Nucleic acids within urinary exosomes/microvesicles are potential biomarkers for renal disease.
Urinary exosomes or microvesicles are being studied intensively to identify potential new biomarkers for renal disease. We sought to identify whether these microvesicles contain nucleic acids. We isolated microvesicles from human urine in the same density range as that previously described for urinary exosomes and found them to have an RNA integrity profile similar to that of kidney tissue, including 18S and 28S rRNA. This profile was better preserved in urinary microvesicles compared with whole cells isolated from urine, suggesting that microvesicles may protect RNA during urine passage. We were able to detect mRNA in the human urinary microvesicles encoding proteins from all regions of the nephron and the collecting duct. Further, to provide a proof of principle, we found that microvesicles isolated from the urine of the V-ATPase B1 subunit knockout mice lacked mRNA of this subunit while containing a normal amount of the B2 subunit and aquaporin 2. The microvesicles were found to be contaminated with extraneous DNA potentially on their surface; therefore, we developed a rapid and reliable means to isolate nucleic acids from within urine microvesicles devoid of this extraneous contamination. Our study provides an experimental strategy for the routine isolation and use of urinary microvesicles as a novel and non-invasive source of nucleic acids to further renal disease biomarker discovery.
Miranda
Kevin C
KC
Division of Nephrology, Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA.
Bond
Daniel T
DT
McKee
Mary
M
Skog
Johan
J
Păunescu
Teodor G
TG
Da Silva
Nicolas
N
Brown
Dennis
D
Russo
Leileata M
LM
D000818:Animals; D015415:Biomarkers / Q000032:analysis*; D055354:Exosomes / Q000737:chemistry*; D006801:Humans; D007674:Kidney Diseases / Q000175:diagnosis* / Q000652:urine*; D051379:Mice; D046529:Microscopy, Electron, Transmission; D009696:Nucleic Acids / Q000032:analysis*; D051381:Rats
D015415:Biomarkers; D009696:Nucleic Acids