Real-time analysis of integrin-mediated chemotactic migration of T lymphocytes within 3-D extracellular matrix-like gels.
We have developed a novel 3-D gel reconstituted with major extracellular matrix (ECM) glycoproteins to follow the dynamics of migration of human T cells locomoting, in real-time, on gradients formed by representative chemoattractants: the C-C chemokine RANTES, and the pro-inflammatory cytokine IL-2. In the absence of chemoattractants, none of the T cells migrated directionally and the levels of random migration or cell polarization were low. However, major fractions of T cells placed in IL-2 and RANTES gradients in the gels polarized immediately after exposure to the chemoattractants. Shortly after polarization, 25% of the T cells migrated, in either a random or directional fashion, towards the sources of the chemoattractants; additional 5-10% of the cells remained polarized but stationary. The number of T cells migrating directionally towards RANTES or IL-2 peaked along with the formation of the chemotactic gradients. The directional migration of T cells was increased by a short pre-exposure to low doses of IL-2, which did not alter the level of expression of the beta1 integrins. The directional migration of T cells towards IL-2 and RANTES was mediated by IL-2R and pertussis toxin-sensitive receptors, respectively, and the directional, and to a lesser degree, the random locomotion of T cells induced by both chemoattractants required intact tyrosine kinase signaling and activities of the alpha4, alpha5, and, to a lesser degree, the alpha2 and alpha6 members the beta1 integrins. Our system enables the real-time tracking of individual locomoting lymphocytes and the analysis of their dynamic interactions with ECM components and cytokines.
Franitza
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The Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.
Alon
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Lider
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D002465:Cell Movement / Q000187:drug effects; D018946:Chemokine CCL5 / Q000502:physiology; D002634:Chemotaxis, Leukocyte / Q000187:drug effects*; D005109:Extracellular Matrix / Q000502:physiology; D005782:Gels; D006801:Humans; D019012:Integrin beta1 / Q000494:pharmacology; D016023:Integrins / Q000502:physiology*; D007376:Interleukin-2 / Q000502:physiology; D010827:Physiology / Q000295:instrumentation; D021382:Protein Sorting Signals / Q000502:physiology; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000166:cytology / Q000187:drug effects / Q000276:immunology*; D013997:Time Factors
D018946:Chemokine CCL5; D005782:Gels; D019012:Integrin beta1; D016023:Integrins; D007376:Interleukin-2; D021382:Protein Sorting Signals