Hy 15886

All the procedures were performed on approval by the Institutional Animal Ethics Committee of Xi'an Jiaotong University and following the Guide for the Care and Use of Laboratory Animals of Xi'an Jiaotong University Animal Experiment Center. Male SD rats were purchased from Xi'an Jiaotong University Experimental Animal Center. All rats were kept on a 12 h light/dark cycle environment and fed ad libitum with a standard diet at temperature of 20 ± 2℃. Briefly, rats (weighing approximately 200‐220 g) were randomly divided into five groups (n = 8 animals/group) and treated as below: CON group: received saline by intraperitoneal (ip) injection on day 1, and then with an equal volume of vehicle (0.9% NaCl) alone for 28 days; MCT group: received a single ip injection of 60 mg/kg MCT (Must Bio‐Technology, Chengdu, China) at day 1 to induce PAH, as previously described²⁷ ; MCT+DMSO group: received vehicle DMSO by daily ip injection; MCT+Glycyrrhizin (GLY) group: received GLY (100 mg/kg, 53956‐04‐0, Santa Cruz, CA, USA) by daily ip injection⁸, ⁹ ; MCT+Mitochondrial division inhibitor (Mdivi‐1) group: received Mdivi‐1 (50 mg/kg, HY‐15886, MedChemExpress, NJ, USA) by twice weekly ip injection ¹² ; MCT+CQ group: received CQ (50 mg/kg, Aladdin, Shanghai, China) by daily gavage tube.¹⁹

For HIF‐1α inhibition, mice were administered PX‐478 (5 mg/kg, HY‐10231, MedChemExpress) in phosphate‐buffered saline (PBS) solution by oral gavage every other day, and the control mice received an equivalent amount of PBS. For NLRP3 inhibition, mice were gavaged with MCC950 (20 mg/kg, HY‐12815, MedChemExpress) or vehicle (.9% NaCl) every day for 5 days/week. For Drp1 inhibition, mice were given mitochondrial division inhibitor 1 (Mdivi‐1) (20 mg/kg, dissolved in dimethyl sulfoxide (DMSO), HY‐15886, MedChemExpress) intraperitoneally, and the control mice were injected with the same volume of DMSO. For ROS scavenging, mice were intraperitoneally injected with mito‐TEMPO (MT, dissolved in PBS, 10 mg/kg, HY‐112879, MedChemExpress) every other day, and the mice in the control group were given an equal volume of PBS intraperitoneally.

UA, CQ, MG-132, and Mdivi-1 (HY-100599, HY-17589A, HY-13259, and HY-15886, respectively) were procured from MedChemExpress (USA) and dissolved in dimethyl sulfoxide (DMSO; Sigma-Aldrich, St. Louis, MO, USA). NIH3T3 cells were exposed to UA (25 μM, 50 μM, 100 μM, or a vehicle) or mdivi-1 (12.5 μM, 25 μM, 50 μM, or a vehicle) for 48 h, or to CQ (25 μM, 50 μM, 100 μM, or a vehicle) or MG132 (5 μM) for 24 h. Subsequently, the cells were harvested for Western blotting analysis.
For the administration of UA via gavage, the rats received a daily gavage of either 20 mg/kg/d UA in PBS or PBS alone at 10 am for a duration of 28 days. After the 28-day period, the rats were euthanized, and their SCN tissues were collected for further analysis.