Abstract

In vitro culture is widely used for characterization of primary human acute myeloid leukemia (AML) cells, but even when using optimized handling and culture conditions the AML cells show spontaneous in vitro apoptosis with a gradual decrease in cell viability during culture. The extent of this stress-induced apoptosis varies between patients, and a high degree of apoptosis is associated with high pre-culture BCL2 levels together with low levels of BAX and Heat Shock Proteins 30 and 90. We compared the global proteomic profiles during ongoing in vitro apoptosis for patients with high and low AML cell viability (i.e., less extensive versus extensive spontaneous apoptosis) after 48 h of culture. We identified 7902 proteins, but only 276 proteins differed significantly between patients with high (i.e., >25% viable cells; 192 upregulated and 84 downregulated peptides) and low viability after in vitro culture. Protein interaction network analysis based on these 276 protein identified three protein networks that included 18 proteins; most of these proteins were localized to the endoplasmic reticulum and several of them are involved in or are altered during the process of endoplasmic reticulum stress/unfolded protein stress response. To conclude, primary AML cells are heterogeneous with regard to degree of apoptosis in response to cellular stress, and this difference in regulation of apoptosis is associated with differences in the induction of and/or response to the unfolded protein stress response.

Highlights

  • Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by bone marrow infiltration of immature leukemia cells [1,2]

  • Cryopreserved primary AML cells derived from 41 patients were thawed and cultured for 48 h before the fractions of viable, early apoptotic and late apoptotic/necrotic cells were determined by flow-cytometric analysis as described previously [8]

  • We did Gene ontology (GO) analyses to compare the three main protein subsets, and these analyses showed that the three clusters differed with regard to proteins involved in endoplasmic reticulum/transport/organelle membrane (Cluster A1), nucleus/transcription, and extracellular/secreted proteins (Cluster B) (Table 5)

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Summary

Introduction

Acute myeloid leukemia (AML) is an aggressive hematological malignancy characterized by bone marrow infiltration of immature leukemia cells [1,2]. In vitro culture of AML cells has been widely used in experimental studies [4,5], and when using cryopreserved leukemic cells from biobanks it is possible to investigate large numbers of patients in repeated experiments and to use the same highly standardized culture conditions for all patients [5]. Such experimental studies have been important for characterization of the hierarchically organized AML cell populations and for identification of new molecular targets.

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