Angiotensin 2 ang 2

Salmon calcitonin (sCT, H-2260; Bachem), pramlintide (SC-476040; Santa Cruz) and liraglutide (gift of NovoNordisk, Denmark) were dissolved in 0.1 M PBS (pH 7.4) for systemic (IP) injections. Amylin (H-9475.100; Bachem), GLP-1 (7–36; H-67951000, Bachem) and Angiotensin II (Ang II: Bachem) were dissolved in artificial cerebrospinal fluid (aCFS; Harvard Apparatus) for central injection. 5′-bromo-2′-deoxyuridine (BrdU; Sigma) was dissolved in double-distilled water and heated to 40–50 °C.

Angiotensin II (Ang II) was purchased from Bachem (Torrance, CA), dihydroethidium (DHE) from Invitrogen (Carlsbad, CA), and 6β-hydroxytestosterone (6β-OHT) from Steraloids (Newport, RI). Phenylephrine, endothelin-1, acetylcholine, the Masson trichrome staining kit, and phosphate-buffered saline were purchased from Sigma (St. Louis, MO).

Peptides were synthesized by Apeptide, Shanghai, China; the peptide sequences have been published previously [32 (link)]. The estimated purity of the peptides, evaluated by mass spectroscopy, was >89%. Lipopolysaccharide (LPS), derived from Escherichia coli 0111:B4 cell membranes and purified by gel-filtration chromatography, was obtained from Sigma-Aldrich (St. Louis, MO, USA, cat # L3012). The Amplex red/ horseradish peroxidase (HRP) assay kit (cat.#A22188) and the carboxy adducts of reduced difluorofluorescein diacetate (DFF-DA, cat.#13293) were purchased from Life Technologies, Grand Island, NY, USA (through Thermo-Fisher Scientific, Waltham, MA, USA). Angiotensin II (Ang II) was obtained from Bachem, Torrance, CA, USA (cat. #4095850.0005). Authentic lipids were purchased from Sigma-Aldrich, St. Louis, MO, USA and liposomes were prepared by evaporation to dryness followed by reconstitution in saline as previously described to reflect the composition of lung surfactant [27 (link)]; the liposome composition was, in mol fraction, 0.5 dipalmitoylphosphatidylcholine (DPPC), 0.25 egg phosphatidylcholine (PC), 0.10 phosphatidylglycerol (PG) and 0.15 cholesterol. PIP-2 when added was 0.15 µg PIP-2/µg lipid.