Long-term neurological deficits due to immature cortical development are emerging as a major challenge in congenital heart disease (CHD). However, cellular mechanisms underlying dysregulation of perinatal corticogenesis in CHD remain elusive. The subventricular zone (SVZ) represents the largest postnatal niche of neural stem/progenitor cells (NSPCs). We show that the piglet SVZ resembles its human counterpart and displays robust postnatal neurogenesis. We present evidence that SVZ-NSPCs migrate to the frontal cortex and differentiate into interneurons in a region-specific manner. Hypoxic exposure of the gyrencephalic piglet brain recapitulates CHD-induced impaired cortical development. Hypoxia reduces proliferation and neurogenesis in the SVZ, which is accompanied by reduced cortical growth. We demonstrate a similar reduction in neuroblasts within the SVZ of human infants born with CHD. Our findings demonstrate that SVZ-NSPCs contribute to perinatal corticogenesis and suggest that restoration of SVZ-NSPCs’ neurogenic potential is a candidate therapeutic target for improving cortical growth in CHD.

Background Neurodevelopmental delays in motor skills and white matter (WM) injury have been documented in congenital heart disease (CHD) and after pediatric cardiac surgery. The lack of a suitable animal model has hampered our understanding of the cellular mechanisms underlying WM injury in these patients. Our aim is to identify an optimal surgical strategy for WM protection to reduce neurological injury in CHD patients. Methods and Results We developed a porcine cardiopulmonary bypass (CPB) model, which displays area dependent WM maturation. In this model, WM injury was identified following CPB-induced ischemia-reperfusion injury. The degree of injury was inversely correlated with the maturation stage, indicating maturation-dependent vulnerability of WM. Within different oligodendrocyte (OL) developmental stages, we show selective vulnerability of O4+ pre-OLs, while OL progenitor cells (OPCs) were resistant to insults. This indicates that immature WM is vulnerable to CPB-induced injury, but has an intrinsic potential for recovery mediated by endogenous OPCs. OPC number decreased with age, suggesting that earlier repair allows successful WM development. OPC proliferation was observed within a few days after CPB-induced ischemia-reperfusion injury; however by four weeks arrested OL maturation and delayed myelination were detected. Logistic model confirmed that maintaining higher oxygenation and reducing inflammation were effective in minimizing the risk of injury at immature stages of WM development. Conclusions Primary repair in neonates and young infants potentially provides successful WM development in CHD patients. Cardiac surgery during this susceptible period should avoid ischemia-reperfusion injury and minimize inflammation to prevent long-term WM-related neurological impairment.

BackgroundWhite matter (WM) injury is common after cardiopulmonary bypass or deep hypothermic circulatory arrest (DHCA) in neonates who have cerebral immaturity secondary to in utero hypoxia. The mechanism remains unknown. We investigated effects of preoperative-hypoxia on DHCA-induced WM-injury using a combined experimental paradigm in rodents.MethodsMice were exposed to hypoxia (Pre-Hypoxia). Oxygen-glucose deprivation (OGD) was performed under three temperatures to simulate brain conditions of DHCA including ischemia-reperfusion/reoxygenation under hypothermia.ResultsWM-injury in Pre-Normoxia was identified after 35°C-OGD. In Pre-Hypoxia, injury was displayed in all groups. Among oligodendrocyte stages, the pre-oligodendrocyte was the most susceptible while the oligodendrocyte progenitor was resistant to insult. When effects of Pre-Hypoxia were assessed, injury of mature oligodendrocytes and oligodendrocyte progenitors in Pre-Hypoxia significantly increased compared with Pre-Normoxia, indicating that mature oligodendrocytes and progenitors which had developed under hypoxia had greater vulnerability. Conversely, damage of oligodendrocyte progenitors in Pre-Hypoxia were not identified after 15°C-OGD, suggesting that susceptible oligodendrocytes exposed to hypoxia are protected by deep hypothermia.DiscussionDevelopmental alterations due to hypoxia result in an increased WM susceptibility to injury. Promoting WM regeneration by oligodendrocyte progenitors after earlier surgery using deep hypothermia is the most promising approach for successful WM development in CHD patients.

Background White matter (WM) injury is common after neonatal cardiopulmonary bypass (CPB). We have demonstrated that the inflammatory response to CPB is an important mechanism of WM injury. Microglia are brain-specific immune cells that respond to inflammation and can exacerbate injury. We hypothesized that microglia activation contributes to WM injury caused by CPB. Methods Juvenile piglets were randomly assigned to one of three CPB-induced brain insults (1: no-CPB, 2: full-flow CPB, 3: CPB/Circulatory-arrest). Neurobehavioral tests were performed. Animals were sacrificed 3-days or 4-weeks post-operatively. Microglia and proliferating cells were immunohistologically identified. Seven analyzed WM regions were further categorized into 3-fiber connections (1: Commissural, 2: Projection, 3: Association fibers). Results Microglia numbers significantly increased on day 3 after CPB/Circulatory-arrest, but not after full-flow CPB. Fiber categories did not affect these changes. On post-CPB week 4, proliferating cell number, blood leukocyte number and IL-6 levels, and neurological scores had normalized. However, both full-flow CPB and CPB/Circulatory-arrest displayed significant increases in the microglia number compared with Control. Thus brain-specific inflammation after CPB persists despite no changes in systemic biomarkers. Microglia number was significantly different among fiber categories, being highest in Association and lowest in Commissural connections. Thus there was a WM fiber-dependent microglia reaction to CPB. Conclusions This study demonstrates prolonged microglia activation in WM after CPB. This brain-specific inflammatory response is systemically silent. It is connection fiber-dependent which may impact specific connectivity deficits observed after CPB. Controlling microglia activation after CPB is a potential therapeutic intervention to limit neurological deficits following CPB.

BackgroundNewly developed white matter (WM) injury is common after cardiopulmonary bypass (CPB) in severe/complex congenital heart disease. Fractional anisotropy (FA) allows measurement of macroscopic organization of WM pathology but has rarely been applied after CPB. The aims of our animal study were to define CPB‐induced FA alterations and to determine correlations between these changes and cellular events after congenital heart disease surgery.Methods and ResultsNormal porcine WM development was first assessed between 3 and 7 weeks of age: 3‐week‐old piglets were randomly assigned to 1 of 3 CPB‐induced insults. FA was analyzed in 31 WM structures. WM oligodendrocytes, astrocytes, and microglia were assessed immunohistologically. Normal porcine WM development resembles human WM development in early infancy. We found region‐specific WM vulnerability to insults associated with CPB. FA changes after CPB were also insult dependent. Within various WM areas, WM within the frontal cortex was susceptible, suggesting that FA in the frontal cortex should be a biomarker for WM injury after CPB. FA increases occur parallel to cellular processes of WM maturation during normal development; however, they are altered following surgery. CPB‐induced oligodendrocyte dysmaturation, astrogliosis, and microglial expansion affect these changes. FA enabled capturing CPB‐induced cellular events 4 weeks postoperatively. Regions most resilient to CPB‐induced FA reduction were those that maintained mature oligodendrocytes.ConclusionsReducing alterations of oligodendrocyte development in the frontal cortex can be both a metric and a goal to improve neurodevelopmental impairment in the congenital heart disease population. Studies using this model can provide important data needed to better interpret human imaging studies.