Basic & Clinical Medicine ›› 2019, Vol. 39 ›› Issue (10): 1404-1409.

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miR-124 targets magnesium transporter 1 to regulate the function exhaustion of activated human T cells

  

  • Received:2019-01-14 Revised:2019-07-02 Online:2019-10-05 Published:2019-09-25

Abstract: Objective To investigate the regulation of miR-124 targeting magnesium transporter 1 on activated function exhaustion of activated T cell. Methods 1) Isolated monocytes from peripheral blood,then treated with IFN-γ, IL-2, anti-CD3 antibody , anti-CD28 antibody and IL-1α to activate T cell. The proportion of CD25 and CD69 subgroups was detected by Flow cytometry. 2) miR -124 / miR-124 sponge lentiviral vector was constructed. After lentivirus infected activated T cell, the expression of miR-124 and MagT1 gene in T cells was detected by RT-qPCR. 3) The targeted sites of miR-124 and MagT1 were analyzed by bioinformatics, and identified by dual luciferase reporter gene system.4) The expression of MagT1 protein and PD-1 protein were detected by Western blot. 5) The proliferation and secretion of TNF-αand TGF-β was detected by CCK8 and ELISA.Results Flow cytometry showed that T cells were activated.RT-qPCR indicated that the construction of lentivirus was successful, and miR-124/miR-124 sponge respectively up-regulated or down-regulated the expression of miR-124 (P<0.01).The dual luciferase reporter system confirmed that miR-124 targeted MagT1 3’UTR and inhibited its expression.Western blot showed that the MagT1 protein level in the miR-124 over-expression group was lower than that in the control group (P<0.05), and the pd-1 protein level was higher than that in the control group (P<0.05). After the inhibition of miR-124, the MagT1 protein level was higher than that in the control group (P<0.05), and the pd-1 protein level was lower than that in the control group (P<0.05). Over-expression of miR-124 significantly decreased the proliferation and secretion of TNF-α of T cells, while inhibiting of miR-124 expression significantly increased the proliferation and secretion of TGF-βof T cells (P<0.01).Conclusion It was confirmed that miR-124 could negatively regulate the expression of the targeted gene MagT1 and play an important regulatory role in the function exhaustion of activated T cells.

Key words: miR-124, magnesium transporter 1, T cell exhaustion