Date of Award

5-31-2018

Document Type

Campus Access Thesis

Degree Name

Master of Science (MS)

Department

Exercise and Health Science

First Advisor

Kai Zou

Second Advisor

Rachel Drew

Third Advisor

Tongjian You

Abstract

Mitochondrial networks are maintained via mitochondrial quality control (MQC) processes, including biogenesis, fusion, fission, and mitophagy. There is evidence suggesting that MQC plays a critical role in regulating mitochondrial function in skeletal muscle, and disruptions of these processes accompany the presence of skeletal muscle insulin resistance and type 2 diabetes (T2D). However, whether disruptions of MQC processes are retained at the muscle cell level is unknown. The purpose of this study was to determine whether the expression of proteins involved in MQC processes and morphologies of mitochondrial networks differed in primary human myotubes derived from severely obese non-diabetic and severely obese T2D humans, and to determine the relationship between MQC protein content and skeletal muscle glucose metabolism. Primary human skeletal muscle cells were isolated from muscle biopsies from ten lean, ten severely obese non-diabetic, and nine severely obese T2D individuals, and were differentiated to myotubes. On day seven of differentiation, myotubes were utilized for measurements of glucose metabolism and immunoblot analysis to assess the expression of proteins involved in MQC processes. Mitochondrial morphology was measured via fluorescence labeling with Mitotracker Red and quantified using Image J mitochondrial network analysis macro. The percent change in glycogen synthesis and glucose oxidation from basal to insulin stimulated conditions was significantly greater in myotubes from both lean and severely obese (P < 0.05), but not in myotubes from T2D humans. Additionally, mitochondrial content (OXPHOS) and biogenesis (PGC-1) were not different between groups. Mitochondrial fusion protein Optic Atrophy Protein 1 (Opa1) was significantly lower in myotubes derived from T2D subjects compared to severely obese non-diabetics (P < 0.05). Mitochondrial networks were significantly smaller in myotubes from T2D subjects compared to lean controls (P < 0.05). No significant differences were found in expression of other MQC proteins. There was a trend toward increased fusion (Opa1) protein content in the severely obese compared to lean group (P = 0.064). These data suggest that mitochondrial quality control is dysregulated in primary myotubes from both severely obese non-diabetic and T2D individuals, presented by mitochondrial hyperfusion that is later followed by network fragmentation with the development of T2D.

Comments

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