Abstract:
MicroRNAs have shown its role in cell proliferation, apoptosis and pathogenesis of acute lymphoblastic leukemia (ALL). Current diagnosis of ALL comprises invasive and inconvenient procedures including bone marrow biopsy. In spite of recent advancements in the treatment of ALL, still there is risk of relapse. MiR-146a-50 and its target gene PBX2 has been reported to have a potential role in ALL. So the present study was designed to evaluate the expression of miR-146a-5p and PBX2 gene in xenograft rabbit model that can be utilized as circulating markers for early diagnosis of ALL. ALL was induced in rabbit model using xenograft method. Whole blood (5 mL) was collected in EDTA tubes from the ALL-induced rabbits and healthy rabbits after approval from animal ethical committee of The University of Lahore. Plasma was isolated and the total RNA including miRNA was extracted using miRVana isolation kit from Thermofischer Scientific (USA). The concentration and purity of total RNA including miRNA was determined by NanoDrop. Reverse transcription of total RNA including miRNA was performed by using SYBR green kit from Thermofischer Scientific (USA) as per to manufacturer's protocol. The quantitative real-time PCR was performed to check the expression of miR-146a-5p in ALL induced and healthy control rabbit samples. Normalized fold expression was calculated by 2−ΔΔCt method using miR-16 as normalizer. The experiment was performed in duplicate and statistical significance of the results was analyzed using SPSS and Graphpad Prism software. The results showed significant up-regulation of circulating miR-146a-5p and PBX2 gene with a mean fold expression of 102±20.79 and 9.18±0.160 respectively, in ALL induced rabbits as compared to healthy controls (P<0.05). miR-146a-5p and PBX2 gene expression levels may be utilized as circulating markers for early diagnosis of ALL. In future, targeting miR-146a-5p and its target gene PBX2 may be utilized as therapeutic target of ALL
