Metabolic Memory in Diabetic Foot Syndrome (DFS): Epigenetic Changes of the Expression of Micro-rnas and Single Nucleotide Polymorphisms (SNPS) Frequency in a Cohort of Diabetic Patients With and Without Foot Ulceration and Correlation With Indices of Endothelial and Adipo-inflammatory Dysfunction
Authors: Cuore Alessandro Del , Pipitone Rosaria Maria , Casuccio Alessandra , Mazzola Marco , Puleo Maria Grazia , Pacinella Gaetano , Riolo Renata , Maida Carlo , Chiara Tiziana , Raimondo Domenico , Zito Rossella , Lupo Giulia , Agnello Luisa , Ciaccio Marcello , Grimaudo Stefania , Tuttolomondo Antonino 
Published: April 2023

Abstract
Diabetic foot is a significant cause of morbidity in diabetic patients, with a rate that is approximately twice that of patients without foot ulcers. There has been much debate in the literature about the role of genetics, particularly epigenetic modifications, in the genesis of the diabetic foot. “Metabolic memory” are all those epigenetic changes induced by chronic hyperglycaemia, despite correction of the glycaemic values themselves. Moreover, these epigenetic modifications would appear to perpetuate the damage caused by persistently elevated glucose levels even in its absence, acting at various levels, mostly affecting the molecular processes of diabetic ulcer healing. Our study aimed to analyze the epigenetic changes induced on miRNAs 126, 305, and 217 and on SNPs of inflammatory molecules, such as IL-6, TNF-alpha, and pro-angiogenic molecules, such as ENOS, VEGF and HIF-1alpha with endothelial dysfunction, assessed noninvasively by RHI and with serum levels of inflammatory molecules and adipokines in a population of people with diabetes with and without lower limb ulcer. Between March 2021 and June 2022, 110 patients were enrolled to the study: 50 diabetic patients with diabetic foot injuries, 40 diabetic patients without ulcerative complications and 20 non-diabetic patients, as control group. Diabetic subjects with lower limb ulcerative lesion exhibited higher values of inflammatory cytokines, such as VEGF (191.40 ± 200 pg/mL vs 98.27 ± 56.92 pg/mL vs 71.01 ± 52.96 pg/mL p = 0.22), HIF-1alpha (40.18 ± 10.80 ng/mL vs 33.50 ± 6.16 ng/mL vs 33.85 ± 6.84 ng/mL p = 0, 10), and Gremlin-1 (1.72 ± 0.512 ng/mL vs 1.31 ± 0.21 ng/mL vs 1.11 ± 0.19 ng/mL p < 0.0005). Furthermore, we observed that miR-217-5p and miR-503-5p were respectively 2.19-fold (p < 0.05) and 6.21-fold (p = 0.001) more expressed in diabetic foot patients than in healthy controls (Table 5, Graphic 1). Additionally, diabetic patients without lower limb ulcer complication showed a 2.41-fold (p = 0) and 2.24-fold (p = 0.029) higher expression of miR-217-5p and miR-503-5p, respectively, than healthy controls. Finally, diabetic patients with and without ulcerative complications of the lower limb showed a higher expression of the VEGFC2578A CC polymorphism (p = 0.001) and a lower expression of the VEGFC2578A AC polymorphism (p < 0.005) compared to the healthy control population. We identified a significant increase in Gremlin-1 in patients with diabetic foot and how this inflammatory adipokine is a predictive marker for the diagnosis of diabetic foot. Our results highlighted that patients with diabetic foot show predominant expression of the VEGF C2578A CC polymorphism and reduced expression of the AC allele. Additionally, we found an overexpression of miR-217-5p and miR-503-5p in diabetic patients with and without diabetic foot syndrome. These results align with the licterature, who evidenced the overexpression of miR-217-5p and miR-503-5p in the context of diabetic foot. These epigenetic modifications could therefore be helpful in the early diagnosis of diabetic foot and the treatment of risk factors. However, only further studies will be able to confirm this hypothesis.