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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/10990/474

Autori: Tissino, Erika
Supervisore afferente all'Università: PUCILLO, CARLO ENNIO MICHELE
Titolo: The Elastin MIcrofibriL INterfacer1 (EMILIN1), is highly expressed in chronic lymphocytic leukemia (CLL)-involved tissues and promotes CLL cell adhesion and survival via CD49d
Abstract (in inglese): CD49d (α4 integrin chain) is a negative prognosticator in chronic lymphocytic leukemia (CLL) with a key role in CLL cell microenvironmental interactions. CD49d triggering by its main ligands Vascular Cell Adhesion Molecule-1 (VCAM-1) and CS-1 fragment of fibronectin, activates signaling pathways delivering pro-survival signals, and promoting resistance to drug-induced apoptosis in CLL. Recently, the globular (g) C1q-like domain of Elastin MIcrofibriL INterfacer1 (EMILIN1), an adhesive extracellular matrix constituent, was described as a new ligand for CD49d operating as a negative modulator of proliferation signals in substrate-adherent non-hematopoietic CD49d+ cells. Here we investigated the distribution of EMILIN1 in normal and CLL-involved tissues, and the effects of CD49d/EMILIN1 interaction in CLL in terms of adhesion and survival. By taking advantage of a specific anti-human EMILIN1 monoclonal antibody, exploratory staining in reactive lymphoid tissues (tonsil) indicated a clear extracellular EMILIN1 specific reactivity in the outer zone of the mantle/marginal areas. When investigated in lymph node tissues from CLL cases (n=3) by both immunohistochemical and immunofluorescence (IF) analysis and in bone marrow biopsies from CLL cases (n=5) by immunohistochemistry, a clear EMILIN1 positive staining was detected intermingled with the neoplastic component and unexpectedly in the cytoplasm of some cells. To verify whether EMILIN1 could promote CLL cells adhesion, in-vitro adhesion assays were performed. Both the CLL-derived CD49d+ Mec-1 cell line and primary CD49d+ CLL cells (n=12) were specifically able to adhere onto EMILIN1 substrates, and the adhesion efficiency was similar to that observed on VCAM-1 and CS-1 fragment as previously demonstrated. Adhesion was specifically blocked by pre-treatment with the anti-CD49d HP1/2 blocking antibody, and absent when a mutant gC1q domain was employed. The effects of CD49d/EMILIN1 interactions in CLL were next investigated performing short-term adhesion onto EMILIN1 substrates. Western blot analysis documented an increased phosphorylation of AKT and ERK1/2, mediators of survival signals, similar to that observed upon CD49d engagement by VCAM-1 and CS-1 fragment. These results were corroborated by IF analysis showing pAKT and pERK up-regulation, and the concomitant increase of pVAV1 and F-actin reorganization, confirming the activation of the integrin signaling pathway. Finally, we verified whether CD49d/EMILIN1 interaction was able to protect CLL cells from spontaneous apoptosis, by culturing purified cells from CLL cases (n=13) on gC1q domain, VCAM-1, or control substrate (1% BSA), and checking cell viability after 5 days by annexinV/7-AAD staining. Both VCAM-1 and EMILIN1 were able to protect CLL cells from spontaneous apoptosis (p=0.009 and p=0.002, respectively), the viability obtained on EMILIN1 was even significantly higher than that observed on VCAM-1 (p=0.004). In conclusion for the first time we showed that EMILIN1 is present in normal and CLL-involved tissues, and it is able to efficiently bind to CD49d, as expressed by CLL cells. At variance of what demonstrated in non-hematopoietic models, EMILIN1 was able to deliver anti-apoptotic/pro-survival signals to circulating CLL cells. These evidences imply a role for CD49d/EMILIN1 interaction in the maintenance of the neoplastic clone in CD49d-expressing CLL.
MIUR : Settore MED/06 - Oncologia Medica
Lingua: eng
Data: 4-apr-2014
Corso di dottorato: Dottorato di ricerca in Scienze biomediche e biotecnologiche
Ciclo di dottorato: 26
Università di conseguimento titolo: Università degli Studi di Udine
Luogo di discussione: Udine
Altre informazioni: Co-supervisori: Valter Gattei, Antonella Zucchetto
Citazione: Tissino, E. The Elastin MIcrofibriL INterfacer1 (EMILIN1), is highly expressed in chronic lymphocytic leukemia (CLL)-involved tissues and promotes CLL cell adhesion and survival via CD49d. (Doctoral Thesis, Università degli Studi di Udine, 2014).
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