Figure 2

Effects of rtTNFα treatment in vitro on apoptosis and cell viability in brown trout preovulatory follicles. A. To determine the incidence of apoptosis, granulosa cells isolated from trout preovulatory follicles incubated in the absence or presence of rtTNFα (100 ng/ml) were stained with propidium iodide (PI) and analyzed by flow cytometry (FACS analysis). Each bar represents the mean ± SE of six independent experiments, each performed with ovarian tissue from a different female (n = 6). The results are expressed with respect to the control group which has been set at 1. Statistically significant (p < 0.05) differences with respect to the control group are indicated by an asterisk (*). B. Granulosa cell viability was assessed by incubating brown trout preovulatory follicles in the absence or presence of 100 ng/ml rtTNFα, or 1% paraformaldehyde (PFA) as a positive control, for 18 h at 15°C, separating the granulosa layers and staining them with PI. Subsequently, sheets of granulosa layers were mounted onto glass slides, counterstained with DAPI and visualized under a fluorescent microscope.