Figure 5. Claudin5 regulates junction protein expression.
(A–C) Area (A) width (B) and intensity (C) of electron dense regions in the ear skin of control and Cldn5 iECKO mice after visualisation by TEM. Right, representative TEM images of junctions in the ear skin of control and Cldn5 iECKO mice. Junctions can be seen within electron dense regions (arrowheads). L, lumen. n≥2 mice, 6 or more fields of view/mouse. (D) Distance of HRP penetrance into EC junctions in the ear skin of control and Cldn5 iECKO mice following systemic histamine stimulation. Right, representative TEM images of HRP penetrance (visualized by electron dense 3,3′-Diaminobenzidine (DAB) reaction precipitate) into EC junctions in the ear skin of control and Cldn5 iECKO mice following systemic histamine stimulation. Dashed regions show areas of disrupted junction into which HRP has penetrated. Note that the typical electron dense area is lacking due to absence of uranyl acetate staining. L, lumen. n≥2 mice, 6 or more fields of view/mouse. (E) Gene expression of AJ- and TJ-associated genes in lung lysates of control and Cldn5 iECKO mice. n≥4 mice. (F) Expression of AJ- and TJ- associated proteins in lung lysates of control and Cldn5 iECKO mice. Right, representative western blots of AJ- and TJ- associated proteins in lung lysates of control and Cldn5 iECKO mice. n≥4 mice. (G) Expression of ZO-1 in ear skin blood vessels of control and Cldn5 iECKO mice. Left, quantification of ZO-1. Right, representative images of ZO-1 in the ear skin of control and Cldn5 iECKO mice. n≥6 mice, 3 or more fields of view/mouse. (H) Expression of VE-Cadherin in ear skin blood vessels of control and Cldn5 iECKO mice. Left, quantification of VE-Cadherin. Right, representative images of VE-Cadherin in the ear skin of control and Cldn5 iECKO mice. n≥9 mice, 3 or more fields of view/mouse. (I) Quantification of ZO-1 (left) and VE-Cadherin (right) in back skin blood vessels of control and Cldn5 iECKO mice. n≥8 mice, 2 or more fields of view/mouse. (J) Quantification of ZO-1 in different vessel subtypes in the ear skin of control and Cldn5 iECKO mice. n≥3 mice, 3 or more fields of view/mouse. (K) Quantification of VE-Cadherin in different vessel subtypes in the ear skin of control and Cldn5 iECKO mice. n≥3 mice, 3 or more fields of view/mouse. Error bars; mean ± SD. Statistical significance: two-tailed paired Student’s t test.
Figure 5—figure supplement 1. Endothelial junction protein expression in Cldn5 iECKO mice.
