Submitted on January 10, 2007
Accepted on September 2, 2007
Induction of group VIA phospholipase A2 activity during in vitro ischemia in C2C12 myotubes is associated with changes in the level of its splice variants
Kristian Arild Poulsen1*, Stine F Pedersen1, Miriam Kolko2, and Ian Henry Lambert1
1 Department of Molecular Biology, University of Copenhagen, Copenhagen, Denmark
2 Eye Pathology Institute, University of Copenhagen, Copenhagen, Denmark
* To whom correspondence should be addressed. E-mail: kapoulsen{at}aki.ku.dk.
The involvement of the group VI iPLA2s (iPLA2-VI) in in vitro ischemia (OGD, oxygen and glucose deprivation) in mouse C2C12 myotubes have been investigated. OGD induced a time-dependent (0-6 h) increase in BEL-sensitive iPLA2 activity which was suppressed by specific siRNA knockdown of iPLA2-VIA. OGD was associated with an increase in iPLA2-VIA protein level whereas the mRNA level was unchanged. The level of iPLA2-VIB mRNA and protein were not increased by OGD. RT-PCR and Western blotting identified a mouse iPLA2-VIA homolog to the catalytically inactive 50 kDa iPLA2-VIA-Ankyrin variants previously identified in human. Both the mRNA and protein levels of this ~ 50 kDa variant were reduced significantly within 1 h following OGD. In C2C12 myoblasts, iPLA2-VIA seemed to predominantly reside at the endoplasmatic reticulum (ER), where it accumulated further during OGD. A time-dependent reduction in cell viability during the early OGD period (3 h) was partially prevented by iPLA2-VIA knockdown or pharmacological inhibition (BEL 10 µM), whereas iPLA2-VIA overexpression had no effect on cell viability. Altogether, these data demonstrate that OGD in C2C12 myotubes is associated with an increase in iPLA2-VIA activity which decrease cell viability. iPLA2-VIA activation may be modulated by changes in the level of active and inactive iPLA2-VIA isoforms.
