Tiron

Physiological Krebs Henseleit buffer of the following composition was used: NaCl 130 mM, KCl 4.7 mM, KH₂PO₄ 1.18 mM, MgSO₄.7H2O 1.17 mM, NaHCO₃ 14.9 mM, EDTA 0.026 mM, CaCl₂.2H₂O 1.6 mM and D‐glucose 5.55 mM. A high concentration of potassium chloride buffer of the following composition was used: NaCl 14 mM, KCl 120 mM, KH₂PO₄ 1.18 mM, MgSO₄.7H₂O 1.17 mM, NaHCO3 14.9 mM, EDTA 0.026 mM, CaCl2.2H2O 1.6 mM and D‐glucose 5.55 mM. The incubations were performed with ET‐1 (1, 10 and 100 nM, Tocris 1160; diluted in 5% BSA + 95% deionized water), MCC950 [1 μM,⁴⁴ Cayman Chemical 17,510; diluted in 5% DMSO and 95% deionized water], tiron (100 μM, Santa Cruz SC253699; diluted in deionized water), BAPTA AM [5 μg/ml,²⁹ Tocris 2787; diluted in deionized water], lipopolysaccharide (LPS) (1 μg/ml; diluted in deionized water), adenosine 5‐triphosphate (ATP) (2 mM, Sigma‐Aldrich A6144; diluted in deionized water), BQ123 (100 nM, 1, 10 μM, Tocris 1188; diluted in deionized water), BQ788 (100 nM, 1, 10 μM, Tocris 1500; diluted in deionized water).

Phenylephrine hydrochloride, acetylcholine chloride, LPS, lucigenin (N,N′-Dimethyl-9,9′-biacridinium dinitrate) and peg-catalase (Catalase-polyethylene glycol) were obtained from Sigma-Aldrich (St. Louis, MO, United States), MCC950 from Avistron^® (Bude, Cornwall, United Kingdom), Tiron from Santa Cruz Biotechnology^® (San Juan, CA, United States) and all reagents used in RT-PCR and murine primers from Invitrogen^® (Carlsbad, CA, United States). Human primers from Sigma-Aldrich^®, All other salts used were obtained from Merck^® (Rio de Janeiro, RJ, Brazil). For cell culture, DMEM was purchased from Sigma^®; FBS and antibiotics (Penicillin/Streptomycin) from Gibco Thermo Fisher Scientific^® (Waltham, MA, United States).

Aldosterone, eplerenone, phenylephrine, and acetylcholine were purchased from Sigma-Aldrich (St. Louis, MO, USA). Tiron (Santa Cruz, Dallas, TX, USA), L-sulforaphane (Cayman, Ann Arbor, MI, USA), and SB 239063 (Tocris, Bristol, UK) were also purchased.