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Frezzato, Federica (2008) Coinvolgimento della proteina HS1 nella sopravvivenza dei linfociti B neoplastici di pazienti con leucemia linfatica cronica. [Tesi di dottorato]

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Abstract (inglese)

B-cell chronic lymphocytic leukemia (B-CLL) is the most common form of leukemia in adults and is characterized by the accumulation of clonal CD5+ B lymphocytes due to uncontrolled growth and resistance to apoptosis. Several protein kinase pathways have been claimed to be involved in the regulation of cell survival. We previously demonstrated that the Src kinase Lyn is overexpressed, constitutively active and anomalously distributed in malignant B cells as compared to normal B
lymphocytes. Our attention was subsequently focused on the 75 kDa HS1 protein, which is one of the major substrate of Lyn kinase upon BCR cross-linking and plays a crucial role in BCR-induced apoptosis in the mouse B lymphoma cell line WEHI-231. In the present study HS1 protein level was measured by western blotting analysis in 43 untreated B-CLL patients and in 26 normal controls. We found a significant difference in HS1 protein level between CLL and normal B cells, being mainly expressed in the leukemic patients with respect to normal controls (p<0.01). When we correlated HS1 protein level with prognostic factors, we observed that patients with more negative prognostic factors had a higher expression of HS1 protein regarding those with a better prognosis. We also analyzed HS1 in 10 CLL patients before and after in vivo therapy with fludarabine and cyclophosphamide; we found a significant reduction of both HS1 protein and mRNA levels in those patients which responded to therapy (n°=7) while non-responder patients (n°=3) did not show any change in HS1 levels.
Using confocal microscopy and subcellular cell fractionation, we observed an abnormal distribution of HS1 in leukemic cells with respect to normal B cells. In particular, the pattern of expression of HS1 appeared with a spotting distribution and a 4-7% aliquot of HS1 was present in the nucleus of leukemic B cells but not in
normal B lymphocytes. This nuclear localization could not be observed following BCR triggering. In other words, after BCR engagement, we observed a redistribution of HS1 that was no longer detectable in the nucleus of stimulated leukemic cells. The pre-incubation of cells with PP2, a Src kinase inhibitor, prevents the IgM-mediated redistribution of HS1; so the disappearance of nuclear HS1 is attributable to Src kinases that act at some level of the signal cascade. Since HS1 can interact with actin through the Arp 2/3 complex we performed additional experiments to investigate whether HS1 could interact with cytoskeletal components. We observed that cytosolic HS1 co-localizes with ß-actin both in normal and leukemic B cells. Moreover, for the first time, we demonstrated that in B-CLL,
but not in normal B cells, HS1 co-localizes with ?-tubulin and, in particular, with the centrosome, suggesting that this protein could play a role in the cytoskeletal reorganization of leukemic B cells.
All these findings seem to suggest a pivotal role for HS1 in the regulation of cell survival of leukemic B cells and hint that this protein might represent a target for the development of new therapeutic approaches.


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Tipo di EPrint:Tesi di dottorato
Relatore:Semenzato, Gianpietro
Correlatore:Trentin, Livio
Dottorato (corsi e scuole):Ciclo 20 > Corsi per il 20simo ciclo > ONCOLOGIA E ONCOLOGIA CHIRURGICA
Data di deposito della tesi:31 Gennaio 2008
Anno di Pubblicazione:31 Gennaio 2008
Parole chiave (italiano / inglese):leucemia linfatica cronica; proteina HS1; apoptosi; chemioterapia; citoscheletro
Settori scientifico-disciplinari MIUR:Area 06 - Scienze mediche > MED/15 Malattie del sangue
Struttura di riferimento:Dipartimenti > Dipartimento di Medicina Clinica e Sperimentale
Codice ID:564
Depositato il:11 Nov 2008
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