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Modeling Encephalopathy of Prematurity Using Prenatal Hypoxia-ischemia with Intra-amniotic Lipopolysaccharide in Rats
Published on: November 20, 2015
Activation of nuclear factor-kappaB signaling pathway by interleukin-1 after hypoxia/ischemia in neonatal rat
Xiaoming Hu1, Olivera Nesic-Taylor, Jingxin Qiu
1Department of Anatomy and Neuroscience, University of Texas Medical Branch, Galveston, Texas, USA.
Insights
Perinatal hypoxia-ischemia (HI) triggers brain damage via Interleukin-1 (IL-1) signaling. Blocking IL-1 with IL-1 receptor antagonist (IL-1Ra) reduces cell death by inhibiting nuclear factor-kappaB (NF-kappaB) activation.
Area of Science:
- Neuroscience
- Molecular Biology
- Developmental Biology
Background:
- Perinatal hypoxia-ischemia (HI) is a leading cause of pediatric neurological deficits.
- Interleukin-1 (IL-1) activity is implicated in HI-induced brain damage, but its precise mechanisms remain unclear.
- Understanding the molecular pathways of IL-1 in HI is crucial for developing targeted therapies.
Purpose of the Study:
- To investigate the role of nuclear factor-kappaB (NF-kappaB) activation in IL-1 signaling during perinatal HI.
- To elucidate the downstream inflammatory and apoptotic pathways influenced by IL-1 in the developing brain post-HI.
- To assess the therapeutic potential of blocking IL-1 signaling in a rat model of HI.
Main Methods:
- A 7-day-old rat model was used to induce HI via carotid artery ligation and hypoxia.
- ELISA, electrophoretic mobility shift assays (EMSA), and Western blotting were employed to measure cell death, caspase 3 activity, IL-1beta expression, NF-kappaB activation (p65/p50), and inflammatory markers (iNOS, COX-2).
- Intracerebroventricular injection of IL-1 receptor antagonist (IL-1Ra) was used to block IL-1 signaling.
Main Results:
- HI significantly increased cell death, caspase 3 activity, and IL-1beta expression in the hippocampus and cortex.
- NF-kappaB activation, evidenced by increased p65/p50 DNA-binding activity and p65 nuclear translocation, was observed 24 hours post-HI.
- IL-1Ra treatment significantly reduced HI-induced cell death, caspase 3 activity, and NF-kappaB activation, as well as the expression of iNOS and COX-2.
- IL-1Ra also attenuated the increase in phosphorylated inhibitor kappaBalpha (IkappaBalpha) and nuclear translocation of Bcl-3.
Conclusions:
- HI stimulates IL-1 signaling, leading to inflammatory and apoptotic neuronal damage in the developing brain.
- NF-kappaB activation plays a critical role in mediating IL-1-induced neuroinflammation and cell death following HI.
- Blocking IL-1 signaling with IL-1Ra represents a promising therapeutic strategy to mitigate brain injury after perinatal HI.
Abstract:
Perinatal hypoxia/ischemia (HI) is a common cause of neurological deficits in children. Interleukin-1 (IL-1) activity has been implicated in HI-induced brain damage. However, the mechanisms underlying its action in HI have not been characterized. We used a 7-day-old rat model to elucidate the role of nuclear factor-kappaB (NF-kappaB) activation in HI stimulation of IL-1 signaling. HI was induced by permanent ligation of the left carotid artery followed by 90 min of hypoxia (7.8% O(2)). Using ELISA assays, we observed increased cell death and caspase 3 activity in hippocampus and cortex 3, 6, 12, 24 and 48 h post-HI. IL-1beta protein expression increased, beginning at 3 h after HI and lasting until 24 h post-HI in hippocampus and 12 h post-HI in cortex. Intracerebroventricular injection of 2 microg IL-1 receptor antagonist (IL-1Ra) 2 h after HI significantly reduced cell death and caspase 3 activity. Electrophoretic mobility shift assay analyses of hippocampus and cortex after HI for NF-kappaB activity showed increased p65/p50 DNA-binding activity at 24 h post-HI. Western blot analyses showed significant nuclear translocation of p65. Protein expression levels of two known inflammatory agents, inducible nitric oxide synthase and cycloxygenase 2, known to be transcriptionally regulated by NF-kappaB, also increased at 24 h after HI. All these HI-induced changes were reversed by IL-1Ra blockade of IL-1 signaling, consistent with IL-1 triggering of inflammatory apoptotic outcomes via NF-kappaB transcriptional activation. The observed increase in cytoplasmic phosphorylated inhibitor kappaBalpha (IkappaBalpha) and nuclear translocation of Bcl-3 24 h after HI was also significantly attenuated by IL-1Ra blockade, suggesting that HI-induced IL-1 activation of NF-kappaB is via both the degradation of IkappaBalpha and the nuclear translocation of Bcl-3.
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