Journal: Cell Death & Disease

Article Title: The pharmalogical reactivation of p53 function improves breast tumor cell lysis by granzyme B and NK cells through induction of autophagy

doi: 10.1038/s41419-019-1950-1

Figure Lengend Snippet: a MDA-MB231 cells express high level of mutated p53 (p53 R280K ) lacking transcriptional activity as shown by the absence of p53-dependent transactivation of its target gene p21 even after γ-irradiation. b , c 24-48 h treatment with CP-31398 reactivates p53 transcriptional activity in MDA-MB231 cells as shown by induction of p21 expression at both mRNA ( b ) and protein ( c ) levels. d – f CP-31398-induced apoptosis and ROS production were evaluated after 48 h treatment using AnnexinV/PI ( d , e ) or DHE ( f ) staining. g , h CP-31398 treatment (48 h) increases MDA-MB231 cell susceptibility to NK-mediated lysis. 51 Cr release assay using NK92 ( d ) or NK cells isolated from a healthy donor ( e ) co-cultured with target cells at different E:T ratios are shown. i CP-31398 treatment increases MDA-MB231 cell susceptibility to NK-mediated lysis in a time dependent manner. A representative 51 Cr release assay (from two independent experiments) using NK cells isolated from a healthy donor is shown. j , k PFT-α (an inhibitor of p53 transcriptional activity) inhibits CP-31398-dependent induction of p21 expression in MDA-MB231 cells at both mRNA ( j ) and protein ( k ) levels. l , m PFT-α inhibits the increase of MDA-MB231 lysis by NK cells observed when CP-31398 is used alone. 51 Cr release assay using NK92 ( l ) or NK cells extracted from a healthy donor ( m ) co-cultured with target cells at the E:T ratio of 50:1 or 30:1, respectively, are shown. n – p The knockdown of mutated p53 expression using siRNA inhibits CP-31398-dependent induction of p21 expression in MDA-MB231 cells at both mRNA ( n ) and protein ( o ) levels and abrogate the CP-31398-dependent increase of MDA-MB231 lysis by NK cells ( p ). Data are representative of at least three independent experiments ( a , c , d , k , o ) or are the mean ± s.d. of three ( e – h , l , m , p ) or five ( b , j , n ) independent experiments. The p values (* P < 0.05; ** P < 0.01) were determined by unpaired two-tailed Student's t test ( b , e , g , h , j , l – n ) or one-way ANOVA ( p )

Article Snippet: From Merck: anti-p21 mouse mAb (clone OP64), anti-mdm2 mouse mAb (clone 2A10).

Techniques: Activity Assay, Irradiation, Expressing, Staining, Lysis, Release Assay, Isolation, Cell Culture, Two Tailed Test

Journal: Cell Death & Disease

Article Title: The pharmalogical reactivation of p53 function improves breast tumor cell lysis by granzyme B and NK cells through induction of autophagy

doi: 10.1038/s41419-019-1950-1

Figure Lengend Snippet: a CP-31398 treatment (48 h) increases p21 and Mdm2 expression by MDA-MB231 cells, does not alter Bax level (used as control) and does not induce the degradation of Bcl-X L , XIAP, Bcl-2, Mcl-1, cIAP-1, IAP-2, or survivin. b The differences in the amount of LC3-II between samples in the presence and absence of chloroquine (CQ), representing the amount of LC3 that is degraded into autolysosomes, and allowing the measurement of the autophagic flux is represented. No additional LC3-II accumulation is observed when CP-31398 (48 h) is used in combination with CQ (2–24 h), showing that the autophagic flux induced by CP-31398 is blocked at the autophagosome-lysosome fusion step. c P62 expression level increases following 48 h treatment with CP-31398. d , e MDA-MB231 cells were transduced with LC3 fusion protein tagged with acid‑sensitive eGFP and acid‑insensitive RFP to monitor the specific loss of eGFP fluorescence upon acidification of the RFP + autophagosome following lysosomal fusion. Serum starvation induces autophagy with the loss of eGFP signal reflecting the fusion of autophagosomes with lysosomes. Fluorescence is seen from both eGFP and RFP channels after CP-31398 treatment, similarly to CQ that blocks autophagy through neutralization of lysosomal pH, demonstrating that CP-31398-induced autophagy flux is blocked at the autophagosomes/lysosomes fusion step. Dashed lines: plasma membrane. Scale bars: 10 μm ( c ). The percentage RFP + or RFP + /GFP + autophagosomes was evaluated in 100 cells after the indicated treatment ( d ). f Representative electron microscopy images of the autophagosomes containing undigested material presents in CP-31398 or chloroquine (CQ)-treated cells. Representative images of multivesicular bodies, mitochondria and endoplasmic reticulum are displayed as control. Data are representative of three independent experiments ( a – d , f ) or are the mean ± s.d. of three independent experiments ( e )

Article Snippet: From Merck: anti-p21 mouse mAb (clone OP64), anti-mdm2 mouse mAb (clone 2A10).

Techniques: Expressing, Transduction, Fluorescence, Neutralization, Electron Microscopy

Journal: Current Health Sciences Journal

Article Title: P21 Immunoexpression in Actinic Keratosis and Cutaneous Carcinomas

doi: 10.12865/CHSJ.43.03.07

Figure Lengend Snippet: p21 immunostaining: A. Actinic keratosis (KIN I), x100; B. well differentiated squamous carcinoma, x100; C. moderate differentiated squamous carcinoma, x100; D. poorly differentiated squamous carcinoma, x100

Article Snippet: In this study we use a monoclonal mouse anti-human anti-p21 antibody (Biocare Medical), clone WA-1, diluted as 1:50, with microwaving in citrate buffer pH 6 as antigen retrieval.

Techniques: Immunostaining

Journal: Current Health Sciences Journal

Article Title: P21 Immunoexpression in Actinic Keratosis and Cutaneous Carcinomas

doi: 10.12865/CHSJ.43.03.07

Figure Lengend Snippet: Distribution of the actinic keratosis cases depending of p21 FSS

Article Snippet: In this study we use a monoclonal mouse anti-human anti-p21 antibody (Biocare Medical), clone WA-1, diluted as 1:50, with microwaving in citrate buffer pH 6 as antigen retrieval.

Techniques:

Journal: Current Health Sciences Journal

Article Title: P21 Immunoexpression in Actinic Keratosis and Cutaneous Carcinomas

doi: 10.12865/CHSJ.43.03.07

Figure Lengend Snippet: Distribution of the squamous carcinomas cases depending on p21 FSS

Article Snippet: In this study we use a monoclonal mouse anti-human anti-p21 antibody (Biocare Medical), clone WA-1, diluted as 1:50, with microwaving in citrate buffer pH 6 as antigen retrieval.

Techniques:

Journal: Current Health Sciences Journal

Article Title: P21 Immunoexpression in Actinic Keratosis and Cutaneous Carcinomas

doi: 10.12865/CHSJ.43.03.07

Figure Lengend Snippet: Distribution of the positive p21 carcinomas depending on differentiation degree (A) and tumor stage (B)

Article Snippet: In this study we use a monoclonal mouse anti-human anti-p21 antibody (Biocare Medical), clone WA-1, diluted as 1:50, with microwaving in citrate buffer pH 6 as antigen retrieval.

Techniques: