Lymphocyte egress from LNs into lymph requires lymphocyte-intrinsic sphingosine-1-phosphate receptor 1 (S1P1), a G protein–coupled receptor (Matloubian et al., 2004
). S1P1 promotes migration into exit structures, the lymphatic vessel endothelial hyaluronan receptor 1+
) cortical sinuses of the LNs, within which lymphocytes may be captured by lymphatic flow and transported to the efferent lymph (Pham et al., 2008
; Grigorova et al., 2009
; Sinha et al., 2009
). S1P is normally low in the lymphoid tissue and abundant in blood and lymph, and disruption of this S1P gradient results in an egress block (Schwab et al., 2005
; Pappu et al., 2007
). However, despite its importance for lymphocyte egress, the cellular source of lymph S1P remains unknown (Schwab and Cyster, 2007
S1P production is dependent on sphingosine kinase (Sphk) 1 and 2, enzymes that are expressed in most eukaryotic cell types (Kono et al., 2008
). Recent work has demonstrated that red blood cells are a major source of plasma S1P, whereas all lymph S1P and ~5% of plasma S1P are supplied by a distinct, radiation-resistant source (Pappu et al., 2007
). In vitro studies have shown that blood endothelial cells (BECs) can act as a source of S1P (Venkataraman et al., 2008
). However, it has not been determined whether endothelial cells are an important source of S1P in vivo.
Lymphatic endothelial cells (LECs) arise from the venous endothelium during embryonic development at around embryonic day (E) 9–9.5, when a subpopulation of endothelial cells of the anterior cardinal vein commit to the lymphatic lineage by turning on Prox1
expression (Karpanen and Alitalo, 2008
). LYVE-1 is the earliest expressing and one of the most specific and widely used markers for LECs (Karpanen and Alitalo, 2008
; Oliver and Srinivasan, 2008
Mice lacking Sphk1
die in utero between E11.5–13.5 because of blood vascular defects (Mizugishi et al., 2005
). In vitro, stimulation of BECs with S1P increases localization of vascular endothelial cadherin (VE-cadherin) at cell–cell junctions and induces tubular morphogenesis (Lee et al., 1999
). Recently, S1P was demonstrated to promote tubular formation of human dermal LECs in vitro and lymphangiogenesis in Matrigel in vivo (Yoon et al., 2008
). However, whether S1P signaling normally plays a role in the development of the lymphatic system is not known.
In this report, by examining mice that lack Sphk2 and have Sphk1 conditionally deleted by a CRE recombinase expressed from the Lyve-1 locus, we provide evidence that LECs are the major source of lymph S1P. Lymphatic Sphk-deficient mice experienced a block of T and B cell egress from LNs. Additionally, lymphatic Sphk-deficient mice displayed altered initial lymphatic vessel morphology and junctional VE-cadherin patterning in the trachea and diaphragm.