Article Title



Seongkyun Lim, David E. Lee, Megan E. Rosa-Caldwell, Jacob L. Brown, Tyrone A. Washington, Nicholas P. Greene.

University of Arkansas, Fayetteville, AR

Type 2 Diabetes Mellitus (T2DM) is a fast-growing epidemic and skeletal muscle insulin resistance may be the onset point in the development of T2DM. Recent data have suggested that microRNAs (miR) may play an important role in T2DM glucose intolerance. Specifically, reduced miR-16 content in muscle has been noted in human and murine models of T2DM. However, regulation of miR-16 and its relation to skeletal muscle insulin resistance is largely unexplored. PURPOSE: To investigate how miR-16 content affects insulin resistance and glucose regulation in myotubes during normal and insulin resistant states. METHODS: This study was performed in two experiments. Experiment 1: To test if miR-16 is necessary for muscle insulin sensitivity C2C12 myoblasts were plated in six well plates and differentiated upon confluence. Cells were transfected with a plasmid to inhibit function of miR-16. Experiment 2: To test if miR-16 is sufficient to ameliorate insulin resistance C2C12 were similarly differentiated and transfected with plasmid to overexpress functional miR-16, cells were then treated with a 1-oleoyl-2-acetyl-sn-glycerol (OAG), a diacylglycerol/DAG analog to stimulate lipid overload-induced insulin resistance. For both experiments cells were stimulated with insulin and measures of glucose uptake and insulin signaling were measured via uptake of 2-NBDG (a fluorescent analog of glucose), and immunoblotting of the insulin signaling cascade. Data were analyzed by ANOVA or t-test as appropriate, significance was denoted at p<0.05. RESULTS: Experiment 1: Insulin-stimulated glucose uptake was ~25% lower in myotubes following miR16 inhibition compared to control (p=0.01). Furthermore, insulin signaling was lower in myotubes with miR16 inhibition compared to control (31%, p=0.002). Experiment 2: OAG-induced insulin resistant myotubes in vitro exhibited significantly lower glucose uptake (p=0.01) compared to control (12%). However, overexpression of miR16 did not improve OAG-induced impaired glucose uptake (p>0.05). CONCLUSION: miR-16 may be necessary for appropriate glucose handling, however miR-16 is not sufficient to rescue glucose regulation during pathological insulin resistance.

ACKNOWLEDGEMENTS: This study was funded by the Arkansas Bioscience Institute and American College of Sports Medicine Research Endowment Grant.

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