Uk5099

A detailed list of all compounds included in the metabolic compound screen is available in supplementary Table 1. mTOR inhibitor rapamycin (S1039, Selleckchem), BH3 mimetic ABT-737 (S1002, Selleckchem) and general caspase inhibitor Z-vad-FMK (550,377 BD biosciences) were dissolved in DMSO according to the manufacturer's instructions. Chemicals for the Seahorse experiments oligomycin A (11,342), trifluoromethoxy carbonylcyanide phenylhydrazone (FCCP, 15,218), antimycin A (19,433), rotenone (13,995), UK5099 (16,980), Etomoxir (11,969) and BPTES (19,284) were all purchased from Cayman Chemical (Massachusetts, USA) and dissolved in DMSO according to the manufacturer's instructions. Doxorubicin and cisplatin in a 0.9% NaCl solution were obtained from the in-house hospital pharmacy.

All animal experiments were carried out according to our previous WIHN model protocol (1 (link), 26 (link)). In short, 21-day-old male and female mice weighing 8 to 12 g with hairs in the first telogen stage were selected. Mice were anesthetized with isoflurane, shaved, and denuded of 1.44 cm² full-thickness skin on the lower center of the back, using sterile procedures. The operation day was defined as WD0, and the treatment was performed at WD3 according to each experimental design. At about WD14, the wound scab would fall off. We defined it as scab-detached day (SD0), which is the time point when hair regeneration began. On day 24 after the procedure (WD24), the number of regenerated hair follicles is measured. We visualized the regenerated hair follicles and quantified them using confocal scanning laser microscopy (CLSM), as described before (26 (link)). In GF mice, we performed the procedure in the bubble of the GF laboratory of the Johns Hopkins School of Public Health. In SPF mice, we performed the procedure in a biological safety hood in the Johns Hopkins School of Medicine. S. aureus (1 × 10⁷/50 μl) was injected under the scab on WD3. We also injected 20 μl of 100 nM LW6 (Selleck Chemicals), 1 mM glutamine (Gibco), 200 nM CB839 (Cayman Chemical), 500 nM FX11 (Sigma-Aldrich), or 200 nM UK5099 (Cayman Chemical) under the scab every other day from WD3 to SD0.

The chemical and biochemical reagents were obtained from the following sources: ascorbic acid, l-buthionine sulfoximine (BSO), cystine, and L-γ-glutamyl-p-nitroanilide (GPNA) were purchased from Wako Pure Chemicals (Osaka, Japan); oligomycin and 3PO were from Calbiochem-Merck (Darmstadt, Germany); antimycin A1, bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES), cilengitide, dimethyl-α-ketoglutarate (DM-αKG), d-glucose, Dulbecco's modified Eagle's medium (DMEM), dimethylsulfoxide, l-glutamine, MEM amino acids solution, MEM non-essential amino acids solution, MEM vitamin solution, N-acetyl-cysteine (NAC), poly 2-hydroxyethyl methacrylate (poly-HEMA), proteinase and phosphatase inhibitor cocktail, sulfasalazine, and 2-deoxy-d-glucose (2DG) were from Sigma-Aldrich (St. Louis, MO, USA); CB839 and UK5099 were from Cayman Chemical (Ann Arbor, MI, USA); etomoxir was from Selleck Chemical (Houston, TX, USA). Antibodies against AMPKα, phosphorylated (p)-AMPKα (Thr172), ASCT2, FAK (Tyr397), p-FAK, IDH1, LKB1, PARP, p-H2AX (Ser139), and xCT were obtained from Cell Signaling (Beverly, MA). Antibodies against GCLC, GCLM, and ME1 were from Abcam (Cambridge, UK). Antibodies against α-tubulin and β-actin were from Wako Pure Chemicals. Antibodies against lamin B1 and Nrf2 were from MBL (Nagoya, Japan) and GeneTex (San Antonio, TX, USA), respectively.