Lymphocytes from chronic lymphocytic leukaemia undergo ABL1-linked amoeboid motility and homotypic interaction as an early adaptive change to ex vivo culture.

Claire V Hutchinson,John Burthem,Suzanne M Johnson,Julie A Adams,Karen S Rees-Unwin,Shiva Natarajan

doi:10.1186/2162-3619-3-7

Abstract

BackgroundThose stimuli that together promote the survival, differentiation and proliferation of the abnormal B-lymphocytes of chronic lymphocytic leukaemia (CLL) are encountered within tissues, where together they form the growth-supporting microenvironment. Different tissue-culture systems promote the survival of the neoplastic lymphocytes from CLL, partly replicating the in vivo tissue environment of the disorder. In the present study, we focussed on the initial adaptive changes to the tissue culture environment focussing particularly on migratory behaviour and cellular interactions.MethodsA high-density CLL culture system was employed to test CLL cell-responses using a range of microscopic techniques and flow cytometric analyses, supported by mathematical measures of cell shape-change and by biochemical techniques. The study focussed on the evaluation of changes to the F-actin cytoskeleton and cell behaviour and on ABL1 signalling processes.ResultsWe showed that the earliest functional response by the neoplastic lymphocytes was a rapid shape-change caused through rearrangement of the F-actin cytoskeleton that resulted in amoeboid motility and promoted frequent homotypic interaction between cells. This initial response was functionally distinct from the elongated motility that was induced by chemokine stimulation, and which also characterised heterotypic interactions between CLL lymphocytes and accessory cells at later culture periods. ABL1 is highly expressed in CLL lymphocytes and supports their survival, it is also recognised however to have a major role in the control of the F-actin cytoskeleton. We found that the cytoplasmic fraction of ABL1 became co-localised with F-actin structures of the CLL lymphocytes and that the ABL1 substrate CRKL became phosphorylated during initial shape-change. The ABL-inhibitor imatinib mesylate prevented amoeboid movement and markedly reduced homotypic interactions, causing cells to acquire a globular shape to rearrange F-actin to a microvillus form that closely resembled that of CLL cells isolated directly from circulation.ConclusionWe suggest that ABL1-induced amoeboid motility and homotypic interaction represent a distinctive early response to the tissue environment by CLL lymphocytes. This response is separate from that induced by chemokine or during heterotypic cell-contact, and may play a role in the initial entry and interactions of CLL lymphocytes in tissues.

Highlights

Those stimuli that together promote the survival, differentiation and proliferation of the abnormal B-lymphocytes of chronic lymphocytic leukaemia (CLL) are encountered within tissues, where together they form the growth-supporting microenvironment
We show that these rearrangements mediate frequent, but transient, homotypic interactions between the neoplastic CLL B-lymphocytes, and that the morphological appearances of the CLL lymphocytes in highdensity culture resemble those of CLL lymphocytes within tissues, and are consistent with the interactions between cells seen in tissues [17]
The present study demonstrates for the first time that activation of ABL1-linked signal pathways is an early response by CLL lymphocytes within the cell culture environment, that is associated with amoeboid-pattern migration and with frequent brief homotypic interactions between the neoplastic cells

Summary

Introduction

Those stimuli that together promote the survival, differentiation and proliferation of the abnormal B-lymphocytes of chronic lymphocytic leukaemia (CLL) are encountered within tissues, where together they form the growth-supporting microenvironment. We focussed on the initial adaptive changes to the tissue culture environment focussing on migratory behaviour and cellular interactions Those stimuli that promote the survival, differentiation and proliferation of the abnormal B-lymphocytes of chronic lymphocytic leukaemia (CLL) are encountered within tissues, where together they form the growth-supporting microenvironment (reviewed in [1]). The enhanced cell survival within these cultures is believed to derive, at least in part, from direct interaction between cells [6] Such interactions resemble the behaviour of normal B-lymphocytes at particular developmental stages in vivo, in which migration through tissues involves repeated brief homotypic and heterotypic cell contacts, as well as more sustained intercellular interactions [12]. We postulated that imatinib might alter F-actin mediated motility or adhesion of CLL lymphocytes

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