Improving type 2 diabetes through a distinct adrenergic signaling pathway involving mTORC2 that mediates glucose uptake in skeletal muscle.

Authors:
Sato M, Dehvari N, Oberg AI, Dallner OS, Sandström AL, Olsen JM, Csikasz RI, Summers RJ, Hutchinson DS, Bengtsson T
In:
Source: Diabetes
Publication Date: (2014)
Issue: Epub: ahead
Cells used in publication:
L6
Species: rat
Tissue Origin: skeletal muscle
Skeletal Muscle Cells, (SkMC) human
Species: human
Tissue Origin: skeletal muscle
Platform:
4D-Nucleofector™ X-Unit
Experiment

L6 Myotubes or myoblasts were detached using trypsin/EDTA, transferred to Eppendorf tubes and centrifuged at 1000rpm for 3 min, resuspended in 20 µl SE Cell Line Nucleofector solution for L6 cells and P1 primary Nucleofector solution for SKMC cells (Lonza, Basel, Swiss) and 100 pmol of siRNA (L6 myotubes) or 0.8 ?g GLUT4mycGFP construct (L6 myoblasts and human SKMC) added. L6 cells were nucleofected by using the DS-137 program and SKMC cells with the DS-138 program in the Lonza 4D nucleofector. Cells were electroporated in 16-well microcuvette stripes (Lonza), pre-warmed RPMI1640 added (80µl), and then transferred to 1.5 ml microcentrifuge tubes or glass bottom slides containing DMEM with 10% FBS for 8h.

Summary of Content: Type 2 diabetes is an increasing worldwide epidemic that poses major health problems. In this paper, the authors describe for the first time that specific activation of ß2-adrenoceptors in skeletal muscle, but not white adipocytes, causes translocation of GLUT4 to increase glucose uptake, by a mechanism dependent on mTORC2. To confirm the involvement of GLUT4, siRNA directed against GLUT4 was nucleofected into L6 myotubes. This caused a markedly reduced ß -adrenoreceptor mediated glucose uptake compared to a scrambled siRNA. Direct demonstration of GLUT4 translocation following ß2-adrenoceptor-stimulated glucose uptake has been shown by transient and stable nucleofection of L6 myotubes and transient nucleofection of human primary skeletal muscle cells with different GLUT4 reporter constructs.

Abstract

Type 2 diabetes is an increasing worldwide epidemic that poses major health problems. We have identified a novel physiological system that increases glucose uptake in skeletal muscle but not in white adipocytes. Activation of this system improves glucose tolerance in Goto-Kakizaki rats or mice fed a high fat diet, which are established models for type 2 diabetes. The pathway involves activation of ß2-adrenoceptors that increase cAMP levels and activate PKA that phosphorylates mammalian target of rapamycin complex 2 (mTORC2) at S2481. The active mTORC2 causes translocation of GLUT4 to the plasma membrane and glucose uptake without the involvement of Akt or AS160. Stimulation of glucose uptake into skeletal muscle following activation of the sympathetic nervous system is likely to be of high physiological relevance since mTOR complex 2 activation was observed at the cellular, tissue and whole animal level in rodent and human systems. This signaling pathway provides new opportunities for the treatment of type 2 diabetes.