Attenuation of hematoma size and neurological injury with curcumin following intracerebral hemorrhage in mice

Abstract

Object: Intracerebral hemorrhage (ICH) is associated with significant morbidity and mortality. Acute hematoma enlargement is an important predictor of neurological injury and poor clinical prognosis; but neurosurgical clot evacuation may not be feasible in all patients and treatment options remain largely supportive. Thus, novel therapeutic approaches to promote hematoma resolution are needed. In the present study, the authors investigated whether the curry spice curcumin limited neurovascular injury following ICH in mice.

Methods: Intracerebral hemorrhage was induced in adult male CD-1 mice by intracerebral administration of collagenase or autologous blood. Clinically relevant doses of curcumin (75-300 mg/kg) were administered up to 6 hours after ICH, and hematoma volume, inflammatory gene expression, blood-brain barrier permeability, and brain edema were assessed over the first 72 hours. Neurological assessments were performed to correlate neurovascular protection with functional outcomes.

Results: Curcumin increased hematoma resolution at 72 hours post-ICH. This effect was associated with a significant reduction in the expression of the proinflammatory mediators, tumor necrosis factor-α, interleukin-6, and interleukin-1β. Curcumin also reduced disruption of the blood-brain barrier and attenuated the formation of vasogenic edema following ICH. Consistent with the reduction in neuroinflammation and neurovascular injury, curcumin significantly improved neurological outcome scores after ICH.

Conclusions: Curcumin promoted hematoma resolution and limited neurological injury following ICH. These data may indicate clinical utility for curcumin as an adjunct therapy to reduce brain injury and improve patient outcome.

Conflict of interest statement

DISCLOSURES

The authors report no conflicts of interest.

Figures

Figure 1. Chemical structure of curcumin

Figure 2. Curcumin promotes hematoma resolution after collagenase-induced ICH

(A) Mice were treated with placebo (left panels) or curcumin (150 mg/kg; right panels) at the time of ICH. At 24h, 48h, or 72h post-ICH, coronal brain sections were prepared and digitally captured. Each panel depicts serial brain sections from a single, representative mouse per group. (B) Determination of hematoma area (mm2) at 48h or 72h post-ICH. Hematoma size was estimated by calculating the area inside a region of interest, as detailed in the Materials and Methods section (C) Hematoma volume was quantified by determining the hemoglobin content of each hemisphere at 72h post-ICH. Data are expressed as mean ± SEM and were analyzed by t-test or One-Way ANOVA (n=8 mice/group) followed by Student Newman-Keul’s post-hoc test (**p<0.01, ***p<0.001).

Figure 3. Curcumin promotes hematoma resolution after autologous blood clot-induced ICH

(A) Mice were treated with placebo (left panel) or curcumin (150 mg/kg; right panel) at the time of ICH. At 72h post-ICH, coronal brain sections were prepared and digitally captured. Each panel depicts a brain sections from a single, representative mouse per group (n=8 mice/group). (B) Hematoma volume was quantified by determining the hemoglobin content of each hemisphere at 72h post-ICH. Data are expressed as mean ± SEM (n=8 mice/group) and were analyzed by One-Way ANOVA followed by Student Newman-Keul’s post-hoc test (**p<0.01).

Figure 4. Establishment of a therapeutic window for curcumin to reduce hematoma volume

(A) Mice were treated with placebo or curcumin (150 mg/kg) at 15 minutes prior to injury or 0.5h, 1h, or 3h following collagenase-induced ICH. Hematoma volume was quantified 72h later using a hemoglobin content assay. Data are expressed as mean ± SEM (8 mice/group) and graphed as “% change from placebo-treated ICH mice”. Groups were analyzed using a One-Way ANOVA followed by Student Newman Keul’s post-hoc test (*p<0.05, **p<0.01, ***p<0.001 vs. ICH). Data are expressed as mean ± SEM (n=8 mice/group) and analyzed using One-Way ANOVA followed by Student Newman-Keul’s post-hoc test (*p<0.05, **p<0.01).

Figure 5. Effect of curcumin on inflammatory gene expression

Mice were treated with curcumin (150 mg/kg) at the time of collagenase-induced ICH following by the quantification of inflammatory gene expression. (A) IL-6, (B) IL-1β, and (C) TNF-α expression was assessed in the peri-hematoma region at 24h post-ICH using qRT-PCR. Data were normalized to RPS3 and expressed as fold change vs. sham-operated mice (mean ± SEM; n=8 mice/group) Data were analyzed using One-Way ANOVA followed by Student Newman-Keul’s post-hoc test (*p<0.05, **p<0.01, ***p<0.001 vs. sham).

Figure 6. Curcumin reduces BBB permeability after ICH

Mice were administered curcumin (150 mg/kg) at the time of collagenase-induced ICH. Evans blue extravasation, a sensitive measure of BBB disruption, was assessed at (A) 6h or (B) 12h or (C) 24h post-ICH. Comparisons between treatments groups were done using a one-way ANOVA followed by Student Newman Keul’s post-hoc test (**p<0.01, ***p<0.001). Data are expressed as mean ± SEM from 10 mice/group.

Figure 7. Curcumin attenuates brain edema following ICH

Mice were administered curcumin (150 mg/kg) at the time of collagenase-induced ICH. Brain water content, a measure of cerebral edema, was assessed in the ipsilateral and contralateral hemispheres at (A) 24h or (B) 72h following ICH. Comparisons within each hemisphere between different treatments groups were done using a one-way ANOVA followed by Student Newman Keul’s post-hoc test (*p<0.05, ***p<0.001). No significant differences were observed between groups in the contralateral hemispheres. Data are expressed as mean ± SEM from 10 mice/group.

Figure 8. Curcumin improves neurological outcomes following ICH

Mice were treated with placebo or curcumin (150 mg/kg) at the time of collagenase-induced ICH. Neurological outcomes were then assessed at 24h, 48h, or 72h following sham injury or ICH (n=10 mice/group. Data are the mean ± SEM and were analyzed using a repeated measures analysis of variance (ANOVA) followed by Bonferroni’s post-hoc test (**p