Semaglutide

Reagents used were 6-hydroxydopamine hydrobromide (6-OHDA, Sigma, USA), pargyline hydrochloride (MedChemExpress, USA), desipramine hydrochloride (MedChemExpress, USA), apomorphine (Absin Bioscience Inc, China), paraformaldehyde (PFA, Boster Biotechnology, China), RIPA lysis buffer (Beyotime Institute of Biotechnology, China), BCA Protein Assay Kit (Boster Biotechnology, China), ECL-enhanced chemoluminescence (Boster Biotechnology, China), Dopamine ELISA kit (Cloud-clone crop, China), TNF-α ELISA kit (Cloud-clone crop, China).
Semaglutide had been purchased from Bachem (Switzerland), and DA5-CH was synthetized by China peptides. The purity of each peptide was analyzed by reversed-phase HPLC and characterized using matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometry, with a purity >97%. Peptides were reconstituted in ultrapure water (Milli-Q) to a concentration of 1 mg/ml, and aliquots were prepared and stored at −20°C.

Treatment was initiated 7 weeks after AAV administration and 6 weeks post-UNx (Figure 1). Randomization and stratification to treatment was based on fed blood glucose and body weight measured one week before treatment start. db/db UNx-ReninAAV mice (n = 14–15 per group) received (q.d.) vehicle (0.5% methyl cellulose, s.c.), semaglutide (30 nmol/kg, s.c., Bachem AG, Bubendorf, Switzerland), or combined semaglutide (30 nmol/kg, s.c.) + lisinopril (30 mg/kg, p.o., MedChemExpress, Monmouth Junction, NJ, USA) for 11 weeks. A dose-escalation scheme was implemented to reduce expected initial effects of semaglutide treatment, as transient GLP-1R-induced discomfort in rodents, including taste aversion and pica behavior, is typically observed within the first 2–3 days of treatment [21 (link)].

D-PBS, DMEM, Trypsin, PBS, penicillin/streptomycin, fetal bovine serum (FBS) were from Gibco (Thermo Fisher Scientific, Waltham, MA, USA). Polyethylenimine (PEI) was purchased from Alfa Aesar (Thermo Fisher Scientific, Waltham, MA, USA). Glucagon was from Sigma Aldrich (Saint Louis, MO, USA). GLP-1 (1-37), GLP-1 (7-36), exenatide, liraglutide, semaglutide, lixisenatide and danuglipron were from either BACHEM (Bubendorf, Switzerland) or MedChemExpress LLC (Monmouth Junction, NJ, USA). Dyngo-4A was from ApexBio (Houston, TX, USA). Coelenterazine 400a was purchased from Nanolight Technologies (Pinetop, AZ, USA).