Fig. 4
Cell morphological effects of knockdown of LAD1. (a) Representative immunofluorescence images in HSC-4 cells on day three after plating; siRNAs against LAD1 (si#1) and negative control (cont). (b) The fractal dimension was obtained by the fractal analysis of immunofluorescence images using FracLac; scale bars, 50 μm (a). In immunofluorescence, the HSC-4 cells (cont) form radially extended filopodia and widely spread lamellae (asterisk), which are composed of thick and elongated actin filaments (white arrows). At the base of the filopodia and lamella, LAD1 is localized along the actin arc. The cells form polygonal and paved shapes when in contact with each other. In contrast, LAD1-knockdown cells appear shrunken with irregularly formed filopodia (white arrowheads) and a lack of lamellar expansion, resulting in a further irregular ruffled cell border shape, in which LAD1 is not localized. The fractal analysis by the box-counting method on auto-segmentation images showed HSC-3 and HSC-4 indicated significantly (p < 0.05) higher average values of fractal dimension in LAD1 knockdown cells by si#1. Inversely, HSC-2 showed a decrease in fractal dimension in si#2.
