Circulating TNFR1 exosome-like vesicles partition with the LDL fraction of human plasma

Abstract

Extracellular type I tumor necrosis factor receptors (TNFR1) are generated by two mechanisms, proteolytic cleavage of TNFR1 ectodomains and release of full-length TNFR1 in the membranes of exosome-like vesicles. Here, we assessed whether TNFR1 exosome-like vesicles circulate in human blood. Immunoelectron microscopy of human serum demonstrated TNFR1 exosome-like vesicles, with a diameter of 27-to 36-nm, while Western blots of human plasma showed a 48-kDa TNFR1, consistent with a membrane-associated receptor. Gel filtration chromatography revealed that the 48-kDa TNFR1 in human plasma co-segregated with LDL particles by size, but segregated independently by density, demonstrating that they are distinct from LDL particles. Furthermore, the 48-kDa exosome-associated TNFR1 in human plasma contained a reduced content of N-linked carbohydrates as compared to the 55-kDa membrane-associated TNFR1 from human vascular endothelial cells. Thus, a distinct population of TNFR1 exosome-like vesicles circulate in human plasma and may modulate TNF-mediated inflammation.

Shedding of Endogenous Interleukin-6 Receptor (IL-6R) Is Governed by A Disintegrin and Metalloproteinase (ADAM) Proteases while a Full-length IL-6R Isoform Localizes to Circulating Microvesicles

“…6B). Full-length IL-6R on microvesicles exhibited a higher electrophoretic mobility than its cellular counterpart in HEK293 cells transiently transfected with a human IL-6R cDNA, which is, however, consistent with a previous report showing less glycosylated TNFRI isoforms on exosomes (44). Moreover, we confirmed the specificity of the employed antibody toward the human IL-6R C terminus using a panel of IL-6R/IL-11R chimeras (Fig.…”

ST6Gal-I Regulates Macrophage Apoptosis via α2-6 Sialylation of the TNFR1 Death Receptor

“…However, to our knowledge, the glycan profile on the native transmembrane form of TNFR1 has not been previously characterized. There are three consensus sequences for N-glycosylation on the human TNFR1 molecule (25,26), but it is not known which of these sites are filled, nor is anything known regarding oligosaccharide composition. In the current study we show for the first time that TNFR1 is ␣2-6 sialylated by ST6Gal-I, and that the degree of TNFR1 ␣2-6 sialylation changes as a consequence of cell differentiation status.…”

Exosomes

“…33 Interestingly, the tumor necrosis factor receptor has also been identified in plasma exosomes. 34 Other molecules important in the cardiovascular system identified in exosomes include proliferator activated receptor-γ, 35 phosphatase and tensin homolog, 36 annexins, 4 dipeptidyl peptidase-4, 4 epidermal growth factor receptor, 37 and a host of metabolic enzymes. 4 The p22 and gp91 subunits of phagocyte-like NADPH oxidase as well as NADPH oxidase activity have been detected in exosomes derived from platelets of septic individuals, although the significance of this is not known.…”