Abstract 17870: Mitochondrial Antioxidant Manganese Superoxide Dismutase (MnSOD) Up-regulated Human Mesenchymal Stem Cells (MSCs) Reduce Inflammation in High Glucose (HG) Exposed Adipocytes

Abstract

Background: Primary MSCs can differentiate into mature adipocytes, osteocytes or chondrocytes. Behavior of MSCs in HG concentrations is largely unknown. HG exposure of MSCs led to increased intracellular lipid and superoxide accumulation in mitochondria that promotes inflammation and adipogenic differentiation which increases CVD risk. We hypothesized that intra-cellular antioxidant upregulation may reduce reactive oxygen species (ROS) accumulation, reduce associated inflammation and adipogenic differentiation of MSC in HG.

Methods: We exposed MSCs to HG (25mM) and normal glucose (NG, 5.5 mM). In HG, increased ROS was noted particularly in mitochondria using Mitosox-Red stain. We interrogated mitochondrial respiration (Seahorse) and complexes 1 & 2 protein by BN-PAGE and SDS-PAGE. Next, we transduced hMSCs with Adenovirus containing MnSOD, CAT or control gene GFP (green florescent protein) genes at 100 MOI, prior to HG exposure. In-vitro co-culture of adipocytes and transduced MSCs (3:1 ratio) were set up in NG and HG.

Results: HG increased lipid accumulation (3.5 fold), increased Superoxide accumulation in both cytosol and mitochondria. HG increased adipogenic gene expression of Leptin (8-fold), Perilipin (4-fold), CREBPα (10-fold), PPARG (16 fold) while it reduced bone formation markers Alkaline Phosphatase (4 fold), osteocalcin (1.6 fold) and osteopontin (4 fold) mRNAs. HG increased TNF (Tumor Necrosis Factor), IL6 (interleukin6) and Endothelin1, inflammatory marker gene expressions, significantly. Mitochondrial complex analyses indicated impaired mitochondrial respiration and suppressed complex 1 protein expression in HG exposed MSCs. MnSOD much more than CAT up-regulation, reduced adipogenic and inflammatory gene expressions and rescued suppressed Complex-1. Mature adipocytes when co-cultured with MnSOD upregulated MSC, reduced fat in adipocytes.

Conclusions: Upregulation of mitochondrial anti-oxidant (MnSOD) much more than cytosolic antioxidant (CAT) reduced inflammation, adipogenic differentiation and improved mitochondrial respiration of stem cells in HG. Therapeutic role of MnSOD upregulated MSC in a setting of HG associated obesity, inflammation and CVD risk in db/db mouse is currently being conducted.