Figure 5
CXCL6 is an important paracrine factor in CPC CM. (a) ELISA evaluation of CXCL6 concentration in CPC (CPC1, CPC3), HDF and MSC CM. Data are shown as mean ± SD (n = 3). (b) Western blot analysis of CXCL6 (11.9 kDa) in CPC (CPC1-3), HDF and MSC CM; β-actin was used as loading control. (c) Western blot analysis of CXCR1 (53 kDa) and CXCR2 (45 kDa) in CPC (CPC1, CPC3), HDF and MSC lysates; β-actin was used as loading control. (d) Immunocytochemical analysis of CXCR1 and CXCR2 expression in CPC (CPC1-3) and MSC; cells were counterstained with laminin (green) and nuclei with DAPI (blue); bars, 20 μm. (e) Role of CXCL6 in pro-repair activity (migration) of CPC CM. Culture with CPC3 CM (1:50 dilution) induced migration of THP-1 cells in comparison with control medium. Coincubation with anti-CXCR1, anti -CXCR2 or anti -CXCL6 antibodies were tested for their ability to interfere with THP-1 cell migration. (f) CXCL6 in the angiogenic activity of CPC CM. HUVEC were incubated with CPC1-CPC3 CM, alone or with anti-CXCL6 antibody (50 nM). After 6 h, cultures were analyzed using ImageJ. Data expressed as mean ± SD (n = 3); black lines indicate p-values (***< 0.002, **< 0.02, *< 0.05; one-way analysis of variance with Bonferroni multiple comparison test).
