Figure 7.

CXCL12 mediates wound closure and induces the accumulation of RhoGTP and F-actin in polarized human model epithelium. CXCL12-treated monolayers demonstrated increased closure and increased TER relative to untreated monolayers. (a, top panel) Wounded polarized T84 monolayers were stimulated with 20 ng/ml CXCL12 and photomicrographs taken directly after wounding (b, 0 h) and daily after that for 4-6 days. Relative area of the wounds was calculated using MetaMorph software and wound closure (a, top) was monitored over time by normalizing wound area to 0 h and shown as a percentage of the initial wound area. (a, bottom panel) TER was measured on the same wounded T84 monolayers as measured in the top panel and demonstrated the parallel increase in barrier integrity with wound closure. (b) Representative T84 monolayer wounds at days 0 and 3 of CXCL12 treatment. Wound closure observed on day 3 outlined and superimposed upon the day 0 wound demonstrated the consistent restitution of CXCL12 treated T84 monolayers. Data are representative of 6-17 independent wounds. (c) Photomicrographs demonstrate the significantly increased accumulation of F-actin and RhoGTP at the leading edge of wounds in CXCL12-treated relative to unstimulated cells (no stim). The length of F-actin and RhoGTP fluorescence from the wound edge was quantified using MetaMorph software as described in Materials and Methods. Cells were examined using fluorescence microscopy at × 100 (insets × 200). Immunofluorescence data are representative of three independent monolayers. Values in (a and c) are the mean±s.d. of three independent monolayers. Asterisks denote statistically significant difference from unstimulated (no stim) monolayers (P≤0.05).