Vesicle-bound and free microRNAs in spent cell culture medium and biological fluids

Abstract

Background: In addition to the control of gene transla- tion intra-cellularly, microRNAs (miRNAs) have been found to exist extracellularly. However, extracellular miRNA identified in previous studies were largely confined to microvesicles. It remains uncertain whether free extracellular miRNA exists. Methods: We quantify a panel of miRNAs (miRNA 200 family, miR-205, and miR-192) in spent culture medium of renal tubular epithelial cells, as well as serum and urine from patients with systemic lupus erythematosus and healthy controls. Microvesicle bound and free miRNA were separated by ultracentrifugation. Results: In spent cell culture medium, we found substantial amount of miR-200a, miR-200b, miR-200c, miR-141, miR-429, miR-205, and miR-192 in microvesicles, as well as the presence of miR-200a, miR-200b, miR-429, miR-205 and miR-192, but not miR-200c or miR-141 as non-vesicle bound free form. In healthy individuals, we found substantial amount of miR-200a, miR-200b, miR-200c, miR-141 and miR-192 in microvesicles from serum and urine, while only miR-141 exists as free form in both serum and urine. There was no significant difference in serum or urinary free miR-141 levels between SLE patients and healthy controls. In SLE patients, urinary free miR-141 level was signifiantly higher than serum. Neither serum nor urinary free miR-141 levels correlated with lupus disease activity. Conclusion: We found miRNA is present extracellularly, both within microvesicles and as free form, in spent cell culture medium, serum and urine. The biological role of extracellular miRNA requires further study.