Journal: Life sciences in space research

Article Title: Comparison of signaling profiles in the low dose range following low and high LET radiation.

doi: 10.1016/j.lssr.2020.02.002

Figure Lengend Snippet: Fig. 1. Average total fold intensity of phospho-protein signal as compared to median control value. Colored bars (dark blue = 0.5 Gy, light blue = 0.1 Gy and green = 0.05 Gy) indicate values significantly different from controls (yellow). Significance bars are shown for all significant differences between doses and 0 (p ≤0.05). Average fold intensity levels are shown at 2 h post radiation for γH2AX (A), pATF2 (B) and pSMC1 (C). Persistent effects are shown at 24 h for γH2AX (D), pATF2 (E) and pSMC1 (F). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: For staining, 0.5 × 106 fixed cells were washed in phosphate buffered saline (PBS), resuspended in blocking buffer (2% FBS/PBS) and incubated for 1 h with primary antibody on ice, with resuspension of the pellet every 15 min. Primary antibodies used in flow cytometry include mouse monoclonal γH2AXSer139 (1:800 dilution) and pSMC1Ser957 (1:800 dilution) from Millipore (Temecula, CA) and rabbit polyclonal pATF2 Ser490/498 (1:1000 dilution) from Rockland, Inc. (Gilbertsville, PA).

Techniques: Control

Journal: Life sciences in space research

Article Title: Comparison of signaling profiles in the low dose range following low and high LET radiation.

doi: 10.1016/j.lssr.2020.02.002

Figure Lengend Snippet: Fig. 3. Average total fold pATF2 intensity over median control level versus fluence for various radiation qualities. Average fold intensity levels are shown at 2 h post radiation for Si ions (A),Fe ions (C) and Ti ions (E). Persistent effects are shown at 24 h for Si ions (B), Fe ions (D) and Ti ions (F).

Article Snippet: For staining, 0.5 × 106 fixed cells were washed in phosphate buffered saline (PBS), resuspended in blocking buffer (2% FBS/PBS) and incubated for 1 h with primary antibody on ice, with resuspension of the pellet every 15 min. Primary antibodies used in flow cytometry include mouse monoclonal γH2AXSer139 (1:800 dilution) and pSMC1Ser957 (1:800 dilution) from Millipore (Temecula, CA) and rabbit polyclonal pATF2 Ser490/498 (1:1000 dilution) from Rockland, Inc. (Gilbertsville, PA).

Techniques: Control

Journal: Life sciences in space research

Article Title: Comparison of signaling profiles in the low dose range following low and high LET radiation.

doi: 10.1016/j.lssr.2020.02.002

Figure Lengend Snippet: Fig. 6. Average total fold intensity of pATF2 signal intensity divided by fluence and graphed versus LET. Average fold intensity levels are shown at 2 h post radiation for 0.05 Gy (A), 0.1 Gy (B) and 0.5 Gy (C). Persistent effects are shown at 24 h for 0.05 Gy (D), 0.1 Gy (E) and 0.5 Gy (F).

Article Snippet: For staining, 0.5 × 106 fixed cells were washed in phosphate buffered saline (PBS), resuspended in blocking buffer (2% FBS/PBS) and incubated for 1 h with primary antibody on ice, with resuspension of the pellet every 15 min. Primary antibodies used in flow cytometry include mouse monoclonal γH2AXSer139 (1:800 dilution) and pSMC1Ser957 (1:800 dilution) from Millipore (Temecula, CA) and rabbit polyclonal pATF2 Ser490/498 (1:1000 dilution) from Rockland, Inc. (Gilbertsville, PA).

Techniques:

Journal: Antioxidants

Article Title: Liproxstatin-1 Attenuates Retinal Ischemia–Reperfusion Injury by Suppressing EGR1-Mediated Ferroptosis

doi: 10.3390/antiox15030391

Figure Lengend Snippet: Lip-1 halts the oxidative stress-to-ferroptosis cascade in retinal I/R injury. ( A ) Immunofluorescence co-staining of Tuj1-positive RGCs (green) with GPX4 (red), FSP1 (red), and ACSL4 (red) and DAPI (blue) at 24 h post-I/R in retinal sections from sham and I/R mice treated with Vehicle or Lip-1. ( B ) Immunofluorescence co-staining of Tuj1-positive RGCs (green) with 4HNE (red), FHC (red), and DHE (red) and DAPI (blue) at 24 h post-I/R in retinal sections from sham and I/R mice treated with Vehicle or Lip-1. ( C ) Immunofluorescence co-staining of FerroOrange (red) with DAPI (blue) at 24 h post-I/R in retinal sections from sham and I/R mice treated with Vehicle or Lip-1. ( D ) Quantification of the relative immunofluorescence intensity of GPX4, FSP1, ACSL4, 4-HNE, FHC, DHE, and FerroOrange from ( A – C ) ( n = 6 biologically independent experiments). ( E ) Immunofluorescence co-staining of C11-BODIPY (green/red) staining with DAPI (blue) at 24 h post-I/R in retinal sections from sham and I/R mice treated with Vehicle or Lip-1. ( F ) Quantification of the C11 fluorescence ratio (green/red) from ( E ) ( n = 6 biologically independent experiments). ( G ) Western blot analysis of GPX4, FSP1, and FHC expression from sham and I/R mice treated with Vehicle or Lip-1. β-Actin served as a loading control. ( H ) Quantification of GPX4, FSP1, and FHC protein levels from ( E ) ( n = 3 biologically independent experiments). Scale bars: 50 μm. Total magnification: 400×. Data were analyzed by two-way ANOVA with Tukey’s post hoc test. All data are shown as mean ± SEM. ns, not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: FSP1 , Rabbit , Proteintech , 20886-AP , 1:1000 (WB).

Techniques: Immunofluorescence, Staining, Fluorescence, Western Blot, Expressing, Control

Journal: Antioxidants

Article Title: Liproxstatin-1 Attenuates Retinal Ischemia–Reperfusion Injury by Suppressing EGR1-Mediated Ferroptosis

doi: 10.3390/antiox15030391

Figure Lengend Snippet: Lip-1 attenuates OGD/R-induced ferroptosis in primary RGCs by restoring antioxidant defenses and suppressing lipid peroxidation. ( A ) Western blot analysis of GPX4 expression in primary RGCs at the indicated time points (3, 6, 12, 24 h) after OGD/R. β-Actin served as a loading control. ( B ) Quantification of GPX4 relative band density from ( A ) ( n = 3 biologically independent experiments). ( C ) Western blot analysis of GPX4, FSP1, and FHC expression in RGCs under different conditions (Control, OGD/R + Vehicle, OGD/R + Lip-1). β-Actin served as a loading control. ( D ) Quantification of GPX4, FSP1, and FHC protein levels from ( C ) ( n = 3 biologically independent experiments). ( E ) Immunofluorescence co-staining of Tuj1-positive RGCs (red) with GPX4 (green) and DAPI (blue) under different conditions. ( F ) Immunofluorescence co-staining of Tuj1-positive RGCs (red) with ferroptosis markers (FSP1, ACSL4, 4-HNE, FHC, all in green) and DAPI (blue) under different conditions. ( G ) Immunofluorescence co-staining of Tuj1-positive RGCs (purple) with FerroOrange staining (red), DCFH-DA (green) and DAPI (blue) under different conditions. ( H ) Quantification of the relative immunofluorescence intensity of GPX4, FSP1, ACSL4, 4-HNE, FHC, FerroOrange, and DCFH-DA from ( E – G ) ( n = 6 biologically independent experiments). ( I ) Immunofluorescence co-staining of Tuj1-positive RGCs (purple) with lipid peroxidation (detected by oxidized C11-BODIPY581/591, green/red), mitochondrial membrane potential (assessed by JC-1 monomer/aggregate ratio, green/red) and DAPI (blue) under different conditions. ( J ) Quantification of the C11 and JC-1 fluorescence ratio (green/red) from ( I ) ( n = 6 biologically independent experiments). ( E – G , I ) Scale bars: 50 μm. Total magnification: 400×. Data were analyzed by two-way ANOVA with Tukey’s post hoc test. All data are shown as mean ± SEM. ns, not significant, * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: FSP1 , Rabbit , Proteintech , 20886-AP , 1:1000 (WB).

Techniques: Western Blot, Expressing, Control, Immunofluorescence, Staining, Membrane, Fluorescence

Journal: Cellular & Molecular Biology Letters

Article Title: The promoting roles of GLP1R and GIPR in stemness maintenance and multiple lineage-specific differentiation of PDLSCs

doi: 10.1186/s11658-026-00867-2

Figure Lengend Snippet: Enhanced cell growth of PDLSCs facilitated by GIPRA/MAPK/ERK axis. a Western blot assay showing the expression of Ki-67, PCNA, and CCND3 affected by GIPRA as well as inhibition of cAMP/PKA/CREB, MAPK/ERK, Wnt/β-catenin, or JAK/STAT3 signaling pathways. The interpretation of bubble colors in the line chart is illustrated in Fig. a. b CCK-8 assay indicating the cell growth enhanced by GIPRA, but suppressed by ERK Inh ( n = 5 in each group). The interpretation of line colors in the line chart is illustrated in Fig. b. c Venn diagram showing the intersection of differentially binding genes by p-CREB, p-ERK1/2, TCF4, and p-STAT3 (log FC > 1, P < 0.05) compared between GIPRA and NC group. d The bubble plot displays the top ten functions of cell division and cell cycle (GO biological process) associated with the 5994 (red color highlighted) specifically binding genes by p-ERK1/2 in c . e – g IGV showing the binding peaks of p-ERK1/2 on MKI67 ( e ), PCNA ( f ), and CCND3 genes ( g ). The differential binding peaks of p-ERK1/2 on these gene promoters are highlighted by blue boxes. h – k Line plot illustrates the distribution of ChIP-seq signal intensity of p-ERK1/2 ( h ), p-CREB ( i ), TCF4 ( j ), and p-STAT3 ( k ) across the transcription start sites (±3 kb) of differentially binding genes in PDLSCs treated with GIPRA as well as GIPRA plus ERK Inh. NC, normal PDLSCs as negative control; Inh., inhibitor

Article Snippet: Immunoprecipitation was carried out overnight at 4 °C using 1 μg IP grade antibodies against p-CREB1 (cat. no. 81871-1-RR, Proteintech), p-ERK1/2 (Thr202/Tyr204), p-STAT3 (Ser727) (same with Western blot), and TCF4 (cat. no. 22337-1-AP, Proteintech).

Techniques: Western Blot, Expressing, Inhibition, Protein-Protein interactions, CCK-8 Assay, Binding Assay, ChIP-sequencing, Negative Control