TY - JOUR
T1 - Domain 1 of the urokinase-type plasminogen activator receptor is required for its morphologic and functional, β2 integrin-mediated connection with actin cytoskeleton in human microvascular endothelial cells
T2 - Failure of association in systemic sclerosis endothelial cells
AU - Margheri, Francesca
AU - Manetti, Mirko
AU - Serratì, Simona
AU - Nosi, Daniele
AU - Pucci, Marco
AU - Matucci-Cerinic, Marco
AU - Kahaleh, Bashar
AU - Bazzichi, Laura
AU - Fibbi, Gabriella
AU - Ibba-Manneschi, Lidia
AU - Del Rosso, Mario
PY - 2006/12
Y1 - 2006/12
N2 - Objective. In systemic sclerosis (SSc) microvascular endothelial cells (MVECs), angiogenesis is blocked by matrix metalloproteinase 12-dependent cleavage of domain 1 of the urokinase-type plasminogen activator receptor (uPAR). Since integrins are associated with the invasive activity of uPAR in angiogenesis, this study was undertaken to show whether full-size and truncated uPAR are differentially associated with integrins and with motor components of the cytoskeleton. Methods. SSc and normal MVECs were isolated from human skin biopsy specimens and studied by confocal laser scanning microscopy and immunoprecipitation to assess the mechanisms of association of truncated and full-size uPAR with integrins and the actin cytoskeleton. The integrin composition of the MVECs was studied by reverse transcription-polymerase chain reaction. Cell migration and capillary morphogenesis were studied on fibrinogen substrates. Involvement of Rac and Cdc42 was evaluated by Western blotting. Results. Only full-size uPAR showed a connection with the actin cytoskeleton in ECs. This connection was mediated by the uPAR-associated αM- and αX-subunits of β2 integrin, and was absent from SSc MVECs. The cleaved uPAR was not associated with β2 integrins or with actin. β3 integrins were associated with both the full-size and cleaved uPAR at focal contacts. The uncoupling of uPAR from β2 integrins in SSc MVECs impaired the activation of Rac and Cdc42 (thus inhibiting their mediation of uPAR-dependent cytoskeletal rearrangements and cell motility) and blocked the integrin-engagement-delivered signals to the actin cytoskeleton. Invasion and capillary morphogenesis on fibrinogen-coated substrates indicated that ligation of uPAR by uPA empowers the β2/β3 integrin-dependent invasion of fibrinogen, and that this system is impaired in SSc MVECs. Conclusion. The reduced angiogenic properties of SSc MVECs can be explained by the effects of uPAR truncation and the subsequent loss of the β2 integrin-mediated connection of uPAR with the actin cytoskeleton in these ECs.
AB - Objective. In systemic sclerosis (SSc) microvascular endothelial cells (MVECs), angiogenesis is blocked by matrix metalloproteinase 12-dependent cleavage of domain 1 of the urokinase-type plasminogen activator receptor (uPAR). Since integrins are associated with the invasive activity of uPAR in angiogenesis, this study was undertaken to show whether full-size and truncated uPAR are differentially associated with integrins and with motor components of the cytoskeleton. Methods. SSc and normal MVECs were isolated from human skin biopsy specimens and studied by confocal laser scanning microscopy and immunoprecipitation to assess the mechanisms of association of truncated and full-size uPAR with integrins and the actin cytoskeleton. The integrin composition of the MVECs was studied by reverse transcription-polymerase chain reaction. Cell migration and capillary morphogenesis were studied on fibrinogen substrates. Involvement of Rac and Cdc42 was evaluated by Western blotting. Results. Only full-size uPAR showed a connection with the actin cytoskeleton in ECs. This connection was mediated by the uPAR-associated αM- and αX-subunits of β2 integrin, and was absent from SSc MVECs. The cleaved uPAR was not associated with β2 integrins or with actin. β3 integrins were associated with both the full-size and cleaved uPAR at focal contacts. The uncoupling of uPAR from β2 integrins in SSc MVECs impaired the activation of Rac and Cdc42 (thus inhibiting their mediation of uPAR-dependent cytoskeletal rearrangements and cell motility) and blocked the integrin-engagement-delivered signals to the actin cytoskeleton. Invasion and capillary morphogenesis on fibrinogen-coated substrates indicated that ligation of uPAR by uPA empowers the β2/β3 integrin-dependent invasion of fibrinogen, and that this system is impaired in SSc MVECs. Conclusion. The reduced angiogenic properties of SSc MVECs can be explained by the effects of uPAR truncation and the subsequent loss of the β2 integrin-mediated connection of uPAR with the actin cytoskeleton in these ECs.
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U2 - 10.1002/art.22263
DO - 10.1002/art.22263
M3 - Article
C2 - 17133606
AN - SCOPUS:33845666597
VL - 54
SP - 3926
EP - 3938
JO - Arthritis care and research : the official journal of the Arthritis Health Professions Association
JF - Arthritis care and research : the official journal of the Arthritis Health Professions Association
SN - 0893-7524
IS - 12
ER -