SAT-LB095 The Role of Type 2 Deiodinase in Interactions Between Muscle Stem Cells and Endothelial Cells

Abstract

Thyroid hormone (TH) is a major muscle target and is required for muscle homeostasis, function, and regeneration. T3, the biologically active TH, is locally produced in muscle from the prohormone T4 by type 2 deiodinase (D2). An increase in D2, and the subsequent T3 generated, is required for normal muscle stem cell (MuSC) differentiation during muscle repair [1]. Interestingly, crosstalk between MuSC and endothelial cells via vascular endothelial growth factor (VEGF) is critical for proper muscle regeneration. It is unknown if local T3 generation by D2 influences interactions between MuSCs and endothelial cells. We hypothesized that D2-deficiency in MuSCs will affect crosstalk with endothelial cells. To study this, we deleted D2 in a myoblast cell line (D2KO C2C12) and explored interactions between C2C12 and human umbilical vein endothelial cells (HUVECs). Initially, we induced differentiation of wild-type (WT) C2C12 and D2KO C2C12 by serum withdrawal and assessed gene expression. We found that VEGF expression increased with differentiation, peaking at day 4. Strikingly, this was 2.3-fold higher in WT than D2KO C2C12 (p< 0.0001). Consistent with mRNA changes, both intracellular and secreted VEGF were increased in a time-dependent manner during differentiation. Notably, secreted VEGF from WT C2C12 was significantly higher than D2KO C2C12 (2.5-fold, p < 0.0001). Further, VEGF expression in differentiated WT C2C12 cells was increased 1.5-fold (p=0.03) with T3-treatment, suggesting VEGF is a T3-responsive gene. VEGF has proangiogenic effects during muscle injury [2]. To assess the functional consequences of decreased VEGF production by D2KO C2C12 cells, HUVECs were incubated with WT and D2KO C2C12-derived conditioned media collected during differentiation. We found that angiogenesis and chemotaxis of HUVECs were enhanced after culture with WT C2C12 conditioned media. In contrast, these parameters were blunted when HUVECS were cultured with D2KO C2C12 conditioned media, consistent with lower VEGF. Our results suggest that local D2-mediated increase in T3 regulates VEGF production during myoblast differentiation, promoting endothelial cell migration and angiogenesis.