FIGURE 1.

Cx43 is expressed in macrophages and localizes to phagosomes. A, Macrophage cell lines, including J774 macrophages (J774), RAW 264.7 (RAW), and primary murine peritoneal macrophages (Mθ) from Cx43 wild-type (+/+) and heterozygous (+/−) mice, were subjected to SDS-PAGE and were assessed for the expression of Cx43 by immunoblotting. Blots were stripped and reprobed with Abs against F-actin. B, RAW264.7 macrophages were allowed to internalize Dynabeads for the indicated periods of time (5, 15, 30, 60, and 90 min) and then washed with copious amounts of PBS, fixed, permeabilized, and immunostained with Abs to Cx43 and imaged by confocal microscopy. Shown are the corresponding images for Cx43 (green) and DIC at each time point indicated. Arrows show the presence of bound (at 5 min) or internalized particles (at all other time points). Representative of three separate experiments. Size bar = 3 μm. C, Phagosomes (Phag) were isolated from J774 macrophages that had been fed a meal of latex beads and immunoblotted alongside J774 cells lysates (Mθ) using Abs against Cx43, LAMP-2, a phagosomal marker, and p38-MAPK (p38), a cytoplasmic protein expectedly absent from phagosomal membranes. Representative of three separate experiments. D, Rate of phagocytosis after 1 h of either Dynabeads (open bars) or latex beads (filled bars) in RAW264.7 macrophages (open bars) or J774 cells (filled bars) that had been either untreated, treated with anti-FcR, or treated with nonspecific IgG.