IGF-I and IGFBP-2 Stimulate AMPK Activation and Autophagy, Which Are Required for Osteoblast Differentiation. Endocrinology 2016 Jan;157(1):268-81
Date
11/12/2015Pubmed ID
26556533Pubmed Central ID
PMC4701891DOI
10.1210/en.2015-1690Scopus ID
2-s2.0-84954482572 (requires institutional sign-in at Scopus site) 83 CitationsAbstract
IGF-I/insulin-like growth factor binding protein 2 (IGFBP-2) coordinately stimulate osteoblast differentiation but the mechanisms by which they function have not been determined. AMP-activated protein kinase (AMPK) is induced during differentiation and AMPK knockout mice have reduced bone mass. IGF-I modulates AMPK in other cell types; therefore, these studies determined whether IGF-I/IGFBP-2 stimulate AMPK activation and the mechanism by which AMPK modulates differentiation. Calvarial osteoblasts and MC-3T3 cells expressed activated AMPK early in differentiation and AMPK inhibitors attenuated differentiation. However, expression of constitutively activated AMPK inhibited differentiation. To resolve this discrepancy we analyzed the time course of AMPK induction. AMPK activation was required early in differentiation (day 3-6) but down-regulation of AMPK after day 9 was also necessary. IGF-I/IGFBP-2 induced AMPK through their respective receptors and blocking-receptor activation blocked AMPK induction. To determine the mechanism by which AMPK functioned we analyzed components of the autophagosome. Activated AMPK stimulated ULK-1 S555 phosphorylation as well as beclin-1 and microtubule-associated protein 1A/1B light-chain phosphatidylethanolamine conjugate (LC3II) induction. Inhibition of AMPK attenuated these changes and direct inhibition of autophagy inhibited differentiation. Conversely, expression of activated AMPK was associated with persistence of these changes beyond day 9 and inhibited differentiation. Blocking AMPK activation after day 9 down-regulated these autophagosome components and rescued differentiation. This allowed induction of mechanistic target of rapamycin and AKT, which suppressed autophagy. The results show that early induction of AMPK in response to IGF-I/IGFBP-2 followed by suppression is required for osteoblast differentiation. AMPK functions through stimulation of autophagy. The findings suggest that these early catabolic changes are important for determining the energy source for osteoblast respiration and down-regulation of these components may be required for induction of glycolysis, which is required during the final anabolic stages of differentiation.
Author List
Xi G, Rosen CJ, Clemmons DRMESH terms used to index this publication - Major topics in bold
AMP-Activated Protein KinasesAmino Acid Substitution
Animals
Autophagy
Cell Line
Cells, Cultured
Enzyme Activation
Humans
Insulin-Like Growth Factor Binding Protein 2
Insulin-Like Growth Factor I
Male
Mice
Mice, Inbred C57BL
Mice, Knockout
Mutant Proteins
Osteoblasts
Osteogenesis
Phagosomes
Phosphorylation
Protein Kinase Inhibitors
Protein Processing, Post-Translational
RNA Interference
