Changes in spectrin organisation in leukaemic and lymphoid cells upon chemotherapy.

The aim of the present study was to investigate changes in spectrin and protein kinase C theta; (PKC theta;) organisation in human lymphoid and leukaemic cells undergoing chemotherapeutically induced apoptosis. An analysis of spectrin arrangement in human peripheral lymphoid (non-Hodgkin lymphoma) and leukaemic (acute lymphoblastic leukaemia) cells before and after chemotherapy revealed radical differences in the distribution of this protein. By using immunofluorescent technique, in lymphocytes isolated before chemotherapy, we found spectrin evenly distributed in the cytoplasm and the plasma membrane, while after the therapy changes in spectrin organisation occurred. Moreover, in lymphocytes after chemotherapy, extraction with buffer containing non-ionic detergent (Triton X-100) revealed presence of an insoluble fraction of spectrin. In normal or malignant cells before chemotherapy spectrin was totally soluble, however it should be mentioned that in total cell extracts and supernatants (but not in pellets) apoptotic fragments of spectrin (in addition to intact alpha and beta chains) were also found. In malignant cells after chemotherapy changes in PKC theta; organisation, similar to this observed in the case of spectrin, were shown by the immunofluorescence technique. In contrast, no differences in the distribution of other isoforms of protein kinase C: betaI and betaII, before and after chemotherapy, were found. Apoptotic phosphatidyloserine (PS) externalisation, as well as cell shrinkage, membrane protrusions and blebbing were observed in lymphocytes after chemotherapy and treatment with cytostatics in vitro. The overall results may suggest that spectrin redistribution/aggregation is the phenomenon involved in programmed cell death (PCD) of normal and neoplastic lymphocytes and lymphoblasts, however molecular basis of this phenomenon should be further investigated.