Interaction of human blood platelets, lymphocytes and monocytes with vascular laminin isoforms

Abstract

Inflammatory and immune responses play a fundamental role in both health and disease, and leukocytes are important actors in these processes. Migration of the leukocytes to sites of injury or inflammation is a crucial component of innate and adaptive immunity. It is currently accepted that leukocyte extravasation is a multistep process. However, a later step in this cascade, namely the interaction of leukocytes with components of extracellular matrices (ECM), such as the vascular endothelial basement membrane (BM) and the interstitium ECM, is poorly understood. There is also limited information concerning the role of vascular BM proteins in hemostasis and/or thrombosis. In this thesis, the interaction of blood platelets, lymphocytes and monocytes with vascular BM components, particularly the endothelial laminin isoforms, has been studied.

Laminins (Lms), major components of all BMs, are a family of heterotrimeric molecules, each composed of á-, â-, and ã-chains. To date, five á-, three â-, and three ã-chains have been identified that associate to form at least 15 Lm isoforms. Lms regulate various cellular functions, such as adhesion, motility, differentiation and proliferation through various integrin and nonintegrin receptors. Lm-411 (á4â1ã1, laminin-8) and Lm-511 (á5â1ã1, laminin-10) are major Lm isoforms of vascular endothelial BMs. These BM components may participate in leukocyte extravasation and, following vascular injury, contribute to hemostasis and/or thrombosis when exposed to circulating platelets.

First, commercially available placenta laminin preparations, often used in functional studies, were characterized. These preparations differed from one another and consisted of highly fragmented proteins, a mixture of laminin isoforms, and/or contaminating fibronectin. They also exhibited major functional differences between batches. In a following study, megakaryocytic cells were found to synthesize and platelets to secrete heterotrimeric á5-Lms. Lm-511 strongly promoted platelet adhesion, but not activation, via á6â1 integrin. Thereafter, the pivotal role of á5-Lm(s), expressed by high endothelial venules, in promoting adhesion and migration of blood lymphocytes via á6â1 integrin was demonstrated. Lm-511 was also able to co-stimulate T cell proliferation, and stimulated blood lymphocytes secreted both á4- and á5-laminins. The lymph node cell number in Lmá4-deficient mice compared to wild type did not differ significantly. Finally, Lm-411 and Lm-511 were found to mediate adhesion and chemokine-induced migration of monocytes via áMâ2 and áXâ2 integrins. Isolated Lmã1, but not Lmâ1, chain reproduced the effect of the Lm heterotrimers. Moreover, endogenous á4-Lm(s) mediated chemokine-induced, áMâ2- and áXâ2-integrin dependent monocyte migration on an albumin substrate.

Altogether, the present studies illustrate the differential effects of laminin isoforms in the biology of platelets, lymphocytes and monocytes, and their potential contribution to hemostasis, and to the generation of immune and inflammatory responses.