Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Airway Epithelial cGAS Is Critical for Induction of Experimental Allergic Airway Inflammation.

doi: 10.4049/jimmunol.1900869

Figure Lengend Snippet: FIGURE 7. Cytosolic dsDNA accumulation was partially inhibited by Mito-TEMPO and colocalized with cGAS in IL-33–treated HBE cells. (A) Representative images of immunofluorescence staining for dsDNA in HBE cells treated with IL-33 and Mito-TEMPO in vitro. The quantitative results of cytosolic dsDNA abundance of three independent experiments in HBE cells were shown (10–20 cells per experiment). (B) Relative mRNA levels of GM-CSF in HBE cells treated with IL-33 and Mito-TEMPO in vitro. Data are the mean 6 SEM of three independent experiments. (C) Representative images of immunofluorescence staining for dsDNA (staining red) and cGAS (staining green) in HBE cells treated with IL-33 in vitro. The quantitative results of cytosolic dsDNA colocalized with cGAS in HBE cells were shown. Data are mean 6 SEM of four independent experiments, with 10–20 cells per experiment. Scale bars, 10 mm. *p , 0.05, ***p , 0.001, ****p , 0.0001.

Article Snippet: Cells are stimulated by IL-33/OVA after transfection for 24 h. Immunofluorescence analysis of dsDNA and cGAS levels in HBE cells and lung tissues of asthmatic mice Sections of lung tissues of mice were immunostained with dsDNA Ab (ab27156; Abcam), CC10 Ab (ab40873; Abcam), or cGAS Ab (26416-1- AP; Proteintech) according to the manufacturer’s instructions.

Techniques: Staining, In Vitro

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: Nature Communications

Article Title: Mycobacterium tuberculosis suppresses host antimicrobial peptides by dehydrogenating L-alanine

doi: 10.1038/s41467-024-48588-4

Figure Lengend Snippet: CFU counts ( A ) and relative intracellular CFU ratio ( B ) in MPMs treated with 1 mM L-alanine, 50 nM rifampicin (RIF) or combination of 1 mM L-alanine and 50 nM RIF followed by H37Rv infection for 3 and 24 h (MOI = 2). CFU counts ( C ) and relative intracellular CFU ratio ( D ) in MPMs treated with 1 mM L-alanine or 50 nM rifampicin (RIF) followed by multidrug-resistant M. tuberculosis (MDR) infection for 3 and 24 h (MOI = 2). E Survival curve of 6-week-old female SCID mice treated with ddH 2 O or 30 mg/ml L-alanine and aerosol infected with roughly 100 CFUs per mouse of H37Rv. CFU assay ( F ), histological score ( G ) and histopathological assay ( H ) with acid-fast staining and haematoxylin and eosin staining in lung tissues of C57BL/6J mice treated with ddH 2 O or 30 mg/ml L-alanine after 28 days of H37Rv infection. Data in ( A – D ) are representative of one experiment with at least three independent biological replicates; ( A – D ) n = 3, each circle represents one technical repeat (mean ± s.e.m); ( F ) n = 6 mice (mean ± s.e.m); ( G ) n = 3 mice (mean ± s.e.m). Two-tailed unpaired Student’s t -test ( A – D ), Log-rank test ( E ) and two-sided Mann-Whitney U -test ( F , G ) were used for statistical analysis. P values are shown in ( A – G ). 1#, 2# and 3# in ( H ) represent lung tissues from 3 mice. Scale bars, 100 μm (top; original magnification, ×400) and 20 μm (bottom; original magnification, ×1,000). Source data are provided as a file.

Article Snippet: To validate the effectiveness of tuberculosis drug regimens, C57BL/6 J mice were treated with RIF (Rifampicin) (10 mg/kg/day), or combination of RIF (10 mg/kg/day) and Rv2780 inhibitor GWP-042 (MedChemExpress, 91%) (10 mg/kg/day) via oral gavage after three weeks of H37Rv aerosol infection.

Techniques: Infection, Aerosol, Colony-forming Unit Assay, Staining, Two Tailed Test, MANN-WHITNEY

Journal: Nature Communications

Article Title: Mycobacterium tuberculosis suppresses host antimicrobial peptides by dehydrogenating L-alanine

doi: 10.1038/s41467-024-48588-4

Figure Lengend Snippet: A Structure of GWP-042. Superimposition image ( B ) of docked pose of the GWP-042 to Rv2780 protein. Ligand-target interaction diagram ( C ) showing interactions of GWP-042 with the active site residues of Rv2780 protein. D Surface plasmon resonance (SPR) assay of the direct interaction of GWP-042 with Rv2780 protein. ( E ) The dehydrogenase activity of Rv2780 was measured by pyruvate concentration in the presence of increasing concentrations of GWP-042. Dose–response curves for IC 50 values were determined by nonlinear regression. F ELISA analysis of Defb4 in MPMs treated with 50 μM GWP-042 followed by H37Rv or H37RvΔRv2780 infection for 0, 24 and 48 h (MOI = 2). CFU counts ( G ) and relative intracellular CFU ratio ( H ) in mice peritoneal macrophages treated with 50 μM GWP-042 followed by H37Rv or H37RvΔRv2780 infection for 3, 6, 12, and 24 h (MOI = 2). CFU assay ( I ), histological score ( J ) and histopathological assay with acid-fast staining and haematoxylin and eosin staining ( K ) in lung tissues of C57BL/6 J mice aerosol-infected with roughly 200 CFUs per mouse of H37Rv for 21 days, and treated with Rifampicin (RIF) (10 mg/kg/day) or GWP-042 (10 mg/kg/day) via oral gavage for another 28 days. Data in ( E–J ) are representative of one experiment with at least three independent biological replicates; ( E – H ) n = 3 samples (mean ± s.e.m), each circle represents one technical repeat in ( F – H ); ( I ) n = 13 mice infected for 28 days with red, blue and white circles denoting separate experiments (mean ± s.e.m); ( J ) n = 3 mice (mean ± s.e.m). Two-tailed unpaired Student’s t -test ( F – H ) and two-sided Mann-Whitney U -test ( I , J ) were used for statistical analysis. P values are shown in ( F–J) . 1#, 2# and 3# in ( K ) represent lung tissues from three mice. Scale bars, 100 μm (top; original magnification, ×400) and 20 μm (bottom; original magnification, ×1000). Source data are provided as a file.

Article Snippet: To validate the effectiveness of tuberculosis drug regimens, C57BL/6 J mice were treated with RIF (Rifampicin) (10 mg/kg/day), or combination of RIF (10 mg/kg/day) and Rv2780 inhibitor GWP-042 (MedChemExpress, 91%) (10 mg/kg/day) via oral gavage after three weeks of H37Rv aerosol infection.

Techniques: SPR Assay, Activity Assay, Concentration Assay, Enzyme-linked Immunosorbent Assay, Infection, Colony-forming Unit Assay, Staining, Aerosol, Two Tailed Test, MANN-WHITNEY