Abstract

The Role of Terminal Complement Components in Plasma miRNAs Composition

Ionita Ghiran, Assistant Professor of Medicine, Beth Israel Deaconess Medical Center

The complement system is comprised of over 20 soluble and membrane bound proteins with critical roles in recognizing, binding, and removal of foreign particles as well as initiating and regulating innate and acquired immune responses. Activation of the complement system occur during both normal (diurnal) and pathological conditions through any of the classical, alternative, or lectine pathways, that leads to insertion of the membrane attack complex (MAC, C5b-9) pore components in the plasma membrane. The result of MAC-induced exocytosis results in the formation and release in the extracellular space of: a) cytoplasmic components, mostly ions and small molecules, and b) plasma membrane derived vesicles into circulation. This mechanism requires Ca++, calpain activation and local disruption of the actin cytoskeleton. We hypothesized that the size of the C5b-9 complex, about 10 nm, is large enough to allow cellular miRNA species located near the plasma membrane to be released extracellularly. Our results showed that when purified human red cells undergo sub-lytic complement activation, there was an increased in the numbers extracellular red cell-derived vesicles, as well as the in concentration RBC-derived free miRNAs, especially mir451 and mir7b, with no measurable increase in free hemoglobin in the supernatant. Also, proteomic analysis on RBC-derived vesicles showed, in addition to C5b-9, increased amounts of complement regulatory proteins, CD55 and CD59 in their plasma membrane, suggesting that miRNAs contained in the extracellular vesicles could “leak” into the plasma long after their cell release. Our results are consistent with previous reports showing diurnal changes in the number and composition of the plasma extracellular vesicles, as well as the disproportionately large concentration of the miRNAs free in plasma compared to extracellular vesicles.