Journal: International Journal of Oncology

Article Title: Anti-silencing function 1B promotes the progression of pancreatic cancer by activating c-Myc

doi: 10.3892/ijo.2022.5456

Figure Lengend Snippet: Silencing of ASF1B inhibits pancreatic cancer cell proliferation, migration and invasion, and induces apoptosis. (A) PANC-1 and SW1990 cells were transfected with ASF1B siRNAs (si-ASF1B-1 and si-ASF1B-2) or siNC. Protein levels of ASF1B were measured by western blotting. (B) Cell proliferation, (C) colony formation, (D) apoptosis, and (E) expression levels of Bax and Bcl-2 were detected by Cell Counting Kit-8 assays, colony formation assays, flow cytometry and western blotting, respectively. (F) PANC-1 and (G) SW1990 cells were transfected with ASF1B siRNAs (si-1 and si-2) or siNC. Cell migration and invasion were detected by transwell assays. Magnification, ×100; scale bar, 100 µ m. The wound distance of (H) PANC-1 and (I) SW1990 cell cultures was detected using a scratch test. Magnification, ×40; scale bar, 200 µ m. Data are presented as the mean ± SD (n=3). * P<0.05, ** P<0.01, *** P<0.001 vs. siNC group. Data in (B) were analyzed using two-way ANOVA followed by Bonferroni's post hoc test. Other data were analyzed using one-way ANOVA followed by Tukey's post hoc test. Brown-Forsythe and Bartlett's tests were used to assess the normality and variance homogeneity. ASF1B, anti-silencing function 1B; NC, negative control; OD, optical density; siRNA/si, small interfering RNA.

Article Snippet: After 48 h, apoptotic cells were stained using the Annexin V-FITC Early Apoptosis Detection Kit (Cell Signaling Technology, Inc.) according to the manufacturer's instructions.

Techniques: Migration, Transfection, Western Blot, Expressing, Cell Counting, Flow Cytometry, Negative Control, Small Interfering RNA

Journal: Journal of Oncology

Article Title: Plumbagin Enhances the Anticancer Effects of PF Chemotherapy via Downregulation of the PI3K/AKT/mTOR/p70S6K Pathway in Human Tongue Squamous Cell Carcinoma

doi: 10.1155/2023/8306514

Figure Lengend Snippet: PB enhanced the proapoptotic effects of PF in TSCC cells. (a) Results of the flow cytometry assay demonstrated apoptosis in Cal27 and Cal27/CDDP cells following the treatment with PB and/or PF. (b-c) A histogram of the levels of apoptosis in Cal27 and Cal27/CDDP cells. (d) Representative blots of the proapoptotic protein Bax and Bad and antiapoptotic protein Bcl-2 and Bcl-xL in Cal27 and Cal27/CDDP cells after the treatment of PB and/or PF. (e) Ratio of Bax/Bcl-2 in Cal27 and Cal27/CDDP cells. (f-g) Histogram of the expression levels of Bcl-xL and Bad. Independent biological experiments were repeated three times, and statistical significance is denoted by ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001 and ∗∗∗∗ P < 0.0001. PB, plumbagin; PF, cisplatin plus 5-fluorouracil; and TSCC, tongue squamous cell carcinoma.

Article Snippet: Cell cycle kits were bought from Lianke Biological Technology Co. AnnexinV-FITC/propidium iodide apoptosis detection kits were purchased from Beibo Co. Primary antibodies against Bcl-2, Bcl-xL, Bax, Bad, PI3K, AKT, phosphorylated (p)-AKT, mTOR, p-mTOR, p70S6K, and p-p70S6K were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Flow Cytometry, Expressing

Journal: Life Science Alliance

Article Title: Trans-differentiation of trophoblast stem cells: implications in placental biology

doi: 10.26508/lsa.202201583

Figure Lengend Snippet: (A, B, C) Quantitative real-time PCR (qRT) analysis of the differentially expressed endothelial cell–specific genes in trophoblast stem cells (TS) and differentiated trophoblast cells (TC). qRT–PCR results are grouped based on functional annotations. (A) Angiogenesis-related genes: Cx3cl1,c-kit , Kdr , Plau , Mmp . (B) Cell adhesion molecules: Cdh5 , Pecam1 , Itg β3, Col18a1 . (C) Apoptosis-related genes: Tnfsf10 , Bcl2 , Cradd , Casp3 . Data are representative of three biological replicates, and error bars represent SEM. * P < 0.5, ** P < 0.01. Source data are available for this figure.

Article Snippet: After 48 and 72 h, co-cultured cells were processed either for both annexin V–PI staining using early apoptosis detection kit (cat no. 6592; Cell Signaling Technology) by flow cytometry or for protein isolation and Western blotting for detection of apoptosis markers.

Techniques: Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Functional Assay

Journal: Frontiers in Immunology

Article Title: Attenuated Salmonella potentiate PD-L1 blockade immunotherapy in a preclinical model of colorectal cancer

doi: 10.3389/fimmu.2022.1017780

Figure Lengend Snippet: PD-1 and PD-L1 are expressed in MC38 tumors. Representative flow cytometric dot plots showing the expression of PD-L1 (A, C) and PD-1 (B, D) on MC38 tumor cells grown in vitro (A, B) or on single cell suspension of dissociated tumor tissues excised from mice 21 post subcutaneous implantation (C, D) . Immunohistochemical staining was also used to illustrate the expression of PD-L1 (E) and PD-1 (F) on MC38 tumor tissue sections. Magnification 400×. Scale bar 20 μm.

Article Snippet: All immunohistochemical studies were done using the described protocol except for cleaved caspase-3 staining which was done following the protocol of IHC Detection Kit of the manufacturer (Cell Signaling Technology; #12692).

Techniques: Expressing, In Vitro, Immunohistochemical staining, Staining

Journal: Frontiers in Immunology

Article Title: Attenuated Salmonella potentiate PD-L1 blockade immunotherapy in a preclinical model of colorectal cancer

doi: 10.3389/fimmu.2022.1017780

Figure Lengend Snippet: Combination treatment increases MC38 tumor infiltration by CD4 + and CD8 + T cells and the expression of their effector molecules. MC38 tumor tissues were resected from non-treated, αPD-L1 and combination-treated mice on day 21 post implantation for further analyses using immunohistochemistry, flow cytometry and qRT-PCR. Representative immunohistochemical images for CD4 (A) and CD8 (C) in tumor tissues are presented for the control and combination-treated groups. Magnification 400×. Scale bar 20 μm. CD4 + (B) and CD8 + (D) cells were quantified in 15 HPF for control, αPD-L1 and combination-treated groups. Each data point represents the average of positive cells/HPF from a single mouse, pooled from 2 independent experiments. Representative flow cytometric dot plots and the combined result analyses for the percentages of CD4 + (E, F) and CD8 + (E, G) cells in MC38 tumors are illustrated. RNA was extracted from total MC38 tumor tissues and gene expression levels were determined using qRT-PCR. The effect of combination treatment on the expression levels of CXCL9 (H) , CXCL10 (I) , IFN-γ (J) and granzyme B (K) was assessed. Representative images for granzyme B staining in tumor tissues are presented for each group (L) . Graph depicts the number of cells/HPF (M) . Magnification 400×. Scale bar 20 μm. Each data point represents a single mouse pooled from two independent experiments. Asterisks denote statistically significant differences from control group, * (P ≤ 0.05), ** (P ≤ 0.01), *** (P ≤ 0.001) and **** (P ≤ 0.0001).

Article Snippet: All immunohistochemical studies were done using the described protocol except for cleaved caspase-3 staining which was done following the protocol of IHC Detection Kit of the manufacturer (Cell Signaling Technology; #12692).

Techniques: Expressing, Immunohistochemistry, Flow Cytometry, Quantitative RT-PCR, Immunohistochemical staining, Staining

Journal: Frontiers in Immunology

Article Title: Attenuated Salmonella potentiate PD-L1 blockade immunotherapy in a preclinical model of colorectal cancer

doi: 10.3389/fimmu.2022.1017780

Figure Lengend Snippet: Combination treatment of Salmonella and αPD-L1 induces apoptosis more efficiently than monotherapy. Representative immunohistochemical images for cleaved caspase-3 in tumor tissues are presented for control, Salmonella , αPD-L1 and combination-treated groups (A) . Magnification 400×. Scale bar 20 μm. Cleaved caspase 3 + cells were quantified in 15 HPF for the different groups (B) . Each data point represents the average of positive cells from a single mouse, pooled from 2 independent experiments for all groups except for the control group, from three independent experiments. Asterisks denote statistically significant differences from control group, * (P ≤ 0.05), ** (P ≤ 0.01), *** (P ≤ 0.001) and ns (no statistical significance, ≥ 0.05).

Article Snippet: All immunohistochemical studies were done using the described protocol except for cleaved caspase-3 staining which was done following the protocol of IHC Detection Kit of the manufacturer (Cell Signaling Technology; #12692).

Techniques: Immunohistochemical staining

Journal: Oxidative Medicine and Cellular Longevity

Article Title: The Protective Effects of Trypsin Inhibitor on Hepatic Ischemia-Reperfusion Injury and Liver Graft Survival

doi: 10.1155/2016/1429835

Figure Lengend Snippet: Hepatocyte apoptosis, Caspase-3 , Bcl-2, and Bax gene expression on liver tissue. (a) Hepatocyte apoptosis TUNEL evaluation revealed a significantly reduced number of hepatocellular apoptotic nuclei treated with UTI at 6 hours, especially in the combined treatment group (48.2 ± 1.9 versus 40.4 ± 2.2 versus 26.6 ± 2.9), ∗ P < 0.05. (b) In terms of percentage calculation, three specimens in each group were stained under a 400x light microscope and 4–10 positive fields were observed in each section. The mean ratio of positive cells (brown-yellow stained cells) accounting for total cell count in each group was taken. (c) Caspase-3 and Bax gene expression on liver tissue was reduced, but Bcl-2 was increased significantly in the UTI-treated group, tested by Western blot. (d) The ratios of Bax/Bcl-2 were 1.5 ± 0.3 versus 0.9 ± 0.2 versus 2.5 ± 0.5, 2.5 ± 0.4 versus 1.8 ± 0.2 versus 3.3 ± 0.2, and 2.3 ± 0.3 versus 1.7 ± 0.1 versus 2.9 ± 0.5. Cleaved-caspase-3/beta-actin was 0.4 ± 0.04 versus 0.2 ± 0.02 versus 0.5 ± 0.02, 0.7 ± 0.04 versus 0.5 ± 0.03 versus 0.8 ± 0.03, and 0.5 ± 0.04 versus 0.3 ± 0.04 versus 0.6 ± 0.03 in the LR + UTI, LR + UTI + RT, and LR groups at 1 h, 6 h, and 18 hours after OLTx, respectively. ( ∗ P < 0.05).

Article Snippet: Rat anti-mouse IL-6, TNF- α , and IFN- γ antibodies were from BD Pharmingen (San Diego, CA, USA), rat anti-mouse IL-10 antibody was from Abcam (Cambridge, UK), Bcl-2, caspase-3, and Bax, β -actin were from Cell Signaling Technology, Inc., (Danvers, MA, USA), and the ApopTag Peroxidase In Situ Apoptosis Detection Kit was from the Millipore Corporation (Billerica, MA, USA).

Techniques: Expressing, TUNEL Assay, Staining, Light Microscopy, Cell Counting, Western Blot

Journal: PLoS ONE

Article Title: The Antidiabetic Drug Ciglitazone Induces High Grade Bladder Cancer Cells Apoptosis through the Up-Regulation of TRAIL

doi: 10.1371/journal.pone.0028354

Figure Lengend Snippet: Mice were inoculated subcutaneously with exponentially growing T24 and RT4 bladder cancer cells (5×10 6 cells). When tumor size reached 40 mm 3 in volume, mice were randomly divided into control and treated groups (n = 10). Intraperitoneal injections of ciglitazone were weekly administered at a dose of 15 mg/kg during three weeks. Control animals received only saline vehicle following an identical schedule. (A) The growth tumor curves were determined by measuring the tumor volume. * P <0.05, significant differences compared with untreated animals with the use of two-way ANOVA test (evaluation of the tumor volume development over time) ; ** P <0.05, significant differences between control and treated mice at each post-graft time with the use of two-tailed unpaired Student's t test. (B) Immunohistochemical staining of representative paraffin-embedded sections of tumors from untreated or ciglitazone-treated mice. Sections were fixed and stained for Ki-67 and active caspase 3. Each panel is representative of 5 sections for each of ten tumors from control and ciglitazone-treated mice. Original magnification, ×20.

Article Snippet: Activated caspase 3 was detected with Apoptosis marker: SignalStain cleaved Caspase 3 (Asp175) IHC Detection kit (Cell Signaling).

Techniques: Two Tailed Test, Immunohistochemical staining, Staining