Journal: Cell Death & Disease

Article Title: PGC-1α inhibits the NLRP3 inflammasome via preserving mitochondrial viability to protect kidney fibrosis

doi: 10.1038/s41419-021-04480-3

Figure Lengend Snippet: A mRNA expression level of Ppargc1a was modulated by Ppargc1a plasmid transfection and siPGC-1α in TGF-β1-treated RTECs. B mRNA expression levels of mitochondrial dynamic-related genes were restored by overexpression of Ppargc1a with plasmid transfection, whereas reversed by siPGC-1α in TGF-β1-treated RTECs. C Protein expression levels of PGC-1α and mitochondrial dynamics were modulated by Ppargc1a plasmid transfection and siPGC-1α in TGF-β1-treated RTECs. D mRNA expression levels of Ppargc1a were increased by metformin in TGF-β1-treated RTECs. E mRNA expression levels of mitochondrial dynamic-related genes were restored by metformin in TGF-β1-treated RTECs. F Protein expression levels of p-AMPK, PGC-1α, and mitochondrial dynamics were restored by metformin in TGF-β1-treated RTECs. G mRNA expression levels of Ppargc1a were increased by metformin in adenine-fed mice. H mRNA expression levels of mitochondrial dynamic-related genes were restored in adenine-fed mice with metformin. I Protein expression levels of p-AMPK, PGC-1α, and mitochondrial dynamics was restored in adenine-fed mice with metformin. J Transmission electron microscopy images of RTECs from adenine-fed mice showed restoration of mitochondrial structures with metformin. Note: * P < 0.05 vs. control; ** P < 0.05 vs. TGF-β1-treated RTECs or adenine-fed mice. p-AMPK phospho-AMP-activated protein kinase; PGC-1α, peroxisomal proliferator-γ coactivator-1α; RTEC renal tubular epithelial cell; Met metformin; Mfn mitofusin; Tfam mitochondrial transcriptional factor A; Drp1 dynamin-related protein 1; Ade adenine; Met metformin.

Article Snippet: Furthermore, the cells were also transfected with Ppargc1a plasmid (1 μg) (Addgene, Cambridge, MA, USA) and Ppargc1a small interfering RNA (siRNA), using Lipofectamine 2000 and Plus reagents (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Plasmid Preparation, Transfection, Over Expression, Transmission Assay, Electron Microscopy, Control

Journal: Cell Death & Disease

Article Title: PGC-1α inhibits the NLRP3 inflammasome via preserving mitochondrial viability to protect kidney fibrosis

doi: 10.1038/s41419-021-04480-3

Figure Lengend Snippet: A – F mRNA and protein expression of fibrotic and apoptotic markers were attenuated in TGF-β1-treated RTECs with overexpression of Ppargc1a and metformin, whereas exacerbated by siPGC-1α. G Degree of kidney fibrosis by Masson’s trichrome staining was attenuated in adenine-fed mice with metformin. Note: * P < 0.05 vs. control; ** P < 0.05 vs. TGF-β1-treated RTECs or adenine-fed mice. PGC-1α peroxisomal proliferator-γ coactivator-1α; RTEC renal tubular epithelial cell; Ade adenine; Met metformin.

Article Snippet: Furthermore, the cells were also transfected with Ppargc1a plasmid (1 μg) (Addgene, Cambridge, MA, USA) and Ppargc1a small interfering RNA (siRNA), using Lipofectamine 2000 and Plus reagents (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Over Expression, Staining, Control

Journal: Cell Death & Disease

Article Title: PGC-1α inhibits the NLRP3 inflammasome via preserving mitochondrial viability to protect kidney fibrosis

doi: 10.1038/s41419-021-04480-3

Figure Lengend Snippet: A mRNA and B protein expression levels of NLRP3 inflammasome pathway were reduced in TGF-β1-treated RTECs with transfection of Ppargc1a plasmid, whereas increased by siPGC-1α. C Concentrations of IL-1β and IL-18 assessed by ELISA were reduced in TGF-β1-treated RTECs with overexpression of Ppargc1a . D mRNA and E protein expression levels of the NLRP3 inflammasome pathway were attenuated in TGF-β1-treated RTECs with metformin. F mRNA and G protein expression levels of the NLRP3 inflammasome pathway and H concentrations of IL-1β and IL-18 assessed by ELISA were attenuated in adenine-fed mice with metformin. Note: * P < 0.05 vs. control; ** P < 0.05 vs. TGF-β1-treated RTECs or adenine-fed mice. PGC-1α peroxisomal proliferator-γ coactivator-1α; NLRP3 NOD-like receptor family, pyrin domain-containing 3; ASC apoptosis-associated speck-like protein containing a caspase recruitment domain; RTEC renal tubular epithelial cell; ELISA enzyme-linked immunosorbent assay; Ade adenine; Met metformin.

Article Snippet: Furthermore, the cells were also transfected with Ppargc1a plasmid (1 μg) (Addgene, Cambridge, MA, USA) and Ppargc1a small interfering RNA (siRNA), using Lipofectamine 2000 and Plus reagents (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Transfection, Plasmid Preparation, Enzyme-linked Immunosorbent Assay, Over Expression, Control

Journal: Cell Death & Disease

Article Title: PGC-1α inhibits the NLRP3 inflammasome via preserving mitochondrial viability to protect kidney fibrosis

doi: 10.1038/s41419-021-04480-3

Figure Lengend Snippet: A – C ASC oligomeric structures assayed by DSS-mediated cross-linking were observed in TGF-β1-treated RTECs, which were attenuated by overexpression of Ppargc1a and metformin, whereas aggravated by siPGC-1α. D , E ASC oligomeric structures were observed in adenine-fed mice, which were attenuated by metformin. Note: * P < 0.05 vs. control; ** P < 0.05 vs. TGF-β1-treated RTECs or adenine-fed mice. PGC-1α peroxisomal proliferator-γ coactivator-1α; NLRP3 NOD-like receptor family, pyrin domain-containing 3; ASC apoptosis-associated speck-like protein containing a caspase recruitment domain; DSS disuccinimidyl suberate; RTEC renal tubular epithelial cell; Ade adenine; Met metformin.

Article Snippet: Furthermore, the cells were also transfected with Ppargc1a plasmid (1 μg) (Addgene, Cambridge, MA, USA) and Ppargc1a small interfering RNA (siRNA), using Lipofectamine 2000 and Plus reagents (Invitrogen, Carlsbad, CA, USA).

Techniques: Over Expression, Control

Journal: Cell Death & Disease

Article Title: PGC-1α inhibits the NLRP3 inflammasome via preserving mitochondrial viability to protect kidney fibrosis

doi: 10.1038/s41419-021-04480-3

Figure Lengend Snippet: A mtDNA copy numbers were decreased in the mitochondrial fraction, while increased in cytosolic fraction in TGF-β1-treated RTECs, which were reversed by Ppargc1a overexpression. B Confocal microscopy analysis with MitoSOX staining revealed increased production of mitochondria-generated ROS in TGF-β1-treated RTECs, which were reduced with overexpression of Ppargc1a and metformin. Conversely, siPGC-1α exacerbated mitochondrial ROS production. C Detection of mitochondria-generated ROS in TGF-β1-treated RTECs by FACS analysis. D Measurement of oxidative stress levels by MDA showed a reduction of oxidative stress levels in TGF-β1-treated RTECs with overexpression of Ppargc1a and metformin, which were increased by siPGC-1α. E The oxidative stress levels measured by MDA were reduced in adenine-fed mice with metformin. F , G mRNA and protein expression levels of TNFAIP3 in TGF-β1-treated RTECs were increased with overexpression of Ppargc1a and metformin, which were reduced by siPGC-1α. H mRNA and protein expression level of TNFAIP3 in adenine-fed mice was increased with metformin. Note: * P < 0.05 vs. control; ** P < 0.05 vs. TGF-β-treated RTECs or adenine-fed mice. PGC-1α peroxisomal proliferator-γ coactivator-1α; mtDNA mitochondrial DNA; NLRP3 NOD-like receptor family, pyrin domain-containing 3; RTEC; renal tubular epithelial cell; ROS reactive oxygen species; MDA malondialdehyde; TNFAIP3 tumor necrosis factor α induced protein 3; FACS Fluorescence-activated cell sorting; Ade adenine; Met metformin.

Article Snippet: Furthermore, the cells were also transfected with Ppargc1a plasmid (1 μg) (Addgene, Cambridge, MA, USA) and Ppargc1a small interfering RNA (siRNA), using Lipofectamine 2000 and Plus reagents (Invitrogen, Carlsbad, CA, USA).

Techniques: Over Expression, Confocal Microscopy, Staining, Generated, Expressing, Control, Fluorescence, FACS

Journal: Science Advances

Article Title: Muscle mitochondrial remodeling by intermittent glucocorticoid drugs requires an intact circadian clock and muscle PGC1α

doi: 10.1126/sciadv.abm1189

Figure Lengend Snippet: Results are shown at 4 hours (ZT4) after a single prednisone pulse in vivo at ZT0. ( A ) Unbiased motif analysis validated the muscle ChIP-seq for GR in both BMAL1-WT and BMAL1-KO quadriceps muscles. ( B ) Muscle GR peak profiles clustered according to genotype and drug. ( C ) Genome-wide GR occupancy of GRE sites was increased by the drug pulse. This effect was strongly limited in the BMAL1-KO muscle. ( D ) Prednisone shifted the muscle GR peaks from distal (>10 kb) to proximal (<10 kb) regions from TSSs, correlating with an enrichment in GR peaks in 5′UTR and promoter regions. These trends were partially blunted in BMAL1-KO muscle. ( E ) BMAL1 ChIP-seq in muscle showed enrichment for E-box motif in signal peaks. Muscle BMAL1 occupancy of E-box sites increased after prednisone pulse. ( F ) In BMAL1-WT muscle, the drug pulse increased the cooccurrence of peaks of GR and BMAL1 in the 0– to 300–base pair (bp) and 300- to 900-bp ranges. ( G and H ) Among genes with a BMAL1-dependent gain of RNA polymerase II (RNApol-II) at TSS with drug pulse, Nampt and Ppargc1a showed enrichment for both GR and BMAL1 signal in the promoter. The drug pulse increased GR, BMAL1, and RNApol-II peaks (arrows) on Nampt and Ppargc1a promoters in BMAL1-WT muscle, but not in BMAL1-KO muscle. N = 3 ♂ per group. * P < 0.05, two-way ANOVA + Sidak. fc, fold change.

Article Snippet: Primary antibodies (all diluted 1:1000 for overnight incubation at 4°C) were as follows: rabbit anti-NAMPT (#A0256, ABclonal), rabbit anti-PGC1a (#A12348, ABclonal), mouse antisarcomeric α-actinin (#A7732, Sigma-Aldrich), rabbit anti–ac-Lys (#2391, ABclonal), rabbit anti–p-Ser (#AP0932, ABclonal), rabbit anti-BMAL1 (A17334), rabbit anti-GR (#A2164, ABclonal), mouse anti-FLAG (also called DDK; #TA50011, Origene), rabbit anti–REV-ERBa (#A20452, ABclonal), rabbit anti-H3 (#A2348, ABclonal), and rabbit anti-HSP90 (#A5027, ABclonal).

Techniques: In Vivo, ChIP-sequencing, Genome Wide