PURPOSE: Liver specific miR-122 has been shown to be potential biomarker for liver injury; however initial production of miR-122 during allograft reperfusion has not been characterized. The aim of this study is to evaluate the effect of ischemic-reperfusion on miR-122 production and whether miR-122 production predicts subsequent allograft injury. METHODS: Serum was sampled immediately prior to reperfusion (time=0) and then every 15-minutes for 90-minutes after reperfusion from 13 patients undergoing liver transplantation. Total micro-RNA was extracted from serum with a micro-RNA specific protocol. Total micro-RNA levels were determined in three patients by micro-fluidics using Small RNA Chips (Agilent Technologies). miR-122 levels were determined in 13 patients by quantitative real time RT-PCR. Copy numbers for miR-122 were obtained from a normalized standard curve using a synthetic miR-122 mimic. Normalization was done with internally spiked C. elegans micro RNA-39. Degree of ischemia-reperfusion injury was measured by determining the peak serum alanine aminotransferase (ALT) level during the first week following liver transplantation. RESULTS: Initial miR-122 levels (at time=0) were lower for Hepatitis C (HCV)-negative patients (n=3) compared to HCV-positive patients (n=9) (5.8±8.4×1010 copies vs. 9.3±2.8×109 copies). miR-122 levels increased initially, peaked 30-60 minutes after reperfusion and then decreased to levels still higher than time=0, except for the one patient with primary non-function (PNF) where miR-122 increased continuously. A significant correlation was observed between initial miR-122 production (90-minute area under the curve (AUC)) and peak post-operative serum ALT (r= 0.94; p=0.0009) for the HCV-positive patients (n=9). Conversely, a significant negative correlation between initial miR-122 production and peak post-operative ALT (r= -0.93; p=0.03) was observed for the HCV negative and PNF patients (n=4). Other micro-RNAs were likely elaborated during this period, as total serum micro-RNA levels did not correlate with miR-122 levels (r= 0.11-0.51; p>0.05). During the analysis of total micro-RNA, a small RNA (approximately 45 nucleotides) was found to increase at all time points from initially undetectable levels and significant negative correlation was found with HCV levels (r: -0.94 – -0.98; p<0.05). CONCLUSION: miR-122 production during initial reperfusion of liver allografts accurately predicts subsequent ischemia-reperfusion injury. The dynamics of miR-122 production differ for HCV infected and un-infected patients and for patients with primary non-function. Allografts also release a significant amount of other micro-RNAs and an unidentified small RNA during reperfusion.

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