miRNAs in Blood Microvesicles Figure 3. miRNA expression from peripheral blood microvesicles and PBMC. Hierarchical cluster analysis for microvesicles and PBMC is shown based on filtering criteria. The (*) denotes the three PBMC samples which clustered with the plasma microvesicle samples. doi:10.1371/journal.pone.0003694.g003 -19b and -20a were among the highest miRNAs detected in PBMC, but not in plasma microvesicles. In contrast, the PBMC samples, shared 6 common miRNAs with plasma microvesicles (miR-223, -484, -191, -146a, -16, and -26a). PBMCs also lacked high expression of miR-222, -24, -126, and -32 in the top expressed miRNAs compared to those found in plasma microvesicles. Based on ranking analysis, miR-223 was the most highly expressed miRNA in both PBMC and plasma microvesicles. We next hypothesized that miRNAs originating from specific organ sites would be detected in the PBMC or plasma microvesicles. Lee et al reported that miR-122a is specifically expressed in liver tissue while miR-216 and miR-217 are preferentially expressed in the pancreas [38]. Interestingly these miRNAs were not identified in the plasma microvesicles (Table S1). We next examined the predicted function of the most highly expressed miRNAs in the plasma microvesicles and PBMC. We examined the top nine expressed miRNAs from the plasma microvesicles and PBMC samples. Using the Sanger miRBase Target v5 and TargetScan v4.2, we found 1578 and 1899 predicted targets, respectively, for the group of highly expressed miRNAs from plasma microvesicles (Table S2). Merging targets from the two databases, we found 94 common predicted targets. These combined targets were subjected to computational analysis to predict pathways that they would collectively regulate. Using Ingenuity Pathway Analysis (IPA) software, we found canonical pathways involved in metabolism, estrogen receptor signaling, and degradation of phospholipids that were predicted to be highly regulated by these miRNAs (Table 2). Of the nine miRNAs examined from the PBMC fraction, we found 1857 and 2072 predicted mRNA targets from Sanger miRBase and TargetScan database, respectively (Table S2) and 89 common targets (Table S2). The top five canonical pathways expected to be regulated by PLoS ONE | www.plosone.org 5 November 2008 | Volume 3 | Issue 11 | e3694