Journal: Radiotherapy and Oncology

Article Title: Regulation of O 2 consumption by the PI3K and mTOR pathways contributes to tumor hypoxia

doi: 10.1016/j.radonc.2014.02.007

Figure Lengend Snippet: Hypoxia is reduced in a 3D spheroid model and is predicted to decline in vivo . (A) EMT6 and FaDu spheroids were treated for 24 h with BEZ235 or BKM120, prior to fixation and sectioning. Sections were DAB-stained for pAKT (Ser473) expression. Representative images are shown for BEZ235 (50 nM) and BKM120 (5 μM). (B) The percentage of DAB-positive pixels in the viable region of the spheroid was reduced with all treatments. (C) Spheroids were treated with 50 nM BEZ235 or 5 μM BKM120 for 24 h, 48 h or 24 h followed by 24 h incubation in drug-free medium (24 h + wash). Hypoxia was assessed by staining central spheroid sections for EF5 (red) with Hoechst as a counterstain (blue). Data shown are representative of two independent experiments, n = 10 spheroids. (D) Mean EF5 fluorescence in spheroids treated for 24 h was reduced in both cell lines tested. (E) To assess whether hypoxia was recoverable, mean EF5 fluorescence in spheroids treated for 24 h was compared with that in spheroids treated for either 48 h or 24 h followed by the 24 h wash in drug-free medium. In cells treated with BEZ235, the 24 h wash period led to a return of the hypoxia signal. (F and G) Simulated distribution of oxygen partial pressure (mmHg) in a representative 1497 μm × 1482 μm vascular network derived from ex vivo samples where oxygen consumption was either 100% (F) or 55% (G) of base level. Highly oxygenated regions are in red, scaling down to poorly oxygenated regions in blue. (H) Average hypoxic fraction (defined as <10 mmHg) calculated across 10 example networks as in F as a function of varying oxygen consumption. All data points represent mean ± 95% Confidence Interval. One-way ANOVAs were performed with Bonferroni-corrected post t -tests. ∗ P < 0.01.

Article Snippet: To assess signaling inhibition in spheroids, sections were stained with anti-pAKT antibody using ImmPRESS™ reagent kit (MP-7401, VectorLabs) and DAB Peroxidase substrate kit (SK-4100, VectorLabs).

Techniques: In Vivo, Staining, Expressing, Incubation, Fluorescence, Derivative Assay, Ex Vivo

Journal: Cell

Article Title: The 7q11.23 Protein DNAJC30 is an Auxiliary Component of ATP Synthase and Links Mitochondria to Brain Development

doi: 10.1016/j.cell.2018.09.014

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Avidin-Biotin blocker , Vector Laboratories , Cat#SP-2001.

Techniques: Recombinant, Protease Inhibitor, Modification, Avidin-Biotin Assay, Plasmid Preparation, Bicinchoninic Acid Protein Assay, Knock-Out, Sequencing, Clone Assay, Software, Microscopy, Digital PCR, Fluorescence, Transmission Assay

Journal: Cell Proliferation

Article Title: Haematopoietic lineage‐committed bone marrow cells, but not cloned cultured mesenchymal stem cells, contribute to regeneration of renal tubular epithelium after HgCl 2 ‐induced acute tubular injury

doi: 10.1111/j.1365-2184.2008.00545.x

Figure Lengend Snippet: Examples of engraftment of haematopoietic lineage marrow cells (HLMCs) and mesenchymal stem cells at 3 days after HgCl2. (a) GFP‐positive control shows proximal tubular cells (PHA‐L, blue colour) co‐stained for GFP (brown colour). (b) GFP‐negative control (wild type) showing no detection of GFP. (c) BMT mouse treated with HgCl2 demonstrating GFP‐positive proximal tubular epithelial cells. Dashed black arrows indicate PHA‐L stained donor HLMC‐derived tubular cells. (d) Male positive control; Y chromosomes were detected by indirect FISH, black arrows point to Y chromosomes (brown dot) in proximal tubular cells histochemically stained with PHA‐L (red colour). (e) Female control showing the lack of Y chromosome detection within proximal tubular cells histochemically stained with PHA‐L. (f) Scattered Y chromosomes (brown dot) in renal interstitial area, not in proximal tubular cells. (g) High magnification of yellow box in (f); a–f, ×400; g, ×600. BMT, bone marrow transplantation; FISH, fluorescence in situ hybridization; PHA‐L, Phaseolus vulgaris leucoagglutinin.

Article Snippet: After GFP staining, tissue sections were microwaved in 10 m m tri‐sodium citrate (pH 6.0) for 10 min and biotin was blocked using a biotin blocking kit (DAKO), and then incubated for 45 min with biotinylated Phaseolus vulgaris leucoagglutinin (PHA‐L) 1/100 (B‐1115, Vector Laboratories, Burlingame, CA, USA) to detect proximal convoluted tubules, or biotinylated peanut agglutinin (PNA) 1/800 (B‐1075, Vector Laboratories) to detect distal convoluted tubules.

Techniques: Positive Control, Staining, Negative Control, Derivative Assay, Transplantation Assay, Fluorescence, In Situ Hybridization

Journal: Cell Proliferation

Article Title: Haematopoietic lineage‐committed bone marrow cells, but not cloned cultured mesenchymal stem cells, contribute to regeneration of renal tubular epithelium after HgCl 2 ‐induced acute tubular injury

doi: 10.1111/j.1365-2184.2008.00545.x

Figure Lengend Snippet: (a) Changes in the abundance of GFP‐positive cells within the PHA‐L stained cell population in control mice and mice treated with HgCl2 (n = 5 per treatment time point). *P < 0.05 versus the same group at day 0; +P < 0.05 versus group CON at the corresponding time point. The percentages of GFP‐positive cells were adjusted based on a correction factor derived from the actual cell count of 41% of PHA‐L stained cells being GFP‐positive in GFP donor mice. (b) Changes in the 3H‐LI of PHA‐L stained proximal tubular cells of: combined indigenous and donor HLMC (left panel); indigenous origin (central panel); donor HLMC origin (right panel). n = 5 per group. *P < 0.05 versus the same group at day 0; +P < 0.05 versus group CON at the corresponding time point. (c–g) Examples of chimerism and proliferation (3H‐thymidine labelling) of GFP+ HLMC‐derived PHA‐L‐stained cells at 3 days after HgCl2. (c) Black arrowheads indicate PHA‐L stained (blue colour in tubular cell apical membrane) donor GFP+ HLMC‐derived tubular cells (brown colour in cytoplasm) under bright field, and black arrows indicate 3H‐thymidine labelling of PHA‐L‐stained HLMC‐derived tubular cells, ×500. (d) The same field under dark field, white arrowheads point to silver grains (3H‐thymidine labelling) and white arrows indicate 3H‐thymidine labelling of PHA‐L‐stained HLMC‐derived tubular cells (×500). (e) Bright field and (f) dark field are the higher magnification of yellow box area in (c). (g) The images in (e) and (f) were combined to help to show silver grains over cells that are donor GFP+ HLMC‐derived PHA‐L‐stained tubular cells. GFP, green fluorescent protein; HLMC, haematopoietic lineage marrow cell; PHA‐L, Phaseolus vulgaris leucoagglutinin.

Article Snippet: After GFP staining, tissue sections were microwaved in 10 m m tri‐sodium citrate (pH 6.0) for 10 min and biotin was blocked using a biotin blocking kit (DAKO), and then incubated for 45 min with biotinylated Phaseolus vulgaris leucoagglutinin (PHA‐L) 1/100 (B‐1115, Vector Laboratories, Burlingame, CA, USA) to detect proximal convoluted tubules, or biotinylated peanut agglutinin (PNA) 1/800 (B‐1075, Vector Laboratories) to detect distal convoluted tubules.

Techniques: Staining, Derivative Assay, Cell Counting

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: Generalized features of apoptosis and pyroptosis.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: Lysis, DNA Laddering

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: Final concentration of apoptosis inducers or marine toxins used for the experiments.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: Concentration Assay

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: Percentage of cell death evaluated through the resazurin assay in hemocytes treated with different concentrations of inducers of apoptosis or marine toxins during 4 h at 25°C. Hemolymph from 10 to 30 oysters was pooled to have a total of 1.5 × 10 6 cells mL −1 . Bars represent mean ± standard deviation of two independent experiments. * P < 0.05. Neg, negative control; Pos, positive control; CPT, camptothecin; STP, staurosporine; STX, saxitoxin; GTX, gonyautoxin; OA, okadaic acid; DTX, dynophysistoxin; PbTx, brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: Resazurin Assay, Standard Deviation, Negative Control, Positive Control

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: In vitro phosphatidylserine translocation to the extracellular leaflet in hemocytes exposed to apoptosis inducers or marine toxins during 4 h at 25°C. (A1) Hemocytes observed by fluorescence, to detect viable or no measurable programmed cell death (PCD, green and red staining negative), (A4) PCD cells (green, annexin V-bound), and (A2 and A3) cells in end stage of PCD and dead (red, propidium iodide stained cells, and green annexin V-bound cells). (B) The graph shows percentages of different stages of cells of (A) . Results are expressed as the mean ± standard deviation. A, annexin V positive; d, dead; v, viable. Scale bar = 5 μm. SS, saline solution; CPT, camptothecin; STP, staurosporine; STX, saxitoxin; GTX, gonyautoxin; OA, okadaic acid; DTX, dynophysistoxin; PbTx, brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: In Vitro, Translocation Assay, Fluorescence, Staining, Standard Deviation

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: In vitro DNA double-strand breakage in hemocytes exposed to apoptosis inducers or marine toxins for 4 h at 25°C. (A) Image of hemocyte nuclei with different DNA damage grades, assessed by neutral comet assay, stained red with propidium iodide. DNA damage categories: undamaged, low damaged, medium damage, high damage, and complete damage, using a scale of 0–4, respectively. Scale bar = 5 μm. (B) Frequency distribution of DNA damage in hemocytes. Data were obtained from 400 scored nuclei. Results are expressed as the mean ± standard deviation. * P < 0.05. SS, saline solution; CPT, camptothecin; STP, staurosporine; STX, saxitoxin; GTX, gonyautoxin; OA, okadaic acid; DTX, dynophysistoxin; PbTx, brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: In Vitro, Neutral Comet Assay, Staining, Standard Deviation

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: In vitro chromatin condensation in nuclei of hemocytes exposed to apoptosis inducers or marine toxins for 4 h at 25°C. (A) DAPI staining (blue) of representative nuclei: observe the normal nuclei (n), hyperchromasia (arrow) characteristic of condensed chromatin, and nuclei with condensation of chromatin in the periphery (arrow head). (B) Percentages of the nuclei with condensed chromatin. In each sample, at least 100 nuclei were counted. Scale bar = 5 μm. Results are expressed as the mean ± standard deviation. * P < 0.05. SS, saline solution; CPT, camptothecin; STP, staurosporine; STX, saxitoxin; GTX, gonyautoxin; OA, okadaic acid; DTX, dynophysistoxin; PbTx, brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: In Vitro, Staining, Standard Deviation

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: In vitro DNA ladder in nuclei of hemocytes exposed to apoptosis inducers or marine toxins for 4 h at 25°C. Electrophoresis was performed on 2% agarose gel. M, kb marker; C+, Positive control (DNA from apoptotic U937 cells); SS, Saline solution; CPT, camptothecin; STP, staurosporine; STX, saxitoxin; GTX, gonyautoxin; OA, okadaic acid; DTX, dynophysistoxin; PbTx, brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: In Vitro, Electrophoresis, Agarose Gel Electrophoresis, Marker, Positive Control

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: Real-time PCR (qRT-PCR) of caspases (Casp) in hemocytes exposed to in hemocytes exposed to apoptosis inducers or marine toxins for 4 h at 25°C. Whisker-box plots of the relative expressions calculated by the REST 2009 Software of the caspase-1, caspase-2, caspase-3, caspase-7, and caspase-8 genes. Boxes represent the interquartile range, or the middle 50% of observations. The dotted line inside the box represents the median gene expression. Whiskers represent the minimum and maximum observations. The proportions over 1 indicate genes that increase expression, while proportions <1 indicate genes decrease in expression. * P < 0.05; ** P < 0.01. Casp, Caspase; CPT, Camptothecin; STP, Staurosporine; STX, Saxitoxin; GTX, Gonyautoxin; OA, Okadaic acid; DTX, Dynophysistoxin; PbTx, Brevetoxin; Vp, Vibrio parahaemolyticus extract; Vc, V. campbellii extract.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Whisker Assay, Software, Expressing

Journal: Frontiers in Immunology

Article Title: In vitro Evaluation of Programmed Cell Death in the Immune System of Pacific Oyster Crassostrea gigas by the Effect of Marine Toxins

doi: 10.3389/fimmu.2021.634497

Figure Lengend Snippet: P -value of relative expression of caspase transcripts in hemocytes of Crassostrea gigas exposed to apoptosis inducers or marine toxins.

Article Snippet: To identify PS exposure on the outer leaflet of the plasma membrane, we used the Annexin V–FITC apoptosis kit (BioVision, K101).

Techniques: Expressing