Studies show that insulin induced activation and assembly of insulin receptor substrate-1 (IRS1) and phosphatidylinositol-3-kinase (PI3K), within remodelled actin structures is critical for GLUT4 translocation to the cell surface in muscle cells. This study identifies the F-actin binding protein, nexilin, as a novel IRS1 binding partner. Insulin stimulates nexilin to dissociate from IRS1 and interact with actin. Nexilin knockdown has no effect on insulin-stimulated IRS1 tyrosine phosphorylation, but does enhance insulin-stimulated IRS1-PI3K interaction, increasing PIP3 formation, PKB activation and glucose uptake. This study also shows that nexilin overexpression may have an inhibitory effect on PKB phosphorylation and glucose uptake in adipocytes. These findings suggest nexilin is a negative regulator of IRS1 action on PI3K and insulin-stimulated dissociation of IRS1-nexilin allows the formation of IRS1-PI3K complexes in cytoskeletal-membrane compartments. Nexilin also specifically associates with the PH domain of IRS1, and not IRS2, suggesting a mechanism for signaling specificity of these isoforms.

Atherosclerosis is a leading cause of morbidity and mortality worldwide and type 2 diabetes and obesity-associated metabolic syndrome, both characterized by insulin resistance, are potent risk factors. These conditions also increase the risk for restenosis after revascularization procedures used for treatment of atherosclerosis. Studies have shown that insulin and resveratrol (RSV), a red wine polyphenol, decrease neointimal growth after vessel injury in models of restenosis, demonstrating a protective effect on the vasculature. However, oral glucose and sucrose were used in insulin studies to maintain normoglycemia, and their effect on neointimal formation was not assessed. Several studies have shown that nitric oxide (NO) production is stimulated by insulin and RSV, and since NO can decrease neointimal growth, the objective of this thesis was to address the mechanism of action of insulin or RSV to protect against restenosis, and determine whether NO production mediates these effects. To examine this, we treated rats with insulin or RSV and performed arterial balloon injury. In Study 1, insulin reduced neointimal area after injury in rats receiving oral glucose but not oral sucrose. Oral glucose alone had no effect on neointimal formation or insulin sensitivity whereas oral sucrose increased neointimal growth and induced insulin resistance. In Study 2, insulin decreased neointimal area and cell migration, and increased re-endothelialization. These effects were abolished by nitric oxide synthase (NOS) inhibition. In addition, insulin increased eNOS protein expression in the vessel. In Study 3, RSV reduced neointimal growth, cell proliferation, and migration after injury, without affecting re-endothelialization. Most of these effects were abolished by NOS inhibition, except for the decrease in cell migration. Insulin sensitivity and systolic blood pressure were not affected by RSV. Together, the results demonstrate that insulin, independent of glycemic effects, and RSV have a protective effect on the vessel against restenosis, which is mediated by NO. Since both insulin and RSV decrease neointimal formation without negatively impacting re-endothelialization, insulin or RSV treatment could provide some advantage over anti-mitogenic agents currently used in drug-eluting stents, which delay re-endothelialization. These studies suggest that insulin or RSV may have clinical potential in the prevention of restenosis after angioplasty.

Metabolic homeostasis is orchestrated by the hypothalamus through the neuroendocrine and the autonomic nervous systems. The hypothalamic nuclei respond to the peptide leptin secreted from adipose tissue to suppress feeding and increase energy expenditure by promoting fat metabolism via sympathetic activity. Another important, but perhaps less appreciated function of central leptin signaling is to elevate peripheral insulin sensitivity. Environmental and genetic risk factors that affect hypothalamic leptin signaling can lead to obesity and type 2 diabetes mellitus (T2DM). Here, we discovered that LIM domain only 4, LMO4, is a novel protein participating in central leptin signaling. In a process strikingly similar to T2DM in humans, CaMKIIα-Cre;LMO4flox/flox mice, which have LMO4 knocked out in the postnatal brain including the hypothalamus, develop visceral adiposity, reduced insulin sensitivity, obesity and diabetes when fed with regular chow. Central leptin signaling was significantly lost in key hypothalamic nuclei of mutant mice. Caloric restriction prevents obesity but not insulin resistance in these mice. Taken together, our results suggest that LMO4 function in the brain is required for central leptin signaling to control fat metabolism and peripheral insulin sensitivity.

<p>Olanzapine is an atypical antipsychotic drug exhibiting a low incidence of extrapyramidal side effects. It is not only effective in treating positive symptoms of schizophrenia, but also more efficacious against negative and depressive symptoms than classical antipsychotics. Olanzapine has been recommended as the first-line drug for the treatment of schizophrenia. Unfortunately, a common side effect of olanzapine, namely weight gain, has also been observed. A comprehensive literature analysis revealed that olanzapine induced higher weight gain than most other antipsychotics, only second to clozapine. The incidence of olanzapine-induced weight gain and related diseases, such as diabetes and cardiovascular diseases, is higher than that of the general population. These unwanted side effects have decreased the adherence to treatment. Many clinical observations and preliminary animal studies have attempted to elucidate the possible mechanism involved. To date, the mechanism for olanzapine-induced weight gain remains unclear.</p><p>This research project utilizes an animal model to investigate the possible mechanism of olanzapine-induced weight gain. The specific research objectives include: 1) does olanzapine affect feeding behavior; 2) can olanzapine influence the levels of glucose and triglyceride; 3) are cytokines, such as insulin, leptin, and TNF-Ñ involved in olanzapine-induced weight gain; 4) how does olanzapine affect adipose tissue?</p><p> An olanzapine-induced weight gain animal model has been established in the present investigation. An increase in food and water intake and increase in fat deposition accompanied with weight gain after treatment were observed. No significant increase in levels of glucose and triglyceride was detected. The changes of insulin and leptin levels in blood suggest that olanzapine may affect the endocrine system. A dramatic morphological alteration of adipose tissue by olanzapine was serendipitously observed. Immunohistochemical staining revealed that olanzapine stimulated collagen VI expression and deposition in the extracellular matrix suggesting that adipocyte differentiation may be enhanced. The effect of olanzapine on fat deposition might play a critical role in olanzapine-induced weight gain. The data from adipose tissue have provided a new clue on future research in understanding the mechanism of olanzapine-induced weight gain. Due to limitation of small number of animals and relatively short term of treatment, a large variation in groups diminished the power of analysis regarding the effects of olanzapine related to weigh gain.

Syfte: att kartlägga hemtjänstpersonalens kunskaper om diabetes och diabetesrelaterade läkemedel, agerandet i vissa specifikt beskrivna sjukdomssituationer samt kunskaper om för verksamheten gällande delegeringsrutiner.Metod: Studien genomfördes med en strukturerad enkät som innehöll frågor om diabetes, diabetesrelaterade läkemedel, hemtjänstpersonalens handlande vid vissa specifikt beskrivna sjukdomsfall samt frågor om för verksamheten gällande delegeringsrutiner. Totalt delades 120 enkäter ut till hemtjänstpersonal i tre olika kommuner. En svarsfrekvens på 70% erhölls. Resultat: De största bristerna i hemtjänstpersonalens kunskaper handlade om läkemedel och insulinpreparat. Personalen hade inte heller helt klart för sig skillnader mellan typ 1 och typ 2 diabetes. De flesta visste hur ett normalt blodsockervärde bör ligga men många visste inte i vilka situationer som ett blodsockervärde var angeläget att kontrollera. De flesta kunde identifiera tecken på lågt blodsocker och visste delvis hur de skulle agera i den aktuella situationen. Det var svårare för personalen att känna igen högt blodsocker där endast en tredjedel kände igen symtomen. Positivt var ändå att fler än de som kände igen symtomen (51 %) handlade korrekt i situationen och kontaktade sjukvård. Delegeringsförfarandet visade sig vara olika. De flesta svarade att de fick teoretisk genomgång inför varje beviljad delegering. Ungefär 46 % ansåg sig ha tillräckliga kunskaper för att ge insulin till vårdtagare.

<p>Insulin secretion is a fundamental process of pancreatic beta-cells required for the maintenance of glucose homeostasis. Fuel-stimulated insulin secretion occurs in proportion to the rate of metabolism of fuel substrates, yet the signals generated by metabolism of these secretagogues are incompletely understood. The increased burden placed on the beta-cell in conditions of obesity and insulin resistance often leads to dysregulation of stimulous-secretion coupling. Therefore, better understanding of the metabolic events required for insulin release is likely to be helpful in development of more effective treatments for diabetes.</p><p>Previous work in our lab revealed a critical role for the pyruvate-isocitrate cycling pathway in glucose-stimulated insulin secretion. It has been our hypothesis that this series of reactions plays a unique role in the beta-cell, and may be responsible for the generation of second-messenger signals critical for insulin secretion in response to increased fuel metabolism. One of the intermediates in the pyruvate/isocitrate cycle is cytosolic 2-oxoglutarate (2OG). In an effort to better understand the components of the pyruvate/isocitrate cycle and the signals that it generates, we initially focused our studies on the transporter protein responsible for the return of 2OG to the mitochondria, the 2-oxoglutarate carrier (OGC).</p><p>OGC was overexpressed in rat insulinoma 832/13 beta-cells and suppressed in both 832/13 cells and islets, and effects on metabolism and insulin secretion were measured. While overexpression of the OGC failed to alter insulin secretion, its siRNA-mediated suppression resulted in decreased insulin secretion in response to glucose, glutamine + BCH, and dimethyl-2-oxoglutarate. Suppression of OGC did not affect core pathways of fuel metabolism such as glucose usage, glucose oxidation or ATP production during glucose-stimulated insulin secretion (GSIS) or glutamine oxidation or ATP production during amino acid-stimulated insulin secretion (AASIS). Similar to previous findings, glucose-induced NADPH production was determined to be decreased in response to OGC suppression, whereas NADPH production during AASIS in untreated cells was already much lower than for GSIS, and suppression of OGC failed to decrease NADPH further.</p><p>As an additional approach to studying the role of 2OG metabolism in insulin secretion, we also investigated the mitochondrial enzyme glutamate dehydrogenase (Glud1). Overexpression of wild-type Glud1 failed to alter insulin secretion in 832/13 cells or in islets; however, suppression of Glud1 decreased both GSIS and AASIS, but did not affect dimethyl-2OG-stimulated insulin secretion. The reduction in AASIS was most likely the result of reduced glutamine oxidation. In contrast, during GSIS, NADPH production was decrease by Glud1 suppression, similar to our observation with the OGC.</p><p>In summary, these data expand our understanding of the metabolic pathways necessary for insulin secretion, and support the idea of a common metabolic pathway required for fuel-stimulated insulin release, including flux through the OGC, Glud1, and ICDc. However, while these data support the hypothesis that NADPH production is necessary for robust GSIS, it plays a less-prominent role during AASIS, and most likely works in concert with additional coupling-factors and signals.</p> / Dissertation

Atomic force microscopy is one of the powerful instruments used to explore the mechanical properties of nanoscale materials. It not only can produce high-resolution images and surface mechanical properties, but also can make use of its probe for surface etching. In this study, we first use atomic force microscopy to measure the Adhesion Map of insulin amyloid fibers, then conduct mechanical lithography on the surface with the probe. In the end, we discuss the effect on insulin amyloid fibrils due to exert different forces and different speeds with the probe. According to Nanoindentation theory and Hertzian model, we can derive the Young's modulus of insulin amyloid fibrils from force-indentation relations. Then we cut the Insulin amyloid fibers with probe. The results showed that when we applied 3.23 nN force by the probe, the insulin amyloid fibers began to break. When we applied 7.07 nN force, insulin amyloid fibers are cut off easily. Therefore, we can bite off insulin amyloid fibers of different lengths and sections, and arrange in the desired pattern by atomic force microscope.

The objective of this study was to more clearly define the relationship between body condition, plasma leptin and insulin-like growth factor-I (IGF-I) in stallions and geldings in moderate (5.0-5.5) versus fleshy (7.0-7.5) body condition. Data analyses of physical measurements showed that there was a difference for BCS (P<0.001) even though the fat group only achieved a mean BCS of 6.3 + 0.2 as compared with a mean BCS of 5.3 + 0.1 for the moderate group. Differences also existed for rump fat (P<0.05) and percent body fat (P<0.05) between BCS groups. Analysis of physical measurements revealed that there was no sex effect as geldings and stallions within each group were not significantly different. Analysis of plasma leptin concurred with previous reports as a difference (P<0.001) existed between the BCS groups. Mean leptin concentrations were 2.13 + 0.1 ng/ml for the fat group and 1.44 + 0.1 ng/ml for the moderate group. After normalization of the data, changes in leptin concentrations still revealed a significant difference (P<0.05) between BCS groups, yet no difference in leptin concentrations between stallions and geldings was seen. Dexamethasone (DEX) treatment on d 0 caused a subsequent 24 h rise in plasma leptin in both groups. Analysis of plasma IGF-I revealed no difference in IGF-I concentrations between BCS groups. Mean plasma IGF-I was 347.2 + 11.4 ng/ml for the fat group and 344.3 + 10.0 ng/ml for the moderate group. There was however a difference (P<0.05) between geldings and stallions. Geldings exhibited an overall mean plasma IGF-I concentration of 360.6 + 9.1 ng/ml with stallions exhibiting a mean IGF-I concentration of 329.1 + 12.1 ng/ml. The post DEX challenge rise seen with leptin was not evident when analyzing the change in plasma IGF-I concentrations. In conclusion, the data presented herein have provided a more accurate profile of circulating concentrations of leptin and IGF-I in stallions and geldings of moderate and fleshy body condition.

Colorectal cancer (CRC) is the 3rd common cancer in the world. Because the five-year survival rate is below 60 % in the patients with CRC, two important respects in CRC researches are early diagnosis and more effective chemotherapeutic drugs. In fact, the studies on molecular pathology of CRC can resolve these two problems. Insulin has a role in the carcinogenesis and developments of CRC, and 5-FU is a standard chemotherapeutic drug for the patients with stage III CRC. As a human securin, PTTG has a major role in many functions. In our studies of 2-D proteomics, PTTG correlates with the invasiveness of CRC. Besides, it is found to be highly expressed in many types of cancer, but the expression of PTTG is, however, low or undetectable in normal tissues. This character of tumor-specific expression is suitable for drug and target therapy. Therefore, we use cell line HT-29 to study the effects of 5-FU and insulin on the expression of PTTG. We have found that insulin in the physiologic level up-regulates PTTG. In normal physiologic level, insulin up-regulates PTTG, in a dose-dependent manner. On the other hand, the induction of PTTG by insulin more than normal physiologic level is decreased. In the studies with 5-FU, PTTG has a higher level after treatment, but not in dose-dependent manner. The up-regulation of PTTG by 5-FU is decreased in a higher dose. In cancer cells, insulin regulated pathways may contribute to the growth, proliferation, and apoptosis of tumors by activating oncogenic molecular targets such as PTTG. We have showed that insulin of bio-physiologic level can up-regulate PTTG in colon cancer HT-29 cell lines. Induction of PTTG by insulin suggests a mechanism by which insulin may contribute to the development and/or progression of colon cancers. To tumor- specific expression of PTTG, induction of PTTG by insulin, consequently, may be a target of colon cancer treatment. In the studies of 5-FU and PTTG, we have found that 5-FU in HT-29 cells can induce PTTG, with a peak effect in a dose around IC50. Interestingly, the induction of PTTG is decreased in a higher dose of 5-FU. This is a new finding in the effect of 5-FU on PTTG. Accordingly, we can realize why PTTG in some studies is suppressed; the others have a higher level. More importantly, the connection of PTTG expression between sister chromatic separation and DNA repair after DNA damage is more reasonable. Based on this finding, we propose that PTTG connects DNA damage response pathways, sister chromatic separation and apoptosis after DNA damage. Therefore, PTTG has a key role after DNA damage, deciding cells to have DNA repair/cell cycle arrest or to progress to apoptosis.

Insulin resistance was thought as the major etiology of hypertension of the metabolic syndrome. Both human and animal studies revealed sympathetic overactivity were present in the metabolic syndrome. Nowadays, most of the studies that examined the etiologies of hypertension of metabolic syndrome were focused on the pathophysiologic effects of insulin resistance on the peripheral vessels. However, there was no study ever examined the insulin resistance in cardiovascular regulatory centers of central nervous system or the pathogenesis of sympathetic overactivity in metabolic syndrome. Our previous study demonstrated that insulin plays a cardiovascular regulatory role in the nucleus tractus solitarii (NTS), one of the cardiovascular regulatory centers in the brain stem. We also demonstrated that the cardiovascular regulatory effects of insulin in the NTS were accomplished through activating PI3K-PKB/Akt-NO signaling pathways. Recently, increases in oxidative stress could raise the incidence rate of diabetes mellitus and cardiovascular diseases had been reported. Besides, it has been reported that there were marked increases in reactive oxidative species (ROS) in various hypertension animal models. It was also reported that elevation of ROS in various tissues may activate the mitogen-activated protein kinase (MAPK) superfamily. Activated MAPKs may phosphorylate insulin receptor substrate 1 (IRS1) on the serine 307 residue. It has been reported that IRS1S307 phosphorylation would inhibit normal insulin signal transduction. The aims of this thesis were to investigate whether the neuronal cells in the NTS would develop insulin resistance in the metabolic syndrome rats, whether development of insulin resistance in the NTS cause hypertension in the metabolic syndrome rats, which signaling molecule in insulin signaling pathway is the key molecule that cause insulin resistance in the NTS, and what the pathogenesis of insulin resistance is in the NTS of metabolic syndrome rats. In the pioneer study, Wistar-Kyoto (WKY) rats were fed with 10% fructose water as their drinking water for 8 weeks. Another group of fructose-fed WKY rats were fed with insulin sensitizer, rosiglitazone, since the 5th week. Blood pressure was measured by tail vein sphygmomanometer every week and venous blood were draw to measure blood sugar and insulin level every other week. Thereafter, all the rats enrolled in this study were fed with 10% fructose water with/without rosiglitazone for 2-3 weeks. My results demonstrated the blood pressure of fructose-fed WKY rats was significantly elevated after 2-week fructose feeding. But at the same time, HOMA-IR did not elevated, which indicated the insulin resistance in the peripheral did not develop yet. Interestingly, at the same time, endogenous insulin in the NTS was significantly elevated in the fructose-fed group. The cardiovascular responses of insulin in the NTS were diminished in the fructose-fed group. While in the rosiglitazone-treated group, the blood pressure and endogenous insulin in the NTS were decreased the baseline level. The cardiovascular responses of insulin in the NTS were restored in the rosiglitazone-treated group. These results indicated insulin resistance do develop in the NTS of fructose-fed rats, and the neuronal insulin resistance in the NTS can induce hypertension. The immunoblotting results demonstrated the phosphorylation of IRS1S307 was significantly elevated in the fructose-fed rats. While the phosphorylation of its downstream molecules, such as AktS473 and eNOSS1177, were significantly decreased as compared with the control group. In the NTS of rosiglitazone-treated group, the phosphorylation of IRS1S307 was decreased, and the phosphorylation of AktS473 and eNOSS1177 were restored. These results indicated that the underline pathogenesis of insulin resistance in the NTS was phosphorylation on the inhibitory serine residue of IRS1, which interfered with the normal insulin signal transduction in the NTS. Increases in ROS in the NTS of fructose-fed rats were demonstrated in the DHE histostaining. Phosphorylation of p38MAPK in the NTS of fructose-fed rats was also detected by immunoblotting. In the NTS of Tempol-treated fructose-fed rats, the phosphorylation of p38MAPK reduced and the nitric oxide production elevated to the basal level. Blood pressure decreased significantly when p38MAPK inhibitor, SB203680, was microinjected into the NTS of fructose-fed rats. These results indicated the pathogenesis of insulin resistance in the NTS is increases in ROS in the NTS, which activate p38MAPK and then phosphorylate IRS1S307. In conclusion, the neuronal cells in the NTS may develop insulin resistance in fructose-fed rats, and the neuronal insulin resistance in the NTS contributes to the hypertension of metabolic syndrome. The mechanism of insulin resistance in the NTS is phosphorylation on the serine 307 residue of IRS1, which interfere with insulin signaling and subsequent NO production in the NTS. The pathogenesis of IRS1S307 phosphorylation is activated p38MAPK which in turn is activated by ROS in the NTS.