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- All Subjects: Obesity
- Creators: Katsanos, Christos
- Member of: Theses and Dissertations
Polarography was used to determine functional differences in isolated SS and IMF mitochondria between lean (37 ± 3 yrs; n = 10) and obese (35 ± 3 yrs; n = 11) subjects during either saline (control) or amino acid (AA) infusions. AA infusion increased ADP-stimulated respiration (i.e., coupled respiration), non-ADP stimulated respiration (i.e., uncoupled respiration), and ATP production rates in SS, but not IMF mitochondria in lean (n = 10; P < 0.05). Neither infusion increased any of the above parameters in muscle SS or IMF mitochondria of the obese subjects.
Using label free quantitative mass spectrometry, we determined differences in proteomes of SM SS and IMF mitochondria between lean (33 ± 3 yrs; n = 16) and obese (32 ± 3 yrs; n = 17) subjects. Differentially-expressed mitochondrial proteins in SS versus IMF mitochondria of obese subjects were associated with biological processes that regulate: electron transport chain (P<0.0001), citric acid cycle (P<0.0001), oxidative phosphorylation (P<0.001), branched-chain amino acid degradation, (P<0.0001), and fatty acid degradation (P<0.001). Overall, these findings show that obesity is associated with redistribution of key biological processes within the mitochondrial reticulum responsible for regulating energy metabolism in human skeletal muscle.
Assessment of DNA methylation was performed on human skeletal muscle and blood using reduced representation bisulfite sequencing (RRBS) for high-throughput identification and pyrosequencing for site-specific confirmation. Sorbin and SH3 homology domain 3 (SORBS3) was identified in skeletal muscle to be increased in methylation (+5.0 to +24.4 %) in the promoter and 5’untranslated region (UTR) in the obese participants (n= 10) compared to lean (n=12), and this finding corresponded with a decrease in gene expression (fold change: -1.9, P=0.0001). Furthermore, SORBS3 was demonstrated in a separate cohort of morbidly obese participants (n=7) undergoing weight-loss induced by surgery, to decrease in methylation (-5.6 to -24.2%) and increase in gene expression (fold change: +1.7; P=0.05) post-surgery. Moreover, SORBS3 promoter methylation was demonstrated in vitro to inhibit transcriptional activity (P=0.000003). The methylation and transcriptional changes for SORBS3 were significantly (P≤0.05) correlated with obesity measures and fasting insulin levels. SORBS3 was not identified in the blood methylation analysis of lean (n=10) and obese (n=10) participants suggesting that it is a muscle specific marker. However, solute carrier family 19 member 1 (SLC19A1) was identified in blood and skeletal muscle to have decreased 5’UTR methylation in obese participants, and this was significantly (P≤0.05) predicted by insulin sensitivity.
These findings suggest SLC19A1 as a potential blood-based biomarker for obese, insulin resistant states. The collective findings of SORBS3 DNA methylation and gene expression present an exciting novel target in skeletal muscle for further understanding obesity and its underlying insulin resistance. Moreover, the dynamic changes to SORBS3 in response to metabolic improvements and weight-loss induced by surgery.
Over 40% of adults in the United States are considered obese. Obesity is known to cause abnormal metabolic effects and lead to other negative health consequences. Interestingly, differences in metabolism and contractile performance between obese and healthy weight individuals are associated with differences in skeletal muscle fiber type composition between these groups. Each fiber type is characterized by unique metabolic and contractile properties, which are largely determined by the myosin heavy chain isoform (MHC) or isoform combination that the fiber expresses. In previous studies, SDS-PAGE single fiber analysis has been utilized as a method to determine MHC isoform distribution and single fiber type distribution in skeletal muscle. Herein, a methodological approach to analyze MHC isoform and fiber type distribution in skeletal muscle was fine-tuned for use in human and rodent studies. In the future, this revised methodology will be implemented to evaluate the effects of obesity and exercise on the phenotypic fiber type composition of skeletal muscle.
Seven human subjects with body mass indices (BMIs) ranging from 19.4 kg/ m2 to 26.7 kg/ m2 and six human subjects with BMIs ranging from 32.1 kg/ m2 to 37.6 kg/ m2 were recruited and subjected to 45-minute bouts of acute exercise to look at the changes in the plasma concentration of the dopamine metabolite homovanillic acid (HVA) in response to acute physical activity. Plasma HVA concentration was measured before exercise and during the last 10 minutes of the exercise bout via competitive ELISA. On average the optical density (OD) of the samples taken from lean subjects decreased from 0.203 before exercise to 0.192 during exercise, indicating increased plasma HVA concentration. In subjects with obesity OD increased from 0.210 before exercise to 0.219 during exercise, indicating reduced plasma HVA concentration. These differences in OD were not statistically significant. Between the lean group and the group with obesity no significant difference was observed between the OD of the plasma samples taken before exercise, but a significant difference (p = 0.0209) was observed between the ODs of the samples taken after exercise. This indicated that there was a significant difference between the percent changes in OD between the lean group and the group with obesity, which suggested that there may be a body weight-dependent difference in the amount of dopamine released in response to exercise. Because of the lack of significance in the changes in OD within the lean group and the group with obesity the results of this study were insufficient to conclude that this difference is not due to chance, but further investigation is warranted.
The prevalence of obesity continues to increase in the United States, along with its risk for other associated cardiovascular and metabolic diseases. Several therapeutic methods are aimed at targeting and reducing obesity, now defined as a state of chronic, low-grade inflammation (in addition to BMI > 30 kg/m2). In an attempt to expand on these therapeutic methods, research on the concept of browning in white adipose tissue (WAT) and brown adipose tissue (BAT) is being conducted. Brown adipose tissue (BAT), and a newly discovered type of adipocyte, beige adipocytes, are heavily involved in thermogenesis with the use of uncoupling protein-1 (UCP-1). This paper focuses on the analysis of common browning genes, ATP-related genes, and metabolic genes in varying biological groups in mice (Chow/High-Fat Diet and Inguinal FAT and Perigonadal Fat) and in humans (Lean/Obese and Subcutaneous WAT (SC) and Omental WAT (OM)) using methods such as RT-PCR and immunohistochemistry. The data obtained shows an increase in browning in the leaner group, specifically in the subcutaneous fat. Further, browning is significantly reduced in the obese groups of subjects and mice tested, in addition to omental/perigonadal versus subcutaneous/inguinal fat depots. Interestingly, two key ATP genes, UCP-1 and COX4I1 are vastly elevated in the OM WAT, indicating that browning may not be as important in the OM, but rather may have a potential role in SC. This is contrary to prior research findings that attempt to exclude mice surrogates in future experimentation of the browning phenomenon. Further experimentation is needed to expand on the findings of this paper.