BACKGROUND: miRNAs are small RNA molecules involved in post-transcription mRNA regulation. A single miRNA may target whole functional pathways. miRNA's have been implicated to play a significant role in immunological, and injury repair mechanisms. The aim of this study was to explore the function of miRNA signaling and impact of their targets in acute rejection. METHODS: The kidney transplant tissue biopsies (TxBx, n=88) were extracted for RNA (Trizol). Samples were chosen to represent the distinct pathology of acute rejection (AR, n=28) or normal (NORM, n=60). miRNAs were selected from the literature (controls) or as predicted targets to acute rejection genes. 25ng of total-RNA was reverse transcribed, followed by target specific pre-amplification using Taqman MegaPlex Pools (ABI). 5ng of pre-amp sample was run on 96.96 microfluidic arrays (Fluidigm), using TaqMan miRNA assays and universal master mix (ABI). Delta-delta Ct method was used with miR191 (internal reference) and universal human total RNA (Ambion, external reference). Students T-test used for significance analysis and Spearmans rank analysis for correlations. Bioinformatics databases were used (mirTarBase, GeneCards, SOURCE, DAVID), with Affy Hu133+2.0 microarrays (mRNA) of matched TxBx used for target expression (n=97). RESULTS: We observed high expression of a number of miRNA species. QPCR identified 7 differentially expressed miRNAs in AR compared to NORM. Specific miRNAs and not whole families were differentially altered. A number of miRNAs correlated with allograft pathology of Banff i, t and ta, indicating that the majority of these miRNA are a consequence of lymphocyte signaling on the renal cells. mRNA targets of the 7 miRNAs were defined using mirTarBase (n=46 targets), and were investigated for altered expression on microarrays of matched TxBx. 11/46 mRNA targets were significantly expressed in the corresponding inverse direction. Gene annotation of altered targets indicated immune signaling, EMT and apoptosis. CONCLUSION: We have identified a number of important miRNA's that are strongly associated with AR on TxBx. These miRNA specific target a number mRNA molecules that appear to play a role in either acute rejection or injury repair mechanisms. We have identified a potential source of infiltrating lymphocyte signaling and renal cell response to injury during the acute rejection.
To cite this abstract in AMA style:Vitalone M, Sigdel T, Khatri P, Hsieh S, Butte A, Sarwal M. Specific miRNA’s and Their Downstream Targets Are Associated with Acute Rejection in Pediatric Renal Allografts [abstract]. Am J Transplant. 2013; 13 (suppl 5). https://atcmeetingabstracts.com/abstract/specific-mirnas-and-their-downstream-targets-are-associated-with-acute-rejection-in-pediatric-renal-allografts/. Accessed November 23, 2020.
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