Journal: Aging (Albany NY)

Article Title: The protective effects of Phoenixin-20 in tumor necrosis factor α (TNF-α)-induced cell senescence of rheumatoid arthritis fibroblast-like synoviocytes (FLS)

doi: 10.18632/aging.205024

Figure Lengend Snippet: Phoenixin-20 downregulated p53 and p21 in TNF-α-treated RA-FLSs. RA-FLSs were stimulated with TNF-α (10 ng/mL) with or without Phoenixin-20 (10, 20 nM) for 7 days. ( A ) mRNA of p53 and p21; ( B ) Protein of p53 and p21 ( ※ P < 0.01 vs. vehicle group; # , ## , P < 0.05, 0.01 vs. TNF-α group, N = 5–6).

Article Snippet: The primary antibodies against p53 (#AB_2860544, 1:2000, Sino Biological, China), p21 (1:1000, #ab109520, Abcam, USA), STAT6 (#ab32108, 1:1500, Abcam, USA), and β-actin (1:10000, #ab6276, Abcam, USA) were imported.

Techniques:

Journal: Aging (Albany NY)

Article Title: The protective effects of Phoenixin-20 in tumor necrosis factor α (TNF-α)-induced cell senescence of rheumatoid arthritis fibroblast-like synoviocytes (FLS)

doi: 10.18632/aging.205024

Figure Lengend Snippet: Overexpression of STAT6 abolished the protective effect of Phoenixin-20 on TNF-α-induced cellular senescence in RA-FLSs. Cells were transduced with lentiviral STAT6, followed by stimulation with TNF-α (10 ng/mL) and Phoenixin-20 (20 nM) for 7 days. ( A ) Western blot analysis revealed successful overexpression of STAT6; ( B ) mRNA of p53 and p21. ( C ) Cellular senescence was assayed using SA-β-gal staining ( ※ P < 0.01 vs. vehicle group; ## P < 0.01 vs. TNF-α group, * P < 0.05 vs. PNX-20 group, N = 5–6).

Article Snippet: The primary antibodies against p53 (#AB_2860544, 1:2000, Sino Biological, China), p21 (1:1000, #ab109520, Abcam, USA), STAT6 (#ab32108, 1:1500, Abcam, USA), and β-actin (1:10000, #ab6276, Abcam, USA) were imported.

Techniques: Over Expression, Transduction, Western Blot, Staining

Journal: bioRxiv

Article Title: Genetic Impairment of Succinate Metabolism Disrupts Bioenergetic Sensing in Adrenal Neuroendocrine Cancer

doi: 10.1101/2022.01.09.475410

Figure Lengend Snippet: Cdk5 in induces senescence–like phenotypic characteristics. Immunoblots quantification showing time dependent effect of Cdk5 in ( i.e MRT3-007) on phosphorylation states or levels of (A) P-S15 p53, (B) p16 INK4a ,(C) p27 Kip , (D) P-S139 H2AX. Values presented as fold change normalized with time=0. n=3, data are means ± S.E,M. (E) Scanning electron microscopy (SEM) images of hPheo1 cell morphology in control vs. Cdk5 in (Indolinone A). (F) Imaging of proliferating cells and those treated with Cdk5 in (MRT3-007, 20 nM, 48 h) for common senescent markers. Representative confocal photomicrographs and quantitation shows phalloidin stain of F-actin, p16 INK4a (inset is senescence-associated-β-galactosidase; β-gal), p27 Kip and P-H2Ax (S139) respectively; scale=136 µM. (G) Representative DNA histogram (left) showing cell cycle profile of KO cells treated with vehicle or Cdk5 in (20 nM); histograms analysis by MODfit LT 3.0 (right) presents cell cycle distribution in G1 (red), early-S (yellow), and late-S/G2/M (green); n=2, Data are means ± SD. (H) Confocal images of Ki67 staining and bar graph show number of Ki67- positive cells.

Article Snippet: Antibodies to the following phosphorylation sites and proteins were used: phospho-Thr172 AMPK (#2535), phospho-Ser79 ACC (#11818), phospho-Ser21/9 GSK3 (#9223), phospho-Ser16 p53 (#82530), p27Kip1(#3686), phospho-Ser139 H2Ax (#80312) from Cell Signaling Technology; anti-p16INK4A (Proteintech#10883-1-AP), anti-spectrin (Millipore Sigma#MAB1622) and, anti- GAPDH (#ZG003), anti-actin (#PA5-78715) from Thermofischer Scientific.

Techniques: Western Blot, Phospho-proteomics, Electron Microscopy, Control, Imaging, Quantitation Assay, Staining

Journal: bioRxiv

Article Title: Genetic Impairment of Succinate Metabolism Disrupts Bioenergetic Sensing in Adrenal Neuroendocrine Cancer

doi: 10.1101/2022.01.09.475410

Figure Lengend Snippet: Inhibition of p53 or AMPK abrogate effects of Cdk5 in (MRT3-007). (A) Representative blots and densitometry detects P-p53 (S16) and p27 Kip upon Cdk5 in treatment in presence or absence of P53 inhibitor PFT-α (10 µM, 10 h pre-treatment). (B) Representative phase contrast microphotographs showing effects of AMPK inhibitor, Compound C (CC, 10 µM) on Cdk5 in induced morphological changes. (C) Representative histograms and quantitation indicating cell cycle profile of KO cells expressing either WT, S65D or S65A phosphomutants of PRKAG2.

Article Snippet: Antibodies to the following phosphorylation sites and proteins were used: phospho-Thr172 AMPK (#2535), phospho-Ser79 ACC (#11818), phospho-Ser21/9 GSK3 (#9223), phospho-Ser16 p53 (#82530), p27Kip1(#3686), phospho-Ser139 H2Ax (#80312) from Cell Signaling Technology; anti-p16INK4A (Proteintech#10883-1-AP), anti-spectrin (Millipore Sigma#MAB1622) and, anti- GAPDH (#ZG003), anti-actin (#PA5-78715) from Thermofischer Scientific.

Techniques: Inhibition, Quantitation Assay, Expressing

Journal: bioRxiv

Article Title: Intracellular accumulation of free cholesterol in macrophages triggers a PARP1 response to DNA damage and PARP1 impairs lipopolysaccharide-induced inflammatory response

doi: 10.1101/2024.07.30.605465

Figure Lengend Snippet: (A) Hallmark pathway analysis of bulk RNA-sequencing datasets from cultured PMφs with and without oxLDL accumulation and 6 h of LPS stimulation. The heatmap includes genes that contributed most to the pathway enrichment and expression values are colored to represent enrichment from low (blue) to high (red). (B) Normalized G6PD activity in PMφs ±oxLDL accumulation and 6 h of LPS stimulation (n = 10). (C) Quantification of extracellular and intracellular metabolites related to pyrimidine salvage synthesis pathways in PMφs ±oxLDL accumulation and 6 h of LPS stimulation. Data were generated by LC-MS metabolomics (n=18). (D) Representative immunoblots and quantification of nuclear p53 protein expression in PMφs ±oxLDL accumulation. A LPS stimulation time-course (0 – 6 h) was performed. Data are normalized to the corresponding Lamin A/C and the 0 h LPS time point of the –oxLDL group (assigned a value of 1, n = 3). (E) Quantification of extracellular and intracellular metabolites related to purine salvage synthesis pathways in PMφs ±oxLDL accumulation and 6 h of LPS stimulation. Data were generated by LC-MS metabolomics (n = 18). The mean ± SEM is plotted in all graphs. Significant differences were determined by unpaired two-way Student’s t tests in (B), (C) and (E) , and a two-way ANOVA with Bonferroni post hoc test in (D) (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001).

Article Snippet: Membranes were incubated with primary antibodies overnight: anti-Actin (Sigma, A2066), anti-p53 (CST#32532), anti-Lamin A/C (CST#2032), anti-Mono/Poly-ADP Ribose (CST#83732), anti-PARP1 (CST#9542), anti-pH2A.X (Ser139) (CST#2577), anti-H2A.X (CST#2595), followed by washing and incubation with HRP-conjugated anti-rabbit IgG (CST#7074) (22°C, 1 h).

Techniques: RNA Sequencing, Cell Culture, Expressing, Activity Assay, Generated, Liquid Chromatography with Mass Spectroscopy, Western Blot

Journal: Journal of proteome research

Article Title: Analysis of Stratifin Expression and Proteome Variation in a Rat Model of Acute Lung Injury.

doi: 10.1021/acs.jproteome.4c00980

Figure Lengend Snippet: Figure 6. Western blotting and qRT-PCR analyses of PAI-1, SFN, and p53. (A) Representative Western blot images of the abundance of PAI-1, SFN, and p53-related proteins in the lung tissue extracts from OA-treated rats. (B, D, F) Graphs depicting the quantitation of phosphorylated p53 at Ser15 (pp53-Ser15, B), PAI-1 (Serpine1, D), and SFN (F) by ImageJ software. The abundance of pp53-Ser15 was normalized with that of total p53, and the abundance values of PAI-1 and SFN were normalized with that of β-actin. Data are shown as mean ± SEM (n = 3), *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 vs 0 h using one-way ANOVA and Dunnett’s posthoc test. (C, E) mRNA levels of PAI-1 (Serpine1, C) and SFN (Sfn, E) in lung tissues of OA-treated rats. mRNA levels were quantified using qRT-PCR and normalized with those of Tbp. Data are shown as mean ± SEM (n = 5 or 6). ***p < 0.001, ****p < 0.0001 vs 0 h using two-way ANOVA and Bonferroni’s posthoc test.

Article Snippet: For detecting each protein, the following rabbit monoclonal antibodies (mAb) or polyclonal antibodies (pAb) were used: rabbit anti-plasminogen activator inhibitor-1 (PAI-1) pAb (#27535, Cell Signaling Technology, Leiden, The Netherlands; 1:1000) rabbit anti-p53 mAb (clone E9B5W [#30313, Cell Signaling Technology; 1:2000]), rabbit anti-phospho-p53 Ser15 pAb (#9284, Cell Signaling Technology; 1:2000), and βactin rabbit pAb (#20536−1-A, Proteintech, Rosemont, IL; 1:5000).

Techniques: Western Blot, Quantitative RT-PCR, Quantitation Assay, Software

Journal: Asian Pacific Journal of Cancer Prevention

Article Title: The Association between p53 Expression and Histopathology Grade of Astrocytoma

doi: 10.31557/apjcp.2025.26.7.2521

Figure Lengend Snippet: Figure 1. Immunohistochemistry Staining of p53 Mutant (High Power View, Magnification 400x). (A). Intense nuclear staining in immunopositive in ³10% of tumor cells was interpreted as positive (p53 mutant). (B). Weak nuclear staining was interpreted as negative as immunonegative (p53 wild-type)

Article Snippet: The research samples comprised Formalin-Fixed Paraffin-Embedded (FFPE) for Hematoxylin-Eosin (HE) and immunohistochemical staining with Anti-p53 mutant (M00001-3, Boster, Pleasanton, CA, USA).

Techniques: Immunohistochemistry, Staining, Mutagenesis