Journal: Molecular Medicine Reports

Article Title: PARP-1 may be involved in hydroquinone-induced apoptosis by poly ADP-ribosylation of ZO-2

doi: 10.3892/mmr.2017.7643

Figure Lengend Snippet: PARP-1 attenuated HQ-induced apoptosis. Empty TK6 cells were used as control cells. Cells were treated with PBS or 10 µM HQ for 72 h. (A) Representative images of flow cytometry using Annexin V-FITC and PI double-stained cells. (B) Cell apoptosis rates are presented as the mean ± standard deviation for at least three independent experiments. (C) Number of cells was counted using Countstar (IC1000). (D) Protein expression levels of the Bcl-2, caspase-3 and Bax proteins were detected using western blot analysis. (E) Caspase-3/7 luminescence values, which indicate caspase-3/7 activity, were detected using Caspase-Glo-3/7 assay kits. (F) ATP luminescence values, which indicate ATP level, were measured using CellTiter-Glo Luminescent Cell Viability assay kits. (G) Reactive oxygen species production was measured using DCFDA. *P<0.05. HQ, hydroquinone; PARP-1, poly (ADP-ribose) polymerase-1; shPARP-1, short hairpin PARP-1; Bcl2, B cell leukemia/lymphoma 2; Bax, BCL2 associated X; DCFDA, 2′,7′-dichlorofluorescin diacetate; FITC, fluorescein isothiocyanate; PI, propidium iodide.

Article Snippet: Rabbit anti-human Bcl-2 (cat. no. ab32124), caspase-3 (cat. no. ab408) primary antibodies and a 2′,7′-dichlorofluorescin diacetate (DCFDA)-Cellular Reactive Oxygen Species Detection assay kit were purchased from Abcam (Cambridge, MA, USA).

Techniques: Flow Cytometry, Staining, Standard Deviation, Expressing, Western Blot, Activity Assay, Caspase-Glo Assay, Cell Viability Assay

Journal: In Vitro Cellular & Developmental Biology. Animal

Article Title: The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm

doi: 10.1007/s11626-024-00944-w

Figure Lengend Snippet: Impact of various antioxidants on fresh bull sperm motility and reactive oxygen species (ROS) levels when exposed to different oxidative stress triggers. ( A ) Experimental design to examine the impact of various antioxidants on sperm motility and levels of reactive oxygen species (ROS) under the two oxidative stress inducers. ( B and C ) Changes in sperm viability and motility under H 2 O 2 ( B ) or antimycin (C). ( D - G ) Effect of antioxidants on the percentages of DCFH-DA and mtSOX deep Red positive sperm under H 2 O 2 or antimycin. ( H - J ) Measurement of mitochondrial activity using the JC-1 kit. ( H ) Representative flow cytometric pattern of sperm that was not treated with any drug (control) stained with JC-1. Effect of antioxidants on the percentages of high mitochondrial membrane potential (hMMP) under the treatment of H 2 O 2 ( I ) and antimycin ( J ). H 2 O 2 or antimycin groups were used as controls for statistical analysis ( p < 0.05).

Article Snippet: Photooxidation-resistant dichloro-dihydro-fluorescein diacetate (DCFH-DA) is a total ROS (H 2 O 2 , superoxide anion, and hydroxyl radical) probe (R253; DOJINDO LABOLATORIES Co., Ltd., Kumamoto, Japan).

Techniques: Activity Assay, Control, Staining, Membrane

Journal: In Vitro Cellular & Developmental Biology. Animal

Article Title: The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm

doi: 10.1007/s11626-024-00944-w

Figure Lengend Snippet: The effect of various antioxidants on frozen-thawed bull sperm motility and ROS levels. ( A ) Experimental design to examine the impact of PQQ, vitamin C, and ergothioneine on frozen-thawed bull sperm motility and ROS levels. Various antioxidants were included in all thawing, washing, and incubation processes using HTF medium as the basic medium. ( B and C ) Effect of antioxidants on the percentages of DCFH-DA ( B ) and mtSOX deep Red ( C ) positive sperm. ( D ) Changes in frozen-thawed sperm viability and kinetics patterns with antioxidants. Asterisks represent significant differences from the control (Ctrl; *: p < 0.05, **: p < 0.01, ***: p < 0.001).

Article Snippet: Photooxidation-resistant dichloro-dihydro-fluorescein diacetate (DCFH-DA) is a total ROS (H 2 O 2 , superoxide anion, and hydroxyl radical) probe (R253; DOJINDO LABOLATORIES Co., Ltd., Kumamoto, Japan).

Techniques: Incubation, Control

Journal: In Vitro Cellular & Developmental Biology. Animal

Article Title: The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm

doi: 10.1007/s11626-024-00944-w

Figure Lengend Snippet: Significant decrease in viability and an increase in ROS levels of fresh bull sperm during the incubation period. ( A ) Tracks of sperm incubated for 0 and 4 h were determined using the CASA system. Red arrows indicate the aggregated sperm, which is the cause of overestimating CASA parameters because it lowers the number of dead sperm. ( B ) Changes in fresh sperm motility single counts implying sperm viability and in kinetics patterns with incubation time. ( C and D ) Time-dependent changes in total ROS in the cytoplasm or mitochondrial superoxide level of fresh bull sperm detected using DCFH-DA or mtSOX deep Red, respectively. Flow cytometric patterns and percentage of positive sperm were shown. Different letters represent significantly different groups ( p < 0.05).

Article Snippet: Photooxidation-resistant dichloro-dihydro-fluorescein diacetate (DCFH-DA) is a total ROS (H 2 O 2 , superoxide anion, and hydroxyl radical) probe (R253; DOJINDO LABOLATORIES Co., Ltd., Kumamoto, Japan).

Techniques: Incubation

Journal: In Vitro Cellular & Developmental Biology. Animal

Article Title: The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm

doi: 10.1007/s11626-024-00944-w

Figure Lengend Snippet: The impact of oxidative stress inducers, H 2 O 2 and antimycin, on fresh bull sperm motility and ROS levels. ( A ) Experimental design to investigate the effect of the oxidative stress inducers on fresh bull sperm. ( B ) Representative images of the sperm fluorescence signal with DCFH-DA and mtSOX deep Red. Left : DCFH-DA fluorescence image of sperm treated with H 2 O 2 . Right : mtSOX deep Red fluorescence image of sperm treated with antimycin. ( C ) Flow cytometric patterns of DCFH-DA. ( D ) Percentages of DCFH-DA positive sperm. ( E ) Sperm kinetics changed by H 2 O 2 . ( F ) Flow cytometric patterns of mtSOX deep Red. ( G ) Percentages of mtSOX deep Red positive sperm. ( H ) Sperm kinetics changed by antimycin. Different letters represent significantly different groups ( p < 0.05).

Article Snippet: Photooxidation-resistant dichloro-dihydro-fluorescein diacetate (DCFH-DA) is a total ROS (H 2 O 2 , superoxide anion, and hydroxyl radical) probe (R253; DOJINDO LABOLATORIES Co., Ltd., Kumamoto, Japan).

Techniques: Fluorescence

Journal: In Vitro Cellular & Developmental Biology. Animal

Article Title: The protective effects of antioxidants against endogenous and exogenous oxidative stress on bull sperm

doi: 10.1007/s11626-024-00944-w

Figure Lengend Snippet: Impact of antioxidants on total ROS and mitochondrial ROS levels in fresh bull sperm under oxidative stress inducers. ( A ) Experimental design to investigate the effect of three antioxidants on induced reactive oxygen species in fresh bull sperm. ( B ) Flow cytometric patterns of H 2 O 2 and antioxidant-treated sperm stained with DCFH-DA. ( C ) Percentages of DCFH-DA positive sperm. ( D ) Flow cytometric patterns of antimycin and antioxidant-treated sperm stained with mtSOX deep Red. ( E ) Percentages of mtSOX deep Red positive sperm. Different letters represent significantly different groups ( p < 0.05).

Article Snippet: Photooxidation-resistant dichloro-dihydro-fluorescein diacetate (DCFH-DA) is a total ROS (H 2 O 2 , superoxide anion, and hydroxyl radical) probe (R253; DOJINDO LABOLATORIES Co., Ltd., Kumamoto, Japan).

Techniques: Staining

Journal: Small (Weinheim an Der Bergstrasse, Germany)

Article Title: Multi‐Functional Polydopamine‐Mucin Hollow Particles Provide Tunable Shell Permeability, ROS Scavenging, Tissue Adhesion, and Lubricity for Biomedical Applications

doi: 10.1002/smll.202503238

Figure Lengend Snippet: Free radical scavenging functionality of the HPs and their effects on HeLa cells. a,b) Chemical structures of DPPH• and ABTS• + before and after their reaction with an antioxidant. c,d) DPPH• and ABTS• + scavenging efficiency of HPs or HPs@Ag at different particle concentrations ( n = 3). The inset shows example images of the corresponding free radical solutions mixed with water (left), with HPs (middle), and with HPs@Ag (right), where the final concentration of the particles is 25 µg mL −1 . e) Images of the HeLa cells stained with the dye DCFH‐DA to visualize intracellular ROS (green signal). The cells are treated with medium only or with medium containing either 400 µ m H 2 O 2 , 400 µ m H 2 O 2 and 100 µg mL −1 HPs, or 400 µ m H 2 O 2 and 100 µg mL −1 HPs@Ag. f) Quantification of the fluorescence intensity of the cell images after ROS staining ( n = 4). g) Live/Dead staining results of HeLa cells treated with medium containing H 2 O 2 (0 or 10 m m ) and particles (0 or 50 µg mL −1 ); living cells appear green and dead cells appear red. h) Percentage of living cells as determined from Live/Dead staining images ( n = 3). i) Schematic illustration of the different effects the HPs and HPs@Ag can have on the cells. The data is expressed as mean ± standard deviation. Asterisks ( * ) mark statistically significant differences determined in two‐tailed t ‐tests based on a p ‐value of 0.05.

Article Snippet: To test the ROS levels in the cells, 2′,7′‐dichlorodihydrofluorescein diacetate (DCFH‐DA; HY‐D0940, MCE) was used to stain the cells.

Techniques: Concentration Assay, Staining, Fluorescence, Standard Deviation, Two Tailed Test