Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: Synthetic Lethal Screens Reveal Co-Targeting FAK and MEK as a Multimodal Precision Therapy for GNAQ -Driven Uveal Melanoma

doi: 10.1158/1078-0432.CCR-20-3363

Figure Lengend Snippet: (A) OMM 1.5 cells expressing Cas9 were infected with the Brunello Human Kinome CRISPR sgRNA KO library at a MOI of 0.3. After selection, cells were treated with vehicle or 0.5μM VS-4718 (FAKi) for 10 days. (B) Left, Cell viability represented as fold change in FAKi-treated cells compared to control. Highlighted significant hits represent synthetic lethal genes with FAKi treatment. Right, KEGG pathways analysis for the top depleted sgRNAs (n=200). (C) 92.1 cell viability after 72h treatment with vehicle, 1μM Go-6983 (PKCi), 1μM VS-4718 or combination of both. (D) Time-course analysis of FAK and ERK phosphorylation in 92.1 cells treated with VS-4718 (1μM), Go-6983 (1μM) or trametinib (MEKi, 10nM). (E) Quantification of pFAK/FAK and pERK/ERK ratios in 92.1 cells treated with 1μM VS-4718 or vehicle for 1h. (F) Left, Cell viability after 72h treatment with VS-4718 (1μM) in 92.1 cells expressing or not MEK-DD (S218/222D). Right, Immunoblot showing pERK levels in 92.1 cells expressing or not MEK-DD (S218/222D). (C, E and F) Data shown represent the mean ± SEM of three independent experiments. ***p<0.001; **p< 0.01; n.s. not significant.

Article Snippet: FAK, phospho-FAK, ERK, phospho-ERK, cleaved PARP1, cleaved Caspase 3, YAP, Cas9, and GAPDH antibodies were purchased from Cell Signaling.

Techniques: Expressing, Infection, CRISPR, Selection, Western Blot

Journal: Open Medicine

Article Title: Long non-coding RNA TUG1 aggravates cerebral ischemia and reperfusion injury by sponging miR-493-3p/miR-410-3p

doi: 10.1515/med-2021-0253

Figure Lengend Snippet: Knockdown of TUG1 hindered OGD/R-mediated decrease in viability and increase in apoptosis in N2a cells in vitro . (a) The transfection efficiency of si-TUG1 (si-TUG1 #1 , si-TUG1 #2 ) was assessed by RT-qPCR in N2a cells. (b) The effect of si-TUG1 #2 on cell viability was evaluated by CCK-8 assay in N2a cells treated by OGD/R. (c) Flow cytometry assay was applied to detect the apoptosis rate in N2a cells treated by OGD/R. (d and e) The proteins levels of apoptosis-related markers (cleaved-cas-3 and cleaved-cas-9) were performed by western blot assay in N2a cells treated by OGD/R. * P < 0.05.

Article Snippet: The antibodies were cleaved-cas-3, cleaved-cas-9, p-JNK and JNK, p-p38, p38 (1:1,000; Cell Signaling, Danvers, MA, USA), GAPDH (1:500; Santa Cruz, Dallas, TX, USA) and HRP-conjugated secondary antibodies (1:5,000; Southern-Biotech, Birmingham, AL, USA).

Techniques: In Vitro, Transfection, Quantitative RT-PCR, CCK-8 Assay, Flow Cytometry, Western Blot

Journal: Open Medicine

Article Title: Long non-coding RNA TUG1 aggravates cerebral ischemia and reperfusion injury by sponging miR-493-3p/miR-410-3p

doi: 10.1515/med-2021-0253

Figure Lengend Snippet: The inhibition of miR-493-3p or miR-410-3p could restore the reverse effects of TUG1 knockdown on OGD/R-mediated decrease in viability and increase in apoptosis in N2a cells in vitro . (a and b) The effect of si-TUG1 #2 and anti-miR-493-3p or anti-miR-410-3p on the expression of miR-493-3p or miR-410-3p was detected by RT-qPCR. (c) The effect of si-TUG1 #2 and anti-miR-493-3p or anti-miR-410-3p on cell viability was evaluated by CCK-8 assay in N2a cells treated by OGD/R. (d) Flow cytometry assay was applied to detect the apoptosis rate in N2a cells, which transfected with si-TUG1 #2 + anti-miR-493-3p or si-TUG1 #2 + anti-miR-410-3p or negative controls, with or without being treated by OGD/R. (e and f) The proteins levels of apoptosis-related markers (cleaved-cas-3 and cleaved-cas-9) were performed by western blot assay in N2a cells, which transfected with si-TUG1 #2 + anti-miR-493-3p or si-TUG1 #2 + anti-miR-410-3p or negative controls, with or without being treated by OGD/R. * P < 0.05.

Article Snippet: The antibodies were cleaved-cas-3, cleaved-cas-9, p-JNK and JNK, p-p38, p38 (1:1,000; Cell Signaling, Danvers, MA, USA), GAPDH (1:500; Santa Cruz, Dallas, TX, USA) and HRP-conjugated secondary antibodies (1:5,000; Southern-Biotech, Birmingham, AL, USA).

Techniques: Inhibition, In Vitro, Expressing, Quantitative RT-PCR, CCK-8 Assay, Flow Cytometry, Transfection, Western Blot