Cell junctions are more sensitive to hypertonic shock in claudin/JAM-A KO cells. (A) Control MDCK II cells had continuous cell junctions indicated by ZO-1 staining. (B) MDCK II cells treated with hypertonic solution showed sporadic junction breakage (B′, arrow). (C) Control claudin/JAM-A KO cells occasionally exhibited discontinuity in ZO-1 staining (asterisk). (D) Claudin/JAM-A KO cells treated with hypertonic solution exhibited numerous discontinuities in ZO-1 staining (D′, arrows). Enlarged images of the regions with yellow brackets are shown in A′–D′. (E) Time-lapse imaging of MDCK II cells expressing ZO1-GFP treated with hypertonic solution. Sporadic junction breakage was observed (arrows). (F) Time-lapse imaging of claudin/JAM-A KO cells expressing ZO1-GFP treated with a hypertonic solution. A marked increase in junction breakage was observed after the application of hypertonic shock (arrows). (G) Quantification of the junction breakage phenotype during treatment with a hypertonic solution. The data represent the numbers of endpoints of ZO-1 staining per unit area at each timepoint and are presented as mean ± SD (n = 4). Scale bars: 10 μm.