Neutralizing anti-interleukin-1β antibodies modulate fetal blood–brain barrier function after ischemia

Abstract

We have previously shown that increases in blood-brain barrier permeability represent an important component of ischemia-reperfusion related brain injury in the fetus. Pro-inflammatory cytokines could contribute to these abnormalities in blood-brain barrier function. We have generated pharmacological quantities of mouse anti-ovine interleukin-1β monoclonal antibody and shown that this antibody has very high sensitivity and specificity for interleukin-1β protein. This antibody also neutralizes the effects of interleukin-1β protein in vitro. In the current study, we hypothesized that the neutralizing anti-interleukin-1β monoclonal antibody attenuates ischemia-reperfusion related fetal blood-brain barrier dysfunction. Instrumented ovine fetuses at 127 days of gestation were studied after 30 minutes of carotid occlusion and 24 hours of reperfusion. Groups were sham operated placebo-control- (n=5), ischemia-placebo- (n=6), ischemia-anti-IL-1β antibody- (n=7), and sham-control antibody- (n=2) treated animals. Systemic infusions of placebo (0.154 M NaCl) or anti-interleukin-1β monoclonal antibody (5.1±0.6 mg/kg) were given intravenously to the same sham or ischemic group of fetuses at 15 minutes and 4 hours after ischemia. Concentrations of interleukin-1β protein and anti-interleukin-1β monoclonal antibody were measured by ELISA in fetal plasma, cerebrospinal fluid, and parietal cerebral cortex. Blood-brain barrier permeability was quantified using the blood-to-brain transfer constant (Ki) with α-aminoisobutyric acid in multiple brain regions. Interleukin-1β protein was also measured in parietal cerebral cortices and tight junction proteins in multiple brain regions by Western immunoblot. Cerebral cortical interleukin-1β protein increased (P<0.001) after ischemia-reperfusion. After anti-interleukin-1β monoclonal antibody infusions, plasma anti-interleukin-1β monoclonal antibody was elevated (P<0.001), brain anti-interleukin-1β monoclonal antibody levels were higher (P<0.03), and interleukin-1β protein concentrations (P<0.03) and protein expressions (P<0.001) were lower in the monoclonal antibody-treated group than in placebo-treated-ischemia-reperfusion group. Monoclonal antibody infusions attenuated ischemia-reperfusion-related increases in Ki across the brain regions (P<0.04), and Ki showed an inverse linear correlation (r = −0.65, P<0.02) with anti-interleukin-1β monoclonal antibody concentrations in the parietal cortex, but had little effect on tight junction protein expression. We conclude that systemic anti-interleukin-1β monoclonal antibody infusions after ischemia result in brain anti-interleukin-1β antibody uptake, and attenuate ischemia-reperfusion-related interleukin-1β protein up-regulation and increases in blood-brain barrier permeability across brain regions in the fetus. The pro-inflammatory cytokine, interleukin-1β, contributes to impaired blood-brain barrier function after ischemia in the fetus.

Interleukin-1β Transfer across the Blood–Brain Barrier in the Ovine Fetus

“…Consistent with our previous findings, brain ischemia and reperfusion did not have a major impact on the systemic hemodynamic and biochemical homeostasis in the fetus. 12,19 The findings of our study show that systemically administered IL-1β is able to cross the BBB in significant amounts to reach the brain during normal homeostatic conditions in the fetus. This finding suggests that conditions such as perinatal inflammation that result in increases in systemic pro-inflammatory cytokines could facilitate cytokines to cross the intact BBB and cause brain injury before birth even under nonischemic conditions.…”

“…[20][21][22] In addition, we have recently shown that BBB dysfunction represents an important component of ischemicreperfusion related brain injury in the fetus, 12 and that the proinflammatory cytokine IL-1β contributes to this barrier dysfunction. 19 Nonetheless, although it has been suggested that systemic cytokines might gain access to the fetal brain by crossing the BBB, 4 there is a paucity of direct experimental evidence to support this concept and no quantitative data measuring BBB permeability with cytokines in the fetal or neonatal brain. Therefore, the objectives of the current study were to determine the ability of systemic circulating IL-1β to cross the intact BBB under homeostatic conditions in the normal fetus, and to determine whether ischemia accentuates the transfer of IL-1β across the BBB in the fetus.…”

“…14,37,40 Moreover, the biologic importance of cytokine transport across the BBB has been established because memory impairment induced by IL-1α depends directly on the ability of this cytokine to cross the BBB. 40 In addition, there are several lines of evidence to support the contention intact cytokines cross the BBB: (1) previous rigorous analysis by Banks et al in several manuscripts established that a number of cytokines cross the adult BBB undegraded; 13,14,26,27,37,[40][41][42] (2) free iodine does not cross the BBB well because of a brain-to-blood efflux system for iodine; 43 (3) we have previously shown that brain levels of cytokines are decreased when antibodies against that specific cytokine were infused into the blood; 19,44 and (4) transport of the labeled ovine cytokines across the BBB is inhibited by unlabeled murine cytokine. 16 This latter finding is particularly important, as unlabeled cytokines would not be expected to inhibit brain uptake of free iodine.…”

“…16 Moreover, we have also reported that the proinflammatory cytokine IL-1β contributes to BBB dysfunction after ischemic injury in the fetus. 19 Pro-inflammatory cytokines produced in the systemic circulation have been postulated to cross the BBB in the fetus and neonate. 4 It has long been suggested that pro-inflammatory cytokines could gain access to the fetal brain after being produced during maternal intrauterine infection or inflammation.…”

Assessing Blood Brain Barrier Permeability in Traumatic Brain Injury Research

“…Briefly, the isotope alpha-[ 14 C]-aminioisobutyric acid is a tracer introduced into the circulation along with a red blood cell labeling agent, technetium-99 m . The amount of tracer that has moved out of the vasculature across the BBB can be described by a transfer constant, Ki by differentiating the parenchymal tracer concentration from the intravascular tracer, using co-localization with red blood cells labeled with technetium-99 m [32,33].…”