Abstract
Circulating microRNAs (miRNAs) are useful biomarkers of hemolysis. Since blood cells are the main origins of circulating miRNAs, we evaluated blood cell-related pre-analytical modification of the miRNA signatures during blood drawing and serum processing. The levels of miRNA before and after ex vivo blood drawing were analyzed with the reverse transcriptase-based polymerase chain reaction method. Furthermore, the changes of miRNA signatures caused by different time-lag between blood drawing and serum preparation by 24 h were evaluated. Finally, we compared the miRNA levels between leftover samples and samples of hemolytic diseases. Blood drawing procedure induced increments of red blood cell (RBC)-related miRNAs (miR-451a, miR-486) about 2-fold. One hour standing of blood samples before serum separation induced almost the same increases in RBC-related miRNAs. To test the clinical usefulness of miR-451a as a biomarker of hemolytic diseases, we analyzed miRNAs of samples from 10 normal subjects, 30 leftover samples in the clinical laboratory, and 20 samples from patients with hemolytic diseases. Serum miR-451a significantly increased in patients with hemolytic anemia more than the levels of pre-analytical modification. In conclusion, the pre-analytical modification of serum miRNAs did not disturb the usefulness of RBC-derived miRNAs as biomarkers of hemolytic diseases.
Highlights
We focused on the pre-analytical modification of serum erythroid-specific miRNAs as biomarkers for hemolytic diseases
2.31-fold by 24 h, respectively. miR-16, which is contained in red blood cell (RBC) as well as in other cell lineages, increased 1.54-fold. miR-223, which is mainly contained in granulocytes, increased 1.81-fold at 24 h after sampling. miR-126, which is abundant in platelets, had a significant increment after 6 h (1.91-fold)
We focused on the pre-analytical steps after blood drawing, namely, the effect of mechanical damage of blood drawing on blood cells and the release of miRNAs from blood cells before serum preparation
Summary
Hemolytic diseases are caused by accelerated RBCs destruction classified as intravascular, extravascular hemolysis, and ineffective erythropoiesis [5]. Hemolytic diseases are classified as intravascular hemolysis, extravascular hemolysis, or ineffective erythropoiesis caused by premature decay of erythroblasts in the bone marrow. The main population of erythroid cells in the bone marrow is erythroblasts which are nucleated and proliferate to produce RBCs [5,6,7]. There are few studies comparing the levels of circulating miRNAs between preanalytical modification and pathophysiological state [10,11,12,13,14,15]. We focused on the pre-analytical modification of serum erythroid-specific miRNAs as biomarkers for hemolytic diseases
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