OBJECTIVES: The liver contains large amounts of microRNA-122 (miR-122), whereas other tissues contain only marginal amounts of this miRNA. MicroRNAs have also been found to circulate in the blood in a cell free form; their potential as readily accessible disease markers is currently evaluated. Here we investigated if the serum levels of miR-122 might be useful as disease parameter in patients with chronic hepatitis C virus (HCV) infection.METHODS: RNA was extracted from sera of patients with chronic HCV infection and healthy controls and was analyzed for miR-122 content by quantitative real-time reversetranscription PCR and for standard parameters of liver function. Liver biopsies from the same patients were examined for the histologic activity index (HAI) and the degree of fibrosis.RESULTS: Sera from patients with chronic HCV infection contained higher levels of miR-122 than sera from healthy controls. Serum miR-122 levels correlated well with markers of liver inflammatory activity, i. e. serum levels of alanine leucine transaminase (ALT) and aspartate transaminase, and the HAI score. In patients with persistently normal ALT levels, serum miR-122 levels did not differ from healthy controls. There was no correlation of serum miR-122 levels with serum albumin, international normalized ratio, liver fibrosis or serum HCV RNA.CONCLUSIONS: The serum level of miR-122 strongly correlates with serum ALT activity and with necroinflammatory activity in patients with chronic HCV infection and elevated ALT levels, but not with fibrosis stage and functional capacity of the liver.

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BackgroundMicroRNAs circulating in the blood, stabilized by complexation with proteins and/or additionally by encapsulation in lipid vesicles, are currently being evaluated as biomarkers. The consequences of their differential association with lipids/vesicles for their stability and use as biomarkers are largely unexplored and are subject of the present study.MethodsThe levels of a set of selected microRNAs were determined by quantitative reverse-transcription PCR after extraction from sera or vesicle- and non-vesicle fractions prepared from sera. The stability of these microRNAs after incubation with RNase A or RNase inhibitor, an inhibitor of RNase A family enzymes was studied.ResultsThe levels of microRNA-1 and microRNA-122, but not those of microRNA-16, microRNA-21 and microRNA-142-3p, declined significantly during a 5-h incubation of the sera. RNase inhibitor prevented the loss of microRNAs in serum as well as the degradation of microRNA-122, a microRNA not expressed in blood cells, in whole blood. Stabilization of microRNA-122 was also achieved by hemolysis. Prolonged incubation of the sera led to enrichment of vesicle-associated relative to non-vesicle-associated microRNAs. Vesicle-associated microRNAs were more resistant to RNase A treatment than the respective microRNAs not associated with vesicles.ConclusionsSerum microRNAs showed differential stability upon prolonged incubation. RNase inhibitor might be useful to robustly preserve the pattern of cell-free circulating microRNAs. In the case of microRNAs not expressed in blood cells this can also be achieved by hemolysis. Vesicle-associated microRNAs appeared to be more stable than those not associated with vesicles, which might be useful to disclose additional biomarker properties of miRNAs.