P1.039: Fetal Brain Injury in Experimental Congenital Diaphragmatic Hernia

Background: It is well known that at least 25% of children born with congenital diaphragmatic hernia (CDH) are diagnosed with neurodevelopmental impairment (NDI). The exact etiology of NDI in children with CDH remains unknown and has traditionally been attributed to postnatal factors, such as prolonged ventilation, use of extracorporeal life support, and surgery. However, there is growing evidence that suggests that the CDH brain may have already sustained injury in utero, as shown by antenatal imaging studies demonstrating alterations of cerebral arterial flow. Beyond the need to understand the pathogenesis of CDH-related brain injury, there is also an urgent need to identify an antenatal therapeutic strategy that could prevent NDI. There is increasing interest in the role of stem cell-derived extracellular vesicles (EVs) in prenatal treatment due to their therapeutic and regenerative potential, their ability to cross the blood brain barrier, their involvement in neurogenesis, and their ability to rescue pulmonary hypoplasia in CDH models. Our objectives were to investigate 1) whether the brain of fetal mice with CDH has changes in progenitor cell populations that are critical for normal neurogenesis; and 2) the rescue potential of amniotic fluid stem cells-extracellular vesicles (AFSC-EVs) as a therapy for CDH-related fetal brain injury.

Methods: Following ethical approval (IACUC#64247), on E8.5 dams were gavaged with either nitrofen/bisdiamine to induce CDH or olive oil to serve as control. On E16.5, fetuses exposed to nitrofen/bisdiamine were intra-amniotically injected with saline or AFSC-EVs. On E19, pups from all three groups (control, CDH+saline, CDH+AFSC-EVs) were delivered and their brain was harvested for analysis. Groups were blindly compared for the expression of OLIG2, a marker of oligodendrocyte lineage (OL-lineage) cells, and SOX2, a marker of neural progenitor cells (NPCs), in the cortex and subventricular zone, respectively.

Results: Compared to normal control, the brain of mice with CDH had a decreased expression of OL-lineage and NPC markers (Figure 1 and 2). Moreover, the brain of pups treated with AFSC-EVs had similar expression levels of OL-lineage and NPC markers to control.

Conclusion: This study shows for the first time an impairment in the density of progenitor cell populations in the brain of fetal mice with CDH. Further studies are underway to characterize the biological processes and molecular pathways responsible for impaired neurogenesis in CDH and to evaluate the potential of an EV-based therapy as a rescue for normal brain development.