Anti pgc1α

The protocol was performed as previously described. The following primary antibodies were used: anti-CXCL15 (ab197016, Abcam, 1:1000); total p65NF-κB (A2547, ABclonal, 1:1000); NF-κB p65-pS536 (3033 T, Cell Signaling Technology, 1:1000)；anti-PGC1α (A19674, ABclonal, 1:1000); anti-UCP1 (A5857, ABclonal, 1:1000); anti-α-Tubulin (66031-1-Ig, Proteintech, 1:1000); anti-LaminB1 (A11495, ABclonal, 1:1000); anti-GAPDH(60004-1-Ig, Proteintech, 1:1000) and anti-Calnexin (2679 T, Cell Signaling Technology, 1:1000).

Proteins were extracted from cells in RIPA buffer (Beyotime, #P0013B) with protease and phosphatase inhibitor. Cell lysates were centrifuged at 12,000 g for 30 min at 4°C, and the protein concentration was measured by BCA Protein Assay Kit (EpiZyme, #ZJ102). Proteins were separated by SDS‐PAGE gel and transferred to 0.45 μm PVDF membranes (Millipore, #IPVH00010), which were then blocked with 5% skim milk for 1 h at room temperature. Primary antibodies were incubated with the membrane at 4°C overnight, including anti‐emerin (1:1000 #PA5‐79201) purchased from Invitrogen; anti‐OXPHOS (1:500, #ab110413) purchased from Abcam; anti‐PGC1α (1:1000, #A12348) purchased from ABclonal Technology; anti‐DRP1 (1:1000, #8570), anti‐MFN2 (1:1000, #9482) and anti‐GAPDH (1:2000 #5174) purchased from Cell Signalling Technology. After washing thrice in TBST, membranes were incubated with horseradish peroxidase (HRP)‐labelled Goat Anti‐Rabbit IgG (Beyotime, #A0208) or Goat Anti‐Mouse IgG (Beyotime, #A0216). Following addition of ECL Prime Western Blotting Detection Reagent (Amersham Bioscience, #RPN2232), immunoblot signal was detected by GeneGnome XRQ Chemiluminescence Imaging System (Syngene). Intensity of bands was quantified using ImageJ software.

CBL was provided by Guangdong Long Fu Pharmaceutical Co., Ltd. (Zhongshan, China). Rabbit anti-phospho-CREB (1:1,000), anti-CREB (1:1,000), anti-PGC-1α (1:1,000), anti-β-actin (1:10,000) antibodies were purchased from the ABclonal company. Rabbit anti-phospho-ERK1/2 (1:1,000), anti-ERK1/2 (1:1,000), anti-phospho-JNK (1:1,000), anti-JNK (1:1,000), anti-phospho-p38 MAPK (1:1,000), anti-p38 MAPK (1:1,000) antibodies were purchased from Cell Signaling Technology (Beverly, MA). Compound 666-15, the inhibitor of CREB was purchased from MedChemExpress (MCE, Shanghai, China). RIPA lysis buffer and LDH kit were purchased from Beyotime Biotechnology (Nanjing, China). Trizol reagent and the cDNA synthesis kit were purchased from Vazyme (Nanjing, China). SYBR Green was purchased from Invitrogen (Camarillo, CA). LPS was purchased from Sigma-Aldrich (St. Louis, USA). Cell culture medium and supplements were purchased from Invitrogen (Carlsbad, CA, USA). TTC (2, 3, 5-triphenyltetrazolium chloride) was bought from Sigma-Aldrich.