Skeletal muscle is responsible for between 50–90% of glucose uptake in humans16. Mechanisms linking skeletal muscle to insulin resistance are numerous. Impaired mitochondrial metabolism leads to increased intramyocellular fat content and inflammation17,18. Signal transduction between IRS-1 and PI3K and other kinases is impaired leading to lower glucose uptake and other effects19. Free fatty acid turnover inhibits PDH activity and lowers glucose oxidation20. Decreased glycogen storage contributes to increased fatty acid uptake and accumulation of lipid intermediates, which impair insulin signaling21. These mechanisms intertwine and overlap making type 2 diabetes a complex disease that remains difficult to manage. The data presented in this manuscript and the published literature on ABA to date, highlight the ability of this compound to intersect the pathogenesis of type 2 diabetes and insulin resistance at multiple levels. ABA treatment, both at the organism levels and in specific muscle cells ex vivo, increases both glucose and fatty acid metabolism in the mitochondria, increases glycogen synthesis, activates PI3K independently of insulin and promotes GLUT4 translocation to the cell membrane.
Endogenous production of ABA has been previously linked to hormonal regulation in vitro. Specifically, GLP-1 increases ABA release from β cells, while ABA has been shown to increase insulin secretion from pancreatic islets and cell lines22. Compelling in vitro, the in vivo data, presented herein, supports the therapeutic efficacy of ABA independent of these mechanisms for glycemic control. Fig extract ABA increases glucose tolerance in IP GTTs. As GLP-1 is primarily released by entero-endocrine cells after oral glucose load, the efficacy of ABA in the context of IP glucose challenge, suggests an ability to function without relying on GLP-1 as a mediator and the potential to synergize with currently available GLP-1 agonists. Further, administration of ABA did not increase plasma insulin levels. In contrast, ABA supplementation provided a small decrease in plasma, suggesting an ability to provide glycemic control in the absence of hyperinsulinemia at doses of 0.125 µg/kg. This indicates increased insulin sensitivity or an ability to support insulin-independent glucose disposal. Either mechanism would serve to alleviate pancreatic stress in type 2 diabetes, during which insulin production is often increased due to diminished responsiveness of cells to insulin stimulus.
Based on previous connections to GLUT4 translocation and the described effects on glycemic control, it is unsurprising that ABA increases the expression of key elements of glucose metabolism, such as Glut4, Hk2, Sdha, and Gys1, in skeletal muscle. Linked to these effects and overall muscle metabolism, is the transcription factor, TFEB. More specifically, TFEB activity in skeletal muscle is increased during periods of exercise, by Pgc1a, and by increase in the intracellular calcium levels23,24. Importantly, TFEB is a main controller of mitochondrial biogenesis and mitophagy, ensuring sufficient mitochondria are present and in good health relative to energetic demands25. In line with this function, we observe increases in important mitochondrial enzymes, Cox5a and Sdha, as well as the main controller of fatty acid oxidation, Cpt1a. As such, ABA treatment likely results in greater mitochondrial mass and enhanced ability of muscle to utilize fatty acids during a resting state. In addition to the increased expression of Tfeb, ABA induces many similar effects to those observed in insulin-independent, exercise-induced glucose uptake by muscle, as well as the mechanisms in which exercise restores insulin sensitivity26, including GLUT4 translocation, activation of AMPK, and induction of calcium release.
Insulin-stimulated glycogen synthesis is decreased in individuals with type 2 diabetes and metabolic syndrome, accounting for most of the differential in whole-body glucose disposal16,27. Activated primarily by PP1 via insulin-stimulation of ISPK1, glycogen synthase is negatively regulated by GSK3. As LANCL2 activation by ABA, leads to insulin-independent upregulation of PI3K/Akt activity, it is likely that this pathway is responsible for the increased glycogen synthesis in human myotubes ex vivo that is further enhanced by the presence of insulin. Alternatively, LANCL2 activation may influence the expression of PP1 regulatory subunits known to bind glycogen leading to downstream regulation of PP1 activity. Notably, significant differences in the distribution of glycogen reserves exist between mice and humans, with a much greater proportion of glycogen stored in muscle in humans and an approximate 10-fold increase in glycogen to muscle mass ratio in humans compared to mice28. The liver remains an important source of glycogen, particularly to avoid hypoglycemia during fasting. In ActaCre mice, an accelerated increase in blood glucose levels after insulin-induced drop could suggest altered liver glycogen storage, potentially as a compensatory mechanism for impaired glucose disposal and glycogen storage in muscle. However, further work on the interplay of muscle and liver glycogen synthesis in the context of ABA treatment is needed.
Beyond direct effects on skeletal muscle, ABA may further support the maintenance of insulin tolerance in muscle through the muscle/adipose axis. The expression profiles of adipocytes are strongly correlated to the function of muscle cells through secreted products, such as IL-629. Perturbed muscle substrate oxidation in obese and diabetic individuals is a contributing factor to impaired glucose homeostasis and reduced insulin sensitivity30,31, and release of inflammatory factors from adipocytes rapidly induce insulin resistance in skeletal muscle using co-culture32. Importantly, exposure to inflammatory cytokines, such as TNF and IL-6, and activation of NF-κB in muscle can decrease oxidative metabolism, induce atrophy, and prevent insulin-stimulated Akt phosphorylation, all of which are factors in the development of type 2 diabetes and metabolic syndrome. Additionally, ABA reduced plasma levels of leptin and resistin, two notable adipokines tied to the diminished uptake of glucose in diabetes and impaired IRS-1 signaling33,34. Therefore, an ability to influence both direct metabolic pathways in skeletal muscle and the underlying inflammation provides a robust platform for the clinical development of ABA.
ABA is a potent insulin-sensitizing compound with the ability to control systemic glycemic responses and skeletal muscle metabolism at oral doses as low as 0.125 µg/kg. In this manuscript, we provide extensive evidence for the efficacy of an ABA-enriched fig extract in DIO and db/db mouse models of insulin insensitivity. Treatment with standardized ABA extract induced greater insulin sensitivity, decreased fasting blood glucose and improved response to oral glucose load. Thus, ABA in combination with diet and exercise may serve as a frontline intervention in early pre-diabetes, as well as type 2 diabetes.