Effects of pathogen reduction systems on platelet microRNAs, mRNAs, activation, and function

Figures & data

Figure 1. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on platelet microRNA levels. The figure shows the median fold change for each individual platelet microRNA level, in each of the four treatment groups (additive solution, Irradiation, mirasol, or intercept) and at each time point (i.e. day 1, 4, or 7 of storage after treatment), compared with its level in the control group at the same time point, which is set at 1.0 (—) (n = 10 donors per group). All comparisons were two-sided. *p < 0.05, namely a statistically significant reduction in the level of the specific microRNA for the corresponding treatment group versus the control group at the particular time point and following correction for multiple comparisons.

Figure 2. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on platelet apoptosis-related mRNAs. The figure shows the absolute number of copies of Bcl-xl (A), CLU (B), and BCL2 (C) mRNAs in platelets isolated from samples collected from PCs at 1.5 and 20 hours after treatment with additive solution, Irradiation, mirasol, intercept, or no treatment (control). For each group and time point, the figure shows the individual data (n = 4–6 donors per group), along with their median. All comparisons were two-sided. *p < 0.05, namely a statistically significant decrease in the number of mRNA copies at 1.5 or 20 hours for the corresponding group, compared with the control group at the same time point).

Figure 3. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on platelet microRNA synthesis. The figure shows the percentage of pre-microRNA substrate cleaved into mature microRNA by platelet Dicer, which is known to mediate platelet microRNA biogenesis [11]. Dicer activity assays were performed by coincubating a ³²P-labeled pre-microRNA substrate (³²P-labeled pre-let-7a-3) with protein extracts from platelets collected from PCs at 1, 4, and 7 days of storage after treatment with additive solution, Irradiation, mirasol, intercept, or no treatment (control). For each group and time point, the figure shows the individual data (n = 8 donors per group), along with their median.

Figure 4. Storage in additive solution or treatment with Irradiation, mirasol, or intercept does not induce formation of cross-linked adducts of endogenous platelet RNAs. The figure shows the lack of cross-linked adducts of endogenous platelet RNAs that can be attributed to treatment with additive solution, Irradiation, mirasol or intercept, as compared with no treatment (control), at any time point (day 1, 4, or 7 of storage) by total RNA extraction, dephosphorylation, ³²P labeling, and analysis by denaturing PAGE. Compare with Supplementary Figure S4, which shows the results without dephosphorylation.

Figure 5. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on the ability of platelets to mediate microRNA function. The figure shows the percentage of miR-223 sensor RNA substrate cleaved by platelet Ago2•miR-223 complexes, which is known to mediate platelet microRNA function [11]. RISC activity assays were performed by coincubating a ³²P-labeled RNA sensor perfectly complementary to miR-223 with protein extracts from platelets collected from PCs at 1, 4, and 7 days of storage after treatment with additive solution, Irradiation, mirasol, intercept, or no treatment (control). For each group and time point, the figure shows the individual data (n = 4 donors per group), along with their median.

Figure 6. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on the activation level of platelets. The figure shows the percentage of platelets expressing CD62P on their surface by flow cytometry analysis of samples collected from the PCs on day 1, 4, or 7 for each group (storage in additive solution, treatment with Irradiation, mirasol or intercept, or control). For each group, the figure shows the individual data (n = 10 donors per group) obtained at each time point, along with the median. All comparisons were two-sided. *p < 0.05, namely a statistically significant decrease in the corresponding treatment (compared with the control) group on the same day of storage.

Figure 7. Effect of storage in additive solution or treatment with Irradiation, mirasol, or intercept on platelet aggregation. The figure shows the aggregation response to 20 µM ADP (A), 40 µM TRAP (B) and 4 µg/ml collagen (C) of samples collected from the PCs on day 1, 4, or 7 for each group (control, additive solution, Irradiation, mirasol, or intercept). Individual results from each PC, expressed as a percentage of maximal transmission (T_max), are shown along with the median (n = 10 donors per group). All comparisons were two-sided. *p < 0.05, namely a statistically significant difference in aggregation in the corresponding treatment (compared with the control) group on the same day of storage.

Landry P, Plante I, Ouellet DL, Perron MP, Rousseau G, Provost P. Existence of a microRNA pathway in anucleate platelets. Nat Struct Mol Biol 2009;16(9):961–966

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