resource source identifier human fetal astrocyte cell applications 882ak 05f human adult astrocyte cell applications 882ak 05a aiw002 2 neuro  (Cell Applications Inc)

Journal: Cell reports

Article Title: Rapid macropinocytic transfer of α-synuclein to lysosomes.

doi: 10.1016/j.celrep.2022.111102

Figure Lengend Snippet: Figure 2. Rapid PFF colocalization with lysosomes in human dopaminergic NPCs, dopaminergic neurons, and astrocytes (A) Dopaminergic NPCs, stained with LysoTracker, were incubated for 0, 2, 10, and 30 min at 37C following the addition of Alexa 488-labeled PFF (white) at 2 mg/mL. The cells were then washed lightly and briefly with trypsin and fixed. Scale bar, 20 mm. (B) NPCs differentiated into dopaminergic neurons were used in the experiments described in (A). Scale bar, 20 mm. (C) Human astrocytes, grown and mounted on coverslips, were given fluorescently labeled PFFs at a 1-mg/mL concentration. Cells were then incubated for 0, 2, 10, and 30 min at 37C. Cells were washed with trypsin, fixed, permeabilized, and stained with LAMP1 antibody. Scale bars, 20 mm and 2.5 mm for the insets. (D and E) Manders’ coefficients were calculated using the JACoP plugin to ascertain the colocalization of LysoTracker with PFFs from experiments described in (A) and (B). n = 6 for NPCs and n = 6 for neurons in each condition (i.e., n = 48 total for NPCs and n = 48 total for neurons), from 3 independent experiments.

Article Snippet: Cellular response to PFF addition was done using human fetal astrocytes (Cell Applications, 882AK-05f).

Techniques: Staining, Incubation, Labeling, Concentration Assay

Journal: Cell reports

Article Title: Rapid macropinocytic transfer of α-synuclein to lysosomes.

doi: 10.1016/j.celrep.2022.111102

Figure Lengend Snippet: Figure 6. PFF trafficking to lysosomes and MVBs (A) Astrocytes administered PFFs for 10 min localize PFFs in larger newly forming MVBs, always in proximity to the vesicular membrane, sometimes at in- vaginations, potentially signifying the early stages of exosome formation. Scale bars, 100 nm for low magnification, 80 nm for moderate magnification, and 50 nm for highest magnification inset. (B) Localization of PFFs outside of vesicles in electron-dense MVBs signifies the progression of MVB maturation. Scale bars, 100 nm and 50 nm for inset. (C) Similar findings regarding the formation of MVB (LAMP1+; white) containing PFF (red) was confirmed using STED microscopy. Scale bars, 20 mm for low- magnification images and 2.5 mm for high-magnification images.

Article Snippet: Cellular response to PFF addition was done using human fetal astrocytes (Cell Applications, 882AK-05f).

Techniques: Membrane, Microscopy

Journal: Pharmaceuticals

Article Title: Transcriptomic Analysis of Fumarate Compounds Identifies Unique Effects of Isosorbide Di-(Methyl Fumarate) on NRF2, NF-kappaB and IRF1 Pathway Genes

doi: 10.3390/ph15040461

Figure Lengend Snippet: RT-PCR analyses and NF-κB luciferase reporter assay. ( A – G ) RT-PCR analyses ( OSGIN1 , IL6 , ICAM1 , MALT1 , TNFAIP3 , IRF1, and CXCL8 ). Average relative expression is shown for each gene (±1 standard error; n = 3 replicates per treatment). Relative expression was calculated using the 2 −∆∆Ct method and ∆Ct values were further normalized to the CTL treatment. Average relative expression is shown for each treatment (bottom margin). Treatments with different letters have significantly different average expression ( p < 0.05, Fisher’s least significant difference). ( H ) NF-κB reporter assay. Human fetal astrocytes were transfected with firefly/Renilla luciferase constructs and treated with TNF-α (20 ng/ml) alone ( n = 6) or TNF-α (20 ng/mL) + IDMF (2.5 µM) ( n = 3) for 6 h. The ratio of luciferase and Renilla (internal control) signals was used to monitor NF-κB induction. The average ratio value is shown (±1 standard error) for each treatment ( p -value: two-sample Wilcoxon rank sum test).

Article Snippet: Experiments were performed using human fetal astrocytes (HA) derived from cerebral cortex (cat no. 882A-05f; Cell Applications, San Diego, CA, USA) grown to subconfluence for 48 h in HA growth medium (cat. no. 820-500, Cell Applications) in a 96-well black wall tissue culture plate.

Techniques: Reverse Transcription Polymerase Chain Reaction, Luciferase, Reporter Assay, Expressing, Transfection, Construct, Control