Metastases are responsible for more than 90% of cancer related mortality. The hematogenous metastatic invasion is a complex process in which the endothelium plays a key role. Extravasation is a dynamic process involving remodeling and change in cell shape and in cytoskeleton whereby a series of strongly dependent interactions between CTCs and endothelium occurs [1]. Talins are proteins regulating focal adhesions and cytoskeleton remodeling. Talin-1 seems to be involved in the aggressiveness, motility, survival and invadopodia formation of cancer cells throughout the entire metastatic cascade [2], being up-regulated in breast cancer cells and mutated in sarcomas. Understand the implication of talin-1 in extravasation could facilitate the design of new therapies and finally fight cancer. AIM: We hypothesized that Talin-1 could be specifically involved in extravasation driving each of its steps. MATERIALS AND METHODS: We developed a human 3D microfluidic model that enables the study of human cancer cell extravasation within a perfusable human microvascularized organ specific environment[3]. For the study of extravasation we applied microfluidic approach through the development of a microfluidic device in which endothelial cells and fibroblasts generated a 3D human functional vascular networks. Microvessel characterization was performed with immunofluorescence and permeability assays. We knocked-down talin-1 in triple negative breast cancer cell line MDA-MB231 and metastatic fibro-sarcoma cell line HT1080 with SiRNA and verified by Western-blot. Cancer cells were then perfused in the vessels and extravasation monitored through confocal imaging. RESULTS: We developed a human vascularized 3D microfluidic device with human perfusable capillary-like structures embedded in fibrin matrix, characterized by mature endothelium markers and physiological permeability (1.5±0.76)×10(-6) cm/s. We focused on the role of Talin-1 in adhesion to endothelium, trans-endothelial migration (TEM) and early invasion. Adhesion to the endothelium, TEM and migration within the ECM were monitored through confocal analyses. We demonstrated that Talin-1 KD significantly reduced the adhesion efficiency and TEM in both cell lines. Early invasion was also strongly and statistically reduced by the SiRNA treatment in both cell lines. CONCLUSIONS: We proved Talin-1 function in driving the extravasation mechanism in a human 3D vascularized environment. We demonstrated that Talin-1 is involved in each part of extravasation significantly affecting adhesion, TEM and the invasion stages. Targeting this protein could thus be an effective strategy to block metastasis

Gilardi, M., Bersini, S., Calleja, A., Kamm, R., Vanoni, M., Moretti, M. (2016). The key role of talin-1 in cancer cell extravasation dissected through human vascularized 3D microfluidic model. In 8th International Conference on Thrombosis and Hemostasis Issues in Cancer (pp.S180-S181). THE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, ENGLAND : PERGAMON-ELSEVIER SCIENCE LTD [10.1016/S0049-3848(16)30145-1].

The key role of talin-1 in cancer cell extravasation dissected through human vascularized 3D microfluidic model

VANONI, MARCO ERCOLE
Penultimo
;
2016

Abstract

Metastases are responsible for more than 90% of cancer related mortality. The hematogenous metastatic invasion is a complex process in which the endothelium plays a key role. Extravasation is a dynamic process involving remodeling and change in cell shape and in cytoskeleton whereby a series of strongly dependent interactions between CTCs and endothelium occurs [1]. Talins are proteins regulating focal adhesions and cytoskeleton remodeling. Talin-1 seems to be involved in the aggressiveness, motility, survival and invadopodia formation of cancer cells throughout the entire metastatic cascade [2], being up-regulated in breast cancer cells and mutated in sarcomas. Understand the implication of talin-1 in extravasation could facilitate the design of new therapies and finally fight cancer. AIM: We hypothesized that Talin-1 could be specifically involved in extravasation driving each of its steps. MATERIALS AND METHODS: We developed a human 3D microfluidic model that enables the study of human cancer cell extravasation within a perfusable human microvascularized organ specific environment[3]. For the study of extravasation we applied microfluidic approach through the development of a microfluidic device in which endothelial cells and fibroblasts generated a 3D human functional vascular networks. Microvessel characterization was performed with immunofluorescence and permeability assays. We knocked-down talin-1 in triple negative breast cancer cell line MDA-MB231 and metastatic fibro-sarcoma cell line HT1080 with SiRNA and verified by Western-blot. Cancer cells were then perfused in the vessels and extravasation monitored through confocal imaging. RESULTS: We developed a human vascularized 3D microfluidic device with human perfusable capillary-like structures embedded in fibrin matrix, characterized by mature endothelium markers and physiological permeability (1.5±0.76)×10(-6) cm/s. We focused on the role of Talin-1 in adhesion to endothelium, trans-endothelial migration (TEM) and early invasion. Adhesion to the endothelium, TEM and migration within the ECM were monitored through confocal analyses. We demonstrated that Talin-1 KD significantly reduced the adhesion efficiency and TEM in both cell lines. Early invasion was also strongly and statistically reduced by the SiRNA treatment in both cell lines. CONCLUSIONS: We proved Talin-1 function in driving the extravasation mechanism in a human 3D vascularized environment. We demonstrated that Talin-1 is involved in each part of extravasation significantly affecting adhesion, TEM and the invasion stages. Targeting this protein could thus be an effective strategy to block metastasis
Si
abstract + poster
Scientifica
microfluidic, 3D human vascularized models, cancer metastases, focal adhesion, Talin-1, breast cancer, extravasation, vascular adhesion, 3D invasion
English
International Conference on Thrombosis and Hemostasis Issues in Cancer April 8-10
Gilardi, M., Bersini, S., Calleja, A., Kamm, R., Vanoni, M., Moretti, M. (2016). The key role of talin-1 in cancer cell extravasation dissected through human vascularized 3D microfluidic model. In 8th International Conference on Thrombosis and Hemostasis Issues in Cancer (pp.S180-S181). THE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, ENGLAND : PERGAMON-ELSEVIER SCIENCE LTD [10.1016/S0049-3848(16)30145-1].
Gilardi, M; Bersini, S; Calleja, A; Kamm, R; Vanoni, M; Moretti, M
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/10281/139623
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