Journal: Cell Death & Disease

Article Title: The activated CD36-Src axis promotes lung adenocarcinoma cell proliferation and actin remodeling-involved metastasis in high-fat environment

doi: 10.1038/s41419-023-06078-3

Figure Lengend Snippet: A PA activated Rac1 in a CD36-Src signal-dependent manner. NCI-H23 and A549 cells were treated with or without 500 μM PA for 0.5 h, or cells were pretreated with 50 μM SSO (a fatty acid analog as CD36 blocker), and 10μM Dasatinib (Src inhibitor) for 15 min, respectively, which was followed by 500 μM PA treatment for another 0.5 h. The GTP binding Rac1, Cdc42, and RhoA were detected with kits as mentioned in Materials and Methods and their expression was examined with antibodies as indicated. B Akt kinase but not ERK regulated Rac1 activity. NCI-H23 and A549 cells were treated with or without 500 μM PA for 0.5 h, or cells were pretreated with MEK/ERK inhibitor U0126 (10 μM), and Akt inhibitor API2 (20 μM) for 15 min respectively, which was followed by 500 μM PA treatment for another 0.5 h. GTP binding Rac1 and its expression were detected. C Rac1 colocalized with the finger like actin remodeling after PA treatment. NCI-H23 and A549 cells were treated with or without 500 μM PA for 0.5 h respectively. Cells were double stained for actin (red) and Rac1 (green) respectively. Bar, 5 μm. The images were representative of three experiments. D Rac1 was required for PA-induced actin remodeling. NCI-H23 and A549 cells were transiently transfected with the HA-tagged wild type Rac1 (Rac1 WT), dominant negative Rac1 (Rac1 DN) and constitutively active Rac1 (Rac1 CA) respectively. After 48 h of transfection, cells were treated with or without 500 μM PA for 0.5 h. Then cells were double stained for actin (red) and HA (green) respectively. Bar, 5 μm. The images were representative of three experiments.

Article Snippet: WB antibodies against CD36 (ab133625) were purchased from Abcam (Cambridge, MA), antibodies against Src (A19119), Rac1 (A5539), Cdc42 (A1188) and RhoA (A15641) were from Abclonal (Wuhan, China), antibodies against p-Src-Tyr416 (6943S), p-Src-Tyr527 (2105S), p-Akt473 (9271S), Akt (9272S), p-ERK1/2 (4370S), ERK1/2 (4695S), MMP-9 (13667S), GAPDH (5174S), anti-mouse (7076) and anti-rabbit (7074) HRP-linked IgGs, were purchased from Cell Signaling (Boston, MA).

Techniques: Binding Assay, Expressing, Activity Assay, Staining, Transfection, Dominant Negative Mutation

Journal: Cell Death & Disease

Article Title: The activated CD36-Src axis promotes lung adenocarcinoma cell proliferation and actin remodeling-involved metastasis in high-fat environment

doi: 10.1038/s41419-023-06078-3

Figure Lengend Snippet: A PA treatment led to redistribution of Arp2. NCI-H23 and A549 cells were treated with or without 500 μM PA for 0.5 h respectively. Cells were double stained for actin (red) and Arp2 (green), respectively. Bar, 5 μm. The images were representative of three experiments. B Rac1 regulated redistribution of Arp2. NCI-H23 and A549 cells stable transfected with Rac1-DN were treated with or without 500 μM PA for 0.5 h respectively. Cells were double stained for actin (red) and Arp2 (green), respectively. Bar, 5 μm. The images were representative of three experiments. C PA regulated MMP-9 expression via Akt and ERK. NCI-H23 and A549 cells were treated with or without 10 μM PA for 24 h, or cells were pretreated with MEK/ERK inhibitor U0126 (10 μM), and Akt inhibitor API2 (20 μM) for 30 min, respectively, which was followed by 10 μM PA treatment for another 24 h. MMP-9 and GAPDH expression were determined by western blot. D , E PA treatment led to redistribution of MMP-9 and cortactin. NCI-H23 and A549 cells were treated with or without 500 μM PA for 0.5 h respectively. Cells were double stained for actin (red), MMP-9 (green) and cortactin (green), respectively. Bar, 5 μm. The images were representative of three experiments.

Article Snippet: WB antibodies against CD36 (ab133625) were purchased from Abcam (Cambridge, MA), antibodies against Src (A19119), Rac1 (A5539), Cdc42 (A1188) and RhoA (A15641) were from Abclonal (Wuhan, China), antibodies against p-Src-Tyr416 (6943S), p-Src-Tyr527 (2105S), p-Akt473 (9271S), Akt (9272S), p-ERK1/2 (4370S), ERK1/2 (4695S), MMP-9 (13667S), GAPDH (5174S), anti-mouse (7076) and anti-rabbit (7074) HRP-linked IgGs, were purchased from Cell Signaling (Boston, MA).

Techniques: Staining, Transfection, Expressing, Western Blot