Delayed administration of interleukin-1 receptor antagonist protects against transient cerebral ischaemia in the rat

Abstract

The cytokine interleukin-1 (IL-1) has been implicated in ischaemic, excitotoxic and traumatic brain damage in rodents. The naturally occurring IL-1 receptor antagonist (IL-1ra) markedly reduces neuronal injury in these conditions. However, the effects of IL-1ra on focal, transient cerebral ischaemia in the rat, which is of major clinical relevance, have not been reported. The objectives of this study were to test the effects of IL-1ra on cell death after temporary cerebral ischaemia, and to investigate the therapeutic time window for IL-1ra treatment. Ischaemia was induced by temporary (60 min) occlusion of the middle cerebral artery (MCAO) in rats, via surgical insertion (and subsequent removal) of a thread into the internal carotid artery. Damage was quantified at various times after MCAO to investigate the temporal progression of damage and establish an appropriate time to assess the effects of IL-1ra on cell death. Cell death was complete 18-24 h after temporary MCAO. Intracerebroventricular injection of IL-1ra (10 microg) at the time of MCAO and 60 min later reduced the lesion volume measured 24 h (57% reduction) or 48 h (52% reduction) after MCAO. Cell death was also significantly reduced when IL-1ra (20 microg) was administered as a single injection, 1 h (47%), 2 h (57%) or 3 h (46%) after MCAO, when compared to vehicle. These data show that IL-1ra markedly reduces cell death even when administration is delayed until 3 h after induction of reversible, focal cerebral ischaemia in the rat, and support our proposal that IL-1ra may be of therapeutic benefit in stroke.

Figures

Figure 1

Lesion development after temporary (60 min) middle cerebral artery occlusion in the rat. Animals were killed at various times after MCAO (2–12 h, n=4 per group, 12 and 18 h n=5 per group) and the areas of total cell death (as indicated by a complete lack of cresyl violet staining) in cortical and subcortical (striatal and thalamic) regions were measured as lesion. *P<0.05, **P<0.01 compared to appropriate group at 2 h, †P<0.05, ††P<01 compared to appropriate group at 4 h. Representative data from 24 h after MCAO (pooled values from experiments in study 3, n=24) are shown for comparison.

Figure 2

Effect of rhIL-1ra on total lesion volume assessed 24 or 48 h after transient (60 min) middle cerebral artery occlusion in the rat. Animals were injected i.c.v. with rhIL-1ra (10 μg) or vehicle (2 μl) immediately after MCAO and immediately after reperfusion (total dose injected=20 μg). Data are presented as mean ±s.d. of a group of animals (24 h: n=6 per group; 48 h: vehicle n=11, rhIL-1ra n=10). *P<0.05, **P<0.01 compared to appropriate vehicle-treated group (Student's t-test).

Figure 3

Representative cresyl violet-stained brain sections showing the lesion 24 h (a) and 48 h (b) after transient (60 min) MCAO and injection of vehicle or rhIL-1ra (10 μg, i.c.v. immediately after MCAO and immediately after reperfusion). Each experimental group is represented by three sections taken from three different anterior–posterior levels (similar levels for each experimental group, approximately +1.4 mm (i), +0.7 mm (ii) and −2.3 mm (iii) relative to bregma). The pattern of injury in vehicle-treated animals was similar 24 and 48 h after MCAO. Damage was smaller in rhIL-1ra-treated animals, most notably in the cortex, and the pattern of damage was similar 24 and 48 h after MCAO.

Figure 4

Effect of delayed administration of rhIL-1ra on (a) total, (b) cortical and (c) subcortical lesion volume assessed 24 h after transient (60 min) middle cerebral artery occlusion. Animals were subjected to MCAO and injected (i.c.v.) with vehicle or rhIL-1ra (20 μg) either 1 h (vehicle n=10, rhIL-1ra n=10), 2 h (vehicle n=7, rhIL-1ra n=8) or 3 h (vehicle n=7, rhIL-1ra n=9) after MCAO. *P<0.05, **P<0.01 compared to vehicle (Student's t-test) ‡P<0.05 compared to vehicle (Student's t-test with Welch correction).