# Diabetic Kidney Disease Progression Alleviated in Mice by ALKBH5‐Mediated UC‐MSCs‐Derived Exosomes That Inhibit TRAF6 m6A Modification and Promote M2 Macrophage Polarisation

## TL;DR

Researchers found that exosomes from modified stem cells can reduce kidney damage in diabetic mice by changing immune cell behavior.

## Contribution

This study shows ALKBH5-modified UC-MSC-derived exosomes alleviate DKD by inhibiting TRAF6 m6A modification and promoting M2 macrophage polarization.

## Key findings

- ALKBH5 overexpression in UC-MSC exosomes promotes M2 macrophage polarization and suppresses M1 phenotype in vitro.
- In diabetic mice, ALKBH5-modified exosomes reduced kidney injury and inflammation by inhibiting TRAF6 m6A methylation.
- These exosomes decreased TRAF6 protein levels and stability, leading to anti-inflammatory M2 macrophage polarization.

## Abstract

Diabetic kidney disease (DKD) is a major diabetes complication with limited treatment options. Exosomes (Exo) from umbilical cord mesenchymal stem cells (UC‐MSCs) have shown therapeutic promise. The role of alkylation repair homologue protein 5 (ALKBH5)‐modified UC‐MSCs Exo in regulating macrophage polarisation and alleviating DKD is investigated.

DKD‐associated inflammation was modelled by Lipopolysaccharide (LPS)/interferon‐gamma (IFN‐γ)‐stimulated RAW264.7 macrophages. RT‐qPCR and western blotting were employed to analyse mRNA and protein expression. Exosomes from ALKBH5‐modified UC‐MSCs were isolated and characterised. Macrophage polarisation (M1/M2) was assessed by flow cytometry, RT‐qPCR, and enzyme‐linked immunosorbent assay (ELISA). Tumor necrosis factor receptor‐associated factor 6 (TRAF6) N6‐methyladenosine (m6A) modification and expression were analysed via methylated RNA immunoprecipitation (MeRIP) and RNA immunoprecipitation (RIP) assays. The DKD model was established using spontaneous diabetic db/db mice. The renal function of mice was evaluated by ELISA and commercial assay kits. Hematoxylin–eosin (HE), periodic acid‐Schiff (PAS), and Masson's trichrome staining were performed to assess the renal histopathology of mice.

ALKBH5 overexpression promoted M2 and inhibited M1 macrophage polarisation. ALKBH5 downregulated TRAF6 via m6A demethylation. ALKBH5‐modified UC‐MSCs Exo enhanced M2 polarisation and suppressed M1 phenotype in vitro. In DKD mice, ALKBH5‐modified UC‐MSCs Exo mitigated renal injury. Moreover, these exosomes enhanced anti‐inflammatory responses and promoted M2 macrophage polarisation in DKD mice.

ALKBH5‐modified UC‐MSCs Exo reduced TRAF6 expression by demethylating its m6A sites, promoting M2 macrophage polarisation and alleviating DKD progression. These findings suggested that ALKBH5‐modified UC‐MSCs Exo might represent a promising therapeutic approach for DKD.

ALKBH5‐modified UC‐MSCs‐derived exosomes (OE‐ALKBH5 Exo) inhibit the expression of M1 markers (iNOS, IL‐1β, TNF‐α, CD86) and enhance the expression of M2 markers (IL‐10, Arg‐1, CD206) by inhibiting the m6A methylation of TRAF6 and reducing its protein level and stability, thereby promoting M2 polarisation and alleviating the progression of DKD.