Orientador: Renée Laufer Amorim / Resumo: O glaucoma é a principal causa global de cegueira irreversível e o número de pessoas com glaucoma em todo o mundo aumentará para 111,8 milhões em 2040, mesmo com os tratamentos vigentes. Deste modo, enseja uma necessidade de desenvolver terapias neuroprotetoras que possam ser usadas para reduzir os efeitos perniciosos do glaucoma, como a morte de células ganglionares da retina (CGR). Avanços na compreensão da fisiopatologia do glaucoma é um fator chave na compreensão da patogênese da neuropatia glaucomatosa. Neste contexto a isquemia retiniana desempenha um papel central em várias doenças da retina. A patogênese da isquemia retiniana envolve alterações temporais da morfologia e morfometria da retina assim como mudanças na expressão gênica e protéica. O Citrato de Sildenafila (SC) mostrou efeito protetor nos modelos de isquemia/reperfusão (I/R) com efeitos neuroprotetores. Contudo, a administração oral de CS, em humanos, encontra inúmeros efeitos colaterais, tais como redução da pressão arterial, dores de cabeça, rubor e congestão nasal são concomitantes, assim como pacientes com moderada a grave doença cardiovascular ou aqueles submetidos a terapia a base de nitrato apresentam riscos secundários aumentados para efeitos cardiovasculares adversos. Deste modo, propomos que a administração tópica de CS pode ser uma alternativa à via oral e também ser igualmente neuroprotetor no glaucoma e podem minimizar os riscos indesejados no uso sistêmico desse fármaco, além de oferecer uma n... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Glaucoma is the leading global cause of irreversible blindness and the number of people with glaucoma worldwide will increase to 111.8 million by 2040, even with current treatments. Thus, there is a need to develop neuroprotective therapies that can be used to reduce the deleterious effects of glaucoma, such as retinal ganglion cell death (RBC). Advances in understanding the pathophysiology of glaucoma are a key factor in understanding the pathogenesis of glaucomatous neuropathy. In this context, retinal ischemia plays a central role in various diseases of the retina. The pathogenesis of retinal ischemia involves temporal changes in retinal morphology and morphometry as well as changes in gene and protein expression. Sildenafil Citrate (SC) showed protective effect in the ischemia / reperfusion (I / R) models with neuroprotective effects. However, oral administration of CS in humans finds numerous side effects such as reduced blood pressure, headaches, flushing and nasal congestion are concomitant, as well as patients with moderate to severe cardiovascular disease or those undergoing nitrate therapy have increased secondary risks for adverse cardiovascular effects. Thus, we propose that the eye drops administration of CS may be an alternative to the oral route and also be equally neuroprotective in glaucoma and may minimize the undesirable risks in the systemic use of this drug, as well as offer a new approach for the therapeutic intervention in the pathogenesis of neuropathy... (Complete abstract click electronic access below) / Doutor

Orientador: Renée Laufer Amorim / Resumo: O glaucoma é a principal causa global de cegueira irreversível e o número de pessoas com glaucoma em todo o mundo aumentará para 111,8 milhões em 2040, mesmo com os tratamentos vigentes. Deste modo, enseja uma necessidade de desenvolver terapias neuroprotetoras que possam ser usadas para reduzir os efeitos perniciosos do glaucoma, como a morte de células ganglionares da retina (CGR). Avanços na compreensão da fisiopatologia do glaucoma é um fator chave na compreensão da patogênese da neuropatia glaucomatosa. Neste contexto a isquemia retiniana desempenha um papel central em várias doenças da retina. A patogênese da isquemia retiniana envolve alterações temporais da morfologia e morfometria da retina assim como mudanças na expressão gênica e protéica. O Citrato de Sildenafila (SC) mostrou efeito protetor nos modelos de isquemia/reperfusão (I/R) com efeitos neuroprotetores. Contudo, a administração oral de CS, em humanos, encontra inúmeros efeitos colaterais, tais como redução da pressão arterial, dores de cabeça, rubor e congestão nasal são concomitantes, / Abstract: Glaucoma is the leading global cause of irreversible blindness and the number of people with glaucoma worldwide will increase to 111.8 million by 2040, even with current treatments. Thus, there is a need to develop neuroprotective therapies that can be used to reduce the deleterious effects of glaucoma, such as retinal ganglion cell death (RBC). Advances in understanding the pathophysiology of glaucoma are a key factor in understanding the pathogenesis of glaucomatous neuropathy. In this context, retinal ischemia plays a central role in various diseases of the retina. The pathogenesis of retinal ischemia involves temporal changes in retinal morphology and morphometry as well as changes in gene and protein expression. Sildenafil Citrate (SC) showed protective effect in the ischemia / reperfusion (I / R) models with neuroprotective effects. However, oral administration of CS in humans finds numerous side effects such as reduced blood pressure, headaches, flushing and nasal congestion are concomitant, as well as patients with moderate to severe cardiovascular disease or those undergoing nitrate therapy have increased secondary risks for adverse cardiovascular effects. Thus, we propose that the eye drops administration of CS may be an alternative to the oral route and also be equally neuroprotective in glaucoma and may minimize the undesirable risks in the systemic use of this drug, as well as offer a new approach for the therapeutic intervention in the pathogenesis of neuropathy... (Complete abstract click electronic access below) / Doutor

Orientador: Carlos Alan Candido Dias Junior / Resumo: As desordens hipertensivas gestacionais são complicações que acometem em torno de 5-10% das gestações. Essas desordens são as maiores causas de morbidade e mortalidade tanto materna quanto fetal. Estudos demonstram redução da biodisponibilidade do óxido nítrico (NO) em doenças hipertensivas gestacionais, tornando uma das condições responsáveis por agravar a disfunção endotelial durante o curso desta doença. Neste sentido, trabalhos demonstram que a administração oral de nitrito de sódio (NaNO2) e a administração de citrato de sildenafil podem reverter a redução da biodisponibilidade de NO e potencializar a via de sinalização NO-GMPc respectivamente. Para realização dos trabalhos foram utilizadas ratas Wistar, no qual, após confirmação da prenhez receberam L-NAME i.p para indução da hipertensão e tratamentos com nitrito de sódio e/ou citrato de sildenafil via oral entre os dias 14-21 de prenhez. A pressão arterial sistólica foi aferida pelo método de pletismografia de cauda. Parâmetros materno-fetais como: peso fetal e placentário, número de fetos viáveis e reabsorvidos foram realizados após a morte das ratas no 21º dia gestacional. O plasma das ratas foram armazenados para dosagens dos níveis plasmáticos de nitrito+nitrato, guanosina monofosfato cíclico (GMPc), mieloperoxidase (MPO), peroxidação lipídica (TBARS), TEAC, MTT sFlt-1 e VEGF. Além disso o plasma das ratas foram incubadas com células endoteliais de cordão umbilical humano (HUVECS) para avaliar a produção de NO endo... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Gestational hypertensive disorders are complications that affect around 5-10% of pregnancies. These disorders are the major causes of both maternal and fetal morbidity and mortality. Studies have demonstrated a reduction in the bioavailability of nitric oxide (NO) in gestational hypertensive diseases, making it one of the conditions responsible for aggravate the endothelial dysfunction during the course of this disease. In this sense, studies demonstrate that oral administration of sodium nitrite (NaNO2) and administration of sildenafil citrate may reverse the reduction of NO bioavailability and potentiate the NO-cGMP signaling pathway, respectively. Wistar rats were used to perform the study, in which, after confirmation of pregnancy, they received L-NAME i.p for induction of hypertension and treatments with sodium nitrite and/or sildenafil citrate orally between days 14-21 of pregnancy. Systolic blood pressure was measured by the tail plethysmography method. Maternal-fetal parameters such as: fetal and placental weight, number of viable and reabsorbed fetuses were performed after the death of the rats on the 21st gestational day. Plasma of the rats were stored for plasma levels of nitrite + nitrate, cGMP, myeloperoxidase (MPO), lipid peroxidation (TBARS), TEAC, MTT sFlt-1 and VEGF. In addition the plasma of the rats were incubated with human umbilical vein endothelial cells (HUVECS) to evaluate endothelial NO production. In both studies, we found reduction of systolic blood... (Complete abstract click electronic access below) / Doutor

Submitted by Victor Hugo Gonçalves Rizzi (victor.rizzi@hotmail.com) on 2018-09-06T18:30:07Z No. of bitstreams: 1 tese victor.pdf: 3098588 bytes, checksum: c82c3535b448e04bc144e33e93ef47f9 (MD5) / Approved for entry into archive by ROSANGELA APARECIDA LOBO null (rosangelalobo@btu.unesp.br) on 2018-09-11T19:49:14Z (GMT) No. of bitstreams: 1 rizzi_vhg_dr_bot.pdf: 3098588 bytes, checksum: c82c3535b448e04bc144e33e93ef47f9 (MD5) / Made available in DSpace on 2018-09-11T19:49:14Z (GMT). No. of bitstreams: 1 rizzi_vhg_dr_bot.pdf: 3098588 bytes, checksum: c82c3535b448e04bc144e33e93ef47f9 (MD5) Previous issue date: 2018-02-26 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / As desordens hipertensivas gestacionais são complicações que acometem em torno de 5-10% das gestações. Essas desordens são as maiores causas de morbidade e mortalidade tanto materna quanto fetal. Estudos demonstram redução da biodisponibilidade do óxido nítrico (NO) em doenças hipertensivas gestacionais, tornando uma das condições responsáveis por agravar a disfunção endotelial durante o curso desta doença. Neste sentido, trabalhos demonstram que a administração oral de nitrito de sódio (NaNO2) e a administração de citrato de sildenafil podem reverter a redução da biodisponibilidade de NO e potencializar a via de sinalização NO-GMPc respectivamente. Para realização dos trabalhos foram utilizadas ratas Wistar, no qual, após confirmação da prenhez receberam L-NAME i.p para indução da hipertensão e tratamentos com nitrito de sódio e/ou citrato de sildenafil via oral entre os dias 14-21 de prenhez. A pressão arterial sistólica foi aferida pelo método de pletismografia de cauda. Parâmetros materno-fetais como: peso fetal e placentário, número de fetos viáveis e reabsorvidos foram realizados após a morte das ratas no 21º dia gestacional. O plasma das ratas foram armazenados para dosagens dos níveis plasmáticos de nitrito+nitrato, guanosina monofosfato cíclico (GMPc), mieloperoxidase (MPO), peroxidação lipídica (TBARS), TEAC, MTT sFlt-1 e VEGF. Além disso o plasma das ratas foram incubadas com células endoteliais de cordão umbilical humano (HUVECS) para avaliar a produção de NO endotelial. Em ambos os trabalhos, nós encontramos redução da pressão arterial sistólica com nitrito e sildenafil, tanto isolado quanto em associação. Somente o sildenafil foi capaz de aumentar o peso fetal, enquanto o peso placentário foi melhorado por ambas as drogas. Tanto nitrito quanto sildenafil foram capaz de reduzir a reabsorção, possivelmente por aumentar a viabilidade fetal. Foi encontrado aumento de NO plasmático em todos os grupo que receberam nitrito, enquanto curiosamente a síntese de NO em HUVECS incubadas com plasma das ratas foi aumentada em ambas as drogas. Ainda em relação a cultura celular, nós encontramos aumento da viabilidade celular em ratas hipertensas tratadas com sildenafil. A concentração plasmática de GMPc estava aumentada em ratas normotensos que receberam sildenafil. Ambas as drogas apresentam efeito antioxidante, mas somente o sildenafil foi capaz de reduzir a atividade plasmática da MPO. Fatores angiogênicos (VEGF) e anti-angiogênicos (sFlt-1) presentes em doenças hipertensivas gestacionais estavam aumentados em ratas que receberam L-NAME, porém o nitrito de sódio foi capaz de reduzir esses fatores. Nosso dados sugerem que o nitrito e o sildenafil, tanto isolado 2 quanto em associação apresentam efeitos anti-hipertensivos e antioxidantes. Portanto, nossos resultados sugerem que a ativação da via NO-GMPc aumentou o fluxo sanguíneo na interface materno-fetal e protegeu contra a hipertensão e a restrição do crescimento fetal induzidas pelo L-NAME. / Gestational hypertensive disorders are complications that affect around 5-10% of pregnancies. These disorders are the major causes of both maternal and fetal morbidity and mortality. Studies have demonstrated a reduction in the bioavailability of nitric oxide (NO) in gestational hypertensive diseases, making it one of the conditions responsible for aggravate the endothelial dysfunction during the course of this disease. In this sense, studies demonstrate that oral administration of sodium nitrite (NaNO2) and administration of sildenafil citrate may reverse the reduction of NO bioavailability and potentiate the NO-cGMP signaling pathway, respectively. Wistar rats were used to perform the study, in which, after confirmation of pregnancy, they received L-NAME i.p for induction of hypertension and treatments with sodium nitrite and/or sildenafil citrate orally between days 14-21 of pregnancy. Systolic blood pressure was measured by the tail plethysmography method. Maternal-fetal parameters such as: fetal and placental weight, number of viable and reabsorbed fetuses were performed after the death of the rats on the 21st gestational day. Plasma of the rats were stored for plasma levels of nitrite + nitrate, cGMP, myeloperoxidase (MPO), lipid peroxidation (TBARS), TEAC, MTT sFlt-1 and VEGF. In addition the plasma of the rats were incubated with human umbilical vein endothelial cells (HUVECS) to evaluate endothelial NO production. In both studies, we found reduction of systolic blood pressure with nitrite and sildenafil, both alone and in combination. Both nitrite and sildenafil were able to reduce reabsorption, possibly by increasing fetal viability. No increase in plasma NO was found in all groups receiving nitrite, while curiously NO synthesis in HUVECS incubated with plasma from rats was increased in both drugs. Still in relation to cell culture, we found increased cell viability in hypertensive rats treated with sildenafil. Plasma cGMP concentration was increased in normotensive rats receiving sildenafil. Both drugs have an antioxidant effect, but only sildenafil was able to reduce plasma myeloperoxidase activity (MPO). Angiogenic (VEGF) and anti-angiogenic (sFlt-1) factors present in gestational hypertensive diseases were increased in rats receiving L-NAME, but sodium nitrite was able to reduce these factors. Our data suggest that both nitrite and sildenafil, both alone and in combination, have antihypertensive and antioxidant effects. Therefore, our results suggest that activation of the NO-cGMP pathway increased blood flow at the maternal-fetal interface and protected against hypertension and fetal growth restriction induced by L-NAME. / FAPESP 16/18782-3 / FAPESP 12/21305-1

Made available in DSpace on 2015-05-14T13:00:11Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1807154 bytes, checksum: 7aebebd024c49081a819ffba28e2ccae (MD5) Previous issue date: 2009-12-21 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Cissampelos sympodialis Eichl. (Menispermaceae) is a plant species used in popular medicine in Northeast of Brazil, whose pharmacological effects shows a great potential in treatment of respiratory diseases. Fosfodiesterase inhibition and smooth muscle relaxant effects are attributed to its leaves extract. Sildenafil, an oral fosfodiesterase type 5 inhibitor, is a drug used in treatment of pulmonary hypertension by vasodilatory effects through a cyclic guanosine 3´,5´-monophosphate (cGMP) dependent mechanism. This study was designed to investigate chronic effects of Cissampelos sympodialis extract in monocrotaline (MCT) induced pulmonary hypertension in rats, using sildenafil as a reference drug. After a single subcutaneous injection of MCT, rats were randomized to receive repeated administration of saline, sildenafil, Cissampelos sympodialis extract or both twice a day for 4 weeks. In the last day of treatment, animals were submitted to a median toracotomy to access pulmonary pressure. Four weeks after MCT injection, there was a significant development of pulmonary hypertension (60.19 ± 6.81), in contrast with SHAM group (32.77 ± 1.80). The increases in pulmonary mean pressure, ratio of right ventricular weight to body weight and thickening of the precapillary artery wall were significantly attenuated in the Cissampelos sympodialis extract (37.40 ± 3.76; 0.63 ± 0.04; 57.33 ± 5.04; respectively) and Sildenafil groups (34.57 ± 3.85; 0.68 ± 0.04; 57.00 ± 4.07; respectively), when compared to control group (52.65 ± 4.05; 1.14 ± 0.07; 89.43 ± 1.29; respectively). Combination therapy (32.16 ± 3.65; 0.86 ± 0.02; 64.62 ± 7.28; respectively) with sildenafil and extract had no additive effects in this model. These results suggest that therapy with oral Cissampelos sympodialis extract and sildenafil attenuates the development of monocrotaline induced pulmonary hypertension and that combination therapy has no additional effects compared with sildenafil or extract given alone. / Cissampelos sympodialis Eichl. (Menispermaceae) é uma planta usada na medicina popular do Nordeste do Brasil, cujos efeitos farmacológicos apresentam um grande potencial no tratamento de doenças respiratórias. Efeitos sobre o relaxamento de músculo liso e inibição de fosfodiesterases são atribuídos ao extrato das folhas de C. sympodialis. Sildenafil, um inibidor da fosfodiesterase do tipo 5, é uma droga usada no tratamento da hipertensão pulmonar pelo seu efeito vasodilatador, causado por um mecanismo dependente de 3´, 5´-monofosfato de guanosina cíclica (GMPc). Este estudo foi desenhado para investigar os efeitos crônicos do extrato de Cissampelos sympodialis em ratos com hipertensão pulmonar induzida pela monocrotalina (MCT), usando o sildenafil como droga de referência. Após uma injeção subcutânea única de MCT, ratos foram randomizados para receber duas doses diárias de salina, sildenafil, extrato de Cissampelos sympodialis ou ambos, durante quatro semanas. No último dia de tratamento, os animais foram submetidos a uma toracotomia mediana para medição das pressões pulmonares. Houve um desenvolvimento significante de hipertensão pulmonar, após quatro semanas, nos ratos submetidos à injeção de MCT (60,19 ± 6,81), em contraste com o grupo SHAM (32,77 ± 1,80). O aumento da pressão arterial pulmonar média, relação entre o peso do ventrículo direito e o peso do animal e espessamento da parede das artérias pré-capilares pulmonares foram significantemente atenuados pelo uso do extrato de Cissampelos sympodialis (37,40 ± 3,76; 0,63 ± 0,04; 57,33 ± 5,04; respectivamente) e sildenafil (34,57 ± 3,85; 0,68 ± 0,04; 57,00 ± 4,07; respectivamente) em relação ao grupo controle (52,65 ± 4,05; 1,14 ± 0,07; 89,43 ± 1,29; respectivamente). A terapia combinada (32,16 ± 3,65; 0,86 ± 0,02; 64,62 ± 7,28; respectivamente) com sildenafil e extrato não provocou efeitos aditivos em relação ao tratamento isolado com cada um deles neste modelo. Estes resultados sugerem que a terapia oral com extrato de Cissampelos sympodialis e sildenafil atenua o desenvolvimento da hipertensão pulmonar induzida pela monocrotalina e que a terapia combinada não traz efeitos adicionais, quando comparados com o emprego do sildenafil ou do extrato de Cissampelos sympodialis isoladamente.

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / INTRODUÃÃO: Recentemente nÃs demonstramos que o sildenafil tem efeito protetor contra a lesÃo gÃstrica induzida por AINEs em ratos, atravÃs de uma diminuiÃÃo na aderÃncia de leucÃcitos e aumento do fluxo sanguÃneo gÃstrico (Santos CL et al, BJP, 2005). O etanol induz lesÃo hemorrÃgica gÃstrica em ratos atravÃs de um aumento na produÃÃo de radicais livres e diminuiÃÃo da concentraÃÃo de glutationa na mucosa. OBJETIVOS: O objetivo deste trabalho foi investigar se o sildenafil possui efeito protetor contra a lesÃo induzida por Ãlcool, e avaliar o papel da via NO/GMPc/KATP no efeito gastroprotetor do sildenafil. MÃTODOS: O sildenafil (0.1, 0.3, 1, ou 3 mg/Kg, v.o) foi administrado 30 min antes do etanol 100% (4 ml/Kg, v.o). Depois de 1 h, os ratos foram sacrificados e os estÃmagos abertos para determinaÃÃo da Ãrea da lesÃo usando planimetria computadorizada. AlÃm disso, fragmentos de tecidos foram removidos para anÃlise microscÃpica e dosagem de glutationa e hemoglobina (teste colorimÃtrico - Bioclin). Os outros grupos foram tratados com L-NAME (1 ou 3 mg/kg, i.p), L-NAME (3 mg/kg, i.p) + L- Arg (200mg/kg, ip), ODQ (10 mg/Kg, v.o), glibenclamida (0.1, 0.3, 1 ou 3 mg/Kg, v.o), glibenclamida (1 mg/Kg) + diazÃxido (3mg/Kg, i.p) ou salina. ApÃs 30 min os ratos receberam sildenafil (1mg/kg) e depois de mais 30 min etanol 100% (4ml/kg, v.o), com o sacrifÃcio ocorrendo 1 h depois. RESULTADOS: O etanol 100% causou lesÃo gÃstrica (158.9 Â 9.3 mm2), hemorragia na mucosa (3787.0 Â 512.9 Âg/100mg) e reduÃÃo da concentraÃÃo de glutationa (78.7 Â 9.5 Âg/g). O Sildenafil protegeu, de forma dose-dependente, a mucosa gÃstrica do efeito do Ãlcool, com o efeito mÃximo na dose de 1 mg/Kg (44.5 Â 7.7 mm2). O sildenafil tambÃm reverteu a diminuiÃÃo da glutationa (143.6 Â 15.7 Âg/g) induzida por etanol. O L-NAME sozinho (151.1 Â 20.9 mm2), o ODQ (137.9 Â 41.6 mm2) e a glibenclamida sozinha (137.1 Â 16,7 mm2) reverteram a proteÃÃo do sildenafil. Mas, nos animais tratados com L-NAME + L-arginina (30.9 Â 10.5 mm2) ou glibenclamida + diazÃxido (60.3 Â 2.0 mm2) nÃo houve mudanÃas no efeito do sildenafil. CONCLUSÃES: O sildenafil protege a mucosa gÃstrica contra a lesÃo induzida por Ãlcool, atravÃs da ativaÃÃo da via NO/GMPc/KATP e por um aumento dos nÃveis de GSH no estÃmago / INTRODUCTION: Recently, we demonstrated that sildenafil has protective effects against NSAID- induced gastric damage in rats, by a decrease in leukocytes adherence and increase in gastric blood flow (Santos CL et al, BJP, 2005). Ethanol induced gastric hemorrhagic damage in rats, by an increase in free radical production and decrease in mucosal glutathione concentration. AIMS: Aim of was work is to investigate if sildenafil has a protective effect against ethanol- induced gastric damage and the role of the way NO/cGMP/KATP in this event. METHODS: Sildenafil (0.1, 0.3, 1, or 3 mg/Kg, p.o) was administrated 30 min before ethanol 100% (4 ml/Kg, p.o). After 1 hour, rats were sacrificed and the stomachs opened along the greater curvature and the mucosal lesion area was measured by computer planimetry program. Furthermore, pieces of gastric mucosal were removed for microscopic analysis and glutathione measure, and hemoglobin concentrations (colorimetric test Bioclin). Other groups had been dealt with L-NAME (1 or 3 mg/kg, i.p), L-NAME (3 mg/kg, i.p) + L- Arg (200mg/kg, i.p), ODQ (10 mg/Kg, p.o), glibenclamide (0.1, 0.3, 1 ou 3 mg/Kg, p.o), glibenclamide (1 mg/Kg) + diazÃxide (3mg/Kg, i.p) ou saline. After 30 min the rats had received sildenafil (1mg/kg), and after more 30 min ethanol 100% (4ml/kg, p.o), with the sacrifice occurring 1 h later. RESULTS: Absolute ethanol induced gastric damage (158.9 Â 9.3 mm2), and gastric mucosal hemorrhage (3787.0 Â 512.9 Âg/100mg) and reduced gastric glutathione concentration (78.7 Â 9.5 Âg/g). Sildenafil protected, in a dose dependent manner, the ethanol- induced gastric damage , with the maximum effect in the dose of 1 mg/Kg (44.5 Â 7.7 mm2). Sildenafil also reversed the decreased in gastric glutathione (143.6 Â 15.7 Âg/g) induced by ethanol. Alone L-NAME (151.1 Â 20.9 mm2), ODQ (137.9 Â 41.6 mm2) and glibenclamide alone (137.1 Â 16,7 mm2) reverted the protection of the sildenafil. But, in the animals trated with L-NAME + L-arginine (30.9 Â 10.5 mm2), or glibenclamide + diazÃxido (60.3 Â 2.0 mm2) did not have changes in the effect of the sildenafil. CONCLUSION: Sildenafil had a gastric protective effect against ethanol- induced gastric damage through the activation of the NO/cGMP/KATP pathway, at least in part through a increase in stomach GSH

The neurobiology of anxiety-related disorders is associated with impaired neuroplasticity. The glutamate/NO/cGMP pathway has been proposed to play a key role in neuroplasticity and neurodevelopment. It was demonstrated in recent reports that chronic co-administration of the phosphodiesterase type 5 (PDE5) inhibitor sildenafil and the antimuscarinic agent atropine exerts antidepressive-like activity in rats, and that this effect is related to PDE5 inhibition, with consequent elevation of cGMP levels and enhanced protein kinase G stimulation. The current study investigated possible anxiolytic effects of the chronic co-administration of sildenafil and atropine in stress-sensitive Flinders Sensitive Line (FSL) rats. FSL rats received vehicle control, fluoxetine (15 mg/kg), atropine (1 mg/kg), sildenafil (10 mg/kg) or sildenafil plus atropine via intraperitoneal administration, either acutely 30 minutes prior to testing (acutely) or daily for 14 days (chronically). FRL control rats received only vehicle. Thereafter anxiety-like behaviour was evaluated in the social interaction test (SIT - acute) and elevated plus maze (EPM - acute and chronic). The current study also compared to different ways to score the EPM, namely the percentage time spend in the open arms of the EPM and both the number of full and half body open arm entries, and also implemented defecation on the EPM as a measure of anxiety. Vehicle-treated FSL rats exhibited more anxiety-like behaviour than FRL rats in both the SIT and EPM following acute treatment, and in the EPM following chronic treatment. Acute treatment with fluoxetine exerted anxiogenic activity in the SIT and EPM, but anxiolytic activity following chronic administration, as observed in the EPM. In acute treatments neither sildenafil nor sildenafil plus atropine yielded any significant effects on anxiety-like behaviour. However, following chronic treatment, sildenafil exerted anxiolytic activity in the EPM by increasing the time spend in the open arms (45.72% ± 9.94% vs. 20.80% ± 9.94%, P<0.001). Atropine exerted a small anxiolytic response (30.71% ± 8.40% vs. 20.80 ± 9.94%), whereas atropine co-administration was additive to sildenafil alone and yielded an enhanced anxiolytic effect in the elevated plus maze (59.56% ± 4.95% vs. 20.80% ± 9.94%, P<0.001), relative to vehicle control. The percentage time spend in the open arms was scored in the EPM, the results suggested that the chronic treatment with sildenafil plus atropine exert an anxiolytic-like effect in FSL rats and the number of fecal droppings did not increase which is also an indication of an anxiolytic-like effects of the treatment. The current study demonstrated that the chronic treatment with sildenafil, alone or in combination with atropine, exhibit an anxiolytic-like action in stress-sensitive rats. In addition, the data support the clinical potential of using PDE5 inhibitors as antidepressant and anxiolytic strategy and warrant further investigation. Furthermore the study supports the previously proposed key role of the glutamate/NO/cGMP pathway in the neurobiology of anxiety-like disorders, and as an important target for drug development. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2011

Hypertensive patients exhibit impaired exercise capacity, a strong independent risk factor for cardiovascular disease, and the mechanisms responsible for this are not fully determined. Potential candidates may include endothelial vasomotor dysfunction and arterial stiffness, both of which are associated with hypertension. Impairment of the nitric oxide-cyclic guanosine monophosphate (NO-cGMP) pathway plays a major role in the development of these abnormalities, suggesting that enhancement of NO-cGMP signalling through phosphodiesterase type 5 (PDE5) inhibition may offer therapeutic potential in arterial hypertension. This thesis investigated the effects of the PDE5 inhibitor sildenafil citrate on exercise-induced vasodilatation, maximal exercise capacity and arterial stiffness in hypertensive patients, using different studies involving local limb and whole body exercise. Preliminary dose-ranging studies were initially performed to investigate the intraarterial (brachial) effects of sildenafil on forearm blood flow (FBF), and to select an appropriate, cGMP-independent, vasodilator to use as a control. On the basis on these studies, it was established that sildenafil, infused at 50μg/min, and verapamil, infused at 5μg/min, had similar vasodilator effect on FBF. Ten untreated hypertensive patients and ten matched normotensive subjects were then studied in a three-way, randomised, single-blind and placebo-controlled FBF study. The aim was to investigate the effects of sildenafil on handgrip exercise-induced vasodilatation, and to compare this response with verapamil and saline (placebo). Preinfusion exercise-induced vasodilatation was significantly reduced in hypertensive compared with normotensive subjects (P<0.001). However, after the infusions, while verapamil did not affect the vasodilator response to exercise in either group, sildenafil substantially enhanced this response in hypertensive patients, but not in normotensive subjects (P<0.05). These results suggested that sildenafil, through an increase in cGMP levels in the vasculature, substantially and selectively improves the vasodilator response to handgrip exercise in hypertensive patients.

Our recent studies have shown that that erectile dysfunction (ED) drugs including Sildenafil (Viagra), Vardenafil (Levitra) and Tadalafil (Cialis) enhance killing of several types of cancer cells by anticancer drug, Doxorubicin (DOX). We observed increased cell death by apoptosis in response to the combined treatment with ED drugs and DOX. However, the mechanism of such enhancement of cell death by combined treatment of ED drugs and DOX is not fully understood. Nuclear factor-κB (NF-κB) is an oxidant-sensitive transcription factor that plays a critical role in the immediate-early activation of a multitude of genes that have been documented to play critical role in programmed cell death (apoptosis). NF-κB activation has been shown to block apoptosis and its inhibition improves existing anti-oncogenic therapy such as chemotherapy. In the present study, we tested the hypothesis whether combined treatment of prostate cancer cells, PC3, with Sildenafil plus DOX would attenuate the activation of NFκB by inhibiting translocation of the p65 and p50 subunits to the nucleus and by phosphorylation of cytosolic IκB In addition, we investigated the effect of DOX and DOX plus Sildenafil on the expression of BCL family of proteins which play critical role in apoptosis. We treated PC3 cells with 1.5 μM DOX with or without 10 µM Sildenafil for 6 hours and 72 hours. The nuclear translocation of p65 and p50 and expression of BCL family of proteins was determined by western blot analysis. Our results show that combined treatment of DOX and Sildenafil significantly reduced the nuclear translocation of p65 and p50 as compared with DOX alone (P < 0.05). This correlated with the significant reduction in the expression of Bcl-2, BclxL and phosphorylation of BAD. These data provide an important mechanism by which Sildenafil treatment augments the apoptotic potential of DOX in PC3 cancer cells.

Background: Ethanol is a cardiotoxic substance that damages the heart by increasing apoptosis, free radical formation and calcium overloading. Consequently, there is an increase in cell death leaving fewer functioning myocytes leading to heart failure. Sildenafil is a phosphodiesterase type-5 (PDE-5) inhibitor approved for treatment of erectile dysfunction. Studies from our lab have demonstrated that PDE-5 inhibition reduces myocardial infarct size and attenuates post-ischemic cardiac dysfunction in both ischemia-reperfusion and permanent coronary artery ligation models. Therefore, in the present study, we hypothesized that treatment with sildenafil will prevent cardiotoxicity associated with acute alcohol exposure by reducing myocyte apoptosis and preserving cardiac function through PKG signaling. Methods and Results: Adult cardiomyocytes were isolated and treated with 100 mM of 100% ethanol ± 10 µM sildenafil. At 24 hours necrosis was assessed via trypan blue exclusion assay, JC-1 staining assessed mitochondrial membrane potential and ROS production was measured by DCF fluorescence. At 48 hours apoptosis was assessed by TUNEL assay. Ethanol increased the rate of necrotic and apoptotic cell death. This was attenuated by co-treatment with sildenafil. Ethanol disrupted the mitochondrial membrane potential and increased ROS production. Sildenafil preserved mitochondrial membrane potential and attenuated ROS production. Treatment of myocytes with 5-HD, a mitochondrial K+atp channel antagonist, blocked the protective effect of sildenafil. Knockdown of PKG using adenoviral siRNA blocked the protective effect of sildenafil, while overexpression of PKG1α conferred protection against ethanol cytotoxicity. To further demonstrate the effect of sildenafil ethanol-cardiotoxicity in vivo, mice were treated with ethanol (3 g/kg/day) with or without sildenafil (0.7 mg/kg) by i.p. injection for three consecutive days. After treatment, the animals were sacrificed and the hearts removed and perfused on a Langendorff system to measure function. After functional analysis, apoptosis and PKG activity was measured in the heart samples. Ethanol decreased the rate-force product and increased myocardial apoptosis. Sildenafil preserved cardiac function and significantly reduced apoptosis. Sildenafil treated myocardium also showed an increase in PKG activity. Conclusion: Sildenafil attenuates the toxic effect of ethanol by reducing apoptosis and maintaining the mitochondrial integrity in cardiomyocytes. Sildenafil also preserved cardiac function in ethanol-treated mice. Protein kinase G-dependent signaling plays a critical role in attenuating cardiotoxic effect of ethanol.