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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
81

Prostaglandin E2 in Brain-mediated Illness Responses

Elander, Louise January 2010 (has links)
We are unceasingly exposed to potentially harmful microorganisms. The battle against threatening infectious agents includes activation of both the innate and of the adaptive immune systems. Illness responses are elicited and include inflammation, fever, decreased appetite, lethargy and increased sensitivity to painful stimuli in order to defeat invaders. While many of these signs of disease are controlled by the central nervous system, it has remained an enigma how signals from the peripheral immune system reach the brain through its blood-brain barrier, which precludes macromolecules, including cytokines, from diffusing into the brain parenchyma. Previous findings indicate the existence of a pathway across the blood-brain barrier, which includes binding of the cytokine interleukin-1 (IL-1) to its receptor in the brain vessels, thereby inducing the production of the prostaglandin E2 (PGE2) synthesizing enzymes cyclooxygenase-2 (Cox-2) and microsomal prostaglandin E synthase-1 (mPGES-1), which ultimately synthesize PGE2. PGE2 subsequently binds to any of the four prostaglandin E2 (EP) -receptors. Previous results from our laboratory have suggested that this pathway plays a critical role in the febrile response to infectious stimuli. The present thesis aims at further investigating the molecular events underlying immune-to-brain signalling, with special emphasis on fever, hypothalamic-pituitary-adrenal (HPA) -axis activation and anorexia and their connection to signalling molecules of the cytokine and prostaglandin families, respectively. In paper I, the molecular processes linking the proinflammatory cytokine interleukin-6 (IL-6) and PGE2 in the febrile response were investigated. Both IL-6 and PGE2 have been shown to be critical players in the febrile response, although the molecular connections are not known, i.e. if IL-6 exerts its effects up- or downstream of PGE2. Mice deficient in IL-6 were unable to respond to bacterial lipopolysaccharide (LPS) with a febrile response, but displayed similar induction of Cox-2 and mPGES-1, and similar concentrations of PGE2 in the cerebrospinal fluid as wild-type mice. Paradoxically, the IL-6 deficient mice responded with a dose-dependent elevation of body temperature in response to intracerebroventricularly injected PGE2. Furthermore, IL-6 per se was not pyrogenic when injected peripherally in mice, and did not cause increased levels of PGE2 in cerebrospinal fluid. IL-6 deficient mice were not refractory to the action of PGE2 because of excess production of some hypothermia-producing factor, since administration of a Cox-2 inhibitor in LPS-challenged IL-6 deficient mice did not unmask any hypothermic response, and neutralization of tumor necrosis factor α (TNFα), associated with hypothermia, did not produce fever in LPS-challenged IL-6 deficient mice. These data indicate that IL-6 rather than exerting its effects up- or down-stream of PGE2 affects some process in parallel to PGE2, perhaps by influencing the diffusion and binding of PGE2 onto its target neurons. In papers II and III, we injected the proinflammatory cytokine IL-1β in free-fed wild-type mice, in mice with a deletion of the gene encoding mPGES-1, or in mice deficient in the EP1, EP2 and EP3. Food intake was continuously measured during their active period, revealing that mPGES-1 deficient mice were almost completely resistant to anorexia induced by IL-1β. However, all of the investigated EP receptor deficient mice exhibited a normal profound anorexic response to IL-1β challenge, suggesting that the EP4 is the critical receptor that mediates IL-1β-induced anorexia. We also investigated the role of mPGES-1 in anorexia induced by lipopolysaccharide (LPS) in mPGES-1 deficient mice. The profound anorexic response after LPS-challenge was similar in mPGES-1 deficient and wild-type mice. To further investigate the anorectic behaviour after LPS injection, we pre-starved the animals for 22 hours before injecting them with LPS. In this paradigm, the anorexia was less profound in mPGES-1 knock-out mice. Our results suggest that while the inflammatory anorexia elicited by peripheral IL-1β seems largely to be dependent on mPGES-1-mediated PGE2 synthesis, similar to the febrile response, the LPS-induced anorexia is independent of this mechanism in free-fed mice but not in pre-starved animals. In papers IV and V, the role of prostanoids for the immune-induced HPA-axis response was investigated in mice after genetic deletion or pharmacological inhibition of prostanoid-synthesizing enzymes, including Cox-1, Cox-2, and mPGES-1. The immediate LPS-induced release of ACTH (adrenocorticotropic hormone and corticosteroids was critically dependent on Cox-1 derived prostanoids and occurred independently of Cox-2 and mPGES-1 derived PGE2. In contrast, the delayed HPA-axis response was critically dependent on immune-induced PGE2, synthesized by Cox-2 and mPGES-1, and occurred independently of Cox-1 derived enzymes. In addition, in the mPGES-1 deficient mice, the synthesis of CRH hnRNA and mRNA was decreased in the paraventricular nucleus of the hypothalamus after LPS-challenge, indicating that the delayed hormone secretion was mediated by PGE2-induced gene-transcription of CRH in the hypothalamus. The expression of the c-fos gene and Fos protein, an index of synaptic activation, was maintained in the paraventricular nucleus and its brainstem afferents both after unselective and Cox-2 selective inhibition as well as in Cox-1, Cox-2, and mPGES-1 knock-out mice. This suggests that the immune-induced neuronal activation of autonomic relay nuclei occurs independently of prostanoid synthesis and that it is insufficient for eliciting stress hormone release.
82

Prostaglandine E2 et mesures du flux mésentérique par Doppler à la suite d’un traitement du canal artériel à l’ibuprofène par voie intraveineuse et entérale chez les bébés prématurés

Dorval, Véronique G 08 1900 (has links)
En dépit du nombre croissant d’études cliniques sur le canal artériel (CA), des failles méthodologiques entretiennent plusieurs incertitudes concernant l’efficacité et la sécurité des traitements chez les bébés nés prématurés. L’objectif de cette recherche était de comparer les concentrations de prostaglandine E2 (PGE2) et les mesures du flux mésentérique par échographie Doppler chez les enfants nés prématurément et ayant un canal artériel traité à l’ibuprofène par voie intraveineuse ou entérale, en utilisant la méthodologie randomisée contrôlée et à double insu. Dans notre étude pilote, 20 nouveau-nés prématurés de moins de 34 semaines ayant un CA symptomatique confirmé par échocardiographie, furent randomisés au traitement à l’ibuprofène par voie intraveineuse ou entérale. La voie d’administration fut maintenue à l’insu de l’équipe traitante, des cardiologues et des investigateurs. Des dosages des prostaglandines plasmatiques ont été mesurés avant le début du traitement ainsi que 3, 24 et 48 h après le début du traitement. Les mesures du flux mésentérique ont été effectuées avant le traitement à l’ibuprofène ainsi que 1 h et 3 h après le traitement. Nous avons démontré à partir de nos observations que les niveaux plasmatiques de prostaglandines E2 diminuent chez les patients ayant répondu au traitement à l’ibuprofène, indépendamment de la voie d’administration. Nous n’avons pas observé de changement dans l’évolution des dosages de PGE2 chez les patients qui n’ont pas répondu au traitement. Les paramètres mesurés par échographie Doppler au niveau de l’artère mésentérique supérieure n’étaient pas affectés par la voie d’administration du traitement à l’ibuprofène, intraveineuse ou entérale. La présente étude suggère ainsi que le traitement du CA par ibuprofène intraveineux ou entéral n’influe pas sur le flux sanguin mesuré par échographie Doppler. La baisse de la prostaglandine E2 coïncide avec la fermeture du CA, et son dosage pourrait jouer un rôle dans la gestion du traitement. Nous avons démontré la faisabilité d’une étude clinique randomisée à double insu dans le traitement du canal artériel; une méthodologie qui devrait désormait être employé dans la recherche clinique sur les traitements de la persistance du CA. / Despite the growing body of research on the patent ductus arteriosus (PDA), issues with clinical research methodology impairs much of our understanding regarding treatment efficacy and safety in the preterm population. The purpose of this study was to determine plasma prostaglandin E2 (PGE2) concentrations in preterm infants with symptomatic persistence of the ductus arteriosus treated with IV and oral ibuprofen, and measure Doppler flow parameters in the superior mesenteric artery, utilizing randomized controlled and double-blind methodology. Twenty patients age < 34 wks with a symptomatic PDA confirmed by echocardiography randomized to oral vs intravenous ibuprofen regimen. Treating physician, cardiologists and study investigators were blinded to treatment allocation. Plasma PGE2 levels were measured prior to ibuprofen treatment and at 3, 24 and 48 h after treatment. Mesenteric Doppler measurements were taken prior to ibuprofen treatment, and 1 h and 3 h after treatment. Our results showed that plasma PGE2 levels decreased over time in patients that exhibited ductal closure after IV or oral ibuprofen treatment; no time-dependent changes in PGE2 were seen in subjects that failed to respond to ibuprofen. Superior mesenteric artery Doppler flow measurements were not affected by ibuprofen treatment (IV or oral), regardless of efficacy on ductal closure and of PGE2 changes. We conclude that treatment with oral or intravenous ibuprofen does not impact on superior mesenteric artery blood flow measured by Doppler ultrasound. Decreases in plasma PGE2 concentrations coincide with ibuprofen efficacy, and may be more cost-effective to monitor than ultrasound. This study also demonstrated the successful use of double blinded randomized controlled research methodology, which should be more strictly applied in future clinical research on PDA treatment.
83

Effects of a New Conjugate Drug in a Rat Model of Postmenopausal Osteoporosis

Liu, Careesa Chang 04 December 2013 (has links)
Postmenopausal osteoporosis is a disease characterized by bone loss and increased risk of fracture, and represents a significant burden on the Canadian health care system. Current treatments lack the ability to simultaneously address the therapeutic needs for promoting bone formation and inhibiting resorption. Our approach employs a novel conjugate drug in which an anabolic agent (EP4 receptor agonist) is reversibly joined with an anti-resorptive agent (alendronate) through a linker. This allows the bone-targeting ability of alendronate to deliver the EP4 agonist to bone sites, thereby mitigating the side effects associated with systemic administration of the EP4 agonist. This study investigated the in vivo efficacy of this drug in a curative experiment to treat postmenopausal osteoporosis using an ovariectomized rat model. Results showed that conjugate treatment dose-dependently stimulated bone formation and restored ovariectomy-induced bone loss, and conjugation between alendronate and the EP4 agonist was crucial to the drug’s anabolic effect.
84

Effects of a New Conjugate Drug in a Rat Model of Postmenopausal Osteoporosis

Liu, Careesa Chang 04 December 2013 (has links)
Postmenopausal osteoporosis is a disease characterized by bone loss and increased risk of fracture, and represents a significant burden on the Canadian health care system. Current treatments lack the ability to simultaneously address the therapeutic needs for promoting bone formation and inhibiting resorption. Our approach employs a novel conjugate drug in which an anabolic agent (EP4 receptor agonist) is reversibly joined with an anti-resorptive agent (alendronate) through a linker. This allows the bone-targeting ability of alendronate to deliver the EP4 agonist to bone sites, thereby mitigating the side effects associated with systemic administration of the EP4 agonist. This study investigated the in vivo efficacy of this drug in a curative experiment to treat postmenopausal osteoporosis using an ovariectomized rat model. Results showed that conjugate treatment dose-dependently stimulated bone formation and restored ovariectomy-induced bone loss, and conjugation between alendronate and the EP4 agonist was crucial to the drug’s anabolic effect.
85

Aspectos imunopatogênicos da leishmaniose cutânea difusa: fatores da leishmania e do hospedeiro

Costa, Jaqueline França January 2013 (has links)
Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2013-11-06T14:31:55Z No. of bitstreams: 1 Jaqueline Franca Costa Aspectos imunopatogênicos da leishmaniose cutanêa...pdf: 1509776 bytes, checksum: 2d3824cc711f84d908e61378ac3d4a8c (MD5) / Made available in DSpace on 2013-11-06T14:31:55Z (GMT). No. of bitstreams: 1 Jaqueline Franca Costa Aspectos imunopatogênicos da leishmaniose cutanêa...pdf: 1509776 bytes, checksum: 2d3824cc711f84d908e61378ac3d4a8c (MD5) Previous issue date: 2013 / Universidade Federal da Bahia. Faculdade de Medicina da Bahia. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil / A progressão crônica da LCD é atribuída à falta da imunidade mediada por células específica para antígeno de Leishmania e predominância de uma resposta do tipo Th2. Neste sentido, tanto fatores do parasita quanto do hospedeiro podem atuar na desativação da resposta imune favorecendo a replicação da Leishmania. Inicialmente avaliamos o papel da exposição de fosfatidilserina na infecção de macrófagos murinos com Leishmania amazonensis isolados de pacientes com LCD. Para isso, macrófagos peritoneais de camundongos F1(BALB/c x C57BL/6) foram infectados com os diferentes isolados obtidos de pacientes com LCD e LCL. Os isolados obtidos de pacientes com LCD apresentaram maior expressão de PS do que os isolados de pacientes com LCL após 24 horas de infecção. Em seguida, avaliamos a infectividade dos diferentes isolados. As amastigotas de pacientes com LCD apresentaram maior porcentagem de macrófagos infectados e índice de infecção, quando comparados com amastigotas de pacientes com LCL. Quanto ao mecanismo, o grupo infectado com os isolados de pacientes com LCD apresentou um aumento na relação TGF-β/TNF-α e IL-10/TNF-α em relação ao grupo LCL. A análise de correlação revelou que a porcentagem de macrófagos infectados, o índice de infecção, os índices de TGF-β/TNF-α e IL-10/TNF-α, bem como o tamanho dos vacúolos estão diretamente associados a maior exposição de PS. Além disso, o número de lesões e o tempo de doença dos pacientes com LCD também estão associados á exposição de PS. O reconhecimento de PS tem como consequência a produção de TGF, IL-10, IL-4 e PGE2, que ativam a via da enzima arginase e consequentemente a produção de poliaminas. Por isso buscamos investigar a participação de tais mediadores em pacientes com LCD. Os níveis da arginase I, ODC e TGF-β no plasma de pacientes com LCD estava elevados quando comparado com os pacientes com LCL ou o controle saudável da área endêmica. Por outro lado, os níveis de TNF-α, IL-12, MCP-1 e CXCL-10 estavam reduzidos no plasma de pacientes com LCD comparado aos pacientes com LCL. Os níveis de arginase apresentaram correlação positiva com ODC, TGF-β e PGE e correlação negativa com TNF-α, IL-12, MCP-1 e CXCL-10. A produção da arginase e ODC também foi avaliada nas lesões dos pacientes através de imunohistoquímica. As lesões dos pacientes com LCD apresentaram uma marcação mais intensa e difusa do que as de LCL. Além disso, a expressão da cicloxigenase 2 também estava aumentada nas lesões de LCD. A expressão do mRNA das enzimas fosfolipase A2, COX-2, prostaglandina sintase, espermina e espermidina sintase apresentaram uma relação positiva com a enzima arginase, indicando que esta interfere diretamente no metabolismo dos mediadores lipídicos e na via de síntese das poliaminas. A inibição das enzimas arginase e ODC com nor-NOHA e DFMO, respectivamente, reduziu a carga parasitária de macrófagos humanos infectados com L. amazonensis após 72 h de infecção. Além disso, os inibidores reduziram a produção de TGF e PGE2 no sobrenadante das culturas. Em conjunto, nossos dados sugerem que a liberação local e sistêmica de prostaglandinas e poliaminas associadas à via da arginase em pacientes com LCD deve estar associada com a inabilidade em montar uma resposta imune eficiente contra a infecção por Leishmania proporcionando um ambiente favorável para a replicação do parasita e disseminação da doença. Nossos resultados mostram também que este ambiente imunossuprimido pode ser induzido pela exposição de PS na superfície de L. amazonensis deflagrando uma resposta anti-inflamatória nos pacientes com LCD. / The chronic progression of DCL is attributed to the lack of specific cell-mediated immunity to Leishmania antigen and predominance of a Th2-type response. In this sense, both factors of the parasite and the host can act in the deactivation of immune response, favoring parasite replication. Initially we evaluate the role of phosphatidylserine exposure in murine macrophages infected with L. amazonensis isolated from patients with DCL. First, peritoneal macrophages of mice F1 (BALB/c x C57BL/6) were infected with different isolates from patients with DCL and LCL. The DCL isolates showed higher PS expression than the LCL isolates after 24 hours of infection.. The DCL-amastigotes patients showed a higher percentage of infected macrophages and the infectivity index when compared with patients with LCL- amastigotes. Regarding the mechanism, the group infected with isolates from patients with LCD showed an increase in TGF/TNF and IL-10/TNF when compared with LCL group. Correlation analysis revealed that the percentage of infected macrophages, the infectivity index, the rate of TGF/TNF and IL-10/TNF as well as the size of the vacuoles are directly associated with higher PS exposure. Moreover, the number of lesions and disease duration of DCL patients are also associated with PS exposure. Recognition of PS results in the production of TGF, IL-10, IL-4 and PGE2, molecules with anti-inflammatory role that activate the enzyme arginase and consequently the polyamines production. Therefore, we investigated the involvement of these mediators in patients with DCL. The plasma of DCL patients showed high levels of arginase, ODC and TGF compared to the LCL patients or healthy control from endemic area. On the other hand, the levels of TNF, IL-12, MCP-1 and CXCL-10 were reduced in the DCL patients plasma compared to patients with LCL. Arginase levels were positively correlated with ODC, TGF and PGE and negatively correlated with TNF, IL-12, MCP-1 and CXCL-10. The production of arginase and ODC was also evaluated in the lesions of patients by immunohistochemistry. The DCL lesions showed a more intense and diffuse staining than LCL lesions. Furthermore, the expression of cyclooxygenase-2 was also increased in lesions of DCL. The mRNA expression of the enzymes phospholipase A2, COX-2, prostaglandin synthase, spermine synthase and spermidine synthase showed a positive relationship with the arginase enzyme, indicating that it directly interferes with the metabolism of lipid mediators and in synthesis of polyamines. The inhibition of the enzyme arginase and ODC with nor-NOHA and DFMO, respectively, reduced the parasite load of L. amazonensis human infected macrophages 72 h after infection. Moreover, NOHA and DFMO reduced TGF and PGE2 production in the supernatant of cultures. Together, local and systemic release of prostaglandins, arginase and polyamines pathways in DCL should be associated with the inability of these patients to mount effective immune response against infection by Leishmania providing a favorable environment for replication and spread of the parasite disease. Our results also show that this immunosuppressed environment can be induced by PS exposure on the L. amazonensis surface triggering anti-inflammatory response in DCL patients.
86

O papel da interleucina-1'beta' produzida no gânglio da raiz dorsal no desenvolvimento da hiperalgesia inflamatória / The role of dorsal root ganglion-produced interleukin-1'beta' in development of inflammatory hyperalgesia

Araldi, Dionéia, 1982- 20 August 2018 (has links)
Orientador: Carlos Amilcar Parada / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-20T20:52:22Z (GMT). No. of bitstreams: 1 Araldi_Dioneia_D.pdf: 2812057 bytes, checksum: ba0136e00ebc1ab271d4e1ed908bcd8c (MD5) Previous issue date: 2012 / Resumo: A liberação de Interleucina-1? (IL-1?) no tecido periférico estimula a síntese de Prostaglandinas (PGs), especialmente, da Prostaglandina-E2 (PGE2), que leva a sensibilização dos nociceptores aferentes primários induzindo a hiperalgesia inflamatória. Recentemente demonstramos que a IL-1? pode ativar diretamente o receptor de Interleucina-1 (IL-1R) do nociceptor aferente periférico e levar a liberação de PGE2 associada ao desenvolvimento da hiperalgesia. A IL-1? também é liberada no Gânglio da Raiz Dorsal (GRD), entretanto a função que a IL-1? desempenha no GRD para o desenvolvimento da hiperalgesia inflamatória ainda não está clara. Portanto, o objetivo deste estudo foi investigar se a liberação de IL-1? e a ativação do Receptor de Interleucina-1 Tipo I (IL-1RI) no GRD estão envolvidos no desenvolvimento da hiperalgesia inflamatória. A administração de IL-1Ra (antagonista natural de receptor IL- 1, 6 ?g) no GRD de ratos preveniu a hiperalgesia mecânica (avaliada por meio do von Frey Eletrônico) induzida pela administração intraplantar (i.pl) de Adjuvante Completo de Freund (CFA, 100 ?L), Carragenina (Cg, 100 ?g) ou IL-1? (0,5 pg), mas não pela administração i.pl de PGE2 (100 ng), avaliadas 3 horas após suas administrações. Além disso, a administração i.pl periférica de CFA ou Cg aumentaram as concentrações de IL-1? (avaliadas por ELISA) no GRD. O tratamento ganglionar (GRD-L5) com oligonucleotídeo (ODN) antisense contra IL-1RI (30 ?g/dia durante 4 dias) reduziu de maneira significativa a expressão de IL-1RI no GRD-L5 e a hiperalgesia mecânica induzida por CFA, Cg e IL-1?, mas não pela PGE2, administradas no tecido periférico da pata. Também verificamos a hipótese de que a prévia ativação do receptor neuronal, IL-1RI, no tecido periférico é importante para a liberação de IL-1? no GRD e para a subsequente hiperalgesia induzida por PGE2. A IL-1? (0,5 pg/pata) co-administrada com a dose sub-limiar de PGE2 (10 ng/pata) em patas traseiras tratadas com indometacina induziu uma proeminente hiperalgesia, que foi prevenida pelo prétratamento com ODN antisense contra IL-1RI ou IL-1Ra (6 ?g) administrados no GRD. Além disso, o IL-1Ra reduziu a expressão de COX-2 em células do GRD. Para confirmar a ativação do IL-1RI em células do GRD, administramos Cg ou CFA no tecido periférico o que levou ao aumentou da expressão de IRAK-1 e IRAK-4 em células do GRD. Os resultados deste estudo sugerem que o desenvolvimento da hiperalgesia inflamatória depende da ativação do receptor IL-1RI neuronal no tecido periférico que, em partes, induz a liberação de IL-1? no GRD e subsequente ativação da COX-2. Os dados aqui apresentados oferecem novas perpectivas sobre a participação das células do GRD nos mecanismos envolvidos na hiperalgesia inflamatória e revelam novos e interessantes alvos para o controle das hiperalgesias inflamatórias / Abstract: The release of Interleukin-1? (IL-1?) in the peripheral tissue stimulates the synthesis of Prostaglandins, specially, Prostaglandin-E2 (PGE2) that ultimately sensitize the peripheral afferent nociceptor inducing inflammatory hyperalgesia. We have recently demonstrated that IL-1? can directly activate IL-1R receptor of peripheral afferent nociceptor to induce release of PGE2 associated to development of hyperalgesia. IL-1? is also released in Dorsal Root Ganglion (DRG), however the role that IL-1? in DRG plays to development of inflammatory hyperalgesia is not yet elucidated. Therefore, the aim of this study was to investigate whether IL-1? released in the DRG and the activation of Interleukin-1 Receptor Type I (IL-1RI) is involved in the development of the inflammatory hyperalgesia. Administration of IL-1Ra (IL-1 receptor antagonist, 6 ?g) in the DRG of rats prevented the mechanical hyperalgesia (measured with Electronic von Frey) induced by intraplantar (i.pl) administration of Complete Freund's Adjuvant (CFA, 100 ?L), Carrageenan (Cg, 100 ?g) or IL- 1? (0.5 pg), but not by PGE2 (100 ng), measured 3 hours after their administrations. Also, peripheral i.pl administration of CFA or Cg induced an increase in IL-1? concentrations (measured by ELISA) in the DRG. Ganglionar (DRG-L5) treatment with oligonucleotides (ODN) antisense against IL-1RI (30 ?g/day for four days) reduced the expression of IL-1RI in the DRG-L5 and the mechanical hyperalgesia induced by CFA, Cg, and IL-1?, but not by PGE2 administered in peripheral tissue. We also verified the hypothesis that previous activation of neuronal IL-1RI in the peripheral tissue is important to the release of IL-1? in the DRG and to the subsequent PGE2-induced hyperalgesia. IL-1? (0.5 pg/paw) co-administrated with a sub-threshold dose of PGE2 (10 ng/paw), in hind paws treated with indomethacin, greatly induces hyperalgesia, which was prevented by pre-treatment with ODN antisense against IL-1RI or IL-1Ra (6 ?g) administrated in DRG. Also, IL-1Ra administrations reduced the COX-2 expression in DRG cells. To confirm IL-1RI activation in DRG cells, it was observed that IRAK-1 and IRAK-4 expression was increased in DRG neurons after administration of Cg or CFA in the peripheral tissue. These findings suggest that the development of inflammatory hyperalgesia depends on neuronal IL-RI activation in the peripheral tissue that, in turn, induces the release of IL- 1? in the DRG and subsequent COX-2 activation. These data provide new insights about the participation of DRG cells in the mechanisms underlying inflammatory hyperalgesia and reveal new interesting targets to control inflammatory hyperalgesia / Doutorado / Fisiologia / Doutor em Biologia Funcional e Molecular
87

Syntéza polysubstituovaných pyrimidinů s potenciálními protizánětlivými vlastnostmi / Synthesis of polysubstituted pyrimidines with potential anti-inflammatory properties

Kalčic, Filip January 2017 (has links)
This thesis is engaged in the synthesis of polysubstituted pyrimidines with anti- inflammatory properties. Such molecules can inhibit production of prostaglandin E2 (PGE2). The aim of this study was to enhance water-solubility and anti-inflammatory efficacy of such derivatives via structural modifications of the lead scaffold. Among applied synthetic tools, the Suzuki-Miyaura cross-coupling was the prevalent reaction, however, many other synthetic procedures (Heck reaction, condensation, borylation, ozonolysis, nucleophilic substitution, etc.) were utilized as well. Overall, 43 final products were prepared. The anti-inflammatory efficacy (inhibition of PGE2 production) was successfully increased as the most potent compound achieved three orders of magnitude higher activity compared to the current lead structure WQE-134. Furthermore, no general influence of the length of the substituent in the C5 position of pyrimidine (C5pyr) on the anti-inflammatory efficacy of synthesized compounds was observed. Significant bioavailability obstacle in future development of the current lead WQE-134 is its poor solubility which was successfully enhanced by introduction of heteroatom bearing moieties to C5pyr. The most water-soluble compound achieved two orders of magnitude higher solubility than WQE-134 while...
88

Differential effects of arachidonic acid and docosahexaenoic acid on cell biology and osteoprotegerin synthesis in osteoblast-like cells

Coetzee, Magdalena 09 March 2006 (has links)
The purpose of the study was to elucidate the mechanisms by which polyunsaturated fatty acids (PUFAs) prevent bone loss. MG-63 human osteoblasts and MC3T3-E1 murine osteoblasts were exposed to the n-6 PUFA arachidonic acid (AA) and the n-3 PUFA docosahexaenoic acid (DHA) as well as oestrogen (E2) and parathyroid hormone (PTH) and the effects thereof tested on a variety of biological parameters characteristic of osteoblasts. These parameters included prostaglandin E2 (PGE2) synthesis, proliferation, differentiation to mature mineralising osteoblasts as well as osteoprotegerin (OPG) and receptor activator of nuclear factor êB ligand (RANKL) secretion. Results showed that AA stimulates PGE2 production significantly in both cell lines. Stimulated PGE2 production by MC3T3-E1 cells however, was significantly higher, which might be attributed to auto-amplification by PGE2 itself in this cell line. Pre-incubation of the MG-63 cells with cyclo-oxygenase (COX)-blockers inhibited PGE2 production significantly, suggesting that both COX enzymes were involved in PGE2 synthesis. The number of functional osteoblasts is important for bone formation therefore in vitro osteoblastic cell proliferation was investigated. In contrast to the hormones E2 and PTH, both AA and DHA inhibited proliferation significantly. The AA-mediated anti-proliferative effect is possibly independent of PGE2 production, as PGE2 per se had little effect on proliferation. DHA inhibited proliferation of MG-63 cells more severely, which might be attributed to the osteosarcoma nature of the MG-63 cells. The anti-proliferative effect of these PUFAs might be attributed to modulation of cell cycle progression or anti-mitotic effects of PUFA peroxidation products. Morphological studies showed apoptotic cells after DHA exposure in MG-63 cells. There is a reciprocal relationship between reduced proliferation and the subsequent induction of cell differentiation in vitro. High basal levels of alkaline phosphatase (ALP) activity, a marker of the mature mineralising osteoblastic phenotype, were detected in MC3T3-E1 cells. Long-term exposure to AA inhibited ALP activity in these cells. This process might be PGE2-mediated. Exposure to PUFAs, however, did not compromise the ability of the MC3T3-E1 cells to differentiate to mature mineralising osteoblasts. In contrast with MC3T3-E1 cells, MG-63 cells demonstrated low basal ALP activity and were unable to differentiate to mature mineralising osteoblasts. In the absence of osteogenic-inducing supplements, PUFAs induced adipocyte-like features that might be due to the expression of high levels of PPARã in this cell line. Lipid-filled vacuoles were absent in the MC3T3-E1 cells suggesting that the MC3T3-E1 cell line may not express PPARã mRNA. The study furthermore demonstrated that PUFAs are able to modulate OPG and RANKL secretion in osteoblasts. AA inhibited OPG secretion dose-dependently in both cell lines, this could be PGE2-mediated. AA dose-dependently stimulated soluble RANKL (sRANKL) secretion in MC3T3-E1 cells thereby affecting the OPG/RANKL ratio in a negative way, supporting various reports that AA and PGE2 do cause bone resorption. No sRANKL could be detected after exposing the MC3T3-E1 cells to DHA suggesting that DHA could be protective to bone. In conclusion, contrary to in vivo evidence, this in vitro study could not indisputably demonstrate protective effects of PUFAs on the osteoblastic cell lines tested. / Thesis (PhD (Physiology))--University of Pretoria, 2006. / Physiology / unrestricted
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Inhalational cough challenges in the assessment of cough

Khalid, Saifudin January 2013 (has links)
Introduction: Cough is the commonest reason for which medical advice is sought. In assessment of chronic cough and in developing anti-tussive medications, inhalational cough challenges with capsaicin and citric acid are commonly employed. However the ability of these inhalational cough challenges to distinguish health and disease is not clear and it is not known which end point is best in making such assessments. Methods: Subjects belonging to five different categories (healthy volunteers, subjects with COPD, asthma, healthy current smokers and chronic cough) were compared with each another by using the standard cough challenges employing Capsaicin and Citric acid and also by using newer inhalational cough challenge agents such as prostaglandin E2 and bradykinin. In addition adaptation to repeated inhalations of tussive agents was also assessed. The relationship between the cough reflex sensitivity as gauged by using inhalational cough challenge tests and objective cough recording was explored in all five groups. Finally the change in C5 in Capsaicin evoked cough by using a substance to block TRPV1 channel and its effect on objective cough recording was assessed in subjects with chronic cough. Results: Different tussive agents have different abilities to distinguish between different diagnostic categories and a combination of inhalational cough challenge tests have a better accuracy of predicting diagnostic groups as compared to one on its own. There are significant differences in the rates of adaptation to repeated inhalations of PGE2 and there is a significant reduction in cough response over period of time in all disease groups. Using the TRPV1 antagonist resulted in a modest increase in the Log C5 concentration of capsaicin but this was not matched by a change in objective cough recording or CQLQ scores. Conclusions: The different abilities of tussive agents to distinguish between different diagnostic categories suggest that the information conveyed by the one inhalational cough challenge test is different from that by another test. The choice of the inhalational cough challenge test should therefore depend on which groups are included in the study. There was no significant difference in the rate of adaptation to prolonged challenge with citric acid or capsaicin and no significant correlation of the magnitude of adaptation with objective cough recording suggesting that this is unlikely to be responsible for the increased cough rates seen in diseases such as chronic cough, COPD or asthma. The TRPV1 antagonist did not result in a significant change in objective cough recording or CQLQ scores. The change in C5 with the TRPV1 antagonist was however modest and this may be reason for this study failing to show a relationship between these different measures.
90

Der Einfluss von Glykosaminoglykanen auf die Bildung und Freisetzung von Prostaglandin E2

Grahl, Katrin 20 October 2015 (has links)
Diese Arbeit verdeutlicht die Wirkung von Chondroitinsulfat auf die Synthese von Prostaglandin E2 in humanen mesenchymalen Stromazellen in Abhängigkeit ihres Sulfatierungsgrades. MSC zeichnen sich durch ihre antiinflammatorischen Eigenschaften aus und haben damit einen modulierenden Effekt auf Wundheilungsprozesse. Als Vorläuferzellen von Osteoblasten sind sie direkt an der Knochenneubildung beteiligt. Eine persistierende Entzündung hat eine kontinuierliche Freisetzung von Zytokinen, wie IL-1 zur Folge. Es konnte gezeigt werden, dass IL-1 in hMSC zu einer Freisetzung von PGE2 führt. Unter kurzzeitiger Wirkung stimuliert PGE2 die Knochenneubildung. Eine langanhaltende Präsenz leitet dagegen die Bildung des Faktors RANKL, einen die Osteoklastogenese stimulierenden Faktor, ein. Seit langem ist der positive Effekt von Chondroitinsulfat in chronischen Entzündungsprozessen, wie Rheumatoider Arthritis, bekannt. Zudem werden sie in aktuellen Studien als Beschichtungsbestandteile von Knochenimplantaten verwendet. Sie führten hier zu einer besseren Bioinduktivität und Biokonduktivität. Bisher ist dennoch der molekulare Wirkmechanismus nicht genau beschrieben. Die Schwierigkeit besteht darin, dass die molekularen Signalkaskaden für die einzelnen Kulturmoldelle Unterschiede aufweisen und kein ubiquitärer Mechanismus dargestellt wird. In hMSC führte die Stimulation mit IL-1 unter vorheriger Zugabe von Chondroitinsulfat zu einer Reduktion der PGE2 Freisetzung. Der Effekt des hochsulfatierten sCS3 war gegenüber dem nativen C4S verstärkt. Die reduzierende Wirkung von C4S setzte verzögert ein. Es ist bereits bekannt, dass die negative Ladung der CS zu einer Bindung von Zytokinen führt. Dadurch wird eventuell die Konzentration der Zytokine, wie IL-1 im Bereich der Zellrezeptoren erniedrigt und führt zu einer verringerten Stimulation der Zelle. Denkbar ist auch die Beeinflussung der intrazellulären Signaltransduktionskaskade durch die Bindung der CS an einen speziellen, bisher unbekannten, Membranrezeptor. Die entscheidenden Enzyme der PGE2 Synthese sind die Cyclooxygenase-2 (Cox-2) und die mikrosomale Prostaglandin E Synthase 1 (mPGES1). Die mRNA beider Enzyme war unabhängig vom Sulfatierungsgrad der CS reduziert. Dieser Effekt konnte auf Protein-ebene nicht belegt werden. Die produzierte Proteinmenge an mPGES1 wird durch IL-1 induziert, bleibt aber auch durch Zugabe von CS unverändert. Somit kann von einer erhöhten Translationseffizient und mRNA Stabilität der mPGES1 RNA ausgegangen werden. MAPK Kinasen sind entscheidende Schnittstellen bei der Regulation der mRNA Stabilität als auch der Aktivität von Transkriptionsfaktoren. In dieser Studie konnte die MAPK p38 als entscheidendes Enzym bei der Wirkung von CS auf die PGE2 Synthese ermittelt werden. Dabei führten sowohl das natürliche C4S als auch das hochsulfatierte sCS3 zu einer verringerten Aktivierung. Der Transkriptionsfaktor NfkB ist einer von mehreren, die an den Promotorbereichen der beiden induzierbaren PGE2 Enzyme, Cox-2 und mPGES1, binden. Es ist anzunehmen, dass die hier aufgezeigte verringerte Aktivität von NfkB als auch die verhinderte Translokation in den Zellkern eine reduzierte Transkription der jeweiligen mRNA bedingten. Abhängig vom untersuchten Modell und den verwendeten Kulturbedingungen können diese Prozesse moduliert sein. Die Erkenntnisse dieser experimentellen Arbeit liefern einen weiteren wichtigen Baustein zum Verständnis der molekularbiologischen Abläufe während entzündlicher Prozesse. Die Verwendung von Chondroitinsulfat, insbesondere hochsulfatiertes CS, in Kombination mit hMSC kann gezielt zu einer Verringerung der Entzündungsreaktion während der Implantateinheilung führen. Die durch PGE2 hervorgerufenen Symptome, wie erhöhte Gefäßpermeabilität, Schwellung und verstärktes Schmerzempfinden begründen diese positiven Effekte.

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