Mitochondrial ADP Sensitivity and Transport: New Insights Into Diet-Induced Mitochondrial Impairments

In the quest to understand the etiology and pathophysiology of insulin resistance and type 2 diabetes (T2DM), a body of evidence representing nearly two decades of research points a finger at mitochondria as organelles of great interest (1–3). The term “mitochondrial dysfunction” was born following early reports that these organelles exhibited altered abundance and morphology (1), attenuated oxidative phosphorylation capacity (4), and an altered redox environment (5) in insulin-resistant individuals. Today, despite great efforts to understand the relationship between mitochondria and metabolic disease, there is little accord to conclusively determine if impairments in skeletal muscle mitochondrial biology are cause or consequence of insulin resistance or simply paraphenomena.

High-fat feeding interventions are used extensively in the quest to understand the etiology and pathophysiology of insulin resistance and T2DM. Such feeding models recapitulate many of the metabolic features along the spectrum of insulin resistance and diabetes (e.g., adiposity, glucose intolerance, metabolic inflexibility) (6). Painstaking efforts have been made to understand the role of mitochondria in the progression of insulin resistance during high-fat feeding, but ambiguity remains. For example, Sparks et al. (7) reported a ∼90% decline in genes associated with oxidative phosphorylation and mitochondrial biogenesis in skeletal muscle of C57BL/6J mice fed a high-fat diet, which aligns with decreased oxidative enzyme activity observed in patients with T2DM (1). Similar declines in maximal ADP-stimulated (state 3) and uncoupled (state 4) oxygen consumption have been reported in isolated mitochondria from skeletal muscle of male Wistar rats fed a high-fat diet (8). Conversely, Hancock et al. (9) documented increased skeletal muscle mitochondrial content in a rat model of high-fat diet–induced insulin resistance, which supports data from human studies revealing preserved mitochondrial function assessed in permeabilized muscle fibers biopsied from patients with T2DM (10). It is important to note that the vast majority of studies assessing mitochondrial respiratory function are performed ex vivo in the presence of saturating ADP concentrations that do not exist under normal physiological conditions in vivo (11). Furthermore, skeletal muscle mitochondria exhibit a ∼150-fold reserve capacity for oxygen uptake during maximal exercise in trained young adults and ∼30–40-fold in patients with insulin resistance (12). In view of that, previously reported deficits in mitochondrial capacity amounting to ∼30–40% in the presence of saturating ADP concentrations (4,13) should be cautiously interpreted in terms of physiological relevance.

In this issue of Diabetes, Miotto et al. (14) provide some fundamental new knowledge that advances our understanding of high-fat diet–induced mitochondrial respiratory dysfunction. Mitochondrial function was evaluated in permeabilized muscle fibers from red gastrocnemius muscles using high-resolution respirometry, enzymatic activity, and protein expression. The authors challenged conventional assessments of mitochondrial function by assessing absolute respiration over a range of increasing, biologically relevant ADP concentrations (i.e., 100–12,000 μmol/L) and determining ADP sensitivity by estimating the apparent K_m in mice fed a control or high-fat diet for 4 weeks. Additional experiments were performed to detect changes in ADP sensitivity mediated by the absence or presence of palmitoyl-CoA, simulating the accumulation of long-chain fatty acid concentrations following high-fat diet consumption, which has been previously shown to inhibit the adenine nucleotide translocase (ANT) and decrease mitochondrial ADP sensitivity (15). Protein expression of multiple ADP transporters was measured to determine the influence of a high-fat diet to alter structural components that could potentially impact ADP sensitivity.

Miotto et al. (14) found that mice exposed to a high-fat diet exhibited insulin resistance, impaired ADP sensitivity, and impaired palmitoyl-CoA–stimulated mitochondrial respiration despite an observed increase in mitochondrial content. It is fascinating to note that measurements of state 2 and state 3 respiration did not reveal any differences between control and high-fat diet mice. However, impaired mitochondrial respiration (∼30%) was present in mice fed a high-fat diet compared with control diet over a range of increasing biologically relevant ADP concentrations (i.e., 100–1,000 μmol/L). Specifically, the high-fat diet decreased absolute respiration at ADP concentrations as low as 100 μmol/L, which more closely represents in vivo ADP concentrations (e.g., 8–37 μmol/L) reported at rest in skeletal muscle of mice (16,17). Furthermore, high-fat diet decreased mitochondrial ADP sensitivity by ∼25%. Impaired ADP sensitivity was also observed in the presence of 60 μmol/L palmitoyl-CoA, suggesting an inhibitory action on ANT imposed by the high-fat diet. Somewhat surprisingly, this study (14) found no effect of high-fat diet on maximal capacity of mitochondrial-derived H₂O₂ production despite evidence of oxidative stress (i.e., 4-hydroxynonenal). Thus, the role of reactive oxygen species as mediators of mitochondrial impairments in high-fat diet–induced insulin-resistant skeletal muscle remains an attractive hypothesis that cannot be supported from data in this article.

Miotto et al. (14) present convincing evidence to support a link between high-fat diet–induced insulin resistance and changes to fundamental bioenergetic properties of skeletal muscle mitochondria. By utilizing physiological concentrations of ADP to elicit mitochondrial respiration, the authors have shed new light on prior studies that employed conventional mitochondrial respiratory assays in the presence of saturating ADP concentrations. Despite these novel findings, several questions remain unanswered. Does impaired ADP sensitivity in skeletal muscle mitochondria play a role in the apparent whole-body metabolic inflexibility following a high-fat diet? It is possible that impaired ADP sensitivity is involved in previously documented “incomplete” fat oxidation in skeletal nmuscle mitochondria under conditions of constrained β-oxidation elicited by a high-fat diet (18). The observed increase in mitochondrial content has been theorized as a compensatory response to preserve ADP sensitivity and attenuate redox stress (14), but an increase in mitochondrial content could compensate for impaired mitochondrial dynamics as a consequence of high-fat diet–induced intracellular stress. From a therapeutic perspective, oxidant scavengers have had mixed effects on improving metabolic homeostasis in animal models of insulin resistance (19). In contrast, prolonged exercise training has been shown to partially reverse mitochondrial impairments in patients with long-standing T2DM (20), although the ability of exercise training to restore mitochondrial ADP sensitivity in insulin-resistant skeletal muscle has not been evaluated. Though several questions remain unsolved regarding the underlying etiology of T2DM, mechanisms influencing the transport of ADP into the mitochondria provide a new focal point for models assessing skeletal muscle alterations associated with the development of insulin resistance.