The aim of this thesis is to investigate different functionalization strategy of graphene nanomaterials for graphene-based different biotechnological applications such as graphene-directed stem cell growth and differentiation and graphene-based biosensors. Chemical functionalization of graphene is required in many biological applications; in this thesis we have focused on exploiting the carboxylic groups available on GO molecules and non-covalent functionalization of graphene. GO has been a promising material for stem cell culture due to high specific surface area, ease of functionalization, its ability to support cell proliferation and to not cause cytotoxicity when stem cells are cultured on its substrate. The impact of biochemical functionalization on stem cell differentiation was not widely researched, and many research groups worldwide have been focusing on GO and rGO surfaces only. The approach of this thesis is to fabricate and characterize different graphene-based substrates to investigate the impact of biochemical functionalization of GO in directing adipose stem cell differentiation and to influence the gene expression pathways of Schwann-like differentiated adipose stem cells. The fabrication of graphene based biosensors is still challenging as biological molecules need to be attached to graphene-based sensors to increase both the specificity and the selectivity of the biosensors. In this thesis, two different chemical functionalization approaches were considered. Firstly, the covalent immobilization of membrane proteins embedded on a lipid nanodisc structure on GO was achieved. Secondly, the feasibility of using dip-pen nanolithography as a tool to locally functionalize graphene arrays with phospholipids was demonstrated. Phospholipid interface layer can act as bioactive layer which can be used for the protein insertion of tail-anchoring recombinant proteins as a new route for a non-covalent biological functionalization of graphene array.

Thesis (M.S.)--University of Delaware, 2007. / Principal faculty advisor: Christopher M. Modlesky, Dept. of Health, Nutrition, and Exercise Sciences. Includes bibliographical references.

Thesis (Ph. D.)--University of North Carolina at Greensboro, 2006. / Advisor: Ron Morrison; submitted to the School of Human Environmental Sciences. Includes bibliographical references.

Pasture feeding depresses adipose tissue development in beef cattle whereas grain feeding, enhances adipogenesis. Therefore, we hypothesized that specific fatty acids would differentially affect lipogenesis in explants of bovine subcutaneous (SC) and intramuscular (IM) adipose tissues. Angus steers were harvested at 12, 14, and 16 mo of age, and IM and SC adipose tissue explants from the 8-11th thoracic rib region were dissected and cultured in media. Media contained no supplemental fatty acids or 40 microM of five fatty acids, stearic acid (18:0), oleic acid (18:1 n-9), trans-11 vaccenic acid (18:1 trans-11), conjugated linoleic acid (CLA, 18:2 trans-10, cis-12), or alpha-linolenic acid (18:3 n-3). After 48 h of culture, lipogenesis using [U-14C]glucose and [1-14C]acetate was measured. Lipogenesis from glucose decreased between 12 and 16 mo of age in SC adipose tissue (from 8.9 to 4.0 nmol per 2 h per 100 mg; P = 0.001) and IM adipose tissue (from 4.4 to 2.7 nmol per 2 h 100 mg ; P = 0.08). Lipogenesis from acetate did not change over time in SC adipose (approximately 56 nmol per 2 h per 100 mg; P = 0.23), but increased over time in IM adipose tissue (from from 11.3 to 17.1 nmol per 2 h 100 mg; P = 0.02). Oleic acid increased lipid synthesis from glucose 125 percent (P = 0.04) in IM adipose tissue, whereas stearic acid and trans-vaccenic acid increased lipogenesis from glucose in SC adipose tissue by approximately 50 percent (P = 0.04). In SC adipose tissue only, trans-vaccenic and increased, lipogenesis from glucose (P < 0.02). Lipogenesis from acetate was depressed by CLA nearly 50 percent in SC adipose tissue. PPARγ gene expression increased between 14 and 16 mo of age in control IM and SC adipocytes. The increase in activity was also observed in AMPK gene expression. C/EBPβ and SCD gene expression did not increase in control samples until 16 mo of age. SC adipose tissue responded to stearic acid by increased GPR43 and AMPK gene expression at 12 mo of age. We conclude that fatty acids differentially affect lipid synthesis in IM and SC adipose tissues, which may account for the effects of pasture and grain feeding on adiposity.

We hypothesized that subcutaneous (s.c.) adipose tissue would differ in monounsaturated (MUFA) and saturated fatty acid (SFA) composition among different depots throughout a beef carcass. To test this, 50 carcasses from a variety of breed types and backgrounds were sampled. External fat samples were collected from eight different carcass locations: round, sirloin, loin, rib, chuck, brisket, plate and flank. Samples were used to provide information on slip points, fatty acid composition and MUFA:SFA ratios. Lipids were extracted from s.c. adipose tissue by a modified chloroform:methanol procedure, and fatty acid composition and slip points were measured. The brisket was significantly lower in palmitic (16:0) and stearic (18:0) acid than the other seven sampling sites (P = 0.001). The brisket demonstrated the highest values of MUFA (P = 0.001) with the exception of possessing the lowest value of transvaccenic (18:1t11) acid (P = 0.002). There were also significant differences in the amounts of PUFA among the eight sampling sites. The lowest values were from the brisket with a mean of 25.1. The flank had the highest slip point with a mean of 39.0 (P < or = 0.001). There was a high negative correlation shown between palmitoleic and stearic acid (R2 = 0.827). The brisket displayed the highest values for MUFA:SFA ratios (P = 0.001), whereas the flank was the lowest. Due to the significant differences amongst fat depots within bovine carcasses in their fatty acid composition we conclude that substantial differences exist across fat depots.

Thesis (Ph. D.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 118-145). Also available in print.

Background and objectives: Circulating endothelial progenitor cells (EPCs) play an essential role in maintaining vascular integrity and preventing endothelial dysfunction. Decreased circulating EPC levels are frequently observed in various cardiovascular risks, including aging and diabetes. Endothelial nitric oxide synthase (eNOS) and adiponectin exert their vasculo-protective effects by directly targeting the key components of the vascular system, such as endothelial cells and smooth muscle cells. Both eNOS and adiponectin have been implicated in the mobilization and in vitro functions of EPCs. However, whether and how circulating EPCs are involved in eNOS and adiponectin-mediated vascular protection remain unclear. The objective of this study is to investigate the role of circulating EPCs in eNOS and adiponectin-mediated regulation of vascular integrity after arterial injury under both physiological and pathophysiological conditions, and to elucidate the underlying mechanisms involved. Key findings: 1. Modulation of eNOS activity in vivo by replacing the serine 1176 (S1176) with an aspartate (S1176D mutation or Dki) to mimic phosphorylation or with an alanine (S1176A mutation or Aki) to render it unphosphorylatable altered reendothelialization and subsequent endothelial function after arterial injury in mice. 2. eNOS S1176D mutation increased the number of circulating EPCs and their incorporation into regenerated endothelium, whereas eNOS S1176A or knockout (KO) impaired the mobilization and reendothelializing capacity of circulating EPCs after injury. 3. eNOS S1176D elevated circulating EPCs by promoting the proliferation and differentiation of bone marrow hematopoietic stem cells (HSCs) into EPCs and by inhibiting apoptosis of circulating EPCs. 4. Adiponectin deficiency in mice resulted in progressive decrease of circulating EPCs with aging. Systemic administration of recombinant adiponectin reversed the decreased EPCs number in adiponectin KO mice. In db(-/-) diabetic mice, adiponectin deficiency further reduced circulating EPCs number and subsequent reendothelialization after injury. Rosiglitazone (Rosi), an antidiabetic drug, induced an upregulation of EPCs number and improved reendothelialization, which were partially abolished in the absence of adiponectin. 5. In cultured EPCs, adiponectin significantly inhibited high glucose-induced premature senescence, whereas its effects on proliferation and apoptosis were not evident. High glucose instigated EPCs senescence by increasing the intracellular accumulation of reactive oxygen species (ROS), activation of p38 MAPK and expression of p16INK4A, whereas all these changes could be abolished by adiponectin through adenosine monophosphate (AMP)-activated protein kinase (AMPK) and cyclic AMP (cAMP)/protein kinase A (PKA)-dependent pathways. 6. Compared to cells from db(-/-) diabetic mice, bone marrow EPCs isolated from db(-/-) plus adiponectin double KO (DKO) mice were more susceptible to high glucose-evoked senescence, which were abrogated by adiponectin in vitro. Importantly, chronic administration of adiponectin or the anti-oxidant N-acetylcysteine (NAC) prevented both aging and diabetes-associated elevation of p16INK4A and decline of circulating EPCs in DKO mice. Conclusions: Collectively, the current study demonstrates that circulating EPCs are actively involved in the vasculo-protective effects of both eNOS and adiponectin under physiological and pathological conditions. These findings enrich our knowledge of the versatile functions of eNOS and adiponectin in vascular protection and provide solid scientific evidence supporting the use of eNOS and adiponectin as possible therapeutic targets for cardiovascular diseases. / published_or_final_version / Medicine / Doctoral / Doctor of Philosophy

Brown adipose tissue (BAT) is a unique organ in existence in mammals. It can induce non-shivering thermogenesis to control body temperature and energy balance through the expression of uncoupling protein 1 (UCP1). In our study, we aimed to evaluate the prevalence of BAT, as detected by fluorine 18-fluorodeoxyglucose (18F-FDG) positron emission tomography combined computer tomography (PET/CT), in a Hong Kong Chinese population. We also assessed the influence of age and sex to BAT in Hong Kong Chinese population. We also determined the factors associated with it, in particular, its relationship with overweight and other metabolic disorders such as diabetes mellitus. We analyzed 1765 consecutive 18F-FDG PET-CT scans of 1442 Chinese for the presence of BAT. Comparison of the variables between positive and negative BAT scans was performed using Student’s t-test. The association between maximum value of standardized uptake value (SUVmax) and variables were explored by Spearman correlation. The predictors of observed BAT were analyzed by multiple logistic regression to determine the significant predictors of positive BAT. The relationship between the monthly numbers of subjects with BAT and the respective mean monthly outdoor temperature was evaluated by Pearson’s correlation co-efficient. P < 0.05 was considered to be statistically significant. Brown adipose tissue was detected in 66 out of 1442 subjects (4.6%). BAT was significantly more commonly found in younger (43.7±13.5 years old vs. 61.4±14.2 years old, P<0.001) and female (59% vs. 46%, P<0.05) subjects. BAT also existed more frequently in subjects with lower body mass index (BMI) (21.2±3.1 kg/m2 vs. 22.4±3.7 kg/m2, P<0.01) and lower blood glucose level (5.9±0.9 mmol/L vs. 6.4±1.6 mmol/L, P<0.01). Also, BAT was detected only in subjects with no history of diabetes meallitus (DM) (0 vs. 10%, P<0.01). Moreover, lower outdoor temperature (21.6±4.6。C vs. 23.4±4.7。C, P<0.005) resulted in higher prevalence of detected BAT. In the multiple logistic regression test, age and mean monthly temperatures were the significant independent predictors of the presence of BAT (P< 0.001 and P=0.001). Age was also significantly correlated to SUVmax (P< 0.001). The monthly prevalence of positive BAT correlated negatively with mean monthly temperature by Pearson’s correlation (r = -0.79; P<0.01). To summarize, BAT was more commonly found in young, female subjects with lower BMI and blood glucose levels, and non-diabetes subjects. Age was the most important factor associated with the prevalence of BAT in humans. Lower outdoor temperature in winter can increase the prevalence of BAT even in Hong Kong’s sub-tropical climates. Also, there was an association of BAT with normal BMI (<=23) and lower blood sugar levels supporting the notion that BAT may potentially be a therapeutic target for obesity and diabetes. / published_or_final_version / Diagnostic Radiology / Master / Master of Philosophy

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