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Long Non-coding RNA ZFAS1 Regulates Fibrosis and Scortosis in the Cell Model of Diabetic Nephropathy Through miR-525-5p/SGK1 Axis

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Abstract

Diabetic nephropathy (DN) is a common clinical syndrome in diabetic patients. Functional characterization of non-coding (ncRNAs) involved in the progression of DN can provide insights into the diagnosis and therapeutic management of DN. Human kidney proximal tubular epithelial cells (HK-2) were challenged by high glucose (HG, 50 mM) as a cell model of DN. The expression level of long non-coding RNA (lncRNA) ZFAS1 was quantified by qRT-PCR. The proteins and cytokines related to fibrosis and scortosis in DN (NLRP3, GSDMD-N, IL-1β and Caspase 1, fibronectin, collagen I, collagen III, IL-1β, and IL-18) were examined by western blot or ELISA. RNA precipitation and luciferase reporter activity experiments were conducted to assess the molecular associations. ZFAS1 and SGK1 were highly induced in HK-2 cells challenged with HG, while miR-525-5p downregulated upon HG treatment. ZFAS1 knockdown attenuated HG-induced fibrosis and scortosis in HK-2 cells by reducing the levels of NLRP3, GSDMD-N, Caspase 1, fibronectin, collagen I/III, IL-1β, and IL-18. Mechanically, ZFAS1 knockdown protected HK-2 cells from HG-induced injury by upregulating miR-525-5p and repressing SGK1 expression. Overall, our results suggest that knocking down ZFAS1 may be formulated as a protective strategy in ameliorating DN progression through regulating miR-525-5p/SGK1 pathway. Targeting ZFAS1 could be further explored as a potential approach for the management of DN.

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Data Availability

The experimental data present in the manuscript can be obtained from the corresponding author upon reasonable request.

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Funding

Specialization in Translational Medicine: Translational Medicine Program of Bengbu Medical College (NO. BYTM2019032). Natural Science Research Project of Anhui Educational Committee 2022AH051468；Scientific Research project of Anhui Provincial Health Commission AHWJ2022b056.

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Authors and Affiliations

  1. Department of Endocrinology, The First Affiliated Hospital of Bengbu Medical College, No. 287, Changhuai Road, Bengbu, 233004, Anhui, China

    Langen Zhuang, Guoxi Jin, Qiong Wang & Xiaoyan Pei

  2. Department of Endocrinology Tongren Hospital Affiliated to Jiaotong University, No. 1111, Xianxia Road, Changning District, Shanghai, 200336, China

    Xiaoxu Ge

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Contributions

Langen Zhuang and Guoxi Jin conceived and designed the experiments; Langen Zhuang, Guoxi Jin, and Qiong Wang conducted the experiments and prepared the manuscript; Langen Zhuang, Xiaoxu Ge, and Xiaoyan Pei analyzed the data. All authors proofread the final draft and agreed with the submission.

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Correspondence to Langen Zhuang.

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Fig. S1

Screening of potential downstream targets of ZFAS1 and miR-525-5p. (A) Starbase prediction of potential binding miRNAs of ZFAS1. (B) RNA-pull down analysis of the interacting miRNAs using biotin-ZFAS1 probe or control oligo in HK-2 cells. Data were normalized to the input sample. (C) Starbase prediction results of mRNA targets of miR-525-5p. (D) RNA-pull down assay of the interacting mRNA targets using biotin-miR-525-5p probe or control oligo in HK-2 cells. Data were normalized to the input sample. *** stands for P < 0.001. (PNG 1523 kb)

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Zhuang, L., Jin, G., Wang, Q. et al. Long Non-coding RNA ZFAS1 Regulates Fibrosis and Scortosis in the Cell Model of Diabetic Nephropathy Through miR-525-5p/SGK1 Axis. Appl Biochem Biotechnol (2023). https://doi.org/10.1007/s12010-023-04721-5

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