Journal: Cell Reports Medicine

Article Title: Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults

doi: 10.1016/j.xcrm.2022.100633

Figure Lengend Snippet: Urolithin A confers a proteomic signature of improved mitochondrial metabolism and mitophagy in human skeletal muscle (A) Venn diagram summarizing pathway enrichment analysis results from proteomics data in vastus lateralis skeletal muscle. Data represent upregulated pathways with an adjusted p value <0.1 in subjects treated with placebo or with UA at 500 and 1,000 mg for 4 months compared with baseline. (B and C) Dot plots showing top enriched pathways (WikiPathways 2019 Human), ranked by protein ratio, in the UA 500- (B) and UA 1,000 mg (C) groups from (A). Dot color and size indicate adjusted p value and protein count, respectively. Significant pathways for the placebo group were filtered out to identify treatment-specific pathways. (D) Western blot analysis of protein lysates from vastus lateralis skeletal-muscle biopsies in subjects treated as above. For each subject, both baseline (Pre) and end-of-the-treatment (Post) samples were run, and membranes were probed for phospho-Parkin, total Parkin, and the mitochondrial proteins ATP5A (belonging to the OXPHOS complex V), UQCRC2 (complex IV), SDHB (complex II), and NDUFB8 (complex I). Tubulin and VDAC were included as markers of total and mitochondrial protein abundance, respectively. Dashed line separates samples from individual subjects. (n = 6 Pre and Post, biologically independent samples). (E) Quantification of phospho-Parkin over Tubulin protein intensity from western blots (WBs) in (D) (n = 6). Two-sided, paired t-test. (F) Quantification of NDUFB8 (left) and SDHB (right) protein intensity, normalized over VDAC from WBs in (D) (n = 6). ∗p < 0.05; ∗∗p < 0.01; two-sided, paired t test.

Article Snippet: The following primary antibodies were incubated overnight diluted in blocking buffer: UBE2N (SantaCruz, sc-376470, 1:3000), Tubulin (Proteintech, 10004185, 1:3000), phospho-Parkin S65 (Cell Signaling, #36866, 1:1000), Parkin (Cell Signaling, #4211, 1:1000), OXPHOS Antibody Cocktail (Abcam, ab110413, 1:2000), VDAC (Proteintech, 10866-1-AP, 1:1000).

Techniques: Western Blot, Quantitative Proteomics

Journal: Cell Reports Medicine

Article Title: Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults

doi: 10.1016/j.xcrm.2022.100633

Figure Lengend Snippet:

Article Snippet: The following primary antibodies were incubated overnight diluted in blocking buffer: UBE2N (SantaCruz, sc-376470, 1:3000), Tubulin (Proteintech, 10004185, 1:3000), phospho-Parkin S65 (Cell Signaling, #36866, 1:1000), Parkin (Cell Signaling, #4211, 1:1000), OXPHOS Antibody Cocktail (Abcam, ab110413, 1:2000), VDAC (Proteintech, 10866-1-AP, 1:1000).

Techniques: Recombinant, RNA Extraction, Software, Protease Inhibitor

Journal: Molecular nutrition & food research

Article Title: Lack of β, β-carotene -9’, 10’-oxygenase 2 leads to hepatic mitochondrial dysfunction and cellular oxidative stress in mice

doi: 10.1002/mnfr.201600576

Figure Lengend Snippet: Functional categorization of proteins differentially expressed in hepatic mitochondria of BCO2 knockout (KO) vs. 129S6 wild type (WT) mice identified by spectrum counting. Values are means, n=3 mice/group with 4 technical replicates/mouse.

Article Snippet: Antibodies against fatty acid synthase (FASN) (catalog # 10624-2-Ap), ATP synthase α subunit 1 (ATP5A1) (catalog # 14676-1-AP), citrate synthase (catalog # 16131-1-AP), hydroxyacyl-coenzyme A dehydrogenase (HADH) (catalog # 19828-1-AP), succinate dehydrogenase α (SDHA) (catalog # , 14865-1-AP), and nicotinamide nucleotide transhydrogenase (NNT) (catalog # 13442-2-AP) were purchased from ProteinTech Group (Chicago, IL, USA); Antibodies against lysosome-associated membrane protein 1 (LAMP1) (catalog # sc-20011), catalase (catalog # sc-50508), NNT (catalog # sc-163154), glutathione reductase (GSR) (catalog # sc-133245) were purchased from Santa Cruz Biotech (Dallas, TX, USA); peroxisome proliferator-activated receptor α (PPARα) antibody (catalog # 101710) was purchased from Cayman (Ann Arbor, MI, USA); Antibodies against β-actin (catalog # 4967), phosphor-Thr172-AMP-activated protein kinase α (pT172-AMPKα) (catalog # 2535), and AMPKα (catalog # 2603) were purchased from Cell Signaling Technology (Danvers, MA, USA); voltage-dependent anion-selective channel protein 1 (VDAC1) antibody (catalog # PA1780) was purchased from Boster Biosciences (Pleasanton, CA, USA).

Techniques: Functional Assay, Knock-Out, Membrane

Journal: Cancer Reports

Article Title: A novel polyethylene glycol ( PEG )‐drug conjugate of Venetoclax, a Bcl‐2 inhibitor, for treatment of acute myeloid leukemia ( AML )

doi: 10.1002/cnr2.1485

Figure Lengend Snippet: Bax expression levels and cytochrome c concentrations in the cytoplasm or mitochondria following treatment with either free VTX or PEG‐VTX in vitro. MV4‐11 cells were seeded into a T25 flask (2.5 × 10 6 cells/flask) and were exposed to either free VTX or PEG‐VTX (0.01, 0.1, or 1 μM as an API VTX concentration). After 5 h of incubation, the cytoplasm and mitochondria fractions were collected. (A) The expression levels of Bax and β‐Actin in cytoplasm fraction and those of Bax and VDAC1/2 in mitochondria fraction were determined using a Wes system. The lower graphs show the relative signal intensity of Bax corrected by that of β‐Actin for cytoplasm fraction or that corrected by VDAX1/2 for mitochondria fraction. The data represent the mean ± SD ( n = 3, ** p < .01, *** p < .001 vs. control). (B) The concentrations of cytochrome c in these fractions was measured via ELISA, and corrected using these protein concentrations. The data represent a typical result from three independent experiments

Article Snippet: The expression of VDAC1/2, as a loading control for mitochondrial fraction, was detected using an anti‐VDAC1/2 primary antibody (10866‐1‐AP; Proteintech) at a dilution of 1:100.

Techniques: Expressing, In Vitro, Concentration Assay, Incubation, Enzyme-linked Immunosorbent Assay