γh2ax  (Bethyl)

Journal: Genes & Diseases

Article Title: Targeting fatty acid synthase to overcome PARP inhibitor resistance and to create an artificial synthetic lethality for triple-negative breast cancer

doi: 10.1016/j.gendis.2025.101817

Figure Lengend Snippet: Combination synergy on DNA damage, NHEJ repair, and apoptosis. (A, B) Western blotting and quantification analyses of FASN, PARP1, γ-H2AX, and actin loading control in MDA-MB-231 and MDA-MB-436 cells treated with 5HLS, talazoparib, or the combination. (C, D) The fraction of maximum γ-H2AX induction derived using the Bliss-compatible scaling formula. (E) Host cell reactivation assay of NHEJ activity in MDA-MB-231 and MDA-MB-436 cells treated with 5HLS, talazoparib, or the combination. (F) Comparison between the observed and expected NHEJ activity inhibition by the combination using the Bliss independence model. (G) Caspasae3/7 activity assay of MDA-MB-231 and MDA-MB-436 cells treated with 5HLS, talazoparib, or the combination. (H) The fraction of maximum caspase 3/7 activation derived using the Bliss-compatible scaling formula. (I) Annexin V staining as an indicator of apoptosis in MDA-MB-231 and MDA-MB-436 cells following treatments with 5HLS, talazoparib, or the combination. (J) The fraction of maximum apoptosis induction derived using the Bliss-compatible scaling formula. n = 3; ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and ∗∗∗∗ P < 0.0001.

Article Snippet: Antibodies against PARP1 (#66520) and phosphorylated histone H2AX (γH2AX) (#613402) were from Proteintech (Rosemont, Illinois, USA) and BioLegend (San Diego, California, USA), respectively.

Techniques: Western Blot, Control, Derivative Assay, Host-Cell Reactivation, Activity Assay, Comparison, Inhibition, Activation Assay, Staining

Journal: Genes & Diseases

Article Title: Targeting fatty acid synthase to overcome PARP inhibitor resistance and to create an artificial synthetic lethality for triple-negative breast cancer

doi: 10.1016/j.gendis.2025.101817

Figure Lengend Snippet: In - vivo effect of 5HLS and talazoparib combination on tumor growth. (A) Tumor volume and body weight of mice treated by vehicle (Veh), 5HLS, talazoparib (Tal), or combination (Comb) of 5HLS and talazoparib. (B, C) Gross anatomy (B) and wet weight (C) of dissected xenograft tumors at the end of the study. (D) Synergy analysis. Tumor growth inhibition (TGI) was derived using the wet weight of tumors. The expected combination inhibition (C (E) ) was calculated from that of 5HLS and talazoparib alone using the Bliss independence model (see Materials and Methods ). C (O) represents the observed combination inhibition. (E) Immunohistochemical analyses of FASN, PARP, and γH2AX. Scale bar, 50 μm. (F) Western blotting analyses of FASN, BRCA1, PARP1, cleaved PARP1 (cPARP1), γH2AX, and actin loading control in xenograft tumors from mice treated with vehicle, 5HLS, talazoparib, and the combination of 5HLS and talazoparib. Each lane represents mixed samples of five tumors in equal proportion within the treatment group. n = 5; ∗ P < 0.05, ∗∗ P < 0.01, and ∗∗∗ P < 0.001.

Article Snippet: Antibodies against PARP1 (#66520) and phosphorylated histone H2AX (γH2AX) (#613402) were from Proteintech (Rosemont, Illinois, USA) and BioLegend (San Diego, California, USA), respectively.

Techniques: In Vivo, Inhibition, Derivative Assay, Immunohistochemical staining, Western Blot, Control

Journal: Clinical and Translational Radiation Oncology

Article Title: Overexpression of JAML in colorectal cancer cells predicts higher radiosensitivity by inactivating ATR pathway

doi: 10.1016/j.ctro.2025.101016

Figure Lengend Snippet: CRC cells with JAML overexpression demonstrate increased apoptosis and DNA damage after irradiation, accompanied by inhibiting the ATR-CHK1 signaling pathway. (A) Representative images of apoptosis of LOVO and DLD-1 cells using flow cytometry after 24 h after X-ray irradiation. (B and C) Quantitative statistics of apoptosis of LOVO and DLD-1 cells. (D) Representative images of fluorescence of γH2AX in LOVO and DLD-1 cells after 12 h of X-ray irradiation or lack of X-ray irradiation. (E and F) Quantitative statistical analysis of the γH2AX-positive rate in LOVO and DLD-1 cells. (G and L) Western blot images of the ATR-CHK1 signaling pathway’s protein expression in LOVO and DLD-1 cells after 24 h of X-ray irradiation/lack of irradiation. (H–K and M−P) Quantitative statistical analysis of the relative ATR-CHK1 signaling pathway’s protein expression in LOVO and DLD-1 cells (*, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; and ****, P ≤ 0.0001).

Article Snippet: In brief, at 12 h after 8-Gy X-ray irradiation, the cells were cultured on 24-well chamber slides, washed with PBS, fixed with 4 % paraformaldehyde (Biosharp, China), permeabilized with 0.1 % TritonX-100 (Solarbio, China), blocked with 3 % bovine serum albumin (BSA), and incubated with an antibody against γH2AX (Cell Signaling Technology, dilution 1:200, USA) at 4 °C overnight.

Techniques: Over Expression, Irradiation, Flow Cytometry, Fluorescence, Western Blot, Protein-Protein interactions, Expressing

Journal: Clinical and Translational Radiation Oncology

Article Title: Overexpression of JAML in colorectal cancer cells predicts higher radiosensitivity by inactivating ATR pathway

doi: 10.1016/j.ctro.2025.101016

Figure Lengend Snippet: LOVO cells with JAML overexpression demonstrate increased apoptosis and DNA damage after irradiation. (A)Representative image and (B) protein quantitative analysis of Western blot analysis of JAML in lentivirus-overexpression-transfected LOVO cells. (C) mRNA expression levels of JAML in lentivirus-overexpression-transfected LOVO cells. (D) Representative images and (E) survival curve of plate cloning of JAML overexpressed in LOVO cells after X-ray (8 Gy) irradiation. (F) Representative images and (G) quantitative analysis of cellular immunofluorescence of γH2AX (red light) of JAML overexpressed in LOVO cells after X-ray (8 Gy) irradiating 12 h. (H) Representative images and (I) quantitative analysis of flow cytometry apoptosis of JAML overexpressed in LOVO cells after X-ray (8 Gy) irradiating 24 h. (*, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001; and ****, P ≤ 0.0001). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: In brief, at 12 h after 8-Gy X-ray irradiation, the cells were cultured on 24-well chamber slides, washed with PBS, fixed with 4 % paraformaldehyde (Biosharp, China), permeabilized with 0.1 % TritonX-100 (Solarbio, China), blocked with 3 % bovine serum albumin (BSA), and incubated with an antibody against γH2AX (Cell Signaling Technology, dilution 1:200, USA) at 4 °C overnight.

Techniques: Over Expression, Irradiation, Western Blot, Transfection, Expressing, Cloning, Immunofluorescence, Flow Cytometry