Journal: Cell Proliferation

Article Title: The inhibitory effects of capillarisin on cell proliferation and invasion of prostate carcinoma cells

doi: 10.1111/cpr.12429

Figure Lengend Snippet: Capillarisin (Cap) modulation of cell cycle and invasion regulatory proteins in DU145 cells. Cells were treated with various concentrations of Cap as indicated for 24 hours. Expression of (A) cyclin D, cyclin A, cyclin B, cyclin E, (C) survivin, (E) matrix metallopeptidase (MMP)2, and MMP9 was determined by immunoblotting assays. The fold‐induction data are expressed as the intensity of the protein bands produced from the target gene/β‐actin (±SE; n = 3) relative to that of the vehicle‐treated group (B,D,F)

Article Snippet: The housekeeping proteins, β‐actin (MAS1501, Millipore, Temecula, CA, USA) and Lamin B (M20, Santa Cruz Biotechnology), were detected as internal controls.

Techniques: Expressing, Western Blot, Produced

Journal: Cell Proliferation

Article Title: The inhibitory effects of capillarisin on cell proliferation and invasion of prostate carcinoma cells

doi: 10.1111/cpr.12429

Figure Lengend Snippet: Capillarisin (Cap) modulation of growth and distribution of LNCaP cells. (A) Cells were treated with various concentrations of Cap at indicated time periods, and cell growth was determined by water‐soluble tetrazolium‐1 (WST‐1) assays. Cell growth of the vehicle‐treated group is set as 100% and data obtained from three independent experiments are expressed as mean ± SE. (B) The cell cycle distribution of LNCaP cells after cells were treated with various concentrations of Cap as indicated for 24 hours. (C) Cells were treated with various concentrations of Cap as indicated for 24 hours. Expression of cyclin D1, cyclin A, p21, and p27 was determined by immunoblotting assays. (D) The fold‐induction data are expressed as the intensity of the protein bands produced from the target gene/β‐actin (±SE; n = 3) relative to that of the vehicle‐treated group

Article Snippet: The housekeeping proteins, β‐actin (MAS1501, Millipore, Temecula, CA, USA) and Lamin B (M20, Santa Cruz Biotechnology), were detected as internal controls.

Techniques: Expressing, Western Blot, Produced

Journal: Cell Proliferation

Article Title: The inhibitory effects of capillarisin on cell proliferation and invasion of prostate carcinoma cells

doi: 10.1111/cpr.12429

Figure Lengend Snippet: Effects of Cap on constitutive and interlukin‐6 (IL‐6) ‐inducible signal transducer and activator of transcription 3 (STAT3) activation in prostate carcinoma DU145 cells. DU145 cells were treated (A) with Cap at indicated concentrations for 24 hours; (B) with 100 μmol L−1 Cap for the indicated period of time. (C) DU145 cells were lysed after treatments with various concentrations of Cap, and then the nuclear and cytoplasmic fractions were separated. The protein levels of pSTAT3 and lamin B in the nuclear fraction were determining by immunoblotting assays. (D) The pSTAT3‐TA‐Luc‐transfected DU145 cells were treated with various concentrations of Cap as indicated for 24 hours. Data are presented as mean percentage ±SE (n = 6) of the luciferase activity in relation to the vehicle‐treated group. (E) DU145 cells were cotreated with 20 ng mL−1 interlukin‐6 (IL‐6) and Cap at indicated concentrations for 24 hours. The protein levels of STAT 3, phosph‐STAT3, matrix metallopeptidase (MMP)2, and MMP9 were analysed by immunoblotting assays. (F) The fold‐induction data are expressed as the intensity of the protein bands produced from the target gene/β‐actin (±SE; n = 3) relative to that of the control group

Article Snippet: The housekeeping proteins, β‐actin (MAS1501, Millipore, Temecula, CA, USA) and Lamin B (M20, Santa Cruz Biotechnology), were detected as internal controls.

Techniques: Activation Assay, Western Blot, Transfection, Luciferase, Activity Assay, Produced

Journal: JID Innovations

Article Title: Unlocking the Mechanisms of Cutaneous Adverse Drug Reactions: Activation of the Phosphatidylinositol 3-Kinase/Protein Kinase B Pathway by EGFR Inhibitors Triggers Keratinocyte Differentiation and Polarization of Epidermal Immune Responses

doi: 10.1016/j.xjidi.2021.100009

Figure Lengend Snippet: Protein array quality check before and after normalization and filtering. ( a ) Box plot depicting the distribution of fluorescence signal before and after QN for the phospho-antibody microarray. ( b ) Unsupervised biclustering for the phosphorylation protein array before and after QN (distance: Pearson’s correlation, aggregation criterion = Ward’s D2). ( c ) Biplots for PCA, before normalization and after normalization and debatching for the phosphorylation protein array. QN followed by the debatching step allows for the capture of time along PC1 and treatment along PC2. AFA, afatinib; h, hour; ID, identification; min, minute; PC, principal component; PCA, principal component analysis; QN, quantile normalization.

Article Snippet: The Phospho Explorer Antibody Array (Full Moon BioSystems, Sunnyvale, CA) was performed by TebuBio (Le Perray-en-Yvelines, France) using 1,330 duplicate spots matching 1,318 proteins (phosphorylated and unphosphorylated), housekeeping proteins (β-actin, GAPDH), negative controls (n = 4), empty spots (n = 4), and positive markers (n = 2).

Techniques: Protein Array, Fluorescence, Microarray, Phospho-proteomics

Journal: The American Journal of Pathology

Article Title: Ketamine-Induced Apoptosis in Normal Human Urothelial Cells

doi: 10.1016/j.ajpath.2015.12.014

Figure Lengend Snippet: Induction of apoptosis in normal (nonimmortalized) human urothelial (NHU) cell cultures by ketamine. A: Western blot analysis of NHU cells separated into cytoplasmic and mitochondrial fractions showed elevated cytochrome c in the cytosolic fractions of cultures treated with 3 mmol/L ketamine for 24 hours. Loading controls were Bcl2 for the mitochondrial fraction and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) for the cytoplasmic fraction. B: Densitometry analysis showed a significant mean 2.4-fold change in cytoplasmic cytochrome c content in three independent NHU cell lines after 24 hours' exposure to 3 mmol/L ketamine. C: Western blot analysis of phospho-Akt (active form), phospho-ERK1/2 (active form), and S9 phospho-glycogen synthase kinase (GSK) 3β (inactive form) showed early depletion of these forms of the kinases in response to 3 mmol/L ketamine. Abundance quantified by densitometry is shown as a percentage of control cells for each time point, normalized to β-actin (combined data). D: Inhibition of GSK3β by SB415286 in ketamine-exposed NHU cells was capable of a slight, but significant, inhibition of toxicity, as assessed by Alamar Blue reduction. E: Inhibition of the mitochondrial permeability transition pore with cyclosporin A (CsA) in ketamine-exposed NHU cells was capable of a small, but significant, inhibition of toxicity, as assessed by Alamar Blue reduction. F: Western blot analysis of caspase 9, caspase 3, and cleaved poly (ADP-ribose) polymerase (PARP) in NHU cells after 72 hours' exposure to 3 mmol/L ketamine. G: The three markers of apoptosis were all significantly increased by more than twofold in densitometry, which was normalized to β-actin. H: Caspase 3/7 activity was assessed in lysates from NHU cell cultures exposed to 0.1 to 6 mmol/L ketamine and normalized to baseline caspase activity in untreated cells. Significant increases in caspase activity were observed after exposure to 3 and 6 mmol/L ketamine. Error bars represent SD ( B , D , E , G , and H ). n = 4 donors ( C ); n = 6 ( D , E , and H ); n = 3 donors ( B and G ). ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001.

Article Snippet: Densitometry was normalized using the intensity of housekeeping proteins β-actin (Sigma-Aldrich; clone AC15, mouse; 1:10,000 dilution) or glyceraldehyde-3-phosphate dehydrogenase (Merck Millipore; clone 6C5, mouse; 1:2000 dilution).

Techniques: Western Blot, Inhibition, Permeability, Activity Assay

Journal: The American Journal of Pathology

Article Title: Ketamine-Induced Apoptosis in Normal Human Urothelial Cells

doi: 10.1016/j.ajpath.2015.12.014

Figure Lengend Snippet: RT-PCR of NMDA receptor transcripts (GRIN isoforms). No expression of GRIN isoforms was detected in normal (nonimmortalized) human urothelial (NHU) cells under either proliferating or differentiated cell culture conditions. Furthermore, no GRIN expression was detected in freshly isolated urothelium after its separation from the underlying stroma. Representative images of one donor (proliferating and differentiated NHU) and one donor P0 urothelium are shown. Uroplakin 2 (UPK2) transcript expression was used to confirm urothelial differentiation, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was included as an internal housekeeping control to confirm RNA integrity. In addition, no products were detected in reverse transcriptase–negative cDNA controls that were generated using each RNA preparation.

Article Snippet: Densitometry was normalized using the intensity of housekeeping proteins β-actin (Sigma-Aldrich; clone AC15, mouse; 1:10,000 dilution) or glyceraldehyde-3-phosphate dehydrogenase (Merck Millipore; clone 6C5, mouse; 1:2000 dilution).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Cell Culture, Isolation, Generated