Journal: Scientific Reports

Article Title: Histological analysis of sleep and circadian brain circuitry in cranial radiation-induced hypersomnolence (C-RIH) mouse model

doi: 10.1038/s41598-022-15074-0

Figure Lengend Snippet: In vitro radiosensitivity of astrocytic cell lines based on region of isolation and treatment timing. ( A ) Photomicrographs of SCN2.2 and CTXTNA2 cells, two astrocytic cell lines with similar morphology. B. Quantification of trypan blue postitive cells to determine cell viability 1, 6 and 24 h after 8 Gy radiation, there were significant main effects for time (F(1,18) = 96.834, p < 0.001), cell type (F(1,18) = 235.937, p < 0.001), and an interaction between time and cell type (F(2,17) = 151.927, < 0.001). Posthoc tests (tukey) were significant if p < 0.05. SCN2.2 cells (green bars) survived at significantly higher level 1 ( p < 0.001) and 6 h ( p = 0.024) after radiation when compared to the cortical astrocytes (purple bars). ( C ) Clonogenic assay to quantify survival fractions from 0 to 8 Gy in SCN and cortical cells. Again SCN2.2 cells (green line) had better overall survival after radiation when compared to CTXTNA2 cells (purple line). There were significant main effects of both cell line (F(1,24) = 33.228, p < 0.001) and radiation dose (F(5,24) = 402.558, p < 0.001), and an interaction between cell line and dose (F(5,24) = 6.172, p = 0.001). The dotted black line across the graph shows the dose modifying fractors (DMF 10 ) levels for both cells lines which was also significantly different between the two cell lines. ( D ) The chronotherapeutic experimental timeline, 1.0 × 10 6 cells were plated on day 0. Cells were then serum shocked with a 50% horse serum solution for 2 h at circadian time(CT) 8 (01:00, Day 2) or 20 (13:00, Day 1) on the subsequent days. All cells were irradiated on Day 2 at 11:00 and then colonies stained and counted after Day 5. ( E ) Clonogenic assay of SCN2.2 cells irradiated at CT8 (dotted line) and CT20 (solid line). Astrocytes irradiated at CT8 were significantly more sensitive to radiotherapy. There were significant main effects of time (F(1,24) = 7.041, p = 0.014) and dose (F(5,24) = 26.995, p < 0.001) but no interaction (F(5,24) = 1.081, p = 0.396). ( F ) Clonogenic assayof CTXTNA2 cells irradiated at CT8 (dotted line) and CT20 (solid line). Again, astrocytes irradiated at CT8 were significantly more sensitive to radiotherapy. There were significant main effect of time (F(1,24) = 14.099, p = 0.001) and dose (F(5,24) = 135.997, p < 0.001) but no interaction (F(5,24) = 1.349, p = 0.278). ( G ) ATM western analysis of both cell lines when samples were collected at two timepoint post-synchronization. Western blot of ATM (blue bands) and control β-actin (teal bands) in both cell lines show higher presence of ATM after serum shock entrainment (Left panel). A graph of the ATM/β-actin relative ratio (Right panel), shows that CTXTNA cells express more of the protein than SCN2.2 cells. Four replicates of each cell line at the 3 timepoints were compared, there were significant main effects of time (F(2,21) = 83.154, p < 0.001) and cell type (F(1,21) = 8.312, p = 0.009) and an interaction (F(2,21) = 5.230, p = 0.014). Posthoc tests (tukey) were significant if p < 0.05. Both cell lines had more protien during CT08 when compared to CT20 but only CTXTNA cells had significant differences between the two ( p = 0.007). Significance was defined as p < 0.05 and indicated by * or a, b, and c with different letters indicating significant differences.

Article Snippet: Membranes were stained using primary antibodies for ATM (1:1000; ab81292, abcam, MA) and β-actin (1:5000; ab8226, abcam, MA) for 24 h at 4°C, followed by 1 h at room temperature in secondary antibodies (1:5000) for IRDye 800CW Goat anti-Mouse (926-32210; Li-Cor Biotechnology, NE) and IRDye 680CW Goat anti-Rabbit (926-68071; Li-Cor Biotechnology, NE).

Techniques: In Vitro, Isolation, IF-P, Clonogenic Assay, Irradiation, Staining, Western Blot, Control

Journal: Cell reports

Article Title: Single-cell analysis of the human pancreas in type 2 diabetes using multi-spectral imaging mass cytometry

doi: 10.1016/j.celrep.2021.109919

Figure Lengend Snippet:

Article Snippet: 154Sm-Beta-actin (Clone 2F1–1) , Fluidigm , Cat#3154021D.

Techniques: Software