miR-192 enhances sensitivity of methotrexate drug to MG-63 osteosarcoma cancer cells
Mohammadreza Bazavar1, Jafar Fazli1, Amir Valizadeh2
1Department of Orthopedic Surgery, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.
Abstract:
Chemo-resistance remains a considerable obstacle encountered in osteosarcoma (OS) therapy. Evidence has implied that a reduction in the expression of microRNAs (miRs/miRNAs) leads to exacerbated chemo-resistance. Hence, to better understand the role of miR-192 in the pathogenesis of OS during methotrexate (MTX) treatment, we restore miR-192 in the MG-63 cells and investigate the mechanisms, which are associated with MTX-resistance in OS. Exogenetic overexpression of miR-192 was established by transfecting miR-192 mimics into MG-63 cells using Lipofectamine. Trypan blue dye exclusion test was performed to evaluate the proliferation of the MG-63 cells. Chemo-resistance to MTX was determined using the MTT method after 48 h. ELISA cell death assay was performed to evaluate the apoptosis rate. The quantitative RT-PCR (RT-qPCR) was applied to determine the mRNA expression levels before and after the transfection. Our results illustrated that miR-192 is down-regulated in OS tumor cells. Transfection of miR-192 noticeably alleviated the mRNA expression levels of MMP9, c-Myc, K-Ras, CXCR-4, and ADAMTS compared with the control groups (P-values< 0.05). MTX Combination treatment with miR-192 noticeably elevated the cytotoxic effect of MTX and alleviated its IC50 (P < 0.05). Moreover, miR-192 significantly increased the apoptotic effect of MTX. These results implied that miR-192 enhances the sensitivity of MG-63 cells to MTX. Collectively, our results elucidated that miR-192 contributes to chemo-sensitizing MG-63 cells to MTX, and could be considered as a promising agent to overcome MTX-resistance in OS.
Insights
Restoring microRNA-192 (miR-192) in osteosarcoma cells enhances sensitivity to methotrexate chemotherapy. This study shows miR-192 can overcome chemo-resistance in osteosarcoma treatment.
Area of Science:
- Oncology
- Molecular Biology
- Biochemistry
Background:
- Chemo-resistance is a major challenge in osteosarcoma (OS) treatment.
- Reduced microRNA (miRNA) expression is linked to increased chemo-resistance in OS.
- The specific role of miR-192 in methotrexate (MTX) resistance in OS requires further investigation.
Purpose of the Study:
- To investigate the role of miR-192 in methotrexate resistance in osteosarcoma (OS).
- To determine if restoring miR-192 expression can overcome MTX resistance in MG-63 OS cells.
Main Methods:
- Overexpression of miR-192 in MG-63 cells using miR-192 mimics and Lipofectamine.
- Assessing cell proliferation via trypan blue dye exclusion.
- Evaluating chemo-resistance to MTX using MTT assay and determining IC50 values.
- Measuring apoptosis rates using ELISA cell death assay.
- Quantifying mRNA expression levels of target genes (MMP9, c-Myc, K-Ras, CXCR-4, ADAMTS) using RT-qPCR.
Main Results:
- miR-192 was found to be downregulated in OS tumor cells.
- Restoring miR-192 significantly reduced the mRNA expression of MMP9, c-Myc, K-Ras, CXCR-4, and ADAMTS.
- Combined treatment with MTX and miR-192 overexpression enhanced MTX's cytotoxic effect and lowered its IC50.
- miR-192 significantly increased MTX-induced apoptosis in MG-63 cells.
Conclusions:
- miR-192 enhances the sensitivity of osteosarcoma cells to methotrexate (MTX).
- Restoring miR-192 expression can overcome MTX resistance in OS.
- miR-192 represents a potential therapeutic agent for improving MTX efficacy in osteosarcoma treatment.
