Role of iron in brain injury after intraventricular hemorrhage

Abstract

Background and purpose: Intraventricular extension of hemorrhage is a predictor of poor outcome in intracerebral hemorrhage, and iron overload contributes to brain injury after intracerebral hemorrhage. The current study investigated the role of iron in ventricular dilatation and neuronal death in a rat model of intraventricular hemorrhage (IVH).

Methods: There were 2 parts in this study. First, male Sprague-Dawley rats had a 200-μL injection of either autologous blood or saline into the right lateral ventricle and were euthanized at different time points. Rats had MRI and brains were used for Western blot analysis, immunohistochemistry, histology, and brain tissue nonheme iron measurements. Second, rats had IVH and were treated with deferoxamine or vehicle, and rats were euthanized 4 weeks later for brain tissue loss and lateral ventricle size measurements.

Results: IVH resulted in brain iron accumulation, bilateral enlargement of the lateral ventricles, and hippocampal brain tissue loss. Iron accumulation was associated with upregulation of heme oxygenase-1 and ferritin. Systemic deferoxamine treatment reduced IVH-induced ventricular enlargement (eg, day 28: 32.7±10.6 vs 43.8±9.7 mm(3) in vehicle-treated group, n=8 to 9; P<0.05) and hippocampal brain tissue loss (hippocampal volume: 89.0±2.7 vs 85.2±4.1 mm(3) in the vehicle-treated group; P<0.05).

Conclusions: Iron has a role in brain injury after IVH. Deferoxamine may be a therapy for patients with IVH or intraventricular extension after intracerebral hemorrhage.

Figure 1

(A) T2 weighted MRI scans (coronal brain sections) at day 1 after saline injection and IVH, and the time-course of lateral ventricular volume changes. Values are mean ±S.D., n=7, #p<0.01 vs. saline group. (B) Measurement of hippocampus volume with T2 weighted MRI four weeks after saline infusion and IVH. The hippocampus is outlined on the images. The hippocampus volumes are expressed as the means ± SD, n=7, # p<0.01 versus saline group.

Figure 2

(A) T2* weighted MRI scans (coronal sections) at day 1 and day 28 after IVH, and the time-course of the T2* lesion (hypodensity) after IVH. Values are expressed as the means ± S.D., n=7. (B) Time course of non-heme brain tissue iron in the left and right hemisphere after IVH. Values are mean ± S.D., n=6, *p<0.05 vs. saline control (same time point) and day 1.

Figure 3

(A) Upper panel: Representative Western blots showing increases of HO-1 levels in the hippocampus 7 days after IVH or saline control. Lower panel: Time course of HO-1 in the hippocampus and in the periventricular area. Values are mean±S.D., n=4, *p<0.05 vs. saline control. (B) Immunoreactivity for HO-1 in hippocampus and periventricular area day 3 after IVH or saline injection. Scale bar = 20 μm. (C, D) Time course of HO-1 expression in the hippocampus (C) and in the periventricular area (D). Values are mean ± SD, n=3~7 in saline group and n=5~7 in IVH group, *p<0.05 and #p<0.01 vs. saline group.

Figure 4

(A) Immunoreactivity of ferritin in the hippocampus and the periventricular area day 28 after IVH or saline control. Scale bar = 20 μm. (B) Time course of ferritin levels in the hippocampus. (C) Time course of ferritin levels in the periventricular area. Values are mean ± SD, n=3~7 in saline group and n=5~7 in IVH group, *p<0.05 and #p<0.01 vs. saline group.

Figure 5

(A, C) Representative Western blots showing ferritin L-chain (A) and ferritin H-chain (C) levels in the hippocampus 7 days after IVH or saline control. (B, D) Time course of ferritin L-chain (B) and ferritin H-chain (D) in the hippocampus and in the periventricular area. Values are mean±S.D., n=4, *p<0.05 vs. saline control. ND=not detected.

Figure 6

Bar graphs showing the time course of lateral ventricular volume (A) and T2* lesion volume (B) over four weeks after IVH in rats treated with and without DFX. Hippocampal volume (C) was determined after 4 weeks. Values are expressed as the means ± SD, n=8–9, * p<0.05 vs. the vehicle-treated group.