Selective inhibitors of cyclooxygenase-2 (prostaglandin-endoperoxide synthase-2; COX-2) augment the rate of hexose uptake in myotubes by recruiting glucose transporter-4 (GLUT-4) to the plasma membrane in an insulin- and AMPKα-independent manner [Alpert E, Gruzman A, Lardi-Studler B, Cohen G, Reich R, Sasson S. Cyclooxygenase-2 (PTGS2) inhibitors augment the rate of hexose transport in L6 myotubes in an insulin- and AMPKα-independent manner. Diabetologia 2006;49:562-70]. We aimed at elucidating the molecular interactions that mediate this effect of COX-2 inhibitors in L6 myotubes. The effects of the inhibitors niflumic acid, nimesulide and rofecoxib on activities and phosphorylation state of key proteins in the insulin transduction pathway were determined. These inhibitors did not induce specific tyrosine phosphorylation in IRS-1, could not assemble a functional IRS-PI3K-PKB/Akt complex and did not activate GSK3α/β, JNK1/2, ERK1/2, p38-MAPK or c-Cbl by site-specific phosphorylation(s). Yet, like insulin, they activated mTOR and induced downstream threonine phosphorylation in p70S6K and 4EBP1. However, rapamycin, which inhibits mTOR enzymatic activity, did not interfere with COX-2 inhibitor-induced stimulation of hexose uptake in myotube. Thus, mTOR activation was not required for COX-2 inhibitor-dependent augmentation of hexose transport in myotubes. Because PKCδ has also been shown to activate mTOR, we asked whether COX-2 inhibitors activate mTOR by a prior activation of PKCδ. Indeed, all three inhibitors induced tyrosine phosphorylation in PKCδ and stimulated its kinase activity. Moreover, pharmacological inhibition of PKCδ or the expression of a dominant-negative form of PKCδ in myotubes completely abolished COX-2 inhibitor-dependent stimulation of hexose uptake. This study shows that selective COX-2 inhibitors activate a unique PKCδ-dependent pathway to increase GLUT-4 abundance in the plasma membrane of myotubes and augment the rate of hexose transport.
Bibliographical noteFunding Information:
We thank Ms. L. Hammer for her assistance in performing the in vitro PKCδ kinase assay. S. Sasson is a member of the David R. Bloom Center for Pharmacy at the Hebrew University of Jerusalem. E. Alpert and A. Gruzman received fellowships from the Hebrew University Center for Diabetes Research. This work was supported by grants from the Chief Scientist of the Israel Ministry of Health, the Yedidut Foundation Mexico and the David R. Bloom Center for Pharmacy at the Hebrew University of Jerusalem.
- Cyclooxygenase-2 inhibitors
- Glucose transport
- Niflumic acid
- Skeletal muscle