No description available.

No description available.

Many obese individuals and type 2 diabetes mellitus (T2DM) patients have elevated levels of insulin. Hyperinsulinemia is a major cancer risk factor in T2DM individuals and activated insulin receptor (IR) has been linked to many types of cancer and poor survival. However, the mechanisms that account for the link between the hyper-active insulin signaling and cancer risk is not well understood. PTEN plays an antagonistic role in the canonical insulin signaling pathway, and is the second most commonly mutated tumour suppressor (after p53) found in human cancers. In many cancers the PTEN gene is not deleted, but instead the protein is lost. Therefore the regulation of PTEN protein in humans is of great importance. Here we hypothesized that the activated insulin signaling down-regulates PTEN. Considering that insulin signaling is highly conserved from C. elegans to human, I used C. elegans as a model and showed that DAF-2, the worm homolog of IR, is a negative regulator of DAF-18, the worm homolog of PTEN. In addition, I showed that DAF-28, the worm homolog of insulin, also negatively regulates DAF-18/PTEN. I used western blot and immunostaining to show that the protein level of DAF-18/PTEN is increased in the daf-2/IR and daf-28/insulin mutants. I further showed that daf-18/Pten is genetically epistatic to daf-2/IR in regulating neuronal development. I then employed human cell culture experiments and reported that this negative regulation is conserved in human cancer cell lines. I showed that knocking-down IR through siRNA up-regulates PTEN, and over-expressing a gain-of-function IR down-regulates PTEN. I also showed that insulin stimulation dramatically decreased PTEN and this decrease is dependent on IR. I further confirmed a physical association between IR and PTEN in both human and C. elegans, and reported that IR could phosphorylate PTEN. To provide mechanistic insight to DAF-18/PTEN regulation, I identified another protein, which is a ubiquitin ligase, that functions in insulin signaling to down-regulate DAF-18/PTEN. Additionally, I also provided evidence that insulin signaling cross talks with Eph receptor signaling. In summary, my findings will be informative for cancer biologists to study the roles of these genes in carcinogenesis. / Thesis (Ph.D, Biology) -- Queen's University, 2013-02-04 14:37:29.376

The notion that insulin binds to a specific site on the cell membrane was first proposed many years ago. However, experimental proof of a membrane bound insulin receptor did not come until the early 1970s when biologically active radiolabelled insulin was used in direct binding studies (Cuatrecasas, 1971). Recent advances in understanding the mechanism of insulin action are the result of studies on the structure and function of the insulin receptor. The membrane receptor would appear to have two functions: firstly, it must bind insulin and secondly, it must couple insulin binding to insulin action. Defects in either of these receptor functions will result in an impaired response to insulin, or insulin resistance (Taylor, 1985). Insulin resistance is a common disorder in a number of disease states in man. For example, non-insulin-dependent diabetes mellitus and obesity are associated with mild insulin resistance (Bar et aZ., 1976). There are also a number of relatively rare syndromes of extreme insulin resistance in which there is either impaired receptor function, or an immunological defect resulting in the development of auto-antibodies against the insulin receptor (Taylor et aZ., 1985).Studies on insulin receptor defects associated with these disease states have led to progress in understanding the molecular mechanisms of insulin action. Ideally when investigating these disease states one should study insulin action on classical target cells such as adipocytes, hepatocytes or muscle. However, it is now well established that the kinetics of insulin binding to its membrane receptor is similar in all human tissue whether or not it is a target for insulin action. This has led to a great deal of research on the more accessible human tissues such as monocytes, erythrocytes, cultured fibroblasts and Epstein-Barr virus (EBV) transformed B-Iymphocytes. The most convenient tissue to study is EBV transformed B-Iymphocytes, as these cells can be taken from individual patients and grown in culture in large quantities, which facilitates biochemical studies. Despite these advantages, it is important to establish that this virus-induced receptor is a true insulin receptor and not an artifact of viral transformation. Studies on B-Iymphocyte proliferation have shown that the insulin receptor appears on the cell membrane during the proliferative phase of B-cell activation. However , this is a transient event and once the cell reaches maturation the insulin receptor is no longer evident (Marchalonis & Galbraith, 1987). The insulin receptor has also been demonstrated in a number of cultured human lymphoblastoid cell lines (Gavin et aL, 1983; Maegawa et aL, 1983). It seems, therefore, that the insulin receptor is normally expressed by blast cells. The purpose of this study was to investigate insulin binding characteristics on a human lymphoblastoid cell line with B-cell characteristics which was originally derived from a patient with Burkitt's lymphoma. These cells, which are known as Raji cells, are unusual in that they carry multiple copies of the EBV genome in their DNA. For this reason they provide a useful model system for studying the insulin receptor in EBV transformed lymphocytes. In addition, studies on the mechanism of insulin action in these cells should give some insight into the function of the insulin receptor during B-cell proliferation. In this study four major characteristics of insulin binding to insulin receptors on Raji cells are described. Firstly, on the basis of kinetic studies a model for insulin-receptor interaction was established. Secondly, processing of insulin and the receptor was investigated to determine whether the receptor is functional. A third aspect was elucidation of the receptor structure and the insulin binding site. Finally, the cross-reaction between insulin and type I IGF receptors was studied, and the cellular response mediated by the insulin receptor and growth factor receptor was determined. / Thesis (M.Med.Sc.)-University of Natal, Durban, 1988.

No description available.

No description available.

Ratten, die intrazerebroventricular (icv) mit Streptozotocin (STZ) behandelt werden, eignen sich gut als Tiermodelle für die sporadische Alzheimererkrankung (sAD). In der hier vorgelegten Arbeit wurden Veränderungen bezüglich der Insulinkonzentration sowie einiger Bestandteile der Insulinrezeptor (IR) – Signalkaskade in Rattengehirnen, welche icv mit STZ behandelt wurden, zu verschiedenen Zeitpunkten untersucht. Die Auswirkungen von STZ auf die zerebrale IR-Signalkaskade wurden dann mit denen von chronisch erhöhten Corticosteronkonzentrationen verglichen. In dieser Studie wurde im Hippocampus eine verminderte mRNA-Expression von Insulin, der IR sowie des insulinabbauenden Enzyms (IDE) nachgewiesen; bezüglich der tau-mRNA-Expression konnten jedoch in diesem Gehirnareal der mit STZ behandelten Ratten keine Veränderungen beobachtet werden. Die Resultate der Insulin-, IR- und IDE-mRNA-Expression fielen bei den mit Corticosteron behandelten Ratten ähnlich aus Im Gegensatz hierzu nahm die tau-mRNA-Expression bei Ratten, die mit Corticosterone behandelt wurden, zu, was auch für eine sAD kennzeichnend ist. Sowohl bei den mit STZ als auch bei den mit Corticosteronen behandelten Ratten konnten Verhaltensanomalien beobachtet werden. Die in dieser Arbeit erzielten Resultate deuten darauf hin, dass viele Merkmale einer sAD experimentell durch eine Beeinträchtigung des Insulin/IR-Signalwegs sowie eine chronische Erhöhung der Corticosteronkonzentration hervorgerufen werden können. Dies untermauert wiederum unsere Hypothese, dass es sich bei sAD um eine neuroendokrine Störung handelt, die mit gehirnspezifischen Fehlfunktionen in der Insulin/IR-Signalkaskade einhergeht, welche zum Teil durch erhöhte Corticosteronkonzentrationen ausgelöst werden können. Auf Grund der in dieser Arbeit erzielten Resultate stellt sich die Frage, ob -Amyloid (A) ein Auslöser oder eine Konsequenz einer sAD darstellt. Die hier vorgelegte Arbeit last den Schlus zu, dass bei sAD-Tiermodellen ein Zusammenhang zwischen primären Fehlfunktionen im zerebralen Insulinsystem und dadwol sekundär ausgeloster A-Pathologie besteht. Weiterfübende Untersuchungen wird aber notwendig um diese Aussagen zu bestätigen. / This research was aimed to evaluate the time-course of changes in the brain insulin and some elements of the insulin receptor (IR) signalling cascade in the streptozotocin-intracerebroventricullarly (STZ-icv) treated rats representing experimental model of sporadic Alzheimer’s disease (sAD) and to compare them with effects of chronically increased corticosterone on the brain insulin system. This study shows down-regulation in mRNA expression of insulin, insulin receptor (IR), and insulin degrading enzyme (IDE) but no changes were observed in the expression of tau mRNA in hippocampus of STZ-icv treated rats. Comparing these results to the ones found in corticosterone treated rats similarities at the level of insulin, IR and IDE mRNA expression can be assumed. In contrast tau mRNA expression in corticosterone treated rats were increased, data which are in line with sAD. Behavioural deficits were found in both STZ-icv and corticosterone treated rats. In conclusion, these results demonstrate that many of the characteristic features of sporadic Alzheimer’s disease (sAD) can be produced experimentally by impairing the insulin/IR signaling pathway combined with a chronic increase of corticosterone. This supports our hypothesis that sAD represents a neuro-endocrine disorder associated with brain-specific disregulation in insulin and IR signaling, caused in part by increased level of corticosterone. In line with that our study puts a question on the classical amyloid β (Aβ) hypothesis, supporting the view of brain insulin system dysfunction as a trigger for the Aβ pathology in an experimental sAD model.

Abstract Mortality of patients requiring intensive care treatment for greater than 5 days has been shown to be about 20% worldwide. Hyperglycaemia is common in critically ill patients. Strict glucose control with insulin in critically ill patients was shown to reduce mortality and morbidity significantly. Several interrelated mechanisms are involved in the development of “stress hyperglycaemia” in critically ill patients. These include dextrose containing intravenous infusions and total parenteral nutrition; the counter regulatory hormones (catecholamines, cortisol, glucagon and growth hormone) which oppose the effects of insulin; nervous system signaling; increased insulin clearance; and excess production of cytokines that interfere with intracellular insulin signaling pathways. Aim of study: To determine if the cytokines TNFα, IL-6 and adiponectin are significant determinants of insulin resistance in critically ill patients. Methods: The study was a prospective observational study conducted in the intensive care unit (ICU) at the Chris Hani Baragwanath hospital. Forty sequential adult ICU admissions that met with the inclusion criteria were enrolled. Blood specimens were drawn for adiponectin, TNF, and IL-6 at the time of ICU admission, on day 3, day 7 and on discharge from the ICU. Demographic data and clinical data were recorded, and body mass index (BMI) and APACHE II scores were calculated on admission. Blood glucose was measured every 2 to 4 hours, recorded and a mean value was calculated over the 24 hour period. Insulin infusions were started when the blood glucose values exceeded 6.0mmol/l. Administration of insulin was according to a fixed sliding scale. The total amount of insulin administered intravenously over that 24 hour period was recorded. Other factors known to be related to insulin sensitivity, such as inflammation (as indicated by C-reactive protein), vii triglycerides, insulin, C-peptide and cortisol levels were also drawn in addition to the blood drawn for routine investigations. Results: Duration of stay in ICU correlated with severity of illness as assessed by the APACHE II score (r = 0.44, p = 0.004). There was no significant difference in the mean 24 hour plasma glucose concentration throughout the duration of stay in ICU, there were however significant differences in the amount of insulin administered to maintain normoglycaemia. The amount of administered insulin required was found to peak on day 3 and decline thereafter. The main determinant of insulin administered was mean glucose (r = 0.79, p < 0.00001). The measured insulin concentrations on admission correlated with mean plasma glucose (r = 0.41, p = 0.009) and C-peptide (r = 0.45, p = 0.004) levels. The main determinants of mean plasma glucose levels on admission were BMI (r = 0.38, p = 0.013) and serum cortisol (r = 0.41, p = 0.008) levels. Serum triglycerides levels showed a significant difference from admission to discharge, with values increasing from admission levels. Adiponectin levels showed a significant increase from admission to discharge. IL-6 levels showed a significant decrease. TNFα levels did not show statistically significant changes. No statistically significant correlations were found between the levels of TNFα or IL-6 and administered insulin. Adiponectin concentrations showed a negative correlation with amount of administered insulin on discharge (r = -0.457, p = 0.0049). There were significant gender differences in BMI, administered insulin on admission, serum cortisol and C-peptide concentrations, with females having higher values than males. BMI was shown to account for the gender differences in administered insulin and C-peptide levels. viii There were significant differences in IL-6 and TNFα concentrations between the survivors and nonsurvivors, with higher levels being seen in non-survivors. Adiponectin levels were lower in nonsurvivors, but this did not reach statistical significance. Conclusion: Although there was a demonstrable change in insulin sensitivity during the stay in ICU, there was no statistically significant association between the cytokines TNFα or IL-6 and insulin administration. There was a negative correlation between adiponectin concentrations and administered insulin on discharge. This data also demonstrates that mortality is associated with increased levels of proinflammatory cytokines.

Objectives. The Pregnancy, Nutrition & Diabetes (PND) study aimed to evaluate the dietary status of urban Black pregnant women attending the Antenatal Clinic at the Charlotte Maxeke (Johannesburg General) Hospital. In addition, the study investigated the effects of maternal dietary intake and hormonal levels during pregnancy on fetal growth, birth size and the early postnatal development of risk factors for future Type 2 diabetes. The study analysis precedes a detailed description of the study population, including its comparison with other populations. Methods. 126 women were enrolled in the study before 24 weeks of gestation. Twice during pregnancy (weeks 20-24 and 30-36, visits V1 and V2) and approximately six months after the delivery (visit V3), volunteers participated in a standard 75 g oral glucose tolerance test (OGTT). Blood pressure, anthropometric and socio-demographic data were taken and a food frequency questionnaire was administered at each visit. The daily maternal intakes of total energy and macronutrients expressed as a % of total energy intake (%E) were calculated and further evaluated. A comparison with the crude nutrients intakes was also performed. During both pregnancy visits, an ultrasound examination was carried out to obtain estimates of fetal biometry and fetal well-being and the postnatal anthropometric parameters were also measured. Blood samples were collected at fasting, 30min, 60min and 120min of the OGTT for the measurements of maternal glucose (GLC), insulin (INS), C-peptide (C-PEP) and proinsulin (PI) and fasting samples for the determination of placental lactogen (HPL), insulinlike growth factor 1 (IGF-1), insulin-like growth factor binding protein 1 (IGF-BP1), free thyroxin (FT4), cortisol (CORT) and leptin (LEPT). Infant fasting blood samples were used for the analysis of the GLC, INS, PI, IGF-BP3 and LEPT. The plasma glucose samples were analysed on the Beckman Glucose Analyser 2, whilst all the other analytes were measured by an immunoassay method on 96-well plates. Results – DESCRIPTIVE DATA. Study participants. The majority of the study participants had completed high school education, however were unemployed and of low and very poor socio-economic levels. In comparison with another study from the developing world, the Pune study in India, the women from the current study were older, heavier with greater body mass index (BMI) and they were also taller. They had higher head and arm circumference. Birth outcomes. The birth size of African babies in the current study, although smaller in all parameters, was still relatively closer to the size of the Caucasian babies seen within the Southampton (Godfrey et al. 1996) or Helsinki (Forsén et al. 1997) studies than to the birth size of the babies from Trivandrum (Jaya et al. 1995) or Pune (Rao et al. 2001) in India. These relatively smaller African babies have comparable (or even larger) maternal placental sizes in relation to the mothers of both European studies. However, in comparison with the small Indian babies, the major difference noted was a substantially smaller placental size of the Pune mothers. Compared to z-scores from the World Health Organization (WHO) child growth standards (WHO Anthropo 2010), the PND study babies were born lighter and shorter with a larger head circumference (HC). A growth delay observed near the neonatal visit was followed by catch up growth in weight for age, however no catch-up growth was observed for length for age by the second postnatal visit. Dietary intakes. The PND study women had pregnancy energy intakes slightly below the national level intakes (Steyn et al. 2006), except for the energy intake at V3, which exceeded the national level. The lower energy intake during pregnancy is attributable to a lower intake of dietary protein and total carbohydrate. The fat intake was substantially higher in the PND study, with levels almost double at the second postnatal visit. This discrepancy may be due to differences between investigated populations or due to increased requirements of energy during lactation. In comparison with the Recommended Dietary Allowances (RDA), during the pregnancy period women in the current study had lesser crude protein and higher carbohydrate intake. Intake of the crude protein after delivery was low in comparison to the RDA for lactating women. Total carbohydrate intake exceeded the RDA. The relation between the intake of macronutrients expressed as a percentage of total energy (%E) and RDA was similar. The current study participants had a lower intake of energy, especially during the pregnancy period, when compared to populations in the developed world. They had a lower intake of protein at all visits in comparison with the national average for women or with that found in the Southampton study or when compared to the usual American diet. The energyadjusted intake of fat and carbohydrate were comparable with the Southampton and American data. This population in our study was therefore more comparable to that of Pune than to American or Southampton populations. However, despite similar total energy intakes as the Pune Study, total protein and fat intakes were higher in the African mothers, which may explain their higher weights and birthweights of their babies. Results – ANALYTICAL DATA Dietary intakes. Maternal dietary intakes showed significant effects on the fetal biometrical measurements, mainly involving the fetal head, femur length and the size of abdomen. The outcomes also show a significant relationship between maternal protein intake and baby’s birth weight. Associations were also found for maternal dietary intakes and the neonatal length and BMI and the markers of the infant β-cell function. Intakes of the plant protein and polyunsaturated fat supported the linear growth. There was no correlation between maternal dietary intake and the fetal growth rate. Effects of the total energy intake and carbohydrate seemed to be direct, while the effect of protein and fat may be delayed, possibly involving metabolic adaptation of the mother and the partitioning of nutrients between the mother, placenta and fetus. (See Table: Associations between the maternal dietary intake and the neonatal INSF1 levels, Pg. xiv). Maternal hormones. The data of the current study show significant relationships between maternal pregnancy hormones and fetal growth rate and the postnatal growth rate, although maternal anthropometry and fetal gender and BMI are also significantly involved. The maternal thyroid hormones seem to play an important role in fetal and postnatal growth and insulin metabolism (N=76, p=0.002, ß=-0.343; AR2=0.106) in the association with the neonatal fasting insulin. Discussion. The outcomes of the current study show that the African pregnant women in the study had lower energy intakes attributable to a lower intake of dietary protein and total carbohydrate. Maternal dietary intakes showed significant effects on the fetal biometrical parameters. Protein dietary intake was positively associated with baby’s birth weight and was shown to be statistically significant. Associations were also found for maternal dietary intakes and the infant postnatal BMI and the markers of the infant β-cell function. Quality of the protein and fat has different effects on fetal/infant growth. Maternal hormones showed correlations with fetal growth rate and the outcomes of the current study also show that maternal hormones can affect neonatal and early postnatal infant glucose levels and β-cell function. They are also linked to the programming of early obesity. The maternal thyroid hormones seem to play an important role. The maternal low energy, protein (especially plant protein) and PUFA intakes during gestation may be the reasons for the lower z-score birth parameters of the infants in comparison with the World Health Organization (WHO) child growth standards (WHO Anthro 2010). The disproportionally larger head of the newborn may be an outcome of the brain sparing effect. A suggestion for an increased maternal dietary intake of energy, protein and PUFA has been made.

Faculty of Science School of Physiology 9808215t romainh@physiology.wits.ac.za / There is a high prevalence of insulin and leptin resistance and increased cortisol concentrations in sleep apnoea patients, independent of obesity. Chronic intermittent hypoxia is used an experimental animal model to simulate the hypoxia occurring in sleep apnoea patients. The aim of this study was to measure plasma insulin and leptin concentrations and hypothalamic-pituitary-axis activity in rats exposed to either intermittent hypoxia (CIH) or sham hypoxia (SH) for fourteen days. To induce CIH plexiglass cylinders were flushed with 100% nitrogen for nine seconds every 90 seconds, seven hours/day. The rats were weighed each day during the exposure period. Venous blood samples for insulin and leptin were collected on days one, three, five, eight and fifteen. Faecal samples were collected to measure glucocorticoid metabolites. There was no significant difference in the daily change in body weight between the rats exposed to CIH compared to the rats exposed to SH (unpaired t-test). Plasma insulin concentrations were not affected by CIH. In both groups of rats plasma leptin concentrations were significantly higher on day fifteen compared to day five (p=0.03, unpaired t-test). Glucocorticoid metabolites were significantly increased in the intermittent hypoxia group on day two (p=0.003 one-way ANOVA). In conclusion, exposing normal weight rats to CIH for fourteen days resulted in a transient iv increase in HPA axis activity on day two and an elevation in plasma leptin levels, in both groups of rats, at day fifteen.