The phospholipid sphingosine-1-phosphate (S1P) has emerged as an important regulator of several cardiovascular functions. Our work has focussed on the microvascular effects of S1P, most notably, its key role for the regulation of myogenic tone. S1P signalling needs to be tightly controlled, since it possesses powerful pro-constrictive effects in resistance arteries. We have identified S1P phosphohydrolase 1 (SPP1) as a functional antagonist of sphingosine kinase 1 (Sk1; the S1P-synthesizing enzyme). Remarkably, despite its intracellular localization to the endoplasmatic reticulum, SPP1 effectively controls responses attributable to extracellular S1P (e.g., myogenic tone, Ca²⁺ sensitivity). We provide evidence that the CFTR transporter in the plasma membrane is the structural prerequisite for this function. CFTR therefore plays a critical role for (i) the degradation of extracellular S1P by SPP1 and (ii) the control of microvascular effects attributable to extracellular S1P. The present study aims to provide additional evidence for this proposed novel role of CFTR within the microcirculation in that it demonstrates: (i) CFTR-dependent uptake of fluorescently-labelled S1P into vascular smooth muscle cells (VSMC); (ii) enhancement of S1P-dependent effects following chemical inhibition of CFTR and (iii) evidence that TNFa enhances S1P-dependent effects via downregulation of CFTR expression. VSMC were isolated from mesenteric arteries of CFTR+/+ or CFTR-/- mice. FACS analysis revealed uptake of S1P-FITC (1mmol/L) in CFTR+/+ VSMC, which was abolished following treatment with a CFTR inhibitor (CFTR-inh(172)). S1P FITC uptake was absent in VSMCs from CFTR-/- mice; forced expression of CFTR in CFTR-/- cells allowed the uptake of S1P-FITC. S1P dose-dependently inhibited VSMC proliferation. The inhibitory effect of 10nmol/L S1P (365%, n=6) was enhanced following treatment with CFTR-inh(172) (628%, n=6). The inhibitory effect of 10nmol/L S1P was larger in VSMCs lacking CFTR (635% inhibition, n=6) compared to wild-type VSMCS. Incubation with 25ng/ml TNFa, a cytokine known to negatively regulate CFTR expression, significantly reduced CFTR mRNA and protein levels. The uptake of FITC-S1P was reduced in VSMCs treated with TNFa. Our results suggest that the uptake and degradation of S1P in VSMCs strongly depends on the presence of CFTR in the plasma membrane. Regulation of CFTR expression (for example, by the cytokine TNFa) can significantly shift the delicate balance between S1P synthesis and degradation towards synthesis, leading to an enhancement of extracellular S1P-dependent effects on VSMCs.

We suggest that CFTR, by virtue of its intimate link with S1P signalling, represents a novel regulator of microvascular tone and even other complex cardiovascular functions.