IGFBP-2 enhances insulin signalling and glucose uptake in human skeletal myotubes. (#86)
Background: Insulin-like growth factor binding protein-2 (IGFBP-2) levels are reduced in obesity and type 2 diabetes mellitus, and IGFBP-2 over-expression protects against these conditions1,2. Mechanisms are unclear.
Objective and hypothesis: To examine the effect of IGFBP-2 on phosphatidylinositol 3-kinase (PI3K)/AKT signalling and glucose uptake in skeletal muscle. Our hypothesis was that IGFBP-2 would directly improve insulin signalling and glucose uptake.
Methods: All experiments utilised in-vitro cultures of fully-differentiated human skeletal myotubes (HSM), held in serum-free media and in the absence of exogenous IGF-1. Treatments were as follows: A) IGFBP-2 (0 or 100ng/ml for 24h) prior to insulin stimulation (0 or 100nM insulin for 30mins); B) IGFBP-2 or insulin stimulation in the absence/presence of Wortmannin (PI3K inhibitor; 100nM); C) Human-specific IGFBP-2 small-interfering RNA (or scrambled siRNA as control) for 24h prior to insulin-stimulation; D) Silencing of IGFBP-2 (as in C), prior to dosing with ‘add-back’ IGFBP-2 for 24h and then insulin stimulation. Outcomes included phosphorylation of AKT(ser473) (pAKT) in cell lysates by Western immunoblotting and glucose uptake using 2-deoxyglucose uptake assays.
Results: A) Basal pAKT was increased fivefold in the presence of IGFBP-2 (p<0.05), while insulin-stimulated increases in pAKT were doubled when IGFBP-2 was present (p<0.05). B) Wortmannin completely ablated insulin-induced (p<0.001), and IGFBP-2-induced (p<0.01) increases in pAKT. C) IGFBP-2 gene silencing reduced insulin-stimulated pAKT by 61% (p<0.01) and glucose uptake by 60% (p<0.001). D) Adding back IGFBP-2 (100ng/ml) completely restored and further enhanced insulin signalling and glucose uptake (300%; p<0.001).Conclusion: These findings indicate that IGFBP-2 directly enhances PI3K/AKT signalling and glucose uptake in HSM.
- Wheatcroft, SB et al. IGF-binding protein-2 protects against the development of obesity and insulin resistance. Diabetes 2007;56:285-94.
- Hedbacker, K et al. Antidiabetic effects of IGFBP2, a leptin-regulated gene. Cell Metabolism 2010;11:11-22.