Background and Purpose Intravascular transplantation of neural stem cells represents a minimally invasive therapeutic approach for the treatment of central nervous system diseases. The cellular biodistribution after intravascular injection needs to be analyzed to determine the ideal delivery modality. We studied the biodistribution and efficiency of targeted central nervous system delivery comparing intravenous and intra-arterial (IA) administration of neural stem cells after brain ischemia. Methods Mouse neural stem cells were transduced with a firefly luciferase reporter gene for bioluminescence imaging (BLI). Hypoxic–ischemia was induced in adult mice and reporter neural stem cells were transplanted IA or intravenous at 24 hours after brain ischemia. In vivo BLI was used to track transplanted cells up to 2 weeks after transplantation and ex vivo BLI was used to determine single organ biodistribution. Results Immediately after transplantation, BLI signal from the brain was 12 times higher in IA versus intravenous injected animals (P<0.0001). After IA injection, 69% of the total luciferase activity arose from the brain early after transplantation and 93% at 1 week. After intravenous injection, 94% of the BLI signal was detected in the lungs (P = 0.004) followed by an overall 94% signal loss at 1 week, indicating lack of cell survival outside the brain. Ex vivo single organ analysis showed a significantly higher BLI signal in the brain than in the lungs, liver, and kidneys at 1 week (P<0.0001) and 2 weeks in IA (P = 0.007). Conclusion IA transplantation results in superior delivery and sustained presence of neural stem cells in the ischemic brain in comparison to intravenous infusion.

Background and Purpose-Intravascular delivery of neural stem cells (NSCs) after stroke has been limited by the low efficiency of transendothelial migration. Vascular cell adhesion molecule-1 is an endothelial adhesion molecule known to be upregulated early after stroke and is responsible for the firm adhesion of inflammatory cells expressing the surface integrin, CD49d. We hypothesize that enriching for NSCs that express CD49d and injecting them into the carotid artery would improve targeted cell delivery to the injured brain. Methods-Mouse NSCs were analyzed for the expression of CD49d by fluorescence activated cell sorting. A CD49d-enriched (CD49d ϩ ) (Ͼ95%) and -depleted (CD49d -; Ͻ5%) NSC population was obtained by cell sorting. C57/Bl6 mice underwent left-sided hypoxia-ischemia surgery and were assigned to receive 3ϫ10 5 CD49d ϩ , CD49d

Background and Purpose The inflammatory response is a critical component of ischemic stroke. In addition to its physiological role, the mechanisms behind transendothelial recruitment of immune cells also offer a unique therapeutic opportunity for translational stem cell therapies. Recent reports have demonstrated homing of neural stem cells (NSC) into the injured brain areas after intravascular delivery. However, the mechanisms underlying the process of transendothelial recruitment remain largely unknown. Here we describe the critical role of the chemokine CCL2 and its receptor CCR2 in targeted homing of NSC after ischemia. Methods Twenty-four hours after induction of stroke using the hypoxia-ischemia model in mice CCR2+/+ and CCR2−/− reporter NSC were intra-arterially delivered. Histology and bioluminescence imaging were used to investigate NSC homing to the ischemic brain. Functional outcome was assessed with the horizontal ladder test. Results Using NSC isolated from CCR2+/+ and CCR2−/− mice, we show that receptor deficiency significantly impaired transendothelial diapedesis specifically in response to CCL2. Accordingly, wild-type NSC injected into CCL2−/− mice exhibited significantly decreased homing. Bioluminescence imaging showed robust recruitment of CCR2+/+ cells within 6 hours after transplantation in contrast to CCR2−/− cells. Mice receiving CCR2+/+ grafts after ischemic injury showed a significantly improved recovery of neurological deficits as compared to animals with transplantation of CCR2−/− NSC. Conclusions The CCL2/CCR2 interaction is critical for transendothelial recruitment of intravascularly delivered NSC in response to ischemic injury. This finding could have significant implications in advancing minimally invasive intravascular therapeutics for regenerative medicine or cell-based drug delivery systems for central nervous system diseases.

Intra-arterial (IA) injection represents an experimental avenue for minimally invasive delivery of stem cells to the injured brain. It has however been reported that IA injection of stem cells carries the risk of reduction in cerebral blood flow (CBF) and microstrokes. Here we evaluate the safety of IA neural progenitor cell (NPC) delivery to the brain. Cerebral blood flow of rats was monitored during IA injection of single cell suspensions of NPCs after stroke. Animals received 1¾106 NPCs either injected via a microneedle (microneedle group) into the patent common carotid artery (CCA) or via a catheter into the proximally ligated CCA (catheter group). Controls included saline-only injections and cell injections into non-stroked sham animals. Cerebral blood flow in the microneedle group remained at baseline, whereas in the catheter group a persistent (15 minutes) decrease to 78% of baseline occurred (P < 0.001). In non-stroked controls, NPCs injected via the catheter method resulted in higher levels of Iba-1-positive inflammatory cells (P = 0.003), higher numbers of degenerating neurons as seen in Fluoro-Jade C staining (P < 0.0001) and ischemic changes on diffusion weighted imaging. With an appropriate technique, reduction in CBF and microstrokes do not occur with IA transplantation of NPCs.

✓ The use of stem cell transplantation to restore neurological function after stroke is being recognized as a potential novel therapy. Before stem cell transplantation can become widely applicable, however, questions remain about the optimal site of delivery and timing of transplantation. In particular, there seems to be increasing evidence that intravascular cell delivery after stroke is a viable alternative to intracerebral transplantation. In this review, the authors focus on the intravascular delivery of stem cells for stroke treatment with an emphasis on timing, transendothelial migration and possible mechanisms leading to neuroprotection, angiogenesis, immunomodulation, and neural plasticity. They also review current concepts of in vivo imaging and tracking of stem cells after stroke.

Background and Purpose— Vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFRs) play important roles during neurovascular repair after stroke. In this study, we imaged VEGFR expression with positron emission tomography (PET) to noninvasively analyze poststroke angiogenesis. Methods— Female Sprague-Dawley rats after distal middle cerebral artery occlusion surgery were subjected to weekly MRI, 18 F-FDG PET, and 64 Cu-DOTA-VEGF 121 PET scans. Several control experiments were performed to confirm the VEGFR specificity of 64 Cu-DOTA-VEGF 121 uptake in the stroke border zone. VEGFR, BrdU, lectin staining, and 125 I-VEGF 165 autoradiography on stroke brain tissue slices were performed to validate the in vivo findings. Results— T2-weighed MRI correlated with the “cold spot” on 18 F-FDG PET for rats undergoing distal middle cerebral artery occlusion surgery. The 64 Cu-DOTA-VEGF 121 uptake in the stroke border zone peaked at ≈10 days after surgery, indicating neovascularization as confirmed by histology (VEGFR-2, BrdU, and lectin staining). VEGFR specificity of 64 Cu-DOTA-VEGF 121 uptake was confirmed by significantly lower uptake of 64 Cu-DOTA-VEGF mutant in vivo and intense 125 I-VEGF 165 uptake ex vivo in the stroke border zone. No appreciable uptake of 64 Cu-DOTA-VEGF 121 was observed in the brain of sham-operated rats. Conclusions— For the first time to our knowledge, we successfully evaluated the VEGFR expression kinetics noninvasively in a rat stroke model. In vivo imaging of VEGFR expression could become a significant clinical tool to plan and monitor therapies aimed at improving poststroke angiogenesis.

Despite state-of-the-art therapy, clinical outcome after stroke remains poor, with many patients left permanently disabled and dependent on care. Stem cell therapy has evolved as a promising new therapeutic avenue for the treatment of stroke in experimental studies, and recent clinical trials have proven its feasibility and safety in patients. Replacement of damaged cells and restoration of function can be accomplished by transplantation of different cell types, such as embryonic, fetal or adult stem cells, human fetal tissue and genetically engineered cell lines. Adult neural stem cells offer the advantage of avoiding the ethical problems associated with embryonic or fetal stem cells and can be harvested as autologous grafts from the individual patients. Furthermore, stimulation of endogenous adult stem cell-mediated repair mechanisms in the brain might offer new avenues for stroke therapy without the necessity of transplantation. However, important scientific issues need to be addressed to advance our understanding of the molecular mechanisms underlying the critical steps in cell-based repair to allow the introduction of these experimental techniques into clinical practice. This review describes up-to-date experimental concepts using adult neural stem cells for the treatment of stroke.

Lewy bodies and Lewy neurites constitute the cardinal neuropathological features of both Parkinson's disease (PD) and Lewy body dementia (LBD). Whereas α-synuclein has been found to be the major component of the Lewy body, the mechanisms by which neurons degenerate, as well as basic mechanisms involved in the formation of α-synuclein-related inclusions, remain obscure. We have suggested previously that potential mechanisms are likely to leave a "molecular signature" or protein adduct within the Lewy body, and have found examples of such signatures in previous studies. In this study, we demonstrate increased FOXO3 in association with Lewy bodies and Lewy neurites in LBD and PD brain tissue. Since FOXO proteins are involved in several pathways responsible for the regulation of cell death, cell proliferation, and cell metabolism, the ectopic localization of FOXO3 to Lewy bodies provides evidence that aberrations in basic cellular biochemistry may contribute to inclusion formation, which is likely more complex than a simple "gain of function" toxicity as is commonly opined. In light of the known interaction of FOXO3 and 14-3-3, basic protein-protein interaction between these proteins and α-synuclein may be key.