METTL14 Promotes HK-2 Cell Injury in Diabetic Nephropathy Through the Regulation of THBS2 Expression in an m6A-YTHDF2-Dependent Manner

Abstract

Objective

To explore how methyltransferase like 14 (METTL14) regulates diabetic nephropathy (DN)-induced cell damage and analyze its underlying mechanisms.

Methods

The gene expression data were obtained from the Gene Expression Omnibus (GEO) database and analyzed for differential expression using the DESeq2 package. The gene expression levels were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting. Cell proliferation and apoptosis were examined through cell counting kit-8 (CCK8) and flow cytometry, respectively. The levels of inflammatory cytokines (interleukin-6 [IL-6], IL-8, and tumor necrosis factor-alpha (TNF-α)) were determined through enzyme-linked immunosorbent assay (ELISA). Levels of ferroptosis indicators, including reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), and ferrous iron (Fe²⁺), were measured using dedicated kits. The m6A modification of mRNA was predicted by the Sequence-based RNA Adenosine Methylation Site Predictor (SRAMP) database and was validated using RNA immunoprecipitation (RIP) assay. The m6A RIP-qPCR was performed to verify the interaction of thrombospondin-2 (THBS2) with METTL14 or YTHN6-methyladenosine RNA binding protein 2 (YTHDF2).

Results

THBS2 was highly expressed in both DN patients and high glucose (HG)-induced HK-2 cells. THBS2 silencing promoted HG-induced HK-2 cell proliferation and decreased apoptosis, fibrosis, inflammation, and ferroptosis. METTL14 stabilized THBS2 in an m6A-YTHDF2-mediated manner. Suppression of METTL14 expression could alleviate the HG-induced damage to HK-2 cells. THBS2 overexpression negated the protective effects of METTL14 knockdown in HK-2 cells.Conclusions: METTL14 exacerbates HG-induced damage in HK-2 cells by modulating THBS2 expression in an m6A-YTHDF2-dependent manner.

Keywords

METTL14 - YTHDF2 - THBS2 - diabetic nephropathy - HG-induced cell damage

Publikationsverlauf

Eingereicht: 22. Januar 2025

Angenommen nach Revision: 23. Mai 2025

Artikel online veröffentlicht:
29. Juli 2025

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