Interleukin-1β accelerates the onset of stroke in stroke-prone spontaneously hypertensive rats

Abstract

High blood levels of inflammatory biomarkers and immune cells in stroke lesions have been recognized as results of stroke. However, recent studies have suggested that inflammation occurs prior to stroke onset. In this study, we aimed to clarify the role of inflammation in stroke onset among stroke-prone spontaneously hypertensive rats (SHRSP). At 4 weeks of age (before stroke onset), the plasma level of IL-1β was significantly higher in SHRSP (153.0 ± 49.7 pg/ml) than in Wistar Kyoto rats (WKY) (7.7 ± 3.4 pg/ml, P < 0.001 versus SHRSP) or spontaneously hypertensive rats (SHR) (28.0 ± 9.1 pg/ml, P < 0.001 versus SHRSP) (n = 6 per strain). Stimulated IL-1β signal was also observed in cerebrovascular endothelial cells of SHRSP. Gene expressions of IL-1β, IL-1 receptors, caspase-1, and downstream genes (MCP-1 and ICAM-1), which associated with immune cell recruitment, were significantly greater in SHRSP than in WKY or SHR, coincident with greater NFκB protein levels in SHRSP compared to WKY or SHR. In addition, continuous administration of IL-1β (2 μg/day) using an osmotic pump slightly increased the incidence of stroke in SHR (P = 0.046) and significantly accelerated the onset of stroke in SHRSP (P = 0.006) compared to each control (n = 10 per group). These results suggest that a stimulated IL-1β signal might be a cause of stroke onset when concomitant with severe hypertension.

Figure 1

IL-1β level in plasma and IL-1β signal-related gene expression in CVECs from each strain. Plasma and CVECs were harvested from WKY, SHR, and SHRSP (male, 4 weeks of age). Total RNA was extracted for quantitative real time PCR. (a) IL-1β level in plasma and (b) IL-1β signal-related gene expression, such as (A) IL-1β, (B) IL-1RI, (C) IL-1RII, (D) caspase-1, (E) MCP-1, and (F) ICAM-1 in CVECs were measured as described in the methods section. Bars show SD, ***P < 0.001 versus WKY, ^††† P < 0.001 versus SHR. n = 5 in each strain. N.D.: not detectable.

Figure 2

NFκB protein levels in CVECs from each strain. CVECs were harvested from WKY, SHR, and SHRSP (male, 4 weeks of age). Whole cell protein was extracted for western blot analysis. Data are presented as mean ± SD of densitometric ratios (WKY was set as 100) of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001 versus WKY, ^††† P < 0.001 versus SHR.

Figure 3

Effect of continuous administration of IL-1β on the incidence of stroke in SHR. SHR (male, 10 weeks of age) were given IL-1β (2 μg/day) or vehicle (phosphate-buffered saline) for 4 weeks using osmotic pumps. Photograph of typical hemorrhage lesion in SHR given IL-1β for 4 weeks.

Figure 4

Effect of continuous administration of IL-1β on the onset of stroke in SHRSP. SHRSP (male, 10 weeks of age) were given IL-1β (2 μg/day) or vehicle (phosphate-buffered saline) for 4 weeks using osmotic pumps. Day of stroke onset was determined as described in the methods. (a) Incidence of stroke. n = 10 in each group. (b) Correlation between plasma IL-1β level (measured at 1 week after administration; before any animals did not show stroke sign) and the day of stroke onset. Open circles, control (n = 9; one rat did not show any stroke sign until 4 weeks); closed circles, IL-1β (n = 10). (c) Lifespan during the 4-week administration period.