The endothelium plays an essential role in the regulation of vascular homeostasis and a state of endothelial dysfunction, which develops in the presence of cardiovascular risk factors, may contribute to the development and progression of cardiovascular disease. As such, the measurement of endothelial function, beyond being an experimental tool, may serve as an important tool to complement current risk assessment algorithms in the identification of high-risk patients. Flow-mediated dilation (FMD) is a non-invasive measure of peripheral conduit artery endothelial function that holds great promise. Presently, FMD suffers from methodological heterogeneity and a poor understanding of the various biological components involved in eliciting the dilatory response to a given shear stimulus. We compared both traditional and alternative methods of arterial diameter characterization with regards to their repeatability, nitric oxide-dependency, and their sensitivity in distinguishing between normal and dysfunctional endothelial responses. Our findings emphasize the importance of continuous arterial diameter measurement and suggest that the time to peak FMD is not a useful adjunctive measure of the FMD response. Given that endothelial dysfunction may be of clinical importance, strategies to correct it or prevent it from occurring may be of benefit. The 3-hydroxy-3-methylglutaryl coenzyme A inhibitors are agents that have demonstrated marked cholesterol-independent, endothelial-protective effects. We investigated the ability of rosuvastatin and atorvastatin to protect against endothelial dysfunction associated with ischemia and reperfusion (IR) injury, and chronic nitrate therapy. Using the FMD technique, we demonstrated, for the first time in humans, that acute rosuvastatin administration protects against IR-induced conduit artery endothelial dysfunction. Additionally, we demonstrated that this effect likely occurred by a cyclooxygenase-2-dependent mechanism, which may provide mechanistic insight into the observed cardio-toxicity with cyclooxygenase-2 inhibitors. In contrast, we observed that this endothelial-protective effect was lost upon sustained rosuvastatin administration, which may have important implications regarding the generation of sustained cardioprotective phenotypes. Finally, we demonstrated that atorvastatin co-administration prevented the development of tolerance and endothelial dysfunction associated with continuous transdermal nitroglycerin therapy in humans, likely through an antioxidant mechanism. Future studies are needed in disease patients to determine whether the concept of nitrate tolerance needs reconsideration in the presence of vascular-protective agents.

No description available.

No description available.

LPA1 lysophosphatidic acid receptors (LPA1Rs) are normally present on the surface of the cell. Our initial findings were that HMG-CoA reductase inhibitors (atorvastatin and mevastatin) induce the sequestration of the G protein-coupled LPA1R in recycling endosomes, most likely by inhibiting the recycling of tonically internalized receptors. Whereas, co-addition of geranylgeranylpyrophosphate (GGPP) or geranylgeraniol (GGOH) prevented atorvastatin-induced sequestration of LPA1Rs, the geranylgeranyltransferase-I inhibitor, GGTI-298, mimicked atorvastatin and induced LPA1R sequestration. This suggested that statin-induced endosomal sequestration was caused by defective protein prenylation. The likely targets of atorvastatin and GGTI-298 are the Rho family GTPases, RhoC and RhoA, since both inhibitors greatly reduced the abundance of these GTPases and since knockdown of endogenous RhoC or RhoA with small interfering RNAs (siRNAs) led to endosomal sequestration of LPA1R. Knockdown of RhoC was much more potent at inducing endosomal sequestration than knockdown of either RhoA or RhoB. In contrast, atorvastatin, GGTI-298, siRNA against RhoA, B, or C did not alter the internalization or recycling of transferrin receptors, indicating that recycling of transferrin receptors is distinct from LPA1Rs. Thus, these results, for the first time, implicate RhoA and RhoC in endocytic recycling of LPA1Rs and identify atorvastatin and GGTI-298 as novel inhibitors of this process. / Per the request of the author and advisor, and with the approval of the Graduate Education office, the following changes were made to this thesis: Replaced original page 1 with Errata Page 2. Replaced original pages 3-28 with Errata Pages 3 – 16. Replaced original pages 69-71 with Errata pages 17 – 19.

Hypercholesterolemia, a condition of high cholesterol levels in the circulation, poses a major risk for developing cardiovascular disease, such as atherosclerosis. A common method of reducing plasma cholesterol levels relies on the administration of drugs that limit cholesterol synthesis or uptake, many of which have undesirable side effects. Thus, some patients are turning to an alternative treatment, namely natural health products. Natural health products are often equally or even more effective at treating illness than synthetic drugs and may produce fewer side effects. The goal of this study was to identify a natural health product that regulates hepatic cholesterol synthesis by inhibiting HMG-CoA reductase, the enzyme which catalyzes the rate-limiting step of the cholesterol synthesis pathway. Several natural compounds were screened using the human hepatoma cell line HepG2. One compound, berberine, showed great potential as a regulator of cholesterol synthesis and so became the subject of this investigation. Berberine inhibited HMG-CoA reductase activity and decreased cellular accumulation of cholesterol. Berberine was shown to regulate HMG-CoA reductase through activation of metabolic regulator AMP-activated protein kinase, which modifies HMG-CoA reductase post-translationally and thereby decreases its activity. In conclusion, this study demonstrates that the natural health product berberine decreases cholesterol synthesis by activating a cellular signalling pathway to bring about post-translational modification of HMG-CoA reductase, and in doing so, inhibits this enzyme. This novel mechanism supports berberine’s potential for a cholesterol-lowering therapy and its role in reducing the risk for cardiovascular disease.

Evidence suggests that there is an immunological response of the recipient to xenograft bioprosthetic heart valves. Information on the impact of HMG-CoA reductase inhibitors (statins) and their anti-inflammatory properties on bioprosthetic valve failure remains limited. We sought to examine the efficacy of statin therapy in a rodent model of bioprosthetic valve implantation. To mimic the human scenario, fresh or glutaraldehyde-fixed aortic valve root conduits from Lewis rats or Hartley guinea pigs were microsurgically implanted intravascularly into the infra-renal aorta of Lewis rats. The syngeneic control group consisted of a fresh rat valve conduit implanted into a rat. The xenogeneic control group consisted of a glutaraldehyde-fixed guinea pig valve conduit implanted into a rat. Treatment groups consisted of xenogeneic groups treated with either daily steroids or statins. Overall, steroid treatment attenuated the inflammatory response observed within the xenogeneic glutaraldehyde-fixed valve conduits. Treatment with statins did not decrease this inflammatory response.

Hypercholesterolemia, a condition of high cholesterol levels in the circulation, poses a major risk for developing cardiovascular disease, such as atherosclerosis. A common method of reducing plasma cholesterol levels relies on the administration of drugs that limit cholesterol synthesis or uptake, many of which have undesirable side effects. Thus, some patients are turning to an alternative treatment, namely natural health products. Natural health products are often equally or even more effective at treating illness than synthetic drugs and may produce fewer side effects. The goal of this study was to identify a natural health product that regulates hepatic cholesterol synthesis by inhibiting HMG-CoA reductase, the enzyme which catalyzes the rate-limiting step of the cholesterol synthesis pathway. Several natural compounds were screened using the human hepatoma cell line HepG2. One compound, berberine, showed great potential as a regulator of cholesterol synthesis and so became the subject of this investigation. Berberine inhibited HMG-CoA reductase activity and decreased cellular accumulation of cholesterol. Berberine was shown to regulate HMG-CoA reductase through activation of metabolic regulator AMP-activated protein kinase, which modifies HMG-CoA reductase post-translationally and thereby decreases its activity. In conclusion, this study demonstrates that the natural health product berberine decreases cholesterol synthesis by activating a cellular signalling pathway to bring about post-translational modification of HMG-CoA reductase, and in doing so, inhibits this enzyme. This novel mechanism supports berberine’s potential for a cholesterol-lowering therapy and its role in reducing the risk for cardiovascular disease.

Evidence suggests that there is an immunological response of the recipient to xenograft bioprosthetic heart valves. Information on the impact of HMG-CoA reductase inhibitors (statins) and their anti-inflammatory properties on bioprosthetic valve failure remains limited. We sought to examine the efficacy of statin therapy in a rodent model of bioprosthetic valve implantation. To mimic the human scenario, fresh or glutaraldehyde-fixed aortic valve root conduits from Lewis rats or Hartley guinea pigs were microsurgically implanted intravascularly into the infra-renal aorta of Lewis rats. The syngeneic control group consisted of a fresh rat valve conduit implanted into a rat. The xenogeneic control group consisted of a glutaraldehyde-fixed guinea pig valve conduit implanted into a rat. Treatment groups consisted of xenogeneic groups treated with either daily steroids or statins. Overall, steroid treatment attenuated the inflammatory response observed within the xenogeneic glutaraldehyde-fixed valve conduits. Treatment with statins did not decrease this inflammatory response.

Conselho Nacional de Desenvolvimento Científico e Tecnológico / O objetivo deste trabalho é realizar uma avaliação econômica analisando o tratamento com atorvastatina e sinvastatina em comparação ao tratamento com placebo, no cenário do Sistema Único de Saúde (SUS), para subgrupos de pacientes classificados como alto risco de doença cardiovascular; avaliando se o custo adicional das estatinas em comparação ao tratamento placebo é justificado pelo ganho clínico esperado, em termos de redução de evento cardiovascular e redução de mortalidade. Utilizou-se o risco de eventos cardiovascular e a taxa de mortalidade como parâmetros de desfecho. Os dados epidemiológicos da doença e eficácia dos agentes terapêuticos foram obtidos de revisão e análise crítica da literatura. Um modelo analítico de decisão (Markov) foi desenhado para estimar a razão de custo-efetividade incremental da atorvastatina 10mg/dia e sinvastatina 40mg/dia em comparação ao placebo. A população analisada foi uma coorte hipotética composta por homens e mulheres, com idade média de 50 anos e com alto risco de doença cardiovascular. O modelo considera apenas custos médicos diretos, obtidos dos Sistemas de Informações Ambulatoriais e Hospitalares e Banco de Preços do Ministério da Saúde e de estudos de dados primários. A análise de custo-efetividade foi realizada através de planilha Excel, para o horizonte de tempo de 5 e 30 anos. O resultado revela que a atorvastatina 10mg/dia em comparação ao placebo tem maior custo e é mais efetiva, tanto no horizonte de tempo de 5 como no de 30 anos. A sinvastatina 40mg/dia mostrou ser uma estratégia de menor custo e maior efetividade, em comparação ao placebo, em ambos os horizontes de tempo analisados. O resultado da análise de impacto no orçamento demonstrou que o uso de sinvastatina 40mg/dia, em pacientes de alto risco de evento cardiovascular, representa minimização do custo anual em comparação ao uso de atorvastatina 10mg/dia. Observa-se que o tratamento com sinvastatina proporciona uma economia de, aproximadamente, 1,1 bilhão de reais em comparação ao tratamento com placebo. Em contrapartida, o tratamento dos pacientes com atorvastatina leva a um gasto adicional de cerca de 118,6 bilhões de reais em comparação ao tratamento com placebo. / The objective of this study is to perform an economic evaluation analyzing the treatment with atorvastatin and sinvastatin in comparison to placebo treatment, within the Brazilian Public Healthcare System (SUS) scenario, for patients with high risk of cardiovascular disease; analyzing if the additional cost related to statin treatment is justified by the clinical benefits expected, in terms of cardiovascular event and mortality reduction. Cardiovascular event risk and mortality risk were used as outcomes. Statin efficacy at LDL-c and cardiovascular events levels lowering data was obtained from a critical literature review. A decision analytic model was developed to perform a cost-effectiveness analysis comparing atorvastatin 10mg/day and sinvastatin 40mg/day to placebo treatment in patients with dyslipidemia in Brazil. The target population of this study was a hypothetic cohort of men and women with a mean age of 50 years old and high risk of cardiovascular disease. The model includes only direct costs obtained from Ambulatory and Hospital Information System and Price Database of Brazilian Ministry of Health. The comparative cost-effectiveness analysis itself was done through Excel spreadsheets covering a 5 or 30-years time horizon. The result shows that atorvastatin 10mg/day in comparison to placebo has higher cost with higher effectiveness in the time horizon of 5 and 30 years. The sinvastatin 40mg/day appears to be a strategy with lower cost and higher effectiveness in comparison to placebo, in both times horizon analyzed. The budget impact analysis shows that the use of sinvastatin 40mg/day, in patients with high risk of cardiovascular disease, leads to a cost minimization in comparison to the use of atorvastatin 10mg/day. The treatment with sinvastatin is responsible for an economy of, approximately, BRL1.1 billion in comparison to treatment with placebo. Otherwise, the treatment with atorvastatin leads to an additional cost of, approximately, BRL118.6 billions in comparison to the treatment with placebo.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Statins are inhibitors of the 3-hydroxy-3-metil-glutaryl coenzyme A reductase, the rate-limiting enzyme in the pathway for cholesterol synthesis. Several studies have shown that statins, particularly atorvastatin, are neuroprotective in several conditions, including stroke, cerebral ischemia, traumatic brain injury and exposure to excitatory amino acids. However, only a few studies have investigated whether statins modulate seizure activity. In the current study we investigated whether atorvastatin or simvastatin alters seizures induced by pentylenetetrazol (PTZ), a classic convulsant agent, GABAA antagonist. Treatment of adult male Wistar rats orally with atorvastatin 10 mg/kg/day for seven days increased the latency to PTZ-induced generalized-seizures. In contrast, when the treatment with atorvastatin was withheld for 24 h (statin withdrawal), seizures were facilitated, evidenced by a decrease in latency for clonic and generalized-seizures. Such effect was not seen with a similar treatment using simvastatin or an acute treatment using a single dose of simvastatin or atorvastatin (10 mg/kg; 30 min before on PTZ). Interestingly, the effects of atorvastatin treatment or withdrawal were not accompanied by changes in plasma or the cerebral cortex cholesterol levels or in the of blood-brain barrier permeability. The atorvastatin levels in plasma and cortex after seven days of treatment were above the IC50 for inhibition of HMG-CoA reductase, whereas atorvastatin was not detectable in the plasma or cortex following 24 hours of the end of treatment. We conclude that treatment with atorvastatin and its withdrawal exert differential effects on PTZ-induced seizures, which are not related to changes in plasma or cerebral cortex levels or in the blood-brain barrier permeability. Additional studies are necessary to evaluate the molecular mechanisms underlying these findings as well as its clinical implications. / As estatinas são fármacos inibidores da enzima 3-hidroxi-3-metil-glutaril coenzima A (HMG-CoA) redutase, enzima marca passo na rota de biossíntese do colesterol. Vários trabalhos têm mostrado que as estatinas, particularmente a atorvastatina, são neuroprotetoras em diversas condições, incluindo isquemia, acidente vascular cerebral, traumatismo crânio-encefálico e exposição a aminoácidos excitatórios. No entanto, poucos estudos têm investigado se as estatinas possuem alguma efeito sobre crises convulsivas. Neste trabalho foi investigado se a atorvastatina ou a sinvastatina alteram as convulsões induzidas por pentilenotetrazol (PTZ), um agente convulsivante clássico, antagonista GABAA. O tratamento de ratos Wistar machos adultos com atorvastatina por via oral durante sete dias (10 mg/kg/dia) aumentou a latência para crises generalizadas induzidas por PTZ (60 mg/kg). Em contraste, o tratamento com atorvastatina durante sete dias (10 mg/kg/dia) diminuiu a latência para convulsões clônicas e generalizadas induzidas por PTZ 24 horas após o término do tratamento (retirada do tratamento com atorvastatina). Tais efeitos não foram vistos com tratamentos similares utilizando sinvastatina. Além disso, o tratamento agudo com sinvastatina ou atorvastatina (10 mg/kg) 30 minutos antes da administração de PTZ não alterou as convulsões induzidas por este agente convulsivante. Curiosamente, a modulação das convulsões por atorvastatina não foi acompanhada de alterações nos níveis de colesterol plasmático ou do córtex cerebral nem na permeabilidade da barreira hemato-encefálica. Os níveis de atorvastatina no plasma e no córtex após sete dias de tratamento estavam acima do IC50 para a inibição da HMG-CoA redutase, enquanto que a atorvastatina não foi detectada tanto no plasma quanto no córtex após 24 horas do término do tratamento. Concluí-se que o tratamento com atorvastatina e a cessação abrupta desse tratamento modulam de maneira diferente as convulsões induzidas por PTZ. Além disso, concluí-se que tais efeitos não estão relacionados com mudanças no colesterol plasmático e do córtex cerebral ou na permeabilidade da barreira hemato-encefálica (BHE). Estudos adicionais são necessários para avaliar os mecanismos moleculares subjacentes a estas descobertas, bem como suas implicações clínicas.