OC 27.5 - Single-cell RNA-Seq reveals age-dependent Mfsd2b expression in megakaryocyte progenitors that regulates thrombosis

Background: Aging is associated with a dramatic increase in thrombotic events, which represent a major cause of morbidity and mortality worldwide. The recent findings demonstrate that the sphingosine-1-phosphate (S1P) transporter, Mfsd2b, is critical for platelet function and thrombosis. However, it is still uncovered whether and how aging affects the diversification and function of Mfsd2b in bone marrow-megakaryocyte progenitor cell (MkP) subpopulations, which thereby regulate platelet hyperreactivity and thrombosis.

Aims: We, therefore, sought to characterize, by single-cell RNA-sequencing (scRNA-seq), BM-MkP aging landscape and age-related Mfsd2b regulation of plasma S1P-Ceramides for studying their effects on platelet hyperreactivity and thrombus formation.

Methods: Here, we use scRNA-seq analysis in old (>24-moths) and young (3-months) mice to characterize age-related bone-marrow MkP heterogeneity. We identify rare subpopulations of MkPs that increases in abundance of Mfsd2b, and we show that these subpopulations regulate plasma S1P and ceramides which thereby modulate platelet hyperreactivity and thrombosis.

Results: Our single-cell transcriptomics reveals that Mfsd2b-expressing cells of specific subpopulations are substantially enriched in bone marrow from old mice. These could explain higher concentrations of S1P 18:1 and ceramides 18:1/16:0 and 18:1/18:0, which are positively correlated with platelet hyperresponsiveness to multiple agonists and heightened thrombus formation in aged mice. Employing Mfsd2b-siRNA, we reveal that MkPs contribute to reduced expression of cellular senescence markers γ-H2A.X (telomere shortening) and p53 (cell cycle arrest), and caspase-3 (apoptosis), and increased expression of Sirt1, which is known to delay cellular senescence and rejuvenate aged cells.

Conclusion(s): Our studies demonstrated that aging affects Mfsd2b expression in specific MkP subpopulations that regulate circulating S1P and ceramides. These findings indicate that modulation of sphingolipids and ceramides by deletion of Mfsd2b strongly regulates aging-related platelet hyperreactivity and thrombus formation, which suggests that targeting Mfsd2b in specific MkP subpopulations could contribute to important clues to future personalized medicine for the prevention of thrombotic disorders in aging.