(P-CB-12) Impact of Leukoreduction on Dysregulation of microRNAs regulating the Genes involved in Apoptosis, Senescence, and Signaling Pathways during Red Cell Storage.

Background/Case Studies: Red Blood cells (RBCs) bring about harmful consequences during storage. MicroRNA (miRNA) dysregulation in stored RBCs could represent potential biomarkers of storage lesions. Although leukoreduction prevents damage to RBCs, it is uncertain whether leukoreduction of RBCs would impact the dysregulation of miRNAs during storage. This study evaluated the potential role of miRNAs for any alteration of leukoreduced (LD) and non-leukoreduced (NLD) RBCs till 21 days of storage. This study further explored the regulation between miRNA and mRNA involved in apoptosis, senescence, and RBC-related signaling pathways.

Study

Design/Methods: The study was a prospective observational study. 30 healthy adult volunteers’ blood was equally divided into leukoreduced RBCs (LD) and non-leukoreduced RBC (NLD) bags and stored till day 21 at 4-6 degree Celsius. RBCs were evaluated to estimate selected microRNAs (miRNAs) and potential target messenger RNAs (mRNAs) on day 0 and day 21 of storage. Comparison of medians values of fold changes between the groups (LD Vs. NLD) was performed using Mann Whitney U test and within the group by two-sided Wilcoxon signed rank test. A p-value < 0.05 was considered to be significant. Further, a bioinformatics tool was used to analyze the selected miRNAs and their predicted target genes (mRNAs) and identify the miRNA-mRNA regulatory relationships.

Results/Findings: A significantly higher fold change values of three miRNAs (miR-96-5p, miR-197-3p, miR-769-3p) were observed in NLD RBCs (p < 0.05). (Table 1) A significantly higher (p < 0.05) expression levels of miR-150-5p and miR-197-3p were observed in NLD RBCs till 21 days of storage. Further, In-Silico analysis revealed the genes involved in the regulatory pathway of RBCs. Correlation with mRNA quantification confirmed the regulatory role of these miRNAs upon functional pathway enrichment analysis.

Conclusions: This study highlighted the higher level of dysregulation of miRNAs in NLD RBCs. Further validation from In-Silico analysis suggested the regulatory role of miRNAs in cell apoptosis, cell senescence, and RBC-related signaling pathways, which indicates that stored LD RBCs will likely have better in vivo survival and function following transfusion. However, a study on the miRNAs in in-vivo RBCs following transfusion will be highly desired to reach conclusive evidence.

Importance of research: Although leukoreduction prevents damage to RBCs, improve recovery, and reduce hemolysis of RBCs, it is uncertain whether leukoreduction of RBCs would impact the dysregulation of miRNAs during RBC storage. Hence, this study was done to unravel the function of miRNAs in stored RBCs and look for any association with the leukoreduction. Bioinformatics analysis was performed to identify the individual miRNAs targeting the mRNA genes involved in apoptosis, senescence, and Red Cell signaling pathway.