier integrity disruption. Endothelial dysfunction, inflammation, and a breach in barrier property play a significant role in contributing to a vicious cycle which promotes brain edema formation and exacerbates neuronal injury post stroke. Data from experimental stroke models and clinical trials suggest that selective sphingosine-1-phosphate receptor type 1 (S1PR1) modulation improves endothelial health and function and, as a result, contributes to improved neurological outcome post ischemic injury. This review highlights the impact of sphingosine-1-phosphate (S1P)/S1PR1 signaling involved in blood brain barrier (BBB) integrity and cerebrovascular inflammation following AIS. We focus on the beneficial actions of S1PR1 signaling during ischemic injury including barrier protection to lessen brain edema formation and reduction in the development and progression of vascular inflammation by attenuating endothelial cell activation resulting in reduced neurovascular inflammation. Potential gaps and future directions related to the role of S1PR during AIS are also discussed.


Assuntos
Barreira Hematoencefálica/metabolismo , Inflamação/metabolismo , AVC Isquêmico/metabolismo , Receptores de Esfingosina-1-Fosfato/metabolismo , Animais , Barreira Hematoencefálica/patologia , Humanos , Inflamação/patologia , AVC Isquêmico/patologia
6.
FASEB J ; 33(10): 10935-10941, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31284754

RESUMO

Sphingosine-1-phosphate receptor (S1PR) modulators provide protection in preclinical and clinical studies for ischemic stroke, but the influences of S1PR modulation on microvascular thrombosis remain poorly understood. This study investigates the impact of a selective S1PR1 modulator RP101075 on microvascular circulation in a mouse model of laser-induced thrombosis. The flow velocity of cortical arterioles in mice was measured in vivo under 2-photon laser scanning microscopy. Thrombosis was induced in cortical arterioles by laser irritation. At 30 min after laser-induced thrombosis, mice were treated with either RP101075 or vehicle. RP101075 did not alter the flow velocity of cortical arterioles under physiologic conditions. Laser-induced thrombosis led to a pronounced reduction of flow velocity in cortical arterioles that persisted for ≥90 min. The reduction of flow velocity in cortical arterioles following thrombosis was significantly attenuated following RP101075 treatment. RP101075 did not significantly affect coagulation time, bleeding time, heart rate, and blood pressure. In addition, RP101075 treatment reduced thrombus volume, which was accompanied by a reduction of leukocyte content in the thrombus. Our findings demonstrate that the selective S1PR1 modulator RP101075 improves microvascular circulation after thrombosis, implying a component of improved microvascular circulation to the benefit of S1PR modulation in cerebral ischemia.-Li, H., Zhou, X., Li, Y., Ma, X., Gonzales, R. J., Qiu, S., Shi, F.-D., Liu, Q. The selective sphingosine 1-phosphate receptor 1 modulator RP101075 improves microvascular circulation after cerebrovascular thrombosis.


Assuntos
Transtornos Cerebrovasculares/tratamento farmacológico , Indanos/uso terapêutico , Microcirculação , Oxidiazóis/uso terapêutico , Moduladores do Receptor de Esfingosina 1 Fosfato/uso terapêutico , Trombose/tratamento farmacológico , Animais , Circulação Cerebrovascular , Masculino , Camundongos , Camundongos Endogâmicos C57BL
7.
Physiol Rep ; 7(7): e14052, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30963722

RESUMO

Anthracycline chemotherapies are effective at reducing disease recurrence and mortality in cancer patients. However, these drugs also contribute to skeletal muscle wasting and dysfunction. The purpose of this study was to assess the impact of chronic doxorubicin (DOX) administration on satellite cell and capillary densities in different skeletal muscles. We hypothesized that DOX would reduce satellite cell and capillary densities of the soleus (SOL) and extensor digitorum longus (EDL) muscles, along with muscle fiber size. Ovariectomized female Sprague-Dawley rats were randomized to receive three bi-weekly intraperitoneal injections of DOX (4 mg∙kg-1 ; cumulative dose 12 mg∙kg-1 ) or vehicle (VEH; saline). Animals were euthanized 5d following the last injection and the SOL and EDL were dissected and prepared for immunohistochemical and RT-qPCR analyses. Relative to VEH, CSA of the SOL and EDL fibers were 26% and 33% smaller, respectively, in DOX (P < 0.05). In the SOL, satellite cell and capillary densities were 39% and 35% lower, respectively, in DOX (P < 0.05), whereas in the EDL satellite cell and capillary densities were unaffected by DOX administration (P > 0.05). Proliferating satellite cells were unaffected by DOX in the SOL (P > 0.05). In the SOL, MYF5 mRNA expression was increased in DOX (P < 0.05), while in the EDL MGF mRNA expression was reduced in DOX (P < 0.05). Chronic DOX administration is associated with reduced fiber size in the SOL and EDL; however, DOX appeared to reduce satellite cell and capillary densities only in the SOL. These findings highlight that therapeutic targets to protect skeletal muscle from DOX may vary across muscles.


Assuntos
Antibióticos Antineoplásicos/administração & dosagem , Capilares/efeitos dos fármacos , Doxorrubicina/administração & dosagem , Músculo Esquelético/efeitos dos fármacos , Células Satélites de Músculo Esquelético/efeitos dos fármacos , Animais , Feminino , Músculo Esquelético/irrigação sanguínea , Ratos , Ratos Sprague-Dawley
8.
Ann Neurol ; 84(5): 717-728, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30295338

RESUMO

OBJECTIVE: The present study was undertaken to determine the efficacy of coadministration of fingolimod with alteplase in acute ischemic stroke patients in a delayed time window. METHODS: This was a prospective, randomized, open-label, blinded endpoint clinical trial, enrolling patients with internal carotid artery or middle cerebral artery proximal occlusion within 4.5 to 6 hours from symptom onset. Patients were randomly assigned to receive alteplase alone or alteplase with fingolimod. All patients underwent pretreatment and 24-hour noncontrast computed tomography (CT)/perfusion CT/CT angiography. The coprimary endpoints were the decrease of National Institutes of Health Stroke Scale scores over 24 hours and the favorable shift of modified Rankin Scale score (mRS) distribution at day 90. Exploratory outcomes included vessel recanalization, anterograde reperfusion, and retrograde reperfusion of collateral flow. RESULTS: Each treatment group included 23 patients. Compared with alteplase alone, patients receiving fingolimod plus alteplase exhibited better early clinical improvement at 24 hours and a favorable shift of mRS distribution at day 90. In addition, patients who received fingolimod and alteplase exhibited a greater reduction in the perfusion lesion accompanied by suppressed infarct growth by 24 hours. Fingolimod in conjunction with alteplase significantly improved anterograde reperfusion of downstream territory and prevented the failure of retrograde reperfusion from collateral circulation. INTERPRETATION: Fingolimod may enhance the efficacy of alteplase administration in the 4.5- to 6-hour time window in patients with a proximal cerebral arterial occlusion and salvageable penumbral tissue by promoting both anterograde reperfusion and retrograde collateral flow. These findings are instructive for the design of future trials of recanalization therapies in extended time windows. Ann Neurol 2018;84:725-736.


Assuntos
Fibrinolíticos/administração & dosagem , Cloridrato de Fingolimode/administração & dosagem , Imunossupressores/administração & dosagem , Acidente Vascular Cerebral/tratamento farmacológico , Ativador de Plasminogênio Tecidual/administração & dosagem , Idoso , Circulação Colateral/efeitos dos fármacos , Quimioterapia Combinada , Feminino , Humanos , Masculino , Recuperação de Função Fisiológica/efeitos dos fármacos , Reperfusão , Acidente Vascular Cerebral/patologia , Tempo para o Tratamento
9.
Stroke ; 49(6): 1471-1478, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29695462

RESUMO

BACKGROUND AND PURPOSE: Autoimmune responses can occur when antigens from the central nervous system are presented to lymphocytes in the periphery or central nervous system in several neurological diseases. However, whether autoimmune responses emerge after brain ischemia and their impact on clinical outcomes remains controversial. We hypothesized that brain ischemia facilitates the genesis of autoimmunity and aggravates ischemic brain injury. METHODS: Using a mouse strain that harbors a transgenic T-cell receptor to a central nervous system antigen, MOG35-55 (myelin oligodendrocyte glycoprotein) epitope (2D2), we determined the anatomic location and involvement of antigen-presenting cells in the development of T-cell reactivity after brain ischemia and how T-cell reactivity impacts stroke outcome. Transient middle cerebral artery occlusion and photothrombotic stroke models were used in this study. We also quantified the presence and status of T cells from brain slices of ischemic patients. RESULTS: By coupling transfer of labeled MOG35-55-specific (2D2) T cells with tetramer tracking, we show an expansion in reactivity of 2D2 T cells to MOG91-108 and MOG103-125 in transient middle cerebral artery occlusion and photothrombotic stroke models. This reactivity and T-cell activation first occur locally in the brain after ischemia. Also, microglia act as antigen-presenting cells that effectively present MOG antigens, and depletion of microglia ablates expansion of 2D2 reactive T cells. Notably, the adoptive transfer of neuroantigen-experienced 2D2 T cells exacerbates Th1/Th17 responses and brain injury. Finally, T-cell activation and MOG-specific T cells are present in the brain of patients with ischemic stroke. CONCLUSIONS: Our findings suggest that brain ischemia activates and diversifies T-cell responses locally, which exacerbates ischemic brain injury.


Assuntos
Lesões Encefálicas/imunologia , Isquemia Encefálica/imunologia , Acidente Vascular Cerebral/imunologia , Linfócitos T/imunologia , Transferência Adotiva/métodos , Animais , Encéfalo/imunologia , Sistema Nervoso Central/imunologia , Humanos , Infarto da Artéria Cerebral Média/imunologia , Camundongos Endogâmicos C57BL , Glicoproteína Mielina-Oligodendrócito/imunologia
10.
FASEB J ; 32(5): 2757-2767, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29401578

RESUMO

Stroke-induced immune suppression predisposes the host to infections and can contribute to high morbidity and mortality in stroke patients. Because ischemic stroke has a profound effect on the systemic immune response, which may explain the increased susceptibility of stroke patients to infection, an urgent need persists for a better understanding of mechanisms associated with immune suppression; new and effective treatments for stroke can then be identified. NK cells play a key role in early host defense against pathogens by killing infected cells and/or producing cytokines such as IFN-γ. Because the phenotype and function of peripheral NK cells have been widely investigated in ischemic stroke, nCounter Inflammation Gene Array Analysis was used to build immune-related gene profiles of NK cells to comprehensively analyze the molecular signature of NK cells after ischemic brain injury. We observed distinct gene expression profiles reflecting different splenic NK-cell phenotypes and functional properties across the time course of transient middle cerebral artery occlusion (MCAO). Based on gene expression and pathway-network analysis, lower expression levels of signal transducer and activator of transcription-3 (STAT3) were observed in animals with MCAO compared with sham control animals. Genetic activation of STAT3 through the introduction of STAT3 clustered regularly interspaced short palindromic repeats (CRISPR) plasmid prevented the loss of NK-cell-derived IFN-γ production after MCAO, together with reduced bacterial burden and mortality. Our data suggest that brain ischemia impairs NK-cell-mediated immune defense in the periphery, at least in part through the JAK-STAT3 pathway, which can be readdressed by modulating STAT3 activation status.-Jin, W.-N., Ducruet, A. F., Liu, Q., Shi, S. X.-Y., Waters, M., Zou, M., Sheth, K. N., Gonzales, R., Shi, F.-D. Activation of JAK/STAT3 restores NK-cell function and improves immune defense after brain ischemia.


Assuntos
Isquemia Encefálica/imunologia , Regulação da Expressão Gênica/imunologia , Interferon gama/imunologia , Janus Quinases/imunologia , Células Matadoras Naturais/imunologia , Fator de Transcrição STAT3/imunologia , Animais , Isquemia Encefálica/genética , Isquemia Encefálica/patologia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Interferon gama/genética , Janus Quinases/genética , Células Matadoras Naturais/patologia , Masculino , Camundongos , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Fator de Transcrição STAT3/genética
11.
Am J Physiol Cell Physiol ; 314(5): C545-C553, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29384693

RESUMO

Vascular basal cyclooxygenase-2 (COX-2) expression and activity can be induced by endotoxin, hypoxia, or ischemia. During vascular pathologies such as atherosclerosis, increases in COX-2 activity result in prostanoid production, a contributor to the development and progression of vascular inflammation leading to unstable atherosclerotic plaques and increased risk for thrombotic events. Recent studies demonstrate that select free fatty acids, such as palmitate, can act as proinflammatory mediators. However, the effect of palmitate on COX-2 expression and activity, and its impact on the development and progression of vascular inflammation, are not well elucidated. We investigated the effect of palmitate on COX-2 expression and function in human vascular smooth muscle cells. Cells were treated with palmitate, COX-2 protein levels were assessed using Western analysis, and activity was assessed via ELISA. We observed that palmitate dose-dependently increased COX-2 levels and specifically enhanced band intensity of the COX-2 74 kDa band (slowest migrating band). This response was attenuated by N-linked glycosylation inhibition, suggesting that palmitate impacts expression of the fully activated glycoform of COX-2. Palmitate-induced increases in COX-2 levels correlated with an increase in prostaglandin E2 production that was also attenuated by a glycosylation inhibitor. Additionally, palmitate altered cell morphology and increased cell density which were reversed by selective COX-2 inhibition. Thus, we conclude that palmitate acts on COX-2 by two separate mechanisms of action in human vascular smooth muscle. It elicits dose-dependent increases in COX-2 protein expression and modulates regulation of COX-2 activity via modification of posttranslational glycosylation.


Assuntos
Ciclo-Oxigenase 2/metabolismo , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Ácido Palmítico/farmacologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Células Cultivadas , Dinoprostona/metabolismo , Relação Dose-Resposta a Droga , Feminino , Glicosilação , Humanos , Masculino , Músculo Liso Vascular/enzimologia , Miócitos de Músculo Liso/enzimologia , Cultura Primária de Células
12.
Med Sci Sports Exerc ; 49(12): 2394-2403, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28767526

RESUMO

PURPOSE: This study aimed to assess the ability for exercise training performed before and during biweekly doxorubicin (DOX) administration to attenuate adverse effects of DOX on skeletal muscle. We hypothesized that DOX treatment would increase REDD1, impair mammalian target of rapamycin (mTOR) signaling, and reduce muscle fiber size, and that exercise training would attenuate these responses. METHODS: Eight-week-old ovariectomized female Sprague-Dawley rats were randomized to one of four treatments: exercise + DOX (Ex-Dox), Ex + vehicle (Ex-Veh), sedentary + DOX (Sed-Dox), and Sed + Veh (Sed-Veh). DOX (4 mg·kg) or vehicle (saline) intraperitoneal injections were performed biweekly for a total of three injections (cumulative dose, 12 mg·kg). Ex animals performed interval exercise (4 × 4 min, 85%-90% V˙O2peak) 5 d·wk starting 1 wk before the first injection and continued throughout study duration. Animals were euthanized ~5 d after the last injection, during which the soleus muscle was dissected and prepared for immunoblot and immunohistochemical analyses. RESULTS: REDD1 mRNA and protein were increased only in Sed-Dox (P < 0.05). The phosphorylation of mTOR and 4E-BP1 and MHC I and MHC IIa fiber size were lower in Sed-Dox versus Sed-Veh (P < 0.05). By contrast, REDD1 mRNA and protein, mTOR, 4E-BP1, and MHC I fiber size were not different between Ex-Dox and Ex-Veh (P > 0.05). LC3BI was higher, and the LC3BII/I ratio was lower in Sed-Dox versus Sed-Veh (P < 0.05) but not between Ex-Dox and Ex-Veh (P > 0.05). CONCLUSION: These data suggest that DOX may inhibit mTORC1 activity and reduce MHCI and MHCIIa fiber size, potentially through elevated REDD1, and that exercise may provide a therapeutic strategy to preserve skeletal muscle size during chronic DOX treatment.


Assuntos
Antibióticos Antineoplásicos/toxicidade , Doxorrubicina/toxicidade , Músculo Esquelético/efeitos dos fármacos , Condicionamento Físico Animal/fisiologia , Animais , Antibióticos Antineoplásicos/administração & dosagem , Autofagia , Proteínas de Transporte/efeitos dos fármacos , Proteínas de Transporte/metabolismo , Tamanho Celular , Doxorrubicina/administração & dosagem , Feminino , Peptídeos e Proteínas de Sinalização Intracelular , Modelos Animais , Fibras Musculares Esqueléticas/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Fosfoproteínas/efeitos dos fármacos , Fosfoproteínas/metabolismo , Fosforilação , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Distribuição Aleatória , Ratos Sprague-Dawley , Proteínas Repressoras/efeitos dos fármacos , Proteínas Repressoras/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição
13.
J Neuroinflammation ; 14(1): 151, 2017 07 28.
Artigo em Inglês | MEDLINE | ID: mdl-28754131

RESUMO

BACKGROUND: Cerebral ischemia is a leading cause of death and disability with limited treatment options. Although inflammatory and immune responses participate in ischemic brain injury, the molecular regulators of neuroinflammation after ischemia remain to be defined. Translocator protein 18 kDa (TSPO) mainly localized to the mitochondrial outer membrane is predominantly expressed in glia within the central nervous system during inflammatory conditions. This study investigated the effect of a TSPO agonist, etifoxine, on neuroinflammation and brain injury after ischemia/reperfusion. METHODS: We used a mouse model of middle cerebral artery occlusion (MCAO) to examine the therapeutic potential and mechanisms of neuroprotection by etifoxine. RESULTS: TSPO was upregulated in Iba1+ or CD11b+CD45int cells from mice subjected to MCAO and reperfusion. Etifoxine significantly attenuated neurodeficits and infarct volume after MCAO and reperfusion. The attenuation was pronounced in mice subjected to 30, 60, or 90 min MCAO. Etifoxine reduced production of pro-inflammatory factors in the ischemic brain. In addition, etifoxine treatment led to decreased expression of interleukin-1ß, interleukin-6, tumor necrosis factor-α, and inducible nitric oxide synthase by microglia. Notably, the benefit of etifoxine against brain infarction was ablated in mice depleted of microglia using a colony-stimulating factor 1 receptor inhibitor. CONCLUSIONS: These findings indicate that the TSPO agonist, etifoxine, reduces neuroinflammation and brain injury after ischemia/reperfusion. The therapeutic potential of targeting TSPO requires further investigations in ischemic stroke.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Infarto da Artéria Cerebral Média/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Oxazinas/uso terapêutico , Receptores de GABA/metabolismo , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Edema Encefálico/tratamento farmacológico , Edema Encefálico/etiologia , Infarto Encefálico/tratamento farmacológico , Infarto Encefálico/etiologia , Citocinas/metabolismo , Modelos Animais de Doenças , Esquema de Medicação , Citometria de Fluxo , Camundongos , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Exame Neurológico , RNA Mensageiro , Traumatismo por Reperfusão/patologia , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismo
14.
J Cereb Blood Flow Metab ; 37(6): 2224-2236, 2017 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28273719

RESUMO

Brain ischemia elicits microglial activation and microglia survival depend on signaling through colony-stimulating factor 1 receptor (CSF1R). Although depletion of microglia has been linked to worse stroke outcomes, it remains unclear to what extent and by what mechanisms activated microglia influence ischemia-induced inflammation and injury in the brain. Using a mouse model of transient focal cerebral ischemia and reperfusion, we demonstrated that depletion of microglia via administration of the dual CSF1R/c-Kit inhibitor PLX3397 exacerbates neurodeficits and brain infarction. Depletion of microglia augmented the production of inflammatory mediators, leukocyte infiltration, and cell death during brain ischemia. Of note, microglial depletion-induced exacerbation of stroke severity did not solely depend on lymphocytes and monocytes. Importantly, depletion of microglia dramatically augmented the production of inflammatory mediators by astrocytes after brain ischemia . In vitro studies reveal that microglia restricted ischemia-induced astrocyte response and provided neuroprotective effects. Our findings suggest that neuroprotective effects of microglia may result, in part, from its inhibitory action on astrocyte response after ischemia.


Assuntos
Isquemia Encefálica/imunologia , Isquemia Encefálica/patologia , Mediadores da Inflamação/metabolismo , Microglia/imunologia , Microglia/patologia , Aminopiridinas/farmacologia , Animais , Isquemia Encefálica/diagnóstico por imagem , Células Cultivadas , Modelos Animais de Doenças , Imageamento por Ressonância Magnética , Masculino , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Neurônios/patologia , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidores , Pirróis/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores
15.
Immunity ; 46(3): 474-487, 2017 03 21.
Artigo em Inglês | MEDLINE | ID: mdl-28314594

RESUMO

Brain ischemia inhibits immune function systemically, with resulting infectious complications. Whether in stroke different immune alterations occur in brain and periphery and whether analogous mechanisms operate in these compartments remains unclear. Here we show that in patients with ischemic stroke and in mice subjected to middle cerebral artery occlusion, natural killer (NK) cells display remarkably distinct temporal and transcriptome profiles in the brain as compared to the periphery. The activation of catecholaminergic and hypothalamic-pituitary-adrenal axis leads to splenic atrophy and contraction of NK cell numbers in the periphery through a modulated expression of SOCS3, whereas cholinergic innervation-mediated suppression of NK cell responses in the brain involves RUNX3. Importantly, pharmacological or genetic ablation of innervation preserved NK cell function and restrained post-stroke infection. Thus, brain ischemia compromises NK cell-mediated immune defenses through mechanisms that differ in the brain versus the periphery, and targeted inhibition of neurogenic innervation limits post-stroke infection.


Assuntos
Isquemia Encefálica/imunologia , Encéfalo/imunologia , Células Matadoras Naturais/imunologia , Baço/imunologia , Idoso , Animais , Isquemia Encefálica/complicações , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Infecções/etiologia , Infecções/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase em Tempo Real , Transcriptoma
16.
J Racial Ethn Health Disparities ; 2(3): 395-402, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26863468

RESUMO

BACKGROUND: Although the population of diverse applicants applying to medical school has increased over recent years (AAMC Diversity in Medical Education: Facts and Figures 2012); efforts persist to ensure the continuance of this increasing trend. Mentoring students at an early age may be an effective method by which to accomplish diversity within the applicant pool. Having a diverse physician population is more likely able to adequately address the healthcare needs of our diverse population. PURPOSE: The purpose of this study is to initiate a pipeline program, called the Medical Student Mentorship Program (MSMP), designed to specifically target high school students from lower economic status, ethnic, or racial underrepresented populations. High school students were paired with medical students, who served as primary mentors to facilitate exposure to processes involved in preparing and training for careers in medicine and other healthcare-related fields as well as research. METHODS: Mentors were solicited from first and second year medical students at the University of Arizona College of Medicine-Phoenix (UACOM-P). Two separate cohorts of mentees were selected based on an application process from a local high school for the school years 2010-2011 and 2011-2012. Anonymous mentee and mentor surveys were used to evaluate the success of the MSMP. RESULTS: A total of 16 pairs of mentees and mentors in the 2010-2011 (Group 1) and 2011-2012 (Group 2) studies participated in MSMP. High school students reported that they were more likely to apply to medical school after participating in the program. Mentees also reported that they received a significant amount of support, helpful information, and guidance from their medical student mentors. Overall, feedback from mentees and mentors was positive and they reported that their participation was rewarding. Mentees were contacted 2 to 3 years post MSMP participation as sophomores or juniors in college, and all reported that they were on a pre-healthcare career track. CONCLUSION: The MSMP may serve as an effective pipeline program to promote future diversity in college and graduate training programs for future careers in science and medicine.


Assuntos
Escolha da Profissão , Ocupações em Saúde , Mentores , Desenvolvimento de Programas , Instituições Acadêmicas/organização & administração , Estudantes de Medicina , Estudantes/psicologia , Adolescente , Arizona , Estudos de Coortes , Diversidade Cultural , Feminino , Previsões , Ocupações em Saúde/tendências , Humanos , Masculino , Inovação Organizacional , Pobreza , Avaliação de Programas e Projetos de Saúde , Estudantes/estatística & dados numéricos
17.
Am J Physiol Regul Integr Comp Physiol ; 305(1): R24-30, 2013 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-23637134

RESUMO

This study evaluated the activity and content of cyclooxygenase (COX)-1 and -2 in response to acute resistance exercise (RE) in human skeletal muscle. Previous work suggests that COX-1, but not COX-2, is the primary COX isoform elevated with resistance exercise in human skeletal muscle. COX activity, however, has not been assessed after resistance exercise in humans. It was hypothesized that RE would increase COX-1 but not COX-2 activity. Muscle biopsies were taken from the vastus lateralis of nine young men (25 ± 1 yr) at baseline (preexercise), 4, and 24 h after a single bout of knee extensor RE (three sets of 10 repetitions at 70% of maximum). Tissue lysate was assayed for COX-1 and COX-2 activity. COX-1 and COX-2 protein levels were measured via Western blot analysis. COX-1 activity increased at 4 h (P < 0.05) compared with preexercise, but returned to baseline at 24 h (PRE: 60 ± 10, 4 h: 106 ± 22, 24 h: 72 ± 8 nmol PGH2·g total protein(-1)·min(-1)). COX-2 activity was elevated at 4 and 24 h after RE (P < 0.05, PRE: 51 ± 7, 4 h: 100 ± 19, 24 h: 98 ± 14 nmol PGH2·g total protein(-1)·min(-1)). The protein level of COX-1 was not altered (P > 0.05) with acute RE. In contrast, COX-2 protein levels were nearly 3-fold greater (P > 0.05) at 4 h and 5-fold greater (P = 0.06) at 24 h, compared with preexercise. In conclusion, COX-1 activity increases transiently with exercise independent of COX-1 protein levels. In contrast, both COX-2 activity and protein levels were elevated with exercise, and this elevation persisted to at least 24 h after RE.


Assuntos
Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Exercício Físico/fisiologia , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Treinamento Resistido , Adaptação Fisiológica/fisiologia , Adulto , Biópsia , Humanos , Masculino , Músculo Esquelético/patologia , Isoformas de Proteínas/metabolismo , Fatores de Tempo , Regulação para Cima/fisiologia
18.
Pflugers Arch ; 465(5): 627-42, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23605065

RESUMO

Sex steroids are commonly known for their contribution to phenotypic as well as biological reproductive sex differences mediated through classical regulation of neuroendocrine loops. However, sex steroids also have considerable impact on physiological function of non-reproductive tissues including the cerebrovasculature. Preclinical studies have shown that endogenous and exogenous administration of sex steroids significantly influences both cerebrovascular tone and brain function under normal conditions and following a pathological insult (e.g., middle cerebral artery occlusion). However, the precise mechanism(s) of how sex steroids modulate vasomotor responses and/or neurological outcomes in vivo is difficult to define since evidence based on both clinical and experimental studies has been shown to be dependent upon several variables including dose, duration of administration, presence of underlying pathologies, species, and sex. While progesterone, testosterone (TEST), and dihydrotestosterone (DHT) have all been investigated for their impact on the cerebral circulation, the effects of 17ß-estradiol (E2) have been best characterized. Since recent reviews have highlighted studies reporting the actions of E2 on cerebral vascular function and health, only key points are included in this review. Conversely, less is known about the effect of androgens on the blood vessel wall, particularly in the cerebral circulation. The few studies that do address a role for androgen's modulation of cerebrovascular function under normal and pathophysiological conditions provide confounding evidence for either beneficial or detrimental effects. Therefore, the focus of this review is to highlight mechanisms associated with TEST, DHT, and its recently recognized androgen metabolite (3ß-diol) on cerebrovascular function during healthy and diseased states.


Assuntos
Androgênios/metabolismo , Encéfalo/irrigação sanguínea , Transtornos Cerebrovasculares/metabolismo , Animais , Encéfalo/metabolismo , Transtornos Cerebrovasculares/fisiopatologia , Feminino , Humanos , Masculino , Caracteres Sexuais
19.
Endocrinology ; 153(12): 5949-60, 2012 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23117931

RESUMO

P-glycoprotein (Pgp), a multiple drug resistance transporter expressed by vascular endothelial cells, is a key component of the blood-brain barrier and has been shown to increase after inflammation. The nonaromatizable androgen, dihydrotestosterone (DHT), decreases inflammatory markers in vascular smooth muscle cells, independent of androgen receptor (AR) stimulation. The principal metabolite of DHT, 5α-androstane-3ß,17ß-diol (3ß-diol), activates estrogen receptor (ER)ß and similarly decreases inflammatory markers in vascular cells. Therefore, we tested the hypothesis that either DHT or 3ß-diol decrease cytokine-induced proinflammatory mediators, vascular cell adhesion molecule-1 (VCAM-1) and cyclooxygenase-2 (COX-2), to regulate Pgp expression in male primary human brain microvascular endothelial cells (HBMECs). Using RT-qPCR, the mRNAs for AR, ERα, and ERß and steroid metabolizing enzymes necessary for DHT conversion to 3ß-diol were detected in male HBMECs demonstrating that the enzymes and receptors for production of and responsiveness to 3ß-diol are present. Western analysis showed that 3ß-diol reduced COX-2 and Pgp expression; the effect on Pgp was inhibited by the ER antagonist, ICI-182,780. IL-1ß-caused an increase in COX-2 and VCAM-1 that was reduced by either DHT or 3ß-diol. 3ß-diol also decreased cytokine-induced Pgp expression. ICI-182,780 blocked the effect of 3ß-diol on COX-2 and VCAM-1, but not Pgp expression. Therefore, in cytokine-stimulated male HBMECs, the effect of 3ß-diol on proinflammatory mediator expression is ER dependent, whereas its effect on Pgp expression is ER independent. These studies suggest a novel role of 3ß-diol in regulating blood-brain barrier function and support the concept that 3ß-diol can be protective against proinflammatory mediator stimulation.


Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/biossíntese , Androstano-3,17-diol/metabolismo , Encéfalo/metabolismo , Ciclo-Oxigenase 2/metabolismo , Regulação da Expressão Gênica , Molécula 1 de Adesão de Célula Vascular/biossíntese , Androstano-3,17-diol/farmacologia , Barreira Hematoencefálica , Células Cultivadas , Di-Hidrotestosterona/farmacologia , Receptor beta de Estrogênio/metabolismo , Humanos , Inflamação , Masculino , RNA Mensageiro/metabolismo
20.
Steroids ; 77(8-9): 835-44, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22542504

RESUMO

Androgens may provide protective effects in the vasculature under pathophysiological conditions. Our past studies have shown that dihydrotestosterone (DHT) decreases expression of cyclooxygenase-2 (COX-2) during cytokine, endotoxin, or hypoxic stimulation in human vascular smooth muscle cells, in an androgen receptor (AR)-independent fashion. Classically DHT is regarded as a pure AR agonist; however, it can be endogenously metabolized to 5α-androstane-3ß, 17ß-diol (3ß-diol), which has recently been shown to be a selective estrogen receptor (ERß) agonist. Therefore, we hypothesized that DHT's anti-inflammatory properties following cytokine stimulation are mediated through ERß. Using primary human brain vascular smooth muscle cells (HBVSMC), we tested whether DHT's effect on IL-1ß induced COX-2 expression was mediated via AR or ERß. The metabolism of DHT to 3ß-diol is a viable pathway in HBVSMC since mRNA for enzymes necessary for the synthesis and metabolism of 3ß-diol [3alpha-hydroxysteroid dehydrogenase (HSD), 3ß-HSD, 17ß-HSD, CYP7B1] was detected. In addition, the expression of AR, ERα, and ERß mRNA was detected. When applied to HBVSMC, DHT (10nM; 18 h) attenuated IL-1ß-induced increases in COX-2 protein expression. The AR antagonist bicalutamide did not block DHT's ability to reduce COX-2. Both the non-selective estrogen receptor antagonist ICI 182,780 (1 µM) and the selective ERß antagonist PHTPP (1 µM) inhibited the effect of DHT, suggesting that DHT actions are ERß-mediated. In HBVSMC and in rat mesenteric arteries, 3ß-diol, similar to DHT, reduced cytokine-induced COX-2 levels. In conclusion, DHT appears to be protective against the progression of vascular inflammation through metabolism to 3ß-diol and activation of ERß.


Assuntos
Encéfalo/citologia , Ciclo-Oxigenase 2/metabolismo , Citocinas/farmacologia , Receptor beta de Estrogênio/metabolismo , Artérias Mesentéricas/efeitos dos fármacos , Artérias Mesentéricas/metabolismo , Músculo Liso Vascular/citologia , Androgênios/farmacologia , Animais , Western Blotting , Células Cultivadas , Receptor beta de Estrogênio/genética , Humanos , Técnicas In Vitro , Masculino , Músculo Liso Vascular/efeitos dos fármacos , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase em Tempo Real , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...