Combination of resveratrol and 5‐azacytydine improves osteogenesis of metabolic syndrome mesenchymal stem cells

Abstract

Endocrine disorders have become more and more frequently diagnosed in humans and animals. In horses, equine metabolic syndrome (EMS) is characterized by insulin resistance, hyperleptinemia, hyperinsulinemia, inflammation and usually by pathological obesity. Due to an increased inflammatory response in the adipose tissue, cytophysiological properties of adipose derived stem cells (ASC) have been impaired, which strongly limits their therapeutic potential. Excessive accumulation of reactive oxygen species, mitochondria deterioration and accelerated ageing of those cells affect their multipotency and restrict the effectiveness of the differentiation process. In the present study, we have treated ASC isolated from EMS individuals with a combination of 5‐azacytydine (AZA) and resveratrol (RES) in order to reverse their aged phenotype and enhance osteogenic differentiation. Using SEM and confocal microscope, cell morphology, matrix mineralization and mitochondrial dynamics were assessed. Furthermore, we investigated the expression of osteogenic‐related genes with RT‐PCR. We also investigated the role of autophagy during differentiation and silenced PARKIN expression with siRNA. Obtained results indicated that AZA/RES significantly enhanced early osteogenesis of ASC derived from EMS animals. Increased matrix mineralization, RUNX‐2, collagen type I and osteopontin levels were noted. Furthermore, we proved that AZA/RES exerts its beneficial effects by modulating autophagy and mitochondrial dynamics through PARKIN and RUNX‐2 activity.

Mitochondrial Dynamics: Fission and Fusion in Fate Determination of Mesenchymal Stem Cells

“…The Parkin is a crucial mitophagy regulator and its activation can build ubiquitin chains to ubiquitinating outer membrane protein on damaged mitochondria to label them for degradation in lysosomes (Bingol and Sheng, 2016). These findings are consistent with previous research by the same team, which showed that increased ASC apoptosis derived from EMS horses resulted in enhanced mitochondrial fission compared with the healthy control (Marycz et al, 2018). Similarly, metabolic syndrome impairs the swine ASC mitochondrial structure featured by an increase in mitochondrial fission and reduction of mitochondrial fusion (Farahani et al, 2020).…”

“…According to the literature, stress often results in fragmentation but not elongation events in mitochondria in MSCs, which is attributed to dramatic enhanced mitochondrial fission with/without inhibited mitochondrial fusion (He et al, 2018;Marycz et al, 2018;Shen et al, 2018a;Ma et al, 2019). In addition, the suppression effect on both mitochondrial fission and fusion was observed in BMSCs when they were treated with high levels of ferric ammonium citrate, a commonly used agent to induce iron overload (Yao et al, 2019).…”

Intra-Vitreal Administration of Microvesicles Derived from Human Adipose-Derived Multipotent Stromal Cells Improves Retinal Functionality in Dogs with Retinal Degeneration

“…Electrophoresis was performed from samples with equal amounts of protein (30 μg). SDS-PAGE and the protein transfer were completed using protocols described previously [34,35]. Primary antibodies against BDNF (SAB2108004, Sigma Aldrich, Poznan, Poland) were prepared at a dilution of 1:250 in Tris/NaCl/Tween buffer (TBST).…”

5-Azacytidine and Resveratrol Enhance Chondrogenic Differentiation of Metabolic Syndrome-Derived Mesenchymal Stem Cells by Modulating Autophagy

“…All flow cytometry analyses were performed after 24 hours of chondrogenic differentiation as described previously [38]. Cells were incubated with anti-5-mC antibody (Abcam, ab73938) and anti-histone H3 (Abcam, ab8898) with 10% goat serum.…”