Figure 1.
Comparison of chemically and HPF fixed umbrella cells of the urothelium.
mFVs are the most prevailing compartments in chemically (A, C) and in HPF fixed umbrella cells (B, D). Regarding the ultrastructure, mFVs are more dilated in chemically fixed (A, C) than in HPF fixed samples (B, D). Regarding immunolabelling with anti-AUM antibody, pattern and density of labelling is comparable on chemically (C) and HPF fixed samples (D). The organization of mFVs into stacks is better preserved in HPF fixed samples (red over-colour in B and D). Bars: 500 nm.
Figure 2.
(A) mFVs are flattened vesicles with two opposing plaques of thickened asymmetric unit membrane (AUM, blue) and slightly dilated, omega shaped hinge region of un-thickened membrane (yellow). (B) AUM and hinge regions can be seen also in the apical plasma membrane. (C) By freeze-fracturing, uroplakin particles are seen concentrated in the centre of the mFV (blue), while thin hinge region contains particle-free membranes (yellow). (D–G) Three-dimensional model of mFV shows that it has the shape of a flattened disk. In D, a slice from a tomogram is shown. In E–G, a 3D model of a mFV in different projections is shown (green). Bars: 100 nm in A, C; 50 nm in B.
Figure 3.
Stacks of mFV in the central cytoplasm of umbrella cells.
(A) Cytoplasm of umbrella cells is divided into two regions: sub-apical (SA) and central (CE). Central cytoplasm contains majority of mFVs and other organelles. The border between the sub-apical and central cytoplasm is shown by a dashed line. (B) In the central cytoplasm, mFVs are often arranged into stacks. (C) Three-dimensional model of a stack shows that mFVs grouped into a stack have the same shape of flattened disk as individually positioned mFVs. Model of four stacked mFVs is presented. (D) Immunolabelling with anti-Rab27b antibody shows positive reactions (arrows) on some mFVs and some iFVs, while there is no labelling seen on the apical plasma membrane (arrowhead). Legend: M – mitochondrion, asterisk – mFV without anti-Rab27b labelling. Bars: 250 nm.
Figure 4.
Orientation of mFVs in the sub-apical cytoplasm changes during bladder stretching.
In umbrella cells of the contracted bladders (A–F), mFVs are oriented mostly perpendicular to the apical plasma membrane, while in the distended bladders (G), mFV are oriented mostly parallel to the apical plasma membrane. Panels B–F show representative slices from the tomogram. In panel D, a contact (arrow) between mFVs and the apical plasma membrane (arrowhead) is seen. White box in D corresponds to the area shown in B, C, E and F. Bars: 250 nm.
Figure 5.
Mature FVs are only present in umbrella cells.
(A) In umbrella cells, some stacks of FVs contain mFVs, which are densely labelled with anti-AUM antibody, and iFVs, which are shorter and more convex. Such stacks are surrounded by rounded uroplakin-positive (short black arrow) and uroplakin-negative (striped arrow) vesicles. (B) In the intermediate cell (ic), mFVs are not present; instead iFVs are seen below the plasma membrane (arrowhead) that borders the basolateral plasma membrane of the umbrella cell (uc). (C) Three-dimensional model of iFVs in the intermediate cell. (D) iFVs of intermediate cell (ic) are less densely labelled with anti-AUM antibody than mFVs of umbrella cell (uc). Bars: 500 nm.