Carnosine increases insulin-stimulated glucose uptake and reduces methylglyoxal-modified proteins in type-2 diabetic human skeletal muscle cells

Abstract

Type-2 diabetes (T2D) is characterised by a dysregulation of metabolism, including skeletal muscle insulin resistance, mitochondrial dysfunction, and oxidative stress. Reactive species, such as methylglyoxal (MGO) and 4-hydroxynonenal (4-HNE), positively associate with T2D disease severity and can directly interfere with insulin signalling and glucose uptake in skeletal muscle by modifying cellular proteins. The multifunctional dipeptide carnosine, and its rate-limiting precursor β-alanine, have recently been shown to improve glycaemic control in humans and rodents with diabetes. However, the precise mechanisms are unclear and research in human skeletal muscle is limited. Herein, we present novel findings in primary human T2D and lean healthy control (LHC) skeletal muscle cells. Cells were differentiated to myotubes, and treated with 10 mM carnosine, 10 mM β-alanine, or control for 4-days. T2D cells had reduced ATP-linked and maximal respiration compared with LHC cells (p = 0.016 and p = 0.005). Treatment with 10 mM carnosine significantly increased insulin-stimulated glucose uptake in T2D cells (p = 0.047); with no effect in LHC cells. Insulin-stimulation increased MGO-modified proteins in T2D cells by 47%; treatment with carnosine attenuated this increase to 9.7% (p = 0.011). There was no effect treatment on cell viability or expression of other proteins. These findings suggest that the beneficial effects of carnosine on glycaemic control may be explained by its scavenging actions in human skeletal muscle.

Keywords: Diabetes; Glycaemia; Metabolism; Prediabetes; Therapeutics.

Fig. 1

Skeletal muscle O₂ consumption measured in human skeletal myotubes using the Seahorse XFe24 Analyser. A, B ATP-linked and maximal respiration for all cell conditions. C Seahorse Mito Stress Test trace for LHC cells. D Maximal respiration for T2D and T2D-GLT cells. E OCR tAUC for all cell conditions, showing the fold-change relative to each control condition (n = 3 independent experiments per condition, n = 6–7 replicates per experiment). *p < 0.05, **p < 0.01. LHC lean healthy control, OCR O₂ consumption rate, T2D-GLT type-2 diabetic glucolipotoxic conditions, tAUC total area-under-the-curve

Fig. 2

Insulin-stimulated glucose uptake and protein adducts in human skeletal myotubes. A Glucose uptake in LHC and T2D; B GLO1 expression in T2D myotubes; C MGO-modified proteins in T2D cells under basal and insulin-stimulated (INS) conditions; D 4-HNE-modified proteins in T2D cells under basal and INS conditions (n = 3 independent experiments per condition, n = 3–4 replicates per experiment). *p < 0.05. β-al β-alanine, Car carnosine, LHC lean healthy control, OCR O₂ consumption rate, T2D type-2 diabetic