Genetic Deficiency of Glycogen Synthase Kinase-3β Corrects Diabetes in Mouse Models of Insulin Resistance
Figure 8
Effects of Inhibition of Gsk-3 Activity on Protein Stability of p27kip1
(A) MIN6 cells were pretreated with either 40 mM lithium chloride (LiCl) or 40 mM NaCl in DMEM with 15% FBS for 1 h and were cotreated with 25 μg/ml cyclohexamide and lithium or NaCl for 4 h. The lysates were subjected to western blot analysis with anti-p27kip1, phospho-glycogen synthase, total Gsk-3β, and α-tubulin. Densitometry of p27kip1 was measured and normalized over α-tubulin.
(B) Islets isolated from 16-wk-old WT mice were deprived of serum for 4 h and were incubated with no addition, or with addition of 100 nM IGF-1, or 40 mM LiCl, or both for 3 h. Lysates were then prepared from islets and were subjected to western blot analysis with anti-p27kip1, anti-phospho glycogen synthase, anti-total Gsk-3β, and β-actin. Representative results of three independent experiments are presented. Densitometry of p27kip1 was measured and normalized over β-actin. Mean protein levels ± S.E.M. are summarized on the graph. A single asterisk (*) indicates p < 0.05; double asterisks (**) indicate p < 0.01.