Glycosphingolipids on Human Myeloid Cells Stabilize E-Selectin-Dependent Rolling in the Multistep Leukocyte Adhesion Cascade

Nandini Mondal, Gino Stolfa, Aristotelis Antonopoulos, Yuqi Zhu, Shuen Shiuan Wang, Alexander Buffone, G. Ekin Atilla-Gokcumen, Stuart M. Haslam, Anne Dell, Sriram Neelamegham

Research output: Contribution to journalArticlepeer-review

32 Scopus citations

Abstract

Objective - Recent studies suggest that the E-selectin ligands expressed on human leukocytes may differ from those in other species, particularly mice. To elaborate on this, we evaluated the impact of glycosphingolipids expressed on human myeloid cells in regulating E-selectin-mediated cell adhesion. Approach and Results - A series of modified human cell lines and primary neutrophils were created by targeting UDP-Glucose Ceramide Glucosyltransferase using either lentivirus-delivered shRNA or CRISPR-Cas9-based genome editing. Enzymology and mass spectrometry confirm that the modified cells had reduced or abolished glucosylceramide biosynthesis. Glycomics profiling showed that UDP-Glucose Ceramide Glucosyltransferase disruption also increased prevalence of bisecting N-glycans and reduced overall sialoglycan expression on leukocyte N- and O-glycans. Microfluidics-based flow chamber studies demonstrated that both the UDP-Glucose Ceramide Glucosyltransferase knockouts and knockdowns display ≈60% reduction in leukocyte rolling and firm adhesion on E-selectin bearing stimulated endothelial cells, without altering cell adhesion to P-selectin. Consistent with the concept that the glycosphingolipids support slow rolling and the transition to firm arrest, inhibiting UDP-Glucose Ceramide Glucosyltransferase activity resulted in frequent leukocyte detachment events, skipping motion, and reduced diapedesis across the endothelium. Cells bearing truncated O- and N-glycans also sustained cell rolling on E-selectin, although their ability to be recruited from free fluid flow was diminished. Conclusions - Glycosphingolipids likely contribute to human myeloid cell adhesion to E-selectin under fluid shear, particularly the transition of rolling cells to firm arrest.

Original languageEnglish (US)
Pages (from-to)718-727
Number of pages10
JournalArteriosclerosis, Thrombosis, and Vascular Biology
Volume36
Issue number4
DOIs
StatePublished - Apr 1 2016
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Cardiology and Cardiovascular Medicine

Keywords

  • cell adhesion molecule
  • endothelial cell
  • flow shear stress
  • inflammation
  • leukocyte

Fingerprint

Dive into the research topics of 'Glycosphingolipids on Human Myeloid Cells Stabilize E-Selectin-Dependent Rolling in the Multistep Leukocyte Adhesion Cascade'. Together they form a unique fingerprint.

Cite this