FIG. 4.

Adherence and phagocytosis of calcium-treated erythrocytes are blocked by annexin V and galactose in an additive manner. (A) Human erythrocytes were preincubated for 24 h with 2.5 mM CaCl₂ in order to induce apoptotic-like surface changes. These cells were washed and then incubated with 7 μM annexin V. Each of the three samples was statistically significantly different from the control (* indicates a P value of <0.05 compared to calcium-treated erythrocytes without d-galactose or annexin V; n = 10). The addition of annexin V to galactose further reduced adherence by 15% when compared to galactose alone (# indicates a P value of <0.01; n = 10). (B) Following calcium treatment (2.5 mM) cells were stained with TAMRA, quenched, and preincubated with 7 μM annexin V. Cells were spun onto amebae and incubated at 37°C for 15 min at a 5:1 erythrocyte-to-ameba ratio. Uningested erythrocytes were lysed in water, and the cells were counted by microscopy. Data are reported as means ± SD. Differences between control cells (calcium-treated cells without annexin or galactose) and all other treatments were statistically significant (* indicates a P value of <0.0001 compared to the control). Also, preincubation with annexin V and addition of galactose were additive (# indicates a P value of <0.002 compared to calcium-treated erythrocytes with d-galactose; n = 5).