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. 2017 Jun 27;8(47):81880–81891. doi: 10.18632/oncotarget.18737

Figure 4. 3T3-A-EXO transferred MMP3 to 3LL tumor cells.

Figure 4

(A) MMP3 and MMP9 protein levels in 20 μg 3T3-EXO, 3T3-A-EXO, 3T3-L1 or 3T3-L1 lysates were detected by Western blot. (B) 3LL cells were pre-treated with 10 μg/ml cytochalasin D for 30 min and then co-cultured with CFSE-labeled 3T3-A-EXO for 4 h. The uptake of 3T3-A-EXO by 3LL cells was detected by confocal microscopy. (C, D) After the pre-treatment with 10 μg/ml cytochalasin D for 30 min, 3LL cells were collected and re-cultured in fresh RPMI 1640 media containing 10% FBS (C) or treated with 30 μg/ml 3T3-A-EXO (D) for 4 h. MMP3 protein levels in these cells were detected by Western blot. (E) After the pre-treatment with 10 μg/ml cytochalasin D for 30 min, 3LL cells were treated with 30 μg/ml 3T3-A-EXO for 4 h, and then the cells were collected and cultured in serum-free RPMI 1640 media for an additional 24 h. The MMP2 and MMP9 activity in the supernatants was detected by gelatin zymography. (F) 3T3-L1 adipocytes were transfected with MMP3 siRNA or negative control (NC) siRNA for 24 h, and then MMP3 protein levels in the cells or 3T3-A-EXO were detected by Western blot. (G) 3LL cells were treated with 30 μg/ml NC 3T3-A-EXO or MMP3KD 3T3-A-EXO for 4 h, and then MMP3 protein levels were detected by Western blot. (H) 3LL cells were treated with 30 μg/ml 3T3-A-EXO in the presence of 10 μg/ml cycloheximide for 4 h, and then MMP3 protein levels were detected by Western blot. The data are representative of three independent experiments. Control indicates 3LL cells treated with PBS. (C-D, E, H) DMSO is solvent control.