5.5.3. Effect of TsHpt-I and -II on Phagocytic Function of Murine Peritoneal Macrophages

Peritoneal macrophages from naïve BALB/c mice (*n* = 6) were obtained as described above (item 5.5.1). Cells were distributed in 24-well culture plates (5 × 105 cells/500 <sup>μ</sup>L/well), with glass coverslips inside, and incubated for 2 h at 37 ◦C and 5% CO2. After incubation, non-adherent cells were discarded, and adherent cells (macrophages) were stimulated in vitro with hypotensins, in duplicate, at a concentration of 100 μg/mL, for 24 h at 37 ◦C and 5% CO2. After this period, the culture supernatants were discarded, and the cells were incubated for 1 h with a suspension of *Saccharomyces cerevisiae* at a concentration of 1.5 × 106 yeasts/mL/well. Coverslips were washed 10 times with PBS, fixed with 0.5% glutaraldehyde and stained by Giemsa. Phagocytosis was evaluated by immersion optical microscopy (1000×) and quantified by counting approximately 200 cells per cover slip. The percentage of phagocytic cells and the average number of internalized yeasts per phagocytic cell were calculated. The phagocytic index was obtained by multiplying the two values (percentage × mean) and represents the total phagocytic capacity of each cell population.
