Long-term magnetic resonance imaging of stem cells in neonatal ischemic injury.

OBJECTIVE: Quantitative magnetic resonance imaging (MRI) can serially and noninvasively assess the degree of injury in rat pup models of hypoxic ischemic injury (HII). It can also noninvasively monitor stem cell migration following iron oxide prelabeling. Reports have shown that neural stem cells (NSCs) may help mediate neuroprotection or stimulate neuroreparative responses in adult and neonatal models of ischemic injury. We investigated the ability of high-field MRI to monitor and noninvasively quantify the migration, proliferation, and location of iron oxide-labeled NSCs over very long time periods (58 weeks) in real time while contemporaneously correlating this activity with the evolving severity and extent of neural damage. METHODS: Labeled clonal murine NSCs (mNSCs) were implanted 3 days after unilateral HII in 10-day-old rat pups into the contralateral striatum or ventricle. We developed methods for objectively quantifying key aspects of dynamic NSC behavior (eg, viability; extent, and speed of migration; degree of proliferation; extent of integration into host parenchyma). MRI images were validated with histological and immunohistochemical assessments. RESULTS: mNSCs rapidly migrated (100 µm/day) to the lesion site. Chains of migrating NSCs were observed in the corpus callosum. In pups subjected to HII, though not in intact control animals, we observed a 273% increase in the MR-derived volume of mNSCs 4 weeks after implantation (correlating with the known proliferative behavior of endogenous and exogenous NSCs) that slowly declined over the 58-week time course, with no adverse consequences. Large numbers of now quiescent mNSCs remained at the site of injury, many retaining their iron oxide label. INTERPRETATION: Our studies demonstrate that MRI can simultaneously monitor evolving neonatal cerebral injury as well as NSC migration and location. Most importantly, it can noninvasively monitor proliferation dynamically for prolonged time periods. To be able to pursue clinical trials in newborns using stem cell therapies it is axiomatic that safety be insured through the long-term real time monitoring of cell fate and activity, particularly with regard to observing unanticipated risks to the developing brain. This study supports the feasibility of reliably using MRI for this purpose.

Albumin reduces blood-brain barrier permeability but does not alter infarct size in a rat model of neonatal stroke.

Human serum albumin therapy confers neurobehavioral and histopathologic neuroprotection in adult stroke models. We investigated whether albumin might also be neuroprotective in ischemic brain injury using a transient filament middle cerebral artery occlusion (tfMCAO) model in 10-d-old rat pups treated with 0.25% albumin or saline 1 h after reperfusion. We performed serial neurobehavioral and magnetic resonance imaging (MRI) assessments immediately after tfMCAO (day 0) and on 1, 3, 7, 14, and 28 d. IgG staining to assess blood-brain barrier (BBB) integrity and standard histology was obtained on 1, 3, and 28 d. Hemispheric infarct volumes from MRI were similar in saline and albumin groups (0 h: 39% and 44%; d 1: 46% and 55%; and d 28:10% and 24%) as were neurobehavioral assessments. IgG staining at 3 d post-ischemia showed loss of BBB integrity that was significantly reduced after albumin. Elevated T2 values suggesting vasogenic edema was seen in albumin compared with saline-treated animals, as was increased water mobility (i.e. increased apparent diffusion coefficient (ADC) reflecting cytotoxic edema. The reasons why albumin was not neuroprotective in neonatal stroke compared with adults remain uncertain. Effective strategies in adult models need to be reassessed in the neonate.

Temporal and regional evolution of aquaporin-4 expression and magnetic resonance imaging in a rat pup model of neonatal stroke.

Edema formation can be observed using magnetic resonance imaging (MRI) in patients with stroke. Recent studies have shown that aquaporin-4 (AQP4), a water channel, is induced early after stroke and potentially participates in the development of brain edema. We studied whether induction of AQP4 correlated with edema formation in a rat pup filament stroke model using high field (11.7-Tesla) MRI followed by immunohistochemical investigation of AQP4 protein expression. At 24 h, we observed increased T2 values and decreased apparent diffusion coefficients (ADC) within injured cortical and striatal regions that reflected the edema formation. Coincident with these MR changes were significant increases in AQP4 expression on astrocytic end-feet in the border regions of injured tissues. Striatal imaging findings were still present at 72 h with a slow normalization of AQP4 expression in the border regions. At 28 d, AQP4 expression normalized in the border while in this region ADC values increased. We show that induction of AQP4 is increased during the period of active edema formation in the border region without regional correlation with edema. Finally, induction of AQP4 on astrocyte end-feet could participate in tissue preservation after ischemia in the immature rat brain.