Enhancement of capillary leakage and restoration of lymphocyte egress by a chiral S1P 1 antagonist in vivo

Sphingosine 1-phosphate (S1P, 1) regulates vascular barrier and lymphoid development, as well as lymphocyte egress from lymphoid organs, by activating high-affinity S1P1 receptors. We used reversible chemical probes (i) to gain mechanistic insights into S1P systems organization not accessible through genetic manipulations and (ii) to investigate their potential for therapeutic modulation. Vascular (but not airway) administration of the preferred R enantiomer of an in vivo‐active chiral S1P1 receptor antagonist induced loss of capillary integrity in mouse skin and lung. In contrast, the antagonist did not affect the number of constitutive blood lymphocytes. Instead, alteration of lymphocyte trafficking and phenotype required supraphysiological elevation of S1P1 tone and was reversed by the antagonist. In vivo two-photon imaging of lymph nodes confirmed requirements for obligate agonism, and the data were consistent with the presence of a stromal barrier mechanism for gating lymphocyte egress. Thus, chemical modulation reveals differences in S1P-S1P1 ‘set points’ among tissues and highlights both mechanistic advantages (lymphocyte sequestration) and risks (pulmonary edema) of therapeutic intervention. Chemical agents provide powerful tools for dissecting complex physiological functions mediated through diverse receptor subtypes. In particular, selective agonist and antagonist pairs that are active in vivo have the distinct advantage of enabling acute, reversible modulation of molecular function while circumventing the developmental compensations that can arise in gene deletion studies. We have targeted this approach to the signaling pathway mediated by S1P and have thereby showed that in vivo–active reversible chemical tools can be used to address a series of mechanistic and therapeutic questions. S1P is a pleiotropic autocrine and paracrine signaling lipid 1 that mediates graded rheostat control of numerous physiological functions through a family of G protein–coupled receptors. Small variations in ligand concentration are amplified by selective high-affinity receptors to acutely regulate vital functions such as heart rate 2,3 ,v ascular and stromal barrier integrity 4 and lymphocyte egress 5 .T he functioning of S1P receptors in the maintenance and modulation of biological barrier activity is of profound biological importance and has therapeutic implications 4 , including prevention of transplant rejection and treatment of multiple sclerosis and perhaps adult respiratory distress syndrome as well 6 .