Hypoxic-ischemic brain injury is the single most important neurologic problem in the perinatal period. In utero hemodynamic abnormalities possibly in association with elevated pro-inflammatory cytokines (cytokines) such as IL-12, IL-6, and TNF-1 predispose to brain injury, particularly in premature neonates. Systemic cytokines produced during maternal infection and/or increases in cytokines after ischemia may accentuate damage to the fetal brain. The neurovascular unit is a privileged site that consists of brain microvascular endothelium, glia, and neurons. Although cytokines are known to cross the adult blood-brain barrier (BBB), evidence to support the hypothesis that systemic cytokines cross the BBB of the fetus and premature neonate is sparse.
Our specific aims test the hypothesis that specific cytokines cross both the intact and injured BBB in the fetus to damage the brain. A consequence of this hypothesis is that blockade of these cytokines would attenuate the ischemia related damage to the neurovascular unit (BBB) and possibly the brain. A multidisciplinary approach will be used to address our hypothesis and will include physiological, biochemical, pathological, immunohistochemical, and molecular methods.
Aim 1 tests the hypothesis that cytokines such as IL-12 &IL-6 cross the BBB in a maturation-dependent manner in ovine fetuses, and that maturation-related changes in barrier permeability to cytokines are primarily related to changes in the composition of the tight junction. BBB permeability will be quantified by the integral technique with 1-aminoisobutyric acid and radiolabeled cytokines. Tight junction (TJ) proteins and mRNA will be measured by Western blot, immunohistochemistry, and Northern blot.
Aim 2 determines whether ischemic injury increases the permeability of the BBB to cytokines as a function of gestational age and tight junction maturation. Brain ischemia is induced by carotid occlusion. BBB permeability and TJ components will be measured as in Aim 1 and brain injury assessed by pathological, immunohistochemical, and molecular methods.
Aim 3 determines whether blocking the effects of cytokines with systemic infusions of neutralizing antibodies attenuates ischemic injury to the fetal neurovascular unit (BBB) and possibly the brain more in preterm than near term fetuses. IL-6 and IL-1 2 blocking antibodies will be infused before ischemia. Brain ischemia will be induced and BBB permeability, TJ components, and brain injury measured as in Aim 2. These studies will provide the first direct evidence whether systemic cytokines cross the intact or injured fetal BBB and whether blocking the effects of cytokines with neutralizing antibodies protect the fetal neurovascular unit (BBB) and brain. This project may provide new insights into novel strategies to prevent brain injury in the human fetus and/or premature infant. Perinatal hypoxic/ischemic brain injury often results in cerebral palsy (CP) and mental retardation. The incidence of CP is 40-148/1,000 in premature and 1-2/1,000 in full term infants. This project could provide new insights into novel strategies to prevent brain injury in the human fetus and/or premature infant. Perinatal hypoxic/ischemic brain injury often results in cerebral palsy (CP) and mental retardation. The incidence of CP is 40-148/1,000 in premature and 1-2/1,000 in full term infants. This project could provide new insights into novel strategies to prevent brain injury in the human fetus and/or premature infant.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD057100-05
Application #
8242876
Study Section
Developmental Brain Disorders Study Section (DBD)
Program Officer
Higgins, Rosemary
Project Start
2008-04-20
Project End
2013-09-30
Budget Start
2012-04-01
Budget End
2013-09-30
Support Year
5
Fiscal Year
2012
Total Cost
$282,680
Indirect Cost
$87,234
Name
Women and Infants Hospital-Rhode Island
Department
Type
DUNS #
069851913
City
Providence
State
RI
Country
United States
Zip Code
02905
Sadowska, G B; Stonestreet, B S (2014) Maternal treatment with glucocorticoids modulates gap junction protein expression in the ovine fetal brain. Neuroscience 275:248-58
Spasova, Mariya S; Sadowska, Grazyna B; Threlkeld, Steven W et al. (2014) Ontogeny of inter-alpha inhibitor proteins in ovine brain and somatic tissues. Exp Biol Med (Maywood) 239:724-36
Threlkeld, Steven W; Gaudet, Cynthia M; La Rue, Molly E et al. (2014) Effects of inter-alpha inhibitor proteins on neonatal brain injury: Age, task and treatment dependent neurobehavioral outcomes. Exp Neurol 261:424-33
Spasova, Mariya S; Sadowska, Grazyna B; Threlkeld, Steven W et al. (2014) Ontogeny of inter-alpha inhibitor proteins in ovine brain and somatic tissues. Exp Biol Med (Maywood) 239:724-36
Virgintino, Daniela; Girolamo, Francesco; Rizzi, Marco et al. (2014) Ischemia/Reperfusion-induced neovascularization in the cerebral cortex of the ovine fetus. J Neuropathol Exp Neurol 73:495-506
Chen, X; Threlkeld, S W; Cummings, E E et al. (2013) In-vitro validation of cytokine neutralizing antibodies by testing with ovine mononuclear splenocytes. J Comp Pathol 148:252-8
Cohen, Susan S; Min, May; Cummings, Erin E et al. (2013) Effects of interleukin-6 on the expression of tight junction proteins in isolated cerebral microvessels from yearling and adult sheep. Neuroimmunomodulation 20:264-73
Mirza, Hussnain; Oh, William; Laptook, Abbot et al. (2013) Indomethacin prophylaxis to prevent intraventricular hemorrhage: association between incidence and timing of drug administration. J Pediatr 163:706-10.e1
Stonestreet, Barbara S; Sadowska, Grazyna B; Hanumara, R Choudary et al. (2012) Comparative effects of glucose- and mannitol-induced osmolar stress on blood-brain barrier function in ovine fetuses and lambs. J Cereb Blood Flow Metab 32:115-26
Kim, Chang-Ryul; Sadowska, Grazyna B; Newton, Stephanie A et al. (2011) Na+,K+-ATPase activity and subunit protein expression: ontogeny and effects of exogenous and endogenous steroids on the cerebral cortex and renal cortex of sheep. Reprod Sci 18:359-73

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