From inflammation to healing: the crucial role of GPR91 activation and SDH inhibition in chronic diabetic wound recovery

Abstract

Background Diabetic foot ulcers (DFU) typically exhibit impaired healing due to dysregulated re-epithelialization and excessive inflammation. Succinate, a key metabolic intermediate, is now understood to regulate inflammation through G Protein-Coupled Receptor 91 (GPR91) and succinate dehydrogenase (SDH), although its role in DFU remains unclear. Methods Co-cultures of M2 macrophages and epithelial cells, along with clinical samples, were used to analyze the expression of GPR91 and SDH. Functional assays were performed using high glucose (HG)-treated M2 macrophages (HG-M2) and an in vivo model. Cytokine and growth factor levels in cell supernatant were measured, and molecular mechanisms were explored via qRT-PCR, flow cytometry, and western blot analysis. Results Elevated glucose concentrations increased succinate levels and disrupted M2 macrophage–epidermal stem cells (EpSCs) interactions. GPR91 knockdown worsened HG-M2 dysfunction, while GPR91 overexpression (OE-GPR91) enhanced anti-inflammatory responses and reduced succinate. OE-GPR91-conditioned medium preserved EpSCs stemness and promoted migration mediated by hepatocyte growth factor (HGF). SDH inhibition (via Dimethyl malonate, DMM) boosted M2 macrophage activity by reducing reactive oxygen species (ROS) and upregulating Gpr91 expression. Mechanistically, GPR91 activated the pAkt/pGSK3β/β-catenin pathway, while DMM enhanced M2 macrophage function via the PI3K-Akt/pERK1/2 pathway. Conclusions GPR91 upregulation and SDH inhibition improve HG-M2 macrophage function, reduce inflammation, and enhance HGF-mediated EpSCs repair. Targeting both pathways may represent a promising approach to promote DFU healing.

1. Introduction

Each year, approximately 18.6 million individuals with diabetes globally suffer from foot ulcers [1]. In China, the prevalence of diabetic foot ulcers (DFU) is alarmingly high, ranging from 5.7 to 8.1%. Understanding the molecular mechanisms underpinning DFU development and developing effective therapeutic strategies is therefore crucial [2–4]. During the advanced stages of DFU, wounds create a microenvironment characterized by elevated glucose levels, along with the release of advanced glycation end products (AGEs), reactive oxygen species (ROS), and inflammatory cytokines [5, 6]. These harmful substances inhibit keratinocyte migration and proliferation, thereby impairing re-epithelialization and wound healing [7, 8]. Studies have shown that inhibiting tumor necrosis factor (TNF) in wounds restores M2 macrophages, leading to improved re-epithelialization [9]. Consequently, the interaction between M2 macrophages and epidermal cells is essential for initiating [10]– [11] and resolving inflammation, as well as progression through the later stages of healing [12–15].

Succinate is a significant pro-inflammatory metabolite [16]. Its regulatory role as a metabolic intermediate in signal transduction, metabolic homeostasis, inflammatory and immune responses, and tissue repair has attracted considerable attention [17, 18]. Elevated succinate levels have been detected in the circulation of obese/diabetic humans and mice [19]. G Protein-Coupled Receptor 91 (GPR91) has been established as the natural ligand for succinate, a crucial tricarboxylic acid (TCA) Cycle intermediate. GPR91 is widely expressed across multiple tissues, particularly in the heart, kidneys, and macrophages [20]. However, the role of GPR91 in pro-inflammatory and anti-inflammatory processes requires further investigation. Littlewood-Evans et al. reported that elevated extracellular succinate could enhance GPR91 expression, thereby amplifying the pro-inflammatory state of macrophages [21]. Conversely, Mette Trauelsen et al.'s study found that GPR91 signaling through the Gq pathway could hyperpolarize human M2 macrophages [22]. Besides, research by Noelia Keiran and colleagues suggests that GPR91 signaling could induce an anti-inflammatory phenotype in adipose tissue macrophages (ATMs) and suppress tissue inflammation following metabolic homeost