Fig. 6

Enhanced survival of IL-22-treated primary astrocytes in insulting conditions. Primary astrocyte cells (HA) were cultured in 24-well plates starting at day −1 and were treated every second day starting at day 0 with astrocyte medium (AM)—representing the optimal culture medium for astrocytes—or with RPMI (poor medium, hereafter referred to as untreated), with or without addition of the following: IL-22 alone, TNFα alone, IL-22 and TNFα together, or still staurosporine (STS) alone, the latter representing a potent inducer of apoptosis. Cells were stained with 7-AAD and Annexin V and analyzed by flow cytometry to assess their survival and apoptotic status. Histograms represent 7-AAD profile of untreated versus IL-22 (a) and of TNFα versus IL-22 + TNFα (b). Survival profile analysis was performed by flow cytometry by following the frequency of living cells (i.e., 7-AAD-negative cells, Fig. 6 c ) and, among those that were not dead (7-AAD-negative cells), by following the proportion of those surviving cells but which underwent apoptosis (d). Each dot represents the median of six replicates, except for AM and STS conditions (three replicates). Orange arrows indicate treatment renewal. Considering STS treatment, 7-AAD kinetic was stopped after 4 days as all recovered cells were dead at this time point.*comparison of IL-22-treated versus untreated cells; †comparison of TNFα- versus TNFα + IL-22-treated cells. The vertical bars determine the 75th percentile of the median. Significance was calculated with unpaired non-parametric Mann–Whitney test. * or †P < 0.05, ** or ††P < 0.01