Journal of Jilin University(Medicine Edition) ›› 2022, Vol. 48 ›› Issue (1): 44-51.doi: 10.13481/j.1671-587X.20220106

• Research in basic medicine • Previous Articles     Next Articles

Expression of miR-150-5p in kidney tissue of diabetic nephropathy model mice and its effect on MPC5 mouse podocyte injury and mechanism

Yu ZHU1,Jingjing WANG2,Fang WU1()   

  1. 1.Department of Traditional Chinese Medicine,Children Hospital,School of Medicine,Zhejiang University,National Clinical Research Center for Child Health,National Children’s Regional Medical Center,Hangzhou 310003,China
    2.Department of Nephrology,Children Hospital,School of Medicine,Zhejiang University,National Clinical Research Center for Child Health,National Children’s Regional Medical Center,Hangzhou 310003,China
  • Received:2021-03-20 Online:2022-01-28 Published:2022-01-17
  • Contact: Fang WU E-mail:6504036@zju.edu.cn

Abstract: Objective

To investigate the expression of miR-150-5p in kidney tissue of the diabetic nephropathy (DN) model mice and the effect of miR-150-5p on the MPC5 mouse podocyte injury induced by high glucose, and to elucidate its mechanism.

Methods

Twenty Kunming mice were randomly divided into control group (n=8, normal feeding) and DN group (n=12, high fat diet feeding combined with intraperitoneal injection of streptozotocin to establish the DN models).The expression levels of miR-150-5p in kidney tissue of the mice in two groups were detected by real-time fluorescence quantitative PCR (RT-qPCR), and the expressions of brain acid soluble protein 1 (BASP1) in kidney tissue of the mice in two groups were observed by immunohistochemical and immunofluorescence methods.The MPC5 cells were cultured in medium without interferon-γ(IFN-γ) for 2 weeks to differentiate and mature, then the cells were divided into control group and high glucose treatment (HG) group. The cells in HG group were treated with 30 mmol·L-1 D-glucose for 12, 24, 48 and 72 h, respectively. The expression levels of miR-150-5p and BASP1 mRNA and protein in the MPC5 cells were detected by RT-qPCR and Western blotting methods, respectively. Bioinformatics and luciferase reporter assays were used to evaluate whether miR-150-5p had a targeting relationship with BASP1.The miR-NC, miR-150-5p mimic, miR-150-5p mimic+pc-DNA3.1-SC and miR-150-5p mimic+pc-DNA3.1-BASP1 were transformed into the mature MPC5 cells, respectively, and they were divided into normal condition+miR-NC (Con+NC) group, HG condition+miR-NC(HG+NC) group, HG condition+miR-150-5p mimic (HG+mimic)group,and HG condition +miR-150-5p mimic+pc-DNA3.1-SC(HG+ mimic+SC) group and HG condition +miR-150-5p mimic+pc-DNA3.1-BASP1 (HG+ mimic+BASP1) group. The viabilities of MPC5 cells in various groups were detected by CCK-8 assay; the reactive oxygen species (ROS) levels and lactate dehydrogenase (LDH) activities of MPC5 cells in various groups were detected by ELISA method, and the apoptotic rates of MPC5 cells in various groups were detected by flow cytometry.

Results

Compared with control group, the expression level of miR-150-5p in kidney tissue of the mice in DN group was decreased (P<0.01), while the expression amount of BASP1 protein was increased, and BASP1 was co-localized with the podocytes. Compared with control group, the expression levels of BASP1 protein in the MPC5 cells in HG group were increased (P<0.01),and the expression levels of miR-150-5p were decreased (P<0.05 or P<0.01) in a time dependent manner. BASP1 was a direct target of miR-150-5p. Compared with Con+NC group, the viability of MPC5 cells in HG+NC group was significantly decreased (P<0.01), while the ROS level, LDH activity and apoptotic rate were significantly increased (P<0.01). Compared with HG+NC group, the viabilities of MPC5 cells in HG+mimic and HG+mimic+SC groups were significantly increased (P<0.01), while the ROS levels, LDH activities and apoptotic rates were significantly decreased (P<0.01).Compared with HG+mimic+SC group, the viability of MPC5 cells in HG+mimic+BASP1 group was significantly decreased (P<0.01), while the ROS level, LDH activity and apoptotic rate were significantly increased (P<0.01).

Conclusion

The decreased expression of miR-150-5p may be one of the mechanisms of podocyte injury in the DN models. Over-expression of miR-150-5p may inhibit the HG-induced podocyte injury by targeting BASP1.

Key words: Diabetic nephropathy, Podocyte injury, MiR-150-5p, Brain acid soluble protein 1, Apoptosis

CLC Number: 

  • R587.1