The Effect of Sphingosine 1‐Phosphate on Endothelial Permeability Is Fluid Flow Dependent
Authors: Akbari Ehsan , Spychalski Griffin B. , Rangharajan Kaushik K. , Prakash Shaurya , Song Jonathan W. 
Published: June 2021

Blood vessel permeability is mediated by the remodeling of endothelial cells (EC) that are connected by specialized junctional complexes. Sphingosine‐1‐phosphate (S1P), a bioactive lipid molecule localized in the blood plasma, has been implicated in regulating endothelial barrier function1, and the biological effect of S1P has been shown to be flow‐dependent. For example, application of laminar shear stress (LSS) upregulates the expression of S1P receptor 1 (S1PR1). However, the role of fluid forces that arise at vessel bifurcations in regulating the effect of S1P remains unclear. To address this gap in knowledge, we developed a novel microfluidic model of a bifurcating vessel (MBV) that reproduces the main fluid forces present in the branching microvasculature, i.e. bifurcating fluid flow (BFF) applied normal and LSS applied tangential on the ECs, to study endothelial hydraulic conductivity (Lp) in response to applied S1P and selective inhibition of S1P receptors 1 and 2 (S1PR2).

The MBV consists of a 1300 μm wide inlet microchannel that bifurcates to two 500 μm wide daughter microchannels (Fig. 1A). In addition, the microdevice includes a central compartment filled with a collagen type I solution that recapitulates the perivascular extracellular matrix (ECM). Polydimethylsiloxane soft lithography was used to fabricate MBV with monolithic features 50 μm in height. The microchannels were lined with human umbilical endothelial cells (HUVECs) (Fig. 1B), and flow rates were controlled with a syringe pump. W123 (10 μM) and JTE‐013 (200 nM) were used to pharmacologically inhibit S1PR1 and S1PR2, respectively. The extravasation rate of 10 kDa FITC‐conjugated Dextran as a fluorescent tracer was quantified to measure Lp under each condition.

Treatment with 500 nM S1P for 6 h under static condition caused a ~7‐fold increase in Lp (Fig.2A). In contrast, treatment with 500 nM S1P after inhibiting S1PR2 with JTE‐013 decreased Lp by ~5‐fold, suggesting that S1PR2 functions to destabilize endothelial barrier function. Interestingly, treatment with 500 nM S1P after inhibition of S1PR1 did not elicit a change in Lp suggesting that functioning S1PR1 is necessary to induce an increase in Lp by S1P (Fig.2A). Application of 500 nM S1P alongside BFF, which is characterized by stagnated fluid flow at the base of the bifurcation point and LSS (~3 dyn/cm2) in the downstream branched vessels decreased Lp by 45% and 52%, respectively, compared to static control condition (Fig.2B). Interestingly, the application of BFF and LSS in absence of S1P elicited similar responses to the case where S1P was present during perfusion (Fig.2B), suggesting a stabilizing role for BFF and LSS independent of S1P.

We developed an in vitro microfluidic model of a branching vessel that enables accurate and quantitative measurements of Lp under the effect of S1P and fluid flow stimuli. The findings from this study can lead to the development of novel therapeutic strategies to restore dysregulated vascular barrier.

Support or Funding Information

Funding was provided by the American Heart Association, and the American Cancer Society.

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.