Figure 2.

M5 cytokine stimulation differentially affects MLC activation in monolayer and RHE cultures

(A) Immunofluorescence staining for vinculin at focal adhesions in M5-stimulated N/TERT, single confocal slice in the basal plane.

(B) Immunofluorescence staining for vinculin at the AJs in M5-stimulated N/TERT, single confocal slice in the apical plane.

(C) Quantitative analysis of the vinculin recruitment to AJs shown in (B), data from four independent experiments. Mean ± SD. Paired t-test, ∗p < 0.05.

(D) Western blotting for active Rho in M5-stimulated non-differentiated N/TERT. One representative experiment out of three independent experiments is shown. Mean ± SD. Paired t-test, ∗p < 0.05.

(E and F) Immunluorescence staining for pMLC (E) and ppMLC (F) in non-differentiated N/TERT, confocal max projections. One representative experiment of three is shown.

(G and H) Quantification of pMLC (G) and ppMLC (H) fluorescence intensity per cell for non-differentiated N/TERT. Mean ± SD. Unpaired t-test, ∗p < 0.05, ∗∗∗∗p < 0.0001.

(I and J) Immunofluorescence staining for pMLC (I) and ppMLC (J) in RHEs from N/TERT, confocal max projections.

(K and L) Quantification of pMLC (K) and ppMLC (L) mean fluorescence intensity per field of view in RHEs from N/TERT cells. Mean ± SD. Unpaired t-test, ∗p < 0.05, ∗∗p < 0.01. The stimulation with M5 was done for 24 h in all panels. Scale bars, 20 μm (A and B), 50 μm (E, F, I, and J).