Modulation of Arteriolar Autophagic Pathways

Isolated adipose and atrial arterioles were incubated for 15-20 hours in endothelial cell growth media containing 5% serum (Lonza), containing either vehicle (DMSO 2% by volume), Bafilomycin A1 (BAFA1; Sigma-Aldrich, 100 nmol/L), trichostatin A (TSA; Cayman Chemical, 100 nmol/L), ATG3 siRNA (Ambion, AM16708; 10 nmol/L), trehalose (Research Products International, 10 mmol/L), BIBR 1532 (Tocris Bioscience; 10 umol/L), or AGS 499 (Ester Priel, PhD, Israel; 20 nmol/L). TSA and trehalose were independently used to induce autophagy in arterioles, while BAFA1 and ATG3 siRNA were used to inhibit or prevent completion of autophagy in arterioles. Previous work from our lab has demonstrated that modulation of TERT with the BIBR 1532 and AGS 499 induce a switch in mechanism of dilation in human atrial and adipose arterioles from NO to H₂O₂, and H₂O₂ to NO, respectively^{12 (link)}. Following incubation, arterioles were cannulated and underwent stimulation of FMD as described above.