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. 2021 Apr 15;11(13):6173–6192. doi: 10.7150/thno.58254

Figure 8.

Figure 8

Effects of the 9-kDa C-terminal SPARC fragment on TNBC cell adhesion, migration, transmigration and invasion. (A) Cell adhesion. MDA-MB-231 cells were let to adhere for 30 min on a fibronectin matrix in the presence of recombinant FL SPARC, recombinant cleaved SPARC fragments (cleaved SPARC), or purified 9-kDa C-terminal SPARC fragment at a final concentration of 240 nM. Left panels, representative images of adherent cells stained with crystal violet. Right panel, adhesion was quantified as described in Figure 6 A. Data are the mean ± SD (n = 3); ns, not significant; ***, p < 0.001, ANOVA and Bonferroni's post hoc test. CTRL, PBS in cleavage buffer and SPARC-immunodepleted supernatant from the 9-kDa SPARC fragment purification. Similar results were obtained in three independent experiments. (B) Cell migration. MDA-MB-231 cells were let to migrate for 16 h on a fibronectin matrix in the absence or presence of FL SPARC, cleaved SPARC fragments, or the 9-kDa C-terminal SPARC fragment at a final concentration of 240 nM. Left panels, representative images of migrating cells stained with crystal violet. Right panel, migration was quantified as described in Figure 6B. Data are the mean ± SD (n = 3); ***, p < 0.001, ANOVA and Bonferroni's post hoc test. CTRL, PBS in cleavage buffer and SPARC immunodepleted supernatant from the 9-kDa SPARC fragment purification. Similar results were obtained in two independent experiments. (C) Endothelial transmigration. MDA-MB-231 cells were let to transmigrate for 16 h through a HUVEC monolayer in the absence or presence of FL SPARC, cleaved SPARC fragments, or the 9-kDa C-terminal SPARC fragment at a final concentration of 240 nM. Left panels, representative images of transmigrating cells. Right panel, transmigrating cells were stained with MTT and quantified by absorbance at 570 nm. Data are the mean ± SD (n = 3); *, p < 0.05, **, p < 0.01, ***, p < 0.001, ANOVA and Bonferroni's post hoc test. CTRL, PBS in cleavage buffer and SPARC-immunodepleted supernatant from the 9-kDa SPARC fragment purification. Similar results were obtained in two independent experiments. (D) Cell invasion. MDA-MB-231 cells were let to invade for 16 h on a Matrigel matrix in the absence or presence of FL SPARC, cleaved SPARC fragments, or the 9-kDa C-terminal SPARC fragment at a final concentration of 240 nM. Left panels, representative images of invading cells stained with crystal violet. Right panel, invading cells were quantified by absorbance at 570 nm. Data are the mean ± SD (n = 3); *, p < 0.05, **, p < 0.01, ***, p < 0.001, ANOVA and Bonferroni's post hoc test. CTRL, PBS in cleavage buffer and SPARC immunodepleted supernatant from the 9-kDa SPARC fragment purification. Similar results were obtained in two independent experiments.