Fig. 2

αSYN accumulation is reduced when microglia and astrocytes are co-cultured. Schematic figure of the study design illustrating that the cells were either treated with αSYN-F constantly (a) or with a 24h pulse (b). Both astrocytes (Iba1- S100B+, filled arrow heads) and microglia (Iba1+, open arrow heads) ingested and accumulated αSYN in the co-culture set-up (c). Image analysis showed that the microglia contained more αSYN signal per cell at 24h and 4 d, compared to the astrocytes (d). Representative images of the αSYN-F deposits at the different time points in the co-culture are shown in e; astrocytes and microglia are indicated with filled respective open arrow heads. Analysis of the total intracellular αSYN in the co-culture revealed that there was an overall reduction of αSYN at d7 in relation to the 24h time point (f). Separate image analysis of the αSYN content in the two cell types of the co-culture confirmed that the decrease in αSYN signal was significantly lower in both microglia (g) and astrocytes (h) at d7. For the astrocytes, the reduction in αSYN was opposite to the pattern in the pure astrocytic culture, suggesting a lower accumulation of αSYN within astrocytes in the co-culture. When the co-cultures were washed at 24h and cultured without αSYN-F for 3d and 6d, a significant reduction was observed over time in the total co-culture (i), which was due to a decrease in both co-cultured microglia (j) and co-cultured astrocytes (k). Scale bars = 20μm