Table 3.
Fluorescence Analyses of Junction Proteins and the Actin Cytoskeleton. Fluorescence intensity measurements were made using the line tool in MetaMorph Image Analysis software at the cell junction. The ratio of junctional to intracellular fluorescent is reported and the fluorescent intensity at the cell junctions are reported ± SEM. Actin staining was quantified at apical z-section stacks that corresponded to the same plane as the tight junction. Fluorescent intensity (arbitrary units) at the cell junction is reported in the lower panel. ANOVA was performed followed by a Bonferroni post test with all comparisons against the control group. A minimum of 50 junctions were analyzed in each group *P < 0.001.
| Fluorescence Intensity Ratio (Junctional/Intracellular) | |||
| Tight Junction Protein | Control | TNFα/IFNγ (10 and 20 ng/ml) | TNFα/IFNγ with U0126 (1 μM) |
| Occludin | 29.12 ± 1.79 | *16.24 ± 0.78 | *21.42 ± 1.15 |
| Claudin-1 | 29.70 ± 2.79 | *11.65 ± 1.53 | *22.08 ± 1.51 |
| Claudin-2 | 29.97 ± 1.76 | *24.57 ± 1.98 | *21.35 ± 1.73 |
| Claudin-3 | 7.27 ± 0.57 | 7.11 ± 0.71 | 9.45 ± 0.88 |
| Fluorescence Intensity (Junctional) | |||
| Actin | 91.55 ± 5.2 | *191.34 ± 10.6 | 116.5 ± 5.0 |
| Occludin | 644.7 ± 25.8 | *917.6 ± 28.4 | *1238.1 ± 30.2 |
| Claudin-1 | 668.3 ± 27.6 | *469.3 ± 22.9 | 557.4 ± 21.7 |
| Claudin-2 | 545.4 ± 24.3 | *364.0 ± 12.7 | *357.4 ± 19.3 |
| Claudin-3 | 204.8 ± 18.2 | 212.7 ± 18.3 | 250.0 ± 18.7 |