During the 1800’s cannabis use was described as a treatment for a variety of metabolic disorders but its recreational use in the twentieth century resulted in laws which made the usage of cannabis illegal despite its medicinal properties. Cannabis usage has been reported to be useful in the treatment of Type 2 diabetes but unfortunately conflicting results are often published and its mechanism of action is still unknown. The aim of this project was to investigate the effect of phytocannabinoid treatment on adipogenesis and lipolysis in 3T3-L1 cells, to unravel their mechanism of action and also to test for potential anti-diabetic properties. The studies showed that phytocannabinoid treatment promoted higher glucose uptake and significantly less fat accumulation when compared to Rosiglitazone. Rosiglitazone is an anti-diabetic drug that has recently been withdrawn from the market since its usage has been associated with severe side effects. It was also found that during the 1800’s cannabis use was described as a treatment for a variety of metabolic disorders but its recreational use in the twentieth century resulted in laws which made the usage of cannabis illegal despite its medicinal properties. Cannabis usage has been reported to be useful in the treatment of Type 2 diabetes but unfortunately conflicting results are often published and its mechanism of action is still unknown. The aim of this project was to investigate the effect of phytocannabinoid treatment on adipogenesis and lipolysis in 3T3-L1 cells, to unravel their mechanism of action and also to test for potential anti-diabetic properties. The studies showed that phytocannabinoid treatment promoted higher glucose uptake and significantly less fat accumulation when compared to Rosiglitazone. Rosiglitazone is an anti-diabetic drug that has recently been withdrawn from the market since its usage has been associated with severe side effects. It was also found that phytocannabinoid treatment was able to reverse the insulin-resistant state of 3T3-L1 cells. The study indicates that the mechanism of action occurs at the mitochondrial level where enzymes such as succinate dehydrogenase and glycerol-3-phosphate dehydrogenase are modulated thereby affecting oxidative phosphorylation involved in the respiratory chain. In addition the effect observed with phytocannabinoid treatment is time dependent and affects the cells differently at different developmental stages. Therefore it can be concluded that phytocannabinoid treatment not only helps to maintain the balance between adipogenesis and lipolysis in 3T3-L1 cells but its use may also be helpful in the treatment of Type 2 diabetes and/or obesity-related insulin resistance.phytocannabinoid treatment was able to reverse the insulin-resistant state of 3T3-L1 cells. The study indicates that the mechanism of action occurs at the mitochondrial level where enzymes such as succinate dehydrogenase and glycerol-3-phosphate dehydrogenase are modulated thereby affecting oxidative phosphorylation involved in the respiratory chain. In addition the effect observed with phytocannabinoid treatment is time dependent and affects the cells differently at different developmental stages. Therefore it can be concluded that phytocannabinoid treatment not only helps to maintain the balance between adipogenesis and lipolysis in 3T3-L1 cells but its use may also be helpful in the treatment of Type 2 diabetes and/or obesity-related insulin resistance.

No description available.

Thesis (Ph. D.)--Ohio State University, 2007. / Includes bibliographical references (p. 139-160).

published_or_final_version / abstract / Physiology / Master / Master of Philosophy

published_or_final_version / Medicine / Master / Master of Philosophy

Background: Regional adiposity and in particular central adiposity is associated with a hazardous metabolic profile, insulin resistance (IR), hypertension and increased cardiovascular mortality whilst gluteo-femoral fat (peripheral fat) is protective. Extreme phenotypes, especially those due to monogenic disorders, are experiments of nature, which have been pivotal in identifying genes involved in disease. The hypothesis tested in this thesis was that investigating individuals with monogenic causes of regional adiposity and IR would identify genes and physiological processes contributing to their phenotype. Methods: Two families with balanced translocations, sixty-nine patients with partial lipodystrophy and eight individuals with lipoma were investigated. Genetic investigations included mapping of balanced translocations using fluorescent in-situ hybridisation, DNA and cDNA sequencing and gene expression studies. In vivo investigations of adipose tissue included microdialysis and measurement of adipose tissue blood flow. Results: Genetic and physiological factors contributing to regional adipose tissue deposition and IR were identified in (1) individuals with lipodystrophy where mutations were identified in eleven families including one novel PPARG mutation V450M. Also a skinfold ratio was developed for the clinical diagnosis of partial lipodystrophy (2) a family with a balanced I translocation, IR and growth retardation, digenic disruption of INSR and CHN2 was identified, which accounted for the clinical phenotype (3) a family with a balanced translocation, central adiposity and peas associated with disruption of KIBRA (4) lipoma tissue relative to adjacent healthy subcutaneous tissue, where differential gene expression profiles were identified, we also demonstrated reduced metabolic flexibility and reduced lipolysis in lipoma relative to healthy adipose tissue despite no change in adipose tissue blood flow. Conclusion: Investigating individuals with extreme phenotypes and monogenic causes of regional adiposity and IR identified key genetic and physiological factors.

by Pik-ying Ng. / Bibliography: leaves 106-117 / Thesis (M.Phil.)--Chinese University of Hong Kong, 1983

Bisphenol A (BPA) is a common component in plastic consumer products and epoxy resin linings. Initially developed in the 1930s-40s as a synthetic hormone treatment, it is now widely considered an endocrine disrupting chemical (EDC). A growing body of epidemiological literature suggests that ubiquitous exposures to BPA may be contributing to the global epidemic of obesity, with children a particularly vulnerable population. Obesity in children, defined by a body mass index (BMI) greater than or equal to the 95th percentile for age and sex, is an epidemic of great concern in the United States. As with other chemicals, the prenatal and early life period are critical windows of exposure to BPA; however, the mechanism by which BPA may influence the development of body size in children remains unclear. Experimental studies have found that BPA influences adipogenesis in both murine and adult human preadipocyte cell lines and BPA is hypothesized to play a role in enhancing adipogenic regulation by nuclear receptors such as peroxisome proliferator-activated receptor gamma (PPARγ). While the timeline of the processes involved in adipogenesis in humans is not universally agreed upon, it is accepted that PPARγ is highly expressed in adipose tissue and considered to be the master regulator of adipogenesis. To answer the question of both timing and developmental origin of BPA effects on adipogensis, we employed both an epidemiological approach, and experimental methodologies using primordial cell lines, mesenchymal stem cells (MSCs). Our study characterizes early life exposures to BPA, explores the adipogenic mechanism of BPA in human MSCs via cellular morphometrics and PPARγ gene expression, and identifies associations between early life exposure to BPA and childhood obesity and adiposity. For our epidemiological assessments, we studied a birth cohort of African American and Dominican mother and child dyads in New York City. BPA was measured in spot urine samples collected during pregnancy and at child ages 3, 5, and 7 years, from mothers and children (n=568 dyads) in the Columbia Center for Children’s Environmental Health (CCCEH). We compared BPA concentrations across paired samples. We explored relationships between BPA and the class of phthalate chemicals, another common plasticizer. BPA was detected in nearly all urine samples from prenatal third trimester and childhood ages 3 years, 5 years and 7 years. Prenatal urinary BPA concentrations were significantly lower than postnatal urinary BPA concentrations (p<0.001). BPA and phthalate metabolites were correlated prenatally and at 3, 5, and 7 years (all p-values < 0.02). BPA concentrations were correlated with phthalate metabolite concentrations prenatally, and at 3, 5 and 7 years(all p-values < 0.05). Geometric means of BPA were higher among African Americans than among Dominicans in prenatal (p<0.01), 5 year (p<0.001) and 7 year (p=0.02) samples. Postnatal BPA concentrations were significantly higher among children with mothers who had never marrried marital status and were significantly higher in summer than in all other seasons (all p-values < 0.05). These findings reveal widespread BPA exposure in an inner-city minority population. Our in vitro experiment was a feasibility study which sought to determine whether exposure to BPA by human umbilical cord mesenchymal stem cells (HUMSC) induces morphological changes and PPARγ gene expression during adipogenesis. An anonymous sample of n=18 umbilical cords was collected at delivery from mothers registered at New York-Presbyterian Sloane Hospital for Women and New York-Presbyterian Allen Hospital in New York City. HUMSCs were harvested from umbilical cords using an adhesion technique. HUMSCs were then induced in culture to differentiate into adipocytes using: a standard differentiation induction mix medium, a negative vehicle control medium, a positive control medium and experimental control media. Differences in cell surface area and cell count in all cultures were assessed using ImageJ software (version 1.49n, 2014). Gene expression of PPARγ in all cultures was evaluated by RT-PCR. Cell morphometric results were based on 11,676 cells from 3 umbilical cord samples. PPARγ1 and PPARγ2 gene expression was assessed during differntiation phase and early terminal phase adipogenesis (0 to 72 hours). Cell morphometrics were assessed during middle to late terminal phase adipogenesis (days 14 and 21). No differences in cell count were observed for experimental conditions compared to standard induction medium. A significant decrease in surface area was seen in cells exposed to 100 μM concentration of BPA as compared to exposure to standard induction medium at day 14 (t=-37.02 p=0.001). Differences in cell surface area were not observed at day 21. A twofold increased expression of PPARγ1 was observed in cells exposed to 10 μM concentration of BPA by 72 hours of adipogenic induction which was higher than the increase in expression observed for cells exposed to the positive control induction medium containing 10 μM concentration of rosiglitazone. All induction media conditions had negligible effects on PPARγ2 expression. As BPA increases expression of PPARγ1 in HUMSCs during the transition into the early terminal differentiation phase of adipogenesis, HUMSCs may be an approximate target tissue for evaluating BPA effects in adipogenesis. Finally, using a longitudinal research design, we analyzed the possible effect of prenatal and postnatal BPA exposures, measured in urine, on childhood anthropometric outcome measures. Participants in the CCCEH have been followed since the third trimester of pregnancy, providing us with anthropometric data on children from birth through the age of seven years. Available anthropometric outcome measures include body mass index z-scores (BMIZ) at 5 and 7 years, as well as fat mass index (FMI), percent body fat (%BF), and waist circumference (WC) at 7 years. Prenatal urinary BPA concentrations were positively associated with child age 7 FMI (beta=0.31 kg/m2, p-value=0.04, [95%CI 0.01, 0.60]), %BF (beta=0.79, p-value=0.04, [95%CI 0.03, 1.55]), and WC (beta=1.29 cm, p-value=0.01, [95%CI 0.29, 2.30]). Child urinary BPA concentrations were not associated with childhood BMI or other anthropometric outcomes. As the prenatal exposures were associated with childhood measures of adiposity, prenatal BPA exposure may have an effect on adiposity as children age that cannot be determined by the use of BMI alone. Our results suggest BPA may contribute to the developmental origins of obesity and adiposity.

Wu, Hoi Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2010. / Includes bibliographical references (leaves 92-104). / Abstracts in English and Chinese. / Thesis/ Assessment Committee --- p.i / Declaration --- p.ii / Acknowledgments --- p.vi / Table of Content --- p.vii / List of Abbreviations --- p.x / List of Figures --- p.xiv / List of Tables --- p.xv / Chapter 1. --- Literature Review --- p.1 / Chapter 1.1 --- Introduction of diabetes mellitus --- p.1 / Chapter 1.1.1 --- Definition and prevalence --- p.1 / Chapter 1.1.2 --- Diagnosis and classification --- p.2 / Chapter 1.1.3 --- Symptoms and complications --- p.4 / Chapter 1.1.4 --- Cause and risk factors --- p.5 / Chapter 1.1.5 --- Prevention and treatment --- p.9 / Chapter 1.2 --- The role of adipose tissue in pathophysiology of T2DM --- p.10 / Chapter 1.2.1 --- Randle's glucose-fatty acid hypothesis --- p.11 / Chapter 1.2.2 --- Ectopic fat storage hypothesis --- p.12 / Chapter 1.2.3 --- Adipose tissue as an endocrine organ --- p.13 / Chapter 1.2.4 --- Low-grade inflammation --- p.15 / Chapter 1.2.5 --- Endoplasmic reticulum (ER) stress --- p.17 / Chapter 1.3 --- Use of berberine in the treatment of T2DM --- p.18 / Chapter 1.3.1 --- Efficacy of berberine in treating diabetes --- p.18 / Chapter 1.3.2 --- Berberine on glucose and lipid metabolism of animals --- p.19 / Chapter 1.3.3 --- Inhibition of adipogenesis --- p.20 / Chapter 1.3.4 --- Activation of AMP-Activated Protein Kinase (AMPK) --- p.20 / Chapter 1.3.5 --- Mitochondrial inhibition --- p.21 / Chapter 1.4 --- Introduction of proteomics --- p.21 / Chapter 1.4.1 --- Why proteomics? --- p.22 / Chapter 1.4.2 --- Gel-based proteomics: Two-Dimensional Gel Electrophoresis --- p.23 / Chapter 1.4.3 --- Gel-free proteomics --- p.25 / Chapter 1.4.4 --- Mass spectrometry --- p.26 / Chapter 1.4.5 --- Proteomics as tool for diabetes research --- p.27 / Chapter 1.5 --- Objectives and significance --- p.32 / Chapter 2. --- Materials and Methods --- p.34 / Chapter 2.1 --- Drug preparation --- p.34 / Chapter 2.2 --- Animal experiment --- p.34 / Chapter 2.3 --- Comparison of proteome of visceral white adipose tissue: obese db/db micevs lean m+/db mice and BBR-treated vs control db/db mice --- p.36 / Chapter 2.3.1 --- Protein sample preparation from adipose tissue --- p.36 / Chapter 2.3.2 --- Protein quantitation --- p.37 / Chapter 2.3.3 --- 2D Gel electrophoresis --- p.37 / Chapter 2.3.4 --- Image analysis --- p.39 / Chapter 2.3.5 --- In-gel digestion and MALDI-ToF MS --- p.39 / Chapter 2.4 --- Cell culture experiment --- p.40 / Chapter 2.5 --- Oil Red O staining --- p.42 / Chapter 2.6 --- Glycerol determination --- p.42 / Chapter 2.7 --- Comparison of proteomes of BBR-treated and control 3T3-L1 adipocytes..… --- p.43 / Chapter 2.7.1 --- Protein sample preparation from 3T3-L1 cells --- p.43 / Chapter 2.7.2 --- Protein quantitation --- p.43 / Chapter 2.7.3 --- 2D Gel electrophoresis --- p.44 / Chapter 2.7.4 --- Image analysis --- p.44 / Chapter 2.7.5 --- In-gel digestion and MALDI-ToF MS --- p.44 / Chapter 2.8 --- Western Immunoblotting --- p.44 / Chapter 2.8.1 --- Protein sample preparation of BBR-treated and control 3T3-L1 --- p.44 / Chapter 2.8.2 --- SDS-PAGE --- p.44 / Chapter 2.8.3 --- Protein blotting --- p.45 / Chapter 2.8.4 --- Membrane blocking and antibody incubations --- p.45 / Chapter 2.8.5 --- Detection of Proteins --- p.46 / Chapter 2.9 --- Statistical analysis --- p.46 / Chapter 3. --- Results --- p.47 / Chapter 3.1 --- Comparison of total protein profiles of visceral adipose tissue of obese db/db and lean m+/db mice --- p.47 / Chapter 3.2 --- Effect of berberine on glucose metabolism of obese db/db mice --- p.53 / Chapter 3.3 --- Comparison of the protein profiles of visceral adipose tissue of BBR-treated and control db/db mice --- p.55 / Chapter 3.4 --- Effect of berberine treatment on 3T3-L1 adipocytes --- p.61 / Chapter 3.4.1 --- Berberine treatment inhibited intracellular triglyceride accumulation in both mature and pre-mature 3T3-L1 adipocytes --- p.61 / Chapter 3.4.2 --- Berberine treatment enhanced lipolysis in mature 3T3-L1 adipocytes but inhibited lipolysis in pre-mature 3T3-L1 adipocytes --- p.65 / Chapter 3.4.3 --- Color change in culture media after berberine treatment --- p.65 / Chapter 3.4.4. --- Comparison of protein profiles between berberine-treated and control 3T3-L1 adipocytes --- p.67 / Chapter 3.4.5 --- Western blotting --- p.73 / Chapter 4. --- Discussion --- p.75 / Chapter 4.1 --- Comparison of total protein profiles of visceral adipose tissue of obese db/db and lean m+/db mice --- p.75 / Chapter 4.2 --- "Berberine lowers body weight, reduces fasting blood glucose level and improves glucose-lowering ability of db/db mice" --- p.78 / Chapter 4.3 --- Comparison of the protein profiles of visceral adipose tissue of BBR-treated and control db/db mice --- p.79 / Chapter 4.4 --- Berberine inhibited lipid accumulation in mature and pre-mature 3T3-L1 adipocytes --- p.84 / Chapter 4.5 --- Berberine enhanced lipolysis in mature 3T3-L1 adipocytes but inhibited lipolysis in pre-mature 3T3-L1 adipocytes --- p.84 / Chapter 4.6 --- Comparison of the protein profiles of BBR-treated and control 3T3-L1 adipocytes --- p.85 / Chapter 4.7 --- Western blotting --- p.88 / Chapter 4.8 --- General discussion --- p.89 / Chapter 5. --- References --- p.92

Thesis (M.S.)--University of North Carolina at Wilmington, 2004. / Includes bibliographical references.