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Session III : Protein kinase cascades as therapeutic targets Poster III, 41
Doxorubicin transitorily triggers Akt/PKB activation in primary cultures of rat
hepatocytes
Rosaura Navarro, Idoia Busnadiego, M. Luisa Hernández, M. Begoña Ruiz-Larrea and
José Ignacio Ruiz-Sanz
Department of Physiology, Medicine and Dentistry School, University of the Basque
Country, 48080-Bilbao, Spain. E-mail: joseignacio.ruizs@ehu.es
The serine-threonine protein kinase Akt/PKB (protein kinase B) has received much interest in
recent years because it is a key mediator of cell proliferation and seems to play an important
role in tumorigenesis and resistance to chemotherapeutic drugs. The kinase suppresses
chemotherapy-triggered apoptosis through interaction with critical molecules that regulate or
execute apoptosis. Although most reports have shown that chemotherapy decreases Akt
activity, the potent anticancer drug doxorubicin can increase Akt activity, depending on the
cell type. Thus, in several cancer cell lines, the drug triggers a transient phosphorylation and
activation of Akt at early time points, prior to the detection of apoptosis. In this work, we
have determined in primary cultures of rat hepatocytes the effects of doxorubicin on the
phosphorylation status of Akt. The levels of Akt phosphorylation were measured by Western
blot analysis with an anti-Ser473-phosphorylated Akt antibody. Doxorubicin (10 µM) had a
biphasic behaviour, increasing (60%) Akt phosphorylation at 30 min and decreasing it (40%)
at 1h. At later times, the Akt phosphorylation status remained similar to control cells (without
doxorubicin). The phosphatidylinositol 3-kinase (PI3K) inhibitor wortmannin prevented Akt
activation. The profile of the phsophorylation levels of Akt with doxorubicin was completely
different from that found for the pro-apoptotic JNK, which increased in a dose- and timedependent
manner.
This work was supported by the Basque Government (Research Project and Predoctoral
Training Grant to R.N.) and the University of the Basque Country (UPV00081.327-E-
15294/2003).
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