Introduction
Neonatal hypoxic-ischemic white matter (WM) damage is a major contributor to chronic neurological dysfunction. Immature oligodendrocytes (OLs) are known to be highly vulnerable to ischemia. As little is known about the loss and restoration of myelin basic protein (MBP) as well as repair mechanisms in neonates, we studied cell outcome in the white matter (cingulum and external capsule) after MCA electrocoagulation and transient homolateral CCA occlusion.
Methods
Seven day-old rat pups were anesthetized with intraperitoneal injection of chloral hydrate (300 mg/kg) and underwent left middle cerebral artery electrocoagulation associated with 50 minutes left common carotid artery occlusion 1 . They were killed at various times (between 6 h and 15 days) after reperfusion. Loss and restoration of MBP were studied by MBP protein and O4 antigen labeling. TUNEL and Fluoro-Jade B are used to label cell death. Proliferation was demonstrated by the Ki-67 marker, a nuclear protein expressed in dividing cells for the entire duration of their mitotic process and expressed in all mammalian species. Microglial activation was followed with Griffonea Simplicifolia I isolectin B4 marker.
Results
In P7-injured rat pups, a marked reduction in MBP immunostaining, corresponded to numerous pyknotic immature OLs as early as 12 hours and a few dying mature OLs. Several GFAP-TUNEL-positive astrocytes were also observed ipsilaterally in the external capsule. In contrast, a substantial restoration of MBP occurred in the cingulum at 48 and 72 hours of recovery. Cell proliferation, demonstrated by the Ki-67 immunostaining, revealed a first peak of new generated cells in the dorsolateral SVZ at 72 hours of reperfusion. However, no double-stained O4 and Ki-67 OLs were detected, suggesting a functional recovery of injured mature OLs. New bipolar OL progenitors are generated between 7 to 15 days of recovery in the dorsolateral SVZ, compared to the contralateral side, indicating cell migration. Microglial-macrophage infiltration occurs over days following ischemia in the cingulum and also in the subcortical white matter where they were shown to engulf immature OLs, leading to impairment of repair. These deleterious events contribute to WM cystic formation.
Conclusion
The overall results suggest that the persistent activation of microglia/macrophages implicates a chronic component of immunoinflammation, which overwhelms repair processes and contributes to cystic growth. The use of drugs that modulate microglial activation can be proposed as candidate therapeutic agents.