Sphingolipid dysregulation in erythrocytes during sickle cell disease contributes to pro-inflammatory microparticle generation and subsequent inflammatory cell activation

Sickle cell disease is a hereditary blood disorder caused by a point mutation in the gene encoding hemoglobin. This mutation causes hemoglobin molecules to polymerize during de-oxygenation of erythrocytes producing rod-shaped polymers that bend and distort the red blood cell membrane, making it more rigid and “sickled”. This sickling causes red blood cells to lose their flexibility and ability to navigate small capillaries and also enhances the production of pro-inflammatory membrane-derived microparticles, leading to chronic inflammation and many complications such as peripheral artery disease, stroke, myocardial infarction, vasculitis and even death. Sphingolipids are a class of lipids containing a backbone of sphingoid bases and are integral components of erythrocyte and microparticle membranes. Many of these lipids are known to mediate biological processes, but their expression, distribution and orientation in erythrocytes during sickle cell disease has never been explored. Sphingomyelin, the most abundant sphingolipid in the red blood cell membrane is hydrolyzed by sphingomyelinase to produce ceramide, which has been shown to alter membrane dynamics and enhance microvessel formation. Additionally, ceramide can be further metabolized to form sphingosine and sphingosine 1-phosphate, which is a bioactive ligand for 5 known G-protein coupled receptors present on most blood and vascular cells that modulates cell motility, proliferation, migration and phenotype. Prior to this work, it was not understood how sphingolipid metabolism contributes to vascular inflammation in sickle cell disease. Together, this body of work has elucidated key enzymatic and lipidomic alterations in sphingolipid metabolism (i.e. the activation of acid sphingomyelinase on red blood cells) that result in the production of sphingolipid-rich erythrocyte-derived microparticles, which enhance inflammatory cell activation. Our work has elucidated novel pharmacological targets to reduce microparticle generation and subsequent vascular inflammation in sickle cell disease.Ph.D

Selective Activation of Sphingosine 1-Phosphate Receptors 1 and 3 Promotes Local Microvascular Network Growth

Proper spatial and temporal regulation of microvascular remodeling is critical to the formation of functional vascular networks, spanning the various arterial, venous, capillary, and collateral vessel systems. Recently, our group has demonstrated that sustained release of sphingosine 1-phosphate (S1P) from biodegradable polymers promotes microvascular network growth and arteriolar expansion. In this study, we employed S1P receptor-specific compounds to activate and antagonize different combinations of S1P receptors to elucidate those receptors most critical for promotion of pharmacologically induced microvascular network growth. We show that S1P1 and S1P3 receptors act synergistically to enhance functional network formation via increased functional length density, arteriolar diameter expansion, and increased vascular branching in the dorsal skinfold window chamber model. FTY720, a potent activator of S1P1 and S1P3, promoted a 107% and 153% increase in length density 3 and 7 days after implantation, respectively. It also increased arteriolar diameters by 60% and 85% 3 and 7 days after implantation. FTY720-stimulated branching in venules significantly more than unloaded poly(D, L-lactic-co-glycolic acid). When implanted on the mouse spinotrapezius muscle, FTY720 stimulated an arteriogenic response characterized by increased tortuosity and collateralization of branching microvascular networks. Our results demonstrate the effectiveness of S1P1 and S1P3 receptor-selective agonists (such as FTY720) in promoting microvascular growth for tissue engineering applications