Journal: Cell Reports Medicine

Article Title: Ongoing evolution of SARS-CoV-2 drives escape from mRNA vaccine-induced humoral immunity

doi: 10.1016/j.xcrm.2024.101850

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Article Snippet: pTwist-SARS-CoV-2 Spike-ΔC18 L452Q (CAA) , This study , Addgene Plasmid ID 212446.

Techniques: Virus, Recombinant, Saline, Modification, Plasmid Preparation, Software, Control

Journal: Frontiers in oncology

Article Title: Identification of Hub Genes Correlated With Poor Prognosis for Patients With Uterine Corpus Endometrial Carcinoma by Integrated Bioinformatics Analysis and Experimental Validation.

doi: 10.3389/fonc.2021.766947

Figure Lengend Snippet: FIGURE 10 | Expression and functional validation of the selected hub mRNAs and lncRNAs in vitro. (A) The expression patterns of these genes in two endometrial cancer cell lines, including Ishikawa (histological grade 1; G1) and KLE (histological grade 3; G3). (B–E) The biological impacts of suppressing AURKA with Alisertib (MLN8237) in UCEC cells in vitro. Alisertib decreased Ishikawa cells viability in a dose-dependent manner, induced G2/M phase arrest and enhanced cellular apoptosis. Furthermore, Alisertib limited the long-term clonogenic survival (1 mM). (F) DUXAP8-shRNA/GFP lenti-virus (versus control virus) was used to knock down DUXAP8. (G, H) Downregulation of DUXAP8 inhibited the colony formation and impaired cells growth. Data are the mean ± SD of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: AURKA Inhibitor Treatment and Cell Viability Alisertib (MLN8237) was purchased from Selleck, dissolved in dimethyl sulfoxide (DMSO) and kept frozen at -20°C.

Techniques: Expressing, Functional Assay, Biomarker Discovery, In Vitro, shRNA, Virus, Control, Knockdown

Journal: Cell Reports Medicine

Article Title: Unraveling AURKB as a potential therapeutic target in pulmonary hypertension using integrated transcriptomic analysis and pre-clinical studies

doi: 10.1016/j.xcrm.2025.101964

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Article Snippet: Mouse anti-AURKA (clone 1F4B10) , Proteintech , Cat# 66757-1-Ig; RRID: AB_2882103.

Techniques: Virus, Plasmid Preparation, Recombinant, Negative Control, Protease Inhibitor, Staining, EdU Assay, TUNEL Assay, Western Blot, SYBR Green Assay, Chromatin Immunoprecipitation, Magnetic Beads, RNA Sequencing Assay, Expressing, Software

Journal: Cell reports

Article Title: BCL-xL inhibition potentiates cancer therapies by redirecting the outcome of p53 activation from senescence to apoptosis

doi: 10.1016/j.celrep.2022.111826

Figure Lengend Snippet: (A) Schematic of the drug screen design. (B) Percentages of dead PI-positive cells in cells treated for 24h with indicated drugs in the presence or absence of BCLi navitoclax. Median values from 6 quantified images are shown per each condition. N=6. (C) Representative microscopy images from the library screen shown in B. Numbers in the top left corners indicate the percentages of apoptotic cells. Scale bar 275μm. (D) Identification of drug screen hits. Drugs that, combined with BCLi, induced at least double the percentage of cell death induced by BCLi alone (indicated by the punctate line) were designated as “BCLi-enhanced.”, and all others as “Unaffected.” Statistics using 2-way ANOVA with Sidak’s multiple comparison test. (E) Pairwise comparison of the percentages of senescent cells with and without BCLi addition. Statistical analysis using paired t-test. (F) Comparison of the percentages of senescent cells induced by BCLi-enhanced and unaffected drugs with and without BCLi. Statistics as in D. (G) Drug categories enriched in BCLi-enhanced and unaffected drug groups. (H) Representative images of crystal violet-stained melanoma cells treated with 1μM navitoclax (BCLi), 1μM alisertib (AURKAi), or both drugs combined for 24 h. Scale bar 200 μm. (I) Cell count from the experiment in H. Data is shown as percentages of cells relative to vehicle control. Statistical comparisons using one-way ANOVA with Dunnett’s post-test. N=3. (J) Representative images of crystal violet-stained A375 cells after 24h treatment with vehicle, 1μM navitoclax (BCLi), 1μM paclitaxel, or both drugs combined for 24 h. Scale bar 50μm. Cell quantification relative to vehicle-treated cells is shown on right. Statistical comparisons using one-way ANOVA with Tukey’s test. N=9 random fields from 3 individual wells (3 images/well). (K) Representative images of crystal violet-stained A375 cells after 24h treatment with 1μM navitoclax (BCL-2/xLi), 1μM venetoclax (BCL-2i), 1μM A-1155463 (BCL-xLi), 1μM A-1210477 (MCL-1i) in the presence or absence of 1μM alisertib (AURKAi). Scale bar 150μm. (L) Relative cell numbers from the experiment in K. Data are presented in percentages relative to untreated cells (Vehicle/-) with statistical analysis using 2-way ANOVA with Sidak’s post-test. Experiment was repeated three times with consistent results. (M) Left: representative images of crystal violet-stained Hs294T cells transfected with AURKA or non-targeting siRNA and treated with 1μM navitoclax (BCL-2/xLi) for 24h. Scale bar 275μm. Experiment was performed with three biological replicates. Middle: quantification from 6 random fields per condition and one-way ANOVA statistical analysis with Sidak’s post-test. Right: efficiency of target knockdown determined by western. (N) Same as M, except BCL-xL-targeting siRNA was used and cells were treated with 1μM alisertib (AURKAi) for 24h. All panels: Ns P > 0.05, * P≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001. When applicable, p-values were adjusted for multiple comparisons. See also Fig. S1.

Article Snippet: Silencer AURKA siRNA , Cell signaling , 8883S.

Techniques: Microscopy, Comparison, Staining, Cell Counting, Control, Transfection, Knockdown, Western Blot

Journal: Cell reports

Article Title: BCL-xL inhibition potentiates cancer therapies by redirecting the outcome of p53 activation from senescence to apoptosis

doi: 10.1016/j.celrep.2022.111826

Figure Lengend Snippet: (A) Western blot of p21 in A375 melanoma cells treated with 1μM navitoclax (BCL-2/xLi), 1μM venetoclax (BCL-2i), 1μM A-1155463 (BCL-xLi), 1μM A-1210477 (MCL-1i) in the presence or absence of 1μM alisertib (AURKAi) for 24hrs. Experiment was repeated three times with consistent results. (B) Real-time PCR of relative CDKN1A (p21) mRNA expression in A375 cells after 24hrs of treatment with vehicle, 1μM navitoclax (BCLi), 1μM alisertib (AURKAi), or combination of both drugs. The experiment was repeated three times with 2 replicates each. Statistical comparison using one-way ANOVA with Tukey’s post- test. (C) Western blot analysis of p21 degradation. A375 melanoma cells were treated as in B for 16hrs followed by the addition of Cycloheximide (150μg/ml) ± MG-132 (1μM). Cell lysates were collected at indicated time points (0.25h – 4h). P21 bands from 3 independent experiments were quantified using densitometry and normalized to corresponding actin bands and plotted on the graph below. The square root transformation was used to correct for heteroscedasticity. Statistical analysis using 2-way ANOVA with Tukey’s post-test. (D) Western blot in A375 cells treated as in B for 24hrs in the absence or presence of pan-caspase inhibitor Z-VAD-FMK (20 μg/ml). Three independent experiments were performed with consistent results. (E) Western blot analysis of p21 knockdown. (F) Representative images of Hs294T cells transfected with p21-targeting or non-targeting siRNA, treated with vehicle or 1μM alisertib (AURKAi) for 24hrs, and stained with Hoechst and PI. Scale bar 50μm. Experiment was performed with 3 biological replicates. Right: data quantification from 6 random fields (n=6) and statistics by 2-way ANOVA with Sidak’s post-test. (G) Western blot analysis of cleaved PARP in Hs294T cells treated as in F. (H) Cristal violet staining in Hs294T cells treated as in F. Statistical analysis as in F. (I) Representative images and quantification of crystal violet-stained A375 cells treated as in B for 24hrs in serum-containing (+ serum, control culture condition) and serum-free (- serum, cell cycle-arresting conditions) media. Scale bar 200μm. Statistical comparisons using one-way ANOVA with Sidak’s post-test. N=3 biological replicates. (J) Western blot analysis of cleaved caspase 3 in A375 cells treated as described in I. Experiment was repeated twice with consistent results. (K) Western blot of BAX protein expression in A375 cells treated with vehicle, 1μM navitoclax (BCLi), 1μM alisertib (AURKAi), or their combination for 16hrs. Three independent experiments were performed with consistent results. (L) Real-time PCR analysis of BAX mRNA in A375 cells treated as in B for 24hrs. Experiment was repeated two times with two replicates each. Statistical analysis using one-way ANOVA with Tukey’s post-test. N=4. (M) Representative images of crystal violet-stained A375 cells transfected with non-targeting and BAX-targeting siRNA. Scale bar 150μm. Two independent experiments were performed with consistent results. Right: quantified cell numbers and statistical comparison using one-way ANOVA with Sidak’s post-test. N=6. (N) Western blot for BAX siRNA efficiency. (O-P) Same as M and N, except BAK-specific siRNA was used. All panels: Ns - P > 0.05, * - P≤ 0.05, ** - P ≤ 0.01, *** - P ≤ 0.001, **** - P ≤ 0.0001. P-values were adjusted for multiple comparisons. See also Fig. S6.

Article Snippet: Silencer AURKA siRNA , Cell signaling , 8883S.

Techniques: Western Blot, Real-time Polymerase Chain Reaction, Expressing, Comparison, Transformation Assay, Knockdown, Transfection, Staining, Control

Journal: Cell reports

Article Title: BCL-xL inhibition potentiates cancer therapies by redirecting the outcome of p53 activation from senescence to apoptosis

doi: 10.1016/j.celrep.2022.111826

Figure Lengend Snippet: (A) RNAseq of tumors shown in Fig. 4A. The heat map of genes modulated by either combined AURKAi and BCLi treatment or corresponding individual treatments. N=3. Rows are centered; unit variance scaling is applied to rows. Both rows and columns are clustered using correlation distance and average linkage. (B) Results of the principal component analysis of gene expression data sin A. (C) Top 10 KEGG pathways enriched within the genes differentially expressed after combo (top panel) and alisertib (bottom panel) treatments. Log10-transformed Benjamini-adjusted p-values were plotted. (D) The heat map shows relative mRNA expression of genes within the KEGG p53 signaling pathway across. “Row max” corresponds to the highest expression value of a protein in this row across all tested samples. “Row min” is expression in the sample with lowest expression value. (E) Western blot analysis of p53 protein in melanoma cells treated with vehicle, 1μM navitoclax (BCLi), 1μM alisertib (AURKAi) or combination for 24 hrs. Three independent experiments were performed with consistent results. (F) Western blot shows the efficiency of p53 knockdown/knockout. (G) Representative images and quantificationof crystal violet-stained Hs294T melanoma cells transfected with p53 siRNA or non-targeting siRNA and treated as described in E. Scale bar 200μm. N=3. Statistical analysis using one-way ANOVA with Sidak’s post- test. Two independent experiments were performed with consistent results. (H) Same as G, except A375 cells were used. (I) Representative images and quantification of crystal violet-stained HCT-116 wild-type and p53-null cells treated as in E. Scale bar 200μm. Statistical analysis as in G. (J) Representative images and quantification of Sk-Mel-28 cells with mutated TP53 treated as in E. Scale bar 200μm. Statistical analysis using one-way ANOVA with Dunnett’s test. (All panels) Ns - P > 0.05, * - P≤ 0.05, ** - P ≤ 0.01, *** - P ≤ 0.001, **** - P ≤ 0.0001. P-values were adjusted for multiple comparisons.

Article Snippet: Silencer AURKA siRNA , Cell signaling , 8883S.

Techniques: Gene Expression, Transformation Assay, Expressing, Western Blot, Knockdown, Knock-Out, Staining, Transfection

Journal: Cell reports

Article Title: BCL-xL inhibition potentiates cancer therapies by redirecting the outcome of p53 activation from senescence to apoptosis

doi: 10.1016/j.celrep.2022.111826

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Silencer AURKA siRNA , Cell signaling , 8883S.

Techniques: Virus, Recombinant, Modification, Staining, Membrane, Enzyme-linked Immunosorbent Assay, cDNA Synthesis, Transfection, Saline, Protein Array, Control, Software

Journal: Molecular Cell

Article Title: BUB1 and CENP-U, Primed by CDK1, Are the Main PLK1 Kinetochore Receptors in Mitosis

doi: 10.1016/j.molcel.2020.10.040

Figure Lengend Snippet:

Article Snippet: anti-pT232 Aurora B rabbit polyclonal , Rockland , Cat#660-401-677; RRID: AB_2061641.

Techniques: Affinity Purification, Virus, Recombinant, Transfection, Labeling, Protease Inhibitor, Chromatography, Staining, Software