S1P and S1PR1 are used by T cells to migrate from thymus across microvascular endothelium to the blood circulation, and across medullary lymphatic endothelium of the lymph node (LN) for egress to efferent lymphatics and the blood circulation. However, whether S1PRs regulate T cell migration from tissues across afferent lymphatic endothelium and into draining LN are unknown. We hypothesized that different S1PRs are utilized by CD4 T cells and lymphatic endothelial cells (LECs) to promote CD4 T cell afferent lymphatic migration.

Methods: MS-1 blood endothelial and SVEC4-10 lymphatic endothelial cell lines and human and murine primary LECs were used in transwell assays. Human and murine CD4 T cells placed in the upper chamber were migrated across endothelium to chemokines, cytokines or S1P.

Results: Human and murine CD4 T cell migration toward S1P was enhanced by T cell-LEC interactions. In contrast, migration to CCL19, CCL21 and several other chemokines and cytokines was not enhanced by T cell-LEC interactions. CD4 T cell migration toward S1P was both chemokinetic and chemotactic. Pretreatment of CD4 T cells but not LEC with the non-specific S1PR inhibitors pertussis toxin, FTY720, or S1P inhibited CD4 T cell migration across LEC, showing that S1PRs played a critical role on CD4 T cell trans-lymphatic endothelial migration. The specific S1PR1, S1PR3, and S1PR4 inhibitors each blocked CD4 T cell function and migration, but did not affect the LEC functional ability to promote CD4 T cell migration. In contrast the S1PR2 inhibitor specifically blocked LEC and their ability to enhance CD4 T cell migration, but did not act on T cells to block migration. S1P upregulated VCAM-1 and VE-cadherin, but not ICAM-1 expression on LECs, and blocking anti-VCAM-1 or anti-VLA-4 mAbs inhibited CD4 T cell migration.

Conclusion: CD4 T cells and LEC utilized distinct S1PRs to regulate CD4 T cell migration across afferent lymphatics in a VCAM-1 and VLA-4 dependent fashion. S1P engages active processes in both T cells and LEC to promote migration. These results demonstrate for the first time unique roles for S1PRs in regulating T cell and LEC functions in migration. These findings suggest new and specific drug targets for regulating lymphatic migration in immunity and tolerance.

CITATION INFORMATION: Xiong Y, Brinkman C, Hippen K, Blazar B, Bromberg J. Sphingosine 1-Phosphate (S1P) Receptors Differentially Regulate Murine and Human CD4 T Cell Migration Across Lymphatic Endothelium. Am J Transplant. 2016;16 (suppl 3).

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