Haniyeh Fooladinezhad1, Hossein Khanahmad2, Mazdak Ganjalikhani-Hakemi3, Abbas Doosti1. 1. a Department of Genetics, Faculty of Sciences, Shahrekord Branch , Islamic Azad University , Shahrekord , Iran. 2. b Department of Genetics, Faculty of Medicine , Isfahan University of Medical Sciences , Isfahan , Iran. 3. c Immunology Department, Faculty of Medicine , Isfahan University of Medical Sciences , Isfahan , Iran .
Abstract
BACKGROUND: Uncontrolled proliferation and accumulation of leukaemic stem cells (LSCs) in bone marrow leads to acute myeloma leukaemia (AML). T cell immunoglobulin and mucine domain (TIM)-3 is a specific surface marker for LSCs and is highly expressed on LSCs compared with normal bone marrow cells, haematopoietic stem cells. Studies have indicated that microRNAs can affect AML progression through targeting different genes expressions like TIM-3. So, based on bioinformatics assessments, we predicted that miR-330-5p may highly inhibit TIM-3 expression. The purpose of the present study was to prove the silencing effect of miR-330-5p on TIM-3 gene expression in AML cell line (HL-60) in vitro. METHODS: HL-60 cells were cultured in RPMI 1640 supplied with 10% FBS. TIM-3 expression was induced in the cells using phorbol myristate acetate (PMA). The cells were transfected with miR-330-5p and then, the gene and protein expression of TIM-3 were measured using q-RT-PCR and flow-cytometry methods, respectively. RESULTS: The results of our bioinformatics surveys revealed that miR-330-5p has high predicted ability to silence TIM-3 gene expression. Accordingly, our experiments confirmed that miR-330-5p is able to strongly silence TIM-3 expression (98.15% silencing) in HL-60 cell line (p = 0.0001). CONCLUSION: According to our results, miR-330-5p has a strong inhibitory effect on TIM-3 expression in AML cell line. Thus, the bioinformatics prediction of Mirwalk and Target Scan softwares for silencing effect of miR-330-5p on TIM-3 is confirmed.
BACKGROUND: Uncontrolled proliferation and accumulation of leukaemic stem cells (LSCs) in bone marrow leads to acute myeloma leukaemia (AML). T cell immunoglobulin and mucine domain (TIM)-3 is a specific surface marker for LSCs and is highly expressed on LSCs compared with normal bone marrow cells, haematopoietic stem cells. Studies have indicated that microRNAs can affect AML progression through targeting different genes expressions like TIM-3. So, based on bioinformatics assessments, we predicted that miR-330-5p may highly inhibit TIM-3 expression. The purpose of the present study was to prove the silencing effect of miR-330-5p on TIM-3 gene expression in AML cell line (HL-60) in vitro. METHODS: HL-60 cells were cultured in RPMI 1640 supplied with 10% FBS. TIM-3 expression was induced in the cells using phorbol myristate acetate (PMA). The cells were transfected with miR-330-5p and then, the gene and protein expression of TIM-3 were measured using q-RT-PCR and flow-cytometry methods, respectively. RESULTS: The results of our bioinformatics surveys revealed that miR-330-5p has high predicted ability to silence TIM-3 gene expression. Accordingly, our experiments confirmed that miR-330-5p is able to strongly silence TIM-3 expression (98.15% silencing) in HL-60 cell line (p = 0.0001). CONCLUSION: According to our results, miR-330-5p has a strong inhibitory effect on TIM-3 expression in AML cell line. Thus, the bioinformatics prediction of Mirwalk and Target Scan softwares for silencing effect of miR-330-5p on TIM-3 is confirmed.
Entities:
Keywords:
AML; HL-60; TIM-3; miR-330-5p; molecular medicine
Authors: Hamid Bolouri; Jason E Farrar; Timothy Triche; Rhonda E Ries; Emilia L Lim; Todd A Alonzo; Yussanne Ma; Richard Moore; Andrew J Mungall; Marco A Marra; Jinghui Zhang; Xiaotu Ma; Yu Liu; Yanling Liu; Jaime M Guidry Auvil; Tanja M Davidsen; Patee Gesuwan; Leandro C Hermida; Bodour Salhia; Stephen Capone; Giridharan Ramsingh; Christian Michel Zwaan; Sanne Noort; Stephen R Piccolo; E Anders Kolb; Alan S Gamis; Malcolm A Smith; Daniela S Gerhard; Soheil Meshinchi Journal: Nat Med Date: 2017-12-11 Impact factor: 53.440