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Figure 3 | Molecular Cancer

Figure 3

From: The role of CCL21/CCR7 chemokine axis in breast cancer-induced lymphangiogenesis

Figure 3

CCL21/CCR7 axis has lymphangiogenic potential in vitro. (A) Real-time PCR of CCR7 and CCL21 mRNA expression in HMVEC-dLy. (B) Western blot of CCR7 and CCL21 protein expression in HMVEC-dLy. GAPDH was used as an internal control. (C) CCL21 protein secretion by HMVEC-dLy as measured by ELISA. Data are represented as mean ± SD (n = 3). (*) indicates significant difference (p < 0.05). (D) HMVEC-dLy proliferation in response to CCL21/6Ckine (0, 100, 200, 350 ng/ml) performed by the measurement of BrdU. Data are presented as mean ± SD (n = 4, p < 0.001). (E) HMVEC-dLy proliferation in response to CCR7 neutralizing antibody (0, 5, 10, 20 μg/ml). Data are presented as mean ± SD (n = 4, p < 0.0003). (F) Representative images of HMVEC-dLy migration (40× magnification). (G) Quantification of HMVEC-dLy cellular migration in response to CCL21/6Ckine (0, 100, 200, and 350 ng/ml). (H) Quantification of HMVEC-dLy migration in response to CCR7 neutralizing antibody (0, 5, 10, 20 μg/ml). Bars in (G, H) represent mean number of migrated cells ± SD (n = 4, p < 0.005). (I) Representative micrographs of HMVEC-dLy tubular network formation in response to CCL21/6Ckine (0, 100, 200, 350 ng/ml). Bar equals 100 μm. (J) Quantification of total length of tubular structures formed by HMVEC-dLy corresponding to (I) as determined by ImageJ. (K) Representative images of inhibition of HMVEC-dLy tubular network formation. Bar equals 100 μm. (L) Quantified HMVEC-dLy tube formation corresponding to images shown in (K). In (J and L) data are presented as mean ± SD (n = 4, p < 0.0001). (M) Quantified HMVEC-dly proliferation, migration, and tube formation in response to treatment with VEGFR-3 neutralizing antibody (0, 1, 2.5, 5 μg/ml). Data are presented as mean ± SD. Different superscripts represent a statistical significant difference.

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