Glucagon-like peptide-1 (GLP-1) synthetic analog therapies are prescribed for type 2 diabetes due to their effects on insulin and glucagon secretion, and glycemic control. Recent studies also suggest that they may have cardiovascular benefits; however, the mechanism responsible for this is unknown. To examine this question, we evaluated the effects of GLP-1 and the GLP-1 synthetic analog, liraglutide on cell signaling and function in human aortic endothelial cells (HAECs). The results indicate that both agents inhibit TNFα and LPS induced cellular adhesion molecule expression and monocyte adhesion. They also show that incubation with 30pM GLP-1 and 100nM liraglutide stimulates an immediate increase in intracellular calcium, which activates calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ). This in turn led to a 2.5 fold increase in the phosphorylation of both AMP-activated protein kinase (AMPK) and calcium/calmodulin-dependent protein kinase 1 (CaMK1) within 5 minutes. In addition both GLP-1 and liraglutide caused a 2-fold increase in the phosphorylation of the downstream AMPK/CaMK1 targets: endothelial nitric oxide synthase (eNOS) and cAMP response element-binding protein (CREB). Inhibition of CaMKKβ with STO-609 (0.5ug/mL) blocked the phosphorylation of both AMPK and CaMK1, confirming its pivotal role. Incubation of the HAECs for three hours with lipopolysaccharide (LPS, 2ug/mL) and TNFα (10ng/mL) increased the expression of vascular cell adhesion molecule-1 (VCAM-1) and E-selectin by 5 and 2 fold, respectively. Comparable increases in THP-1 monocyte adhesion to the HAECs, a putative initiating event in atherogenesis, also occurred. Pre-incubation for one hour with either GLP-1 or liraglutide inhibited these events. Likewise, pre-incubation with the CaMKK inhibitor STO-609, or use of lentivirus shRNA to knock down AMPK, blocked the inhibitory effects of both GLP-1 and liraglutide on monocyte adhesion. These results suggest that the recently observed cardiovascular benefits of GLP-1 and liraglutide could be mediated by their effects on CaMKKβ, AMPK and CaMK1 activation, which lead to decreased adhesion molecule expression and monocyte adhesion in endothelial cells. The finding that these effects occur at concentrations of GLP-1 (30pM) and liraglutide (100nM) observed in vivo also suggests they are physiologically relevant.

Made available in DSpace on 2014-06-12T17:40:16Z (GMT). No. of bitstreams: 2 arquivo7616_1.pdf: 2419909 bytes, checksum: faa7cec4b100ec2dd55f49cd985a9a83 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2006 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A produção industrial de gesso (CaSO4. 1/2H2O) no Brasil é realizada pelo método de desidratação (calcinação) da gipsita natural (CaSO4. 2H2O), utilizando calor de queima essencialmente de combustíveis sólidos (lenha local, coque de petróleo) e líquidos (óleo BPF, óleo alternativo). Aumento constante no preço da matriz energética tem motivado as calcinadoras buscarem combustíveis mais baratos. Por exemplo, em função do elevado custo de combustíveis de origem fosseis recentemente as calcinadoras estão utilizando lenha do Semi-Árido no sertão Nordestino brasileiro e contribuindo para o aumento da desertificação da região. O uso de combustíveis sólidos tais como coque de petróleo, resulta em emissão de gases nocivos que podem causam um grande impacto ambiental devido emissão de SOx, NOx. Entretanto, devido ao envolvimento de vários parâmetros de calcinação, o controle é de alta complexidade, muitas vezes gerando produtos de qualidade não homogênea. Foi realizado um estudo da viabilidade econômica e tecnológica para a combustão de gases (GN+GLP) visando à geração de calor em regiões remotas para a calcinação da gipsita para produção de gesso, calculando as propriedades térmicas da combustão da mistura do Gás Natural e GLP em diferentes proporções utilizando um software comercial Acomb5 (IPT) e medição experimental de PCSs de combustíveis líquidos e sólidos utilizados no pólo gesseiro. Estes resultados auxiliarão na redução do impacto ambiental minimizando o avanço da desertificação devido a derruba de árvores locais para queima visando à geração de calor, a queima dos gases em contato direto com o produto a aquecer decorrente de uma combustão limpa, com isto, não haveria contaminação do produto, gerando assim uma produção de gesso com qualidade controlada

La valutazione del comportamento ambientale del metabolita di un pesticida è stato suddiviso in due studi indipendenti ma complementari tra loro utilizzando il metabolita radiomarcato con 14C: 1. Valutazione della degradazione aerobica del 14C-metabolita nel suolo; 2. Valutazione dell'assorbimento e desadsorbimento del 14C-metabolita nel suolo. L'attività di studio delle normative e dei regolamenti concernenti le Buone Pratiche di Laboratorio nell'ambito di studi multisito ed è poi sfociato nella gestione di uno studio di validazione di un metodo di analisi per la determinazione dei residui di glyphosate in frutta ed ortaggi. / The behaviour evaluation of a pesticide metabolite was divided in two different but complementary studies, using the radiolabeled metabolite: 1. evaluation of aerobic degradation of 14C-metabolite in soil; 2. evaluation of sorption of 14C-metabolite in soil. The study of GLP (Good Laboratory Practices) for multisite studies lead to the management of validation study for determination of glyphosate residues in fruits and vegetables.

Mechanisms to augment the cellular function and mass of beta-cells may be effective means of treating type 2 diabetes. Important in the physiological control of beta-cell function and nutrient disposal are factors released from gut enteroendocrine cells during nutrient digestion. In enteroendocrine L-cells, post-translational processing of proglucagon gives rise to a number of proglucagon-derived peptides. One such peptide, glucagon-like peptide-1 (GLP-1), acts via its own receptor (GLP-1R) to stimulate beta-cell insulin secretion, proliferation and survival. Another, oxyntomodulin (OXM), weakly activates the GLP-1R and inhibits food intake in a GLP-1R-dependent manner in rodents, which led us to hypothesize that OXM modulates GLP-1R-dependent glucoregulation. While OXM did not mimic the inhibitory effect of GLP-1 on gastric emptying in mice, OXM stimulated insulin secretion, beta-cell survival and improved glucose tolerance in a GLP-1R-dependent manner. In a similar manner to GLP-1, glucose-dependent insulinotropic polypeptide (GIP), secreted from enteroendocrine K-cells, physiologically stimulates insulin secretion via a distinct GIP receptor (GIPR) in beta-cells. Beyond the beta-cell, GIP and GLP-1 appear to exert divergent actions for the control of glucose homeostasis. Moreover, I illustrate that physiological and pharmacological GLP-1R signalling may be comparatively more important for the preservation of beta-cell mass and glucose homeostasis in murine streptozotocin-induced diabetes. Lastly, studies in rodents and humans have showed that metformin increases circulating levels of GLP-1, leading us to hypothesize that GIP and GLP-1 may be involved in the glucoregulatory effects of metformin. Interestingly, transcripts for the Glp1r and Gipr were significantly increased within islets of metformin-treated mice, and metformin treatment enhanced the sensitivity of cultured beta-cells to GIP and GLP-1. In summary, these studies illustrate mechanisms by which enteroendocrine peptides compare and contrast with respect to beta-cell survival and function and the control of glucose homeostasis.

Abstract Bachelor’s degree thesis in Business Administration School of Economics and Management, University of Växjö, FEN 330, Spring 2006 Authors: Malin Jansson and Mirja Wynn-Williams Supervisor: Stig Malm How does Good Laboratory Practice improve quality? Background: The quality systems that steer manufacturing of pharmaceutical products from the testing phase to commercial manufacturing are the national and international regulatory frameworks and legislation. Good Laboratory Practice (GLP) is a quality system concerned with the organizational process and the conditions under which non-clinical health and environmental safety studies are planned, performed, monitored, recorded, archived and reported. In Sweden the Medical Products Agency monitors and regulates compliance with the principles of Good Laboratory Practice. Aim: The aim of our thesis is to explore the reasons why companies/laboratories adopt Good Laboratory Practice. We shall do this by identifying the advantages and disadvantages of adopting GLP principles for companies/laboratories, and how quality is improved by adopting GLP principles. We have summarized our aim in one principal question: How does Good Laboratory Practice function as a tool for quality improvement? Limitations: We will focus on GLP, and this thesis will not consider other Good Practice procedures such as Good Manufacturing Practice or Good Clinical Practice. Only laboratories in Sweden monitored by the Medical Products Agency are targeted in the empirical part of this work. Method: Literature research and interviews with GLP contact persons listed by MPA. Results and conclusions: Though costumers’ needs and legislation seem to be the motivations for companies deciding to comply with GLP, quality improvement seems to be the biggest advantage of validation. Proposal for further research: Do laboratory assistants and technical employees differ from management in their experience and views on GLP?

<p>Abstract</p><p>Bachelor’s degree thesis in Business Administration</p><p>School of Economics and Management, University of Växjö, FEN 330, Spring 2006</p><p>Authors: Malin Jansson and Mirja Wynn-Williams</p><p>Supervisor: Stig Malm</p><p>How does Good Laboratory Practice improve quality?</p><p>Background: The quality systems that steer manufacturing of pharmaceutical products from the testing phase to commercial manufacturing are the national and international regulatory frameworks and legislation. Good Laboratory Practice (GLP) is a quality system concerned with the organizational process and the conditions under which non-clinical health and environmental safety studies are planned, performed, monitored, recorded, archived and reported. In Sweden the Medical Products Agency monitors and regulates compliance with the principles of Good Laboratory Practice.</p><p>Aim: The aim of our thesis is to explore the reasons why companies/laboratories adopt Good Laboratory Practice. We shall do this by identifying the advantages and disadvantages of adopting GLP principles for companies/laboratories, and how quality is improved by adopting GLP principles. We have summarized our aim in one principal question: How does Good Laboratory Practice function as a tool for quality improvement?</p><p>Limitations: We will focus on GLP, and this thesis will not consider other Good Practice procedures such as Good Manufacturing Practice or Good Clinical Practice. Only laboratories in Sweden monitored by the Medical Products Agency are targeted in the empirical part of this work.</p><p>Method: Literature research and interviews with GLP contact persons listed by MPA.</p><p>Results and conclusions: Though costumers’ needs and legislation seem to be the motivations for companies deciding to comply with GLP, quality improvement seems to be the biggest advantage of validation.</p><p>Proposal for further research: Do laboratory assistants and technical employees differ from management in their experience and views on GLP?</p>

Cardiovascular disease is the most common cause of morbidity and mortality in type 2 diabetes. Short-term studies of glucagon-like peptide-1 (GLP-1)-targeted therapies suggest potential beneficial effects on cardiovascular outcomes. The mechanism behind this unexpectedly rapid effect is not known. In this study, full-length human GLP-1 receptor (GLP-1R) mRNA was cloned and sequenced from a human megakaryocyte cell line. Quantitative RT-PCR results showed that expression levels were comparable to other GLP-1R expressing tissues. Furthermore, incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in these cells. As megakaryocytes are the cellular precursors of platelets, the effect of GLP-1 and exenatide were studied in gel-filtered human platelet aggregation, where they were both shown to have an inhibitory effect on thrombin-stimulated platelet aggregation. Platelet inhibition by GLP-1 and GLP-1R agonists presents a potential mechanism for the reduced incidence of atherothrombotic events thought to be associated with GLP-1-targeted therapies.

Cardiovascular disease is the most common cause of morbidity and mortality in type 2 diabetes. Short-term studies of glucagon-like peptide-1 (GLP-1)-targeted therapies suggest potential beneficial effects on cardiovascular outcomes. The mechanism behind this unexpectedly rapid effect is not known. In this study, full-length human GLP-1 receptor (GLP-1R) mRNA was cloned and sequenced from a human megakaryocyte cell line. Quantitative RT-PCR results showed that expression levels were comparable to other GLP-1R expressing tissues. Furthermore, incubation with GLP-1 and the GLP-1R agonist exenatide elicited a cAMP response in these cells. As megakaryocytes are the cellular precursors of platelets, the effect of GLP-1 and exenatide were studied in gel-filtered human platelet aggregation, where they were both shown to have an inhibitory effect on thrombin-stimulated platelet aggregation. Platelet inhibition by GLP-1 and GLP-1R agonists presents a potential mechanism for the reduced incidence of atherothrombotic events thought to be associated with GLP-1-targeted therapies.

Mechanisms to augment the cellular function and mass of beta-cells may be effective means of treating type 2 diabetes. Important in the physiological control of beta-cell function and nutrient disposal are factors released from gut enteroendocrine cells during nutrient digestion. In enteroendocrine L-cells, post-translational processing of proglucagon gives rise to a number of proglucagon-derived peptides. One such peptide, glucagon-like peptide-1 (GLP-1), acts via its own receptor (GLP-1R) to stimulate beta-cell insulin secretion, proliferation and survival. Another, oxyntomodulin (OXM), weakly activates the GLP-1R and inhibits food intake in a GLP-1R-dependent manner in rodents, which led us to hypothesize that OXM modulates GLP-1R-dependent glucoregulation. While OXM did not mimic the inhibitory effect of GLP-1 on gastric emptying in mice, OXM stimulated insulin secretion, beta-cell survival and improved glucose tolerance in a GLP-1R-dependent manner. In a similar manner to GLP-1, glucose-dependent insulinotropic polypeptide (GIP), secreted from enteroendocrine K-cells, physiologically stimulates insulin secretion via a distinct GIP receptor (GIPR) in beta-cells. Beyond the beta-cell, GIP and GLP-1 appear to exert divergent actions for the control of glucose homeostasis. Moreover, I illustrate that physiological and pharmacological GLP-1R signalling may be comparatively more important for the preservation of beta-cell mass and glucose homeostasis in murine streptozotocin-induced diabetes. Lastly, studies in rodents and humans have showed that metformin increases circulating levels of GLP-1, leading us to hypothesize that GIP and GLP-1 may be involved in the glucoregulatory effects of metformin. Interestingly, transcripts for the Glp1r and Gipr were significantly increased within islets of metformin-treated mice, and metformin treatment enhanced the sensitivity of cultured beta-cells to GIP and GLP-1. In summary, these studies illustrate mechanisms by which enteroendocrine peptides compare and contrast with respect to beta-cell survival and function and the control of glucose homeostasis.

The incidence of obesity is on the rise globally and is associated with many comorbidities, especially type 2 diabetes mellitus (T2DM). Bariatric surgery is the most effective intervention for weight loss and reducing obesity-associated morbidity. The most common bariatric surgeries are roux-en-Y gastric bypass (RYGB) and sleeve gastrectomy (SG). RYGB and SG are equally efficacious at long-term reduction of weight in obese individuals and amelioriation of T2DM. Interestingly, the improvement of glucose regulation is noted before weight loss is observed. The most likely mechanism underlying glucose homeostasis after bariatric surgery is hormonal changes in the intestine. Enteroendrocrine changes favorable of an anti-diabetic profile are noted after only a few days of receiving either RYGB or SG surgery. Most consistently, elevated postprandial GLP-1, a potent regulator of appetite and glucose control, is observed in post-bariatric surgery patients. However, data is limited regarding post-prandial GLP-1 levels beyond two years after surgery. This study will address the gap in literature by assessing postprandial elevations of GLP-1 following RYGB or SG for up to five years. We will recruit obese type-2 diabetics from an outpatient bariatric surgery clinic at Boston Medical Center scheduled to receive RYGB or SG and periodically assess postprandial GLP-1 levels to determine if they remain elevated after 5 years. Additionally, we will provide evidence if there is a correlation among changes in postprandial GLP-1, weight loss, and hemoglobin A1c at five years. Our proposed study will help direct researchers to develop safer and more efficacious interventions for obesity and T2DM.