Enhanced sphingosine-1-phosphate receptor 2 expression underlies female CNS autoimmunity susceptibility
Lillian Cruz-Orengo, … , Seth D. Crosby, Robyn S. Klein
Lillian Cruz-Orengo, … , Seth D. Crosby, Robyn S. Klein
Published May 8, 2014
Citation Information: J Clin Invest. 2014;124(6):2571-2584. https://doi.org/10.1172/JCI73408.
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Research Article Autoimmunity

Enhanced sphingosine-1-phosphate receptor 2 expression underlies female CNS autoimmunity susceptibility

Abstract

Multiple sclerosis (MS) is an inflammatory disease of the CNS that is characterized by BBB dysfunction and has a much higher incidence in females. Compared with other strains of mice, EAE in the SJL mouse strain models multiple features of MS, including an enhanced sensitivity of female mice to disease; however, the molecular mechanisms that underlie the sex- and strain-dependent differences in disease susceptibility have not been described. We identified sphingosine-1-phosphate receptor 2 (S1PR2) as a sex- and strain-specific, disease-modifying molecule that regulates BBB permeability by destabilizing adherens junctions. S1PR2 expression was increased in disease-susceptible regions of the CNS of both female SJL EAE mice and female patients with MS compared with their male counterparts. Pharmacological blockade or lack of S1PR2 signaling decreased EAE disease severity as the result of enhanced endothelial barrier function. Enhanced S1PR2 signaling in an in vitro BBB model altered adherens junction formation via activation of Rho/ROCK, CDC42, and caveolin endocytosis-dependent pathways, resulting in loss of apicobasal polarity and relocation of abluminal CXCL12 to vessel lumina. Furthermore, S1PR2-dependent BBB disruption and CXCL12 relocation were observed in vivo. These results identify a link between S1PR2 signaling and BBB polarity and implicate S1PR2 in sex-specific patterns of disease during CNS autoimmunity.

Authors

Lillian Cruz-Orengo, Brian P. Daniels, Denise Dorsey, Sarah Alison Basak, José G. Grajales-Reyes, Erin E. McCandless, Laura Piccio, Robert E. Schmidt, Anne H. Cross, Seth D. Crosby, Robyn S. Klein

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Figure 7

S1PR2 signaling dysregulates CNS endothelial barrier structure and function through Rho/ROCK, CDC42, and caveolin endocytosis-dependent pathways.

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S1PR2 signaling dysregulates CNS endothelial barrier structure and funct...
(A–F) Paracellular permeability of in vitro BBB cultures was assessed by electrode recording of TEER, reported in Ω/cm2. (A) TEER after treatment of in vitro BBB cultures with 10, 100, or 1,000 nM exogenous S1P for 4 hours or (B and C) 1,000 nM S1P for 4 hours with or without (B) 1,000 nM JTE-013 at 2 hours or (C) the S1PR1-specific antagonist W146 (1,000 nM) at 2 hours. (D–F) TEER after 1,000 nM S1P treatment of BBB cultures pretreated for 2 hours with (D) the caveolin endocytosis inhibitor MBCD (10 mM), (E) Rho-associated protein kinase (ROCK) inhibitor H1152P (10 nM), or (F) CDC42 inhibitor ML141 (100 nM). TEER values are mean ± SEM of 6 to 9 replicates of 2 to 3 independent experiments. ***P < 0.001, repeated-measures 2-way ANOVA. (G–L) Immunocytochemical staining of AJs in HCMEC/D3 cells via labeling of VE-cadherin (red, left and middle panels; scale bar: 30 μm) and confocal z-stack reconstruction of HCMEC/D3 cells (right panels; scale bar: 15 μm), demonstrating polarized expression of canonical apical marker GGT-1 (green) and basolateral CXCL12 (red) after treatment with (G) vehicle or 1,000 nM S1P for 4 hours followed by treatment with (H) JTE, (I) W146, (J) MBCD, (K) H1152P, or (L) ML141 treatment at 2 hours. Inhibitor concentrations in H–L are identical to those in B–F. Immunocytochemical images are representative images of 2 to 3 independent experiments. IC, isotype controls.