Visit our Expo - Redox and Inflammation signaling 2012

Session XVII : Cell signaling in health and disease Poster XVII, 3
Hypoxic responses in acute myeloid leukaemia (AML)
Siv Lise Bedringaas, Kimberley Hatfield, Gry Sjøholt, Lars Helgeland#, Jørn Skavland,
Olav Dahl§, Øystein Bruserud, Bjørn Tore Gjertsen
Hematology section, Institute of Medicine, University of Bergen and Department of
Internal Medicine, Haukeland University Hospital, N-5021 Bergen. #Department of
Pathology, The Gade Institute, University of Bergen, Haukeland University Hospital.
§Section of Oncology, Institute of Medicine and Department of Oncology, Haukeland
University Hospital, Bergen, Norway.
Acute myeloid leukaemia (AML) is a haematological malignancy characterized with block in
differentiation and numerous, but also recurrent, chromosomal translocations. The most
common recognized mutation (approx. 30%) is in the class III receptor FMS-related tyrosine
kinase 3 (Flt3/Stk1/Flk2). Even though AML blasts are in a normoxic environment in the
peripheral blood, our hypotheses is that excessive growth of malignant blasts in the bone
marrow create a hypoxic environment. Tumour hypoxia is known to promote chemo
resistance and aggressive disease.
In this study we have investigated the hypoxic effect in cryopreserved primary AML cells and
AML cell lines (HL-60, NB4) by Western blot, ELISA, flow cytometry, two-dimensional gel
electrophoresis (2DE) and mass spectrometric identification of proteins, as well as the twodimensional
differential gel electrophoresis (DIGE). In particular the protein levels of
Hypoxia Inducible Factor-1" (HIF-1"), vascular epithelial growth factor (VEGF),
osteopontin (OPN), cyclin-dependent kinase inhibitor 1A (CDKN1A/p21/CIP1), and Flt3
were also investigated in the patient samples, and HDM2 in cell lines were examined.
We investigated AML cells in hypoxia (1% O2) and Co2+ that mimics hypoxic conditions.
Preliminary results show that 30% of patients (10/30) and both cell lines responded to
hypoxia with increased protein level of HIF-1". More than 50% of the patients responded
with VEGF production (17/30), but a significant fraction of the VEGF responders were not
HIF-1" responders (7/17). The VEGF non-responders had non-detectable levels of VEGF
before and during hypoxia. Approximately 50% of the patients responded with OPN increase.
CDKN1A/p21 and Flt3 showed no response to hypoxia in any patients. The level of HDM2
was reduced in both cell lines tested during hypoxia.
Preliminary results suggest that DIGE is more sensitive than conventional 2DE in detection of
protein modulation induced by hypoxia (1% O2) and Co2+. Ongoing experiments are aimed
to map the protein networks that differ between hypoxic HIF-1" responsive and nonresponsive
AML.
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