Abstract
Insulin action in adipose tissue is crucial for whole-body glucose homeostasis, with insulin resistance being a major risk factor for metabolic diseases such as type 2 diabetes. Recent studies have proposed mitochondrial oxidants as a unifying driver of adipose insulin resistance, serving as a signal of nutrient excess. However, neither the substrates for nor sites of oxidant production are known. Because insulin stimulates glucose utilization, we hypothesized that glucose oxidation would fuel respiration, in turn generating mitochondrial oxidants. This would impair insulin action, limiting further glucose uptake in a negative feedback loop of "glucose-dependent" insulin resistance. Using primary rat adipocytes and cultured 3T3-L1 adipocytes, we observed that insulin increased respiration, but notably this occurred independently of glucose supply. In contrast, glucose was required for insulin to increase mitochondrial oxidants. Despite rising to similar levels as when treated with other agents that cause insulin resistance, glucose-dependent mitochondrial oxidants failed to cause insulin resistance. Subsequent studies revealed a temporal relationship whereby mitochondrial oxidants needed to increase before the insulin stimulus to induce insulin resistance. Together, these data reveal that (a) adipocyte respiration is principally fueled from nonglucose sources; (b) there is a disconnect between respiration and oxidative stress, whereby mitochondrial oxidant levels do not rise with increased respiration unless glucose is present; and (c) mitochondrial oxidative stress must precede the insulin stimulus to cause insulin resistance, explaining why short-term, insulin-dependent glucose utilization does not promote insulin resistance. These data provide additional clues to mechanistically link nutrient excess to adipose insulin resistance.
Highlights
Insulin action in adipose tissue is crucial for whole-body glucose homeostasis, with insulin resistance being a major risk factor for metabolic diseases such as type 2 diabetes
We hypothesised that insulin resistance was caused by mitochondrial oxidants (mitoROS) generated from increased glucose metabolism
Since mitoROS is generated by enzymes that participate in mitochondrial respiration, we reasoned that glucose would fuel insulinstimulated respiration to generate mitoROS
Summary
We sought to understand the role of glucose metabolism in adipose insulin resistance. We confirmed that galactose blunted insulin-responsive lactate levels (Fig 1D) With this tool, we found that insulin slightly increased respiration (Fig 1E), but this did not require glucose except in the presence of the mitochondrial uncoupler, FCCP (Fig 1F). MitoROS is sensitive to insulin-stimulated glucose uptake These data are consistent with the notion that glucose-dependent mitoROS is an endogenous signal to cause insulin resistance under conditions of excess nutrient supply (Fig 3A). Contrary to our model (Fig 3A), we did not observe any detectable inhibition of insulin action by DCA treatment, with only mPQ causing insulin resistance within this acute time-frame (Fig 3D) This concurred with the presence of glucose and/or DCA having no consistent effect on insulinstimulated signalling as measured by Akt and its substrate, AS160 (Fig 2C,E). Our data suggest that mitoROS only leads to insulin resistance if mitoROS increases prior to insulin stimulation (Fig 4C)
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