Fig. 1

Cellular response following exposure to hemorrhagic CSF. a–h Representative images are from mixed glial cell cultures following exposure to 3% hemorrhagic CSF (containing 7 μM cell-free Hb) from preterm human infants with GM-IVH. Images illustrate the detected IF labeling of Iba1 (red; b, d, f, h), F-actin cytoskeletal protein (phalloidin, green; c, d, g, h) and DAPI nuclear staining (blue; a, d, e, h), in the wells of mixed glial cells exposed to culture media only (control; a–d) and in wells exposed to hemorrhagic CSF (e–h). d, h Merged images of DAPI, Iba1, and phalloidin. Scale bar in h indicates 200 μm and is representative for a–h. i ROS production was investigated (by performing the DCFDA assay as described in materials and method) following a 24 h exposure of mixed glial cells to 3% hemorrhagic CSF (containing 7 µM cell-free Hb). j-m TNFα and HO-1 mRNA expression were analyzed at 8 and 24 h after exposure to 3% hemorrhagic CSF (containing 7 µM cell-free Hb). Hemorrhagic CSF (dark gray bars, n = 6), co-administration with Hp2-2/2-1 (10 µM, light gray bars, n = 6) and control cells (white bars, n = 6). Results are presented as box plots displaying medians and 25th and 75th percentiles. Differences between hemorrhagic CSF vs. control and hemorrhagic CSF with co-administered Hp vs. control were analyzed using the Kruskal–Wallis test followed by pairwise comparison with significance values adjusted for multiple comparisons. *P < 0.05, **P < 0.01