Figure 6

Compromised NRF2-HO-1 axis accounts for redox dysregulation in SIRT6-deficient hMSCs. (A) FACS analyses of ROS level in hMSCs transduced with lentiviral vector encoding luciferase (control) or HO-1. (B) Lactate dehydrogenase (LDH) detection in the indicated hMSCs transduced with lentiviral vector encoding luciferase (control) or HO-1 in the presence of 50 μM PX-12 treatment. Data were presented as mean ± SEM, n = 3, *P < 0.05. (C) FACS analyses of PX-12-induced cytotoxicity in hMSCs transduced with lentiviral vector encoding luciferase (control) or HO-1 (left panel). Cells were treated with vehicle (DMSO) or 50 μM PX-12 for 24 h. Statistical analysis of apoptotic cells (right panel) was presented as mean ± SEM. n = 6, **P < 0.01. (D) Measurement of luciferase activity in immunodeficient mice with IVIS. SIRT6-deficient hMSCs overexpressing GFP plus luciferase (control group, left) and SIRT6-deficient hMSCs overexpressing HO-1 plus luciferase (right) were implanted into the TA muscles of mice. Six days after implantation, mice were intraperitoneally injected with 20 mg/kg PX-12 for 24 h, and then luciferase activity was measured. Data were presented as mean ± SEM, n = 4, *P < 0.05. (E) A putative model for SIRT6-mediated redox regulation in hMSCs. In WT hMSCs, SIRT6 is a key regulator of the cellular redox homeostasis by co-activating NRF2 antioxidant pathway. SIRT6 associates with NRF2 and deacetylates H3K56 at the promoter of NRF2 target genes (i.e., HO-1), which is required for the recruitment of RNAP II complex and subsequent transactivation of NRF2. In SIRT6-deficient hMSCs, SIRT6 deficiency causes increased level of H3K56Ac and impaired recruitment of RNAP II complex to HO-1 promoter, resulting in decrease in HO-1 expression and compromised cellular redox homeostasis.