JingMei Ren

Functional magnetic resonance imaging of reorganization in rat brain after stroke

Functional recovery after stroke has been associated with brain plasticity; however, the exact relationship is unknown. We performed behavioral tests, functional MRI, and histology in a rat stroke model to assess the correlation between temporal changes in sensorimotor function, brain activation patterns, cerebral ischemic damage, and cerebrovascular reactivity. Unilateral stroke induced a large ipsilateral infarct and acute dysfunction of the contralateral forelimb, which significantly recovered at later stages. Forelimb impairment was accompanied by loss of stimulus-induced activation in the ipsilesional sensorimotor cortex; however, local tissue and perfusion were only moderately affected and cerebrovascular reactivity was preserved in this area. At 3 days after stroke, extensive activation-induced responses were detected in the contralesional hemisphere. After 14 days, we found reduced involvement of the contralesional hemisphere, and significant responses in the infarction periphery. Our data suggest that limb dysfunction is related to loss of brain activation in the ipsilesional sensorimotor cortex and that restoration of function is associated with biphasic recruitment of peri- and contralesional functional fields in the brain.

Intracisternal osteogenic protein-1 enhances functional recovery following focal stroke

OSTEOGENIC protein-1 (OP-1, BMP-7) is a member of the transforming growth factor-β(TGF-β) superfamily that selectively induces dendritic outgrowth from cultured neurons. We injected human recombinant OP-1 (1 or 10 μg) or vehicle into the cisterna magna of mature male Sprague—Dawley rats 1 and 4 days after focal cerebral infarction induced by middle cerebral artery (MCA) occlusion. OP-1 treatment was associated with a marked enhancement of recovery of sensorimotor function of the impaired forelimb and hindlimb (contralateral to infarcts) as assessed by limb placing tests. This effect appeared to be dose dependent. There was no difference in infarct volume between OP-1 and vehicle-treated rats. The mechanisms of enhanced recovery by intracisternal OP-1 may include promotion of dendritic sprouting in the intact uninjured brain.

Time window of intracisternal osteogenic protein-1 in enhancing functional recovery after stroke.

Osteogenic protein-1 (OP-1, BMP-7) is a member of the bone morphogenetic protein subfamily of the TGF-ss superfamily that selectively stimulates dendritic neuronal outgrowth. In previous studies, we found that the intracisternal injection of OP-1, starting at one day after stroke, enhanced sensorimotor recovery of the contralateral limbs following unilateral cerebral infarction in rats. In the current study, we further explored the time window during which intracisternal OP-1 enhances sensorimotor recovery, as assessed by limb placing tests. We found that intracisternal OP-1 (10 microg) given 1 and 3 days, or 3 and 5 days, but not 7 and 9 days after stroke, significantly enhanced recovery of forelimb and hindlimb placing. There was no difference in infarct volume between vehicle- and OP-1-treated animals. The mechanism of OP-1 action might be stimulation of new dendritic sprouting in the remaining uninjured brain.

Postischemic Infusion of Cu/Zn Superoxide Dismutase or SOD:Tet451 Reduces Cerebral Infarction Following Focal Ischemia/Reperfusion in Rats ☆

Oxygen-free radicals play a major role in neuronal cell injury following cerebral ischemia/reperfusion. The free-radical scavenging enzyme, Cu/Zn superoxide dismutase (SOD-1), ameliorates various types of brain injury resulting from temporary CNS ischemia. We have compared the cerebroprotective properties of human SOD-1 (hSOD-1) with a novel recombinant SOD-1 hybrid protein, SOD:Tet451, composed of hSOD-1 linked to the neuronal binding fragment of tetanus toxin (TTxC). Following 2 h of temporary middle cerebral artery occlusion, rats infused with equivalent activities of either hSOD-1 or SOD:Tet451 for the initial 3 h of reperfusion showed reductions in cerebral infarct volume of 43 and 57%, respectively, compared to saline-treated controls (P < 0.01). Serum hSOD-1 concentrations in rats receiving SOD:Tet451 were seven-fold higher than those in rats receiving the native enzyme. Animals treated with SOD:Tet451 also demonstrated an extended persistence of hSOD-1 in the bloodstream during drug washout as compared to animals given free enzyme. Immunohistochemical examination of brain sections from an SOD:Tet451-treated ischemic rat showed positive immunoreactivity in the ipsilateral cerebral cortex using either anti-TTxC or anti-human SOD-1 antibodies. Our results document that both hSOD-1 and SOD:Tet451 significantly reduce brain infarct volume in a model of transient focal ischemia/reperfusion in rats. Additionally, our findings suggest that the cerebroprotective effects of SOD-1 may be enhanced by neuronal targeting as seen with the hybrid protein SOD:Tet451.

Time window of infarct reduction by intravenous basic fibroblast growth factor in focal cerebral ischemia

Basic fibroblast growth factor (bFGF) is a heparin-binding polypeptide with potent trophic and protective effects on brain neurons, glia and endothelia. In previous studies, we showed that intravenously administered bFGF reduced the volume of cerebral infarcts following permanent occlusion of the middle cerebral artery in rats. In the current study, we examined the time dependence of bFGF infusion on infarct reduction, and the effect of co-infusion of bFGF with heparin. We found a significant reduction in infarct volume when the bFGF infusion (50 microg/kg per h for 3 h) was begun up to 3 h, but not 4 h after the onset of ischemia. The infarct reducing effects of bFGF were not altered by co-infusion of heparin. These results are potentially important in light of the ongoing clinical trials of intravenous bFGF in acute stroke.

Human umbilical cord blood cells differentiate into muscle in sjl muscular dystrophy mice.

Limb girdle muscular dystrophy type 2B form (LGMD‐2B) and Miyoshi myopathy (MM) are both caused by mutations in the dysferlin (dysf) gene. In this study, we used dysferlin‐deficient sjl mice as a mouse model to study cell therapy for LGMD‐2B and MM. A single‐blind study evaluated the therapeutic potential of human umbilical cord blood (HUCB) as a source of myogenic progenitor stem cells. Three groups of donor cells were used: unfractionated mononuclear HUCB cells, HUCB subfractionated to enrich for cells that were negative for lineage surface markers (LIN−) and substantially enriched for the CD34 surface marker (CD34+), and irradiated control spleen cells. We administrated 1 × 106 donor cells to each animal intravenously and euthanized them at different time points (1–12 weeks) after transplantation. All animals were immunosuppressed (FK506 and leflunomide) from the day before the injection until the time of euthanasia. Immunohistochemical analyses documented that a small number of human cells from the whole HUCB and LIN−CD34+/−‐enriched HUCB subgroups engraft in the recipient muscle to express both dysferlin and human‐specific dystrophin at 12 weeks after transplantation. We conclude that myogenic progenitor cells are present in the HUCB, that they can disseminate into muscle after intravenous administration, and that they are capable of myogenic differentiation in host muscle.

Intracisternal Antisense Oligonucleotide to Growth Associated Protein-43 Blocks the Recovery-Promoting Effects of Basic Fibroblast Growth Factor after Focal Stroke

Focal infarction (stroke) of the lateral cerebral cortex of rats (including the sensorimotor cortex) produces deficits in sensorimotor function of the contralateral limbs that recover partially over time. In previous studies, we found that the intracisternal injection of basic fibroblast growth factor (bFGF), a potent neurotrophic growth factor, starting at 1 day after stroke, significantly enhanced recovery of sensorimotor function of the contralateral forelimb and hindlimb. Moreover, immunoreactivity (IR) for growth-associated protein-43 (GAP-43), a molecular marker of new axonal growth, was increased in the intact contralateral sensorimotor cortex following bFGF treatment. In the current study, we found that the intracisternal administration of antisense, but not missense, oligonucleotide to GAP-43 blocked the recovery-enhancing effects of bFGF and blocked the increase in GAP-43 IR in the contralateral cortex. These results suggest that upregulation of GAP-43 expression and consequent enhanced axonal sprouting in intact uninjured parts of the brain are likely mechanisms for the recovery-promoting effects of bFGF.

Glial growth factor 2 promotes functional recovery with treatment initiated up to 7 days after permanent focal ischemic stroke.

Neuregulins are a family of growth factors essential for normal cardiac and nervous system development. The EGF-like domain of neuregulins contains the active site which binds and activates signaling cascades through ErbB receptors. A neuregulin-1 gene EGF-like fragment demonstrated neuroprotection in the transient middle cerebral artery occlusion (MCAO) stroke model and drastically reduced infarct volume (Xu et al., 2004). Here we use a permanent MCAO rat model to initially compare two products of the neuregulin-1 gene and also assess levels of recovery with acute versus delayed time to treatment. In the initial study full-length glial growth factor 2 (GGF2) and an EGF-like domain fragment were compared with acute intravenous delivery. In a second study GGF2 only was delivered starting at 24h, 3 days or 7 days after permanent ischemia was induced. In both studies daily intravenous administration continued for 10 days. Recovery of neurological function was assessed using limb placing and body swing tests. GGF2 had similar functional improvements compared to the EGF-like domain fragment at equimolar doses, and a higher dose of GGF2 demonstrated more robust functional improvements compared to a lower dose. GGF2 improved sensorimotor recovery with all treatment paradigms, even enhancing recovery of function with a delay of 7 days to treatment. Histological assessments did not show any associated reduction in infarct volume at either 48 h or 21 days post-ischemic event. Neurorestorative effects of this kind are of great potential clinical importance, given the difficulty of delivering neuroprotective therapies within a short time after an ischemic event in human patients. If confirmed by additional work including additional data on mechanism(s) of improved outcome with verification in other stroke models, one can make a compelling case to bring GGF2 to clinical trials as a neurorestorative approach to improving outcome following stroke injury.

Dalfampridine Improves Sensorimotor Function in Rats With Chronic Deficits After Middle Cerebral Artery Occlusion

Background and Purpose— Stroke survivors often have permanent deficits that are only partially addressed by physical therapy. This study evaluated the effects of dalfampridine, a potassium channel blocker, on persistent sensorimotor deficits in rats with treatment initiated 4 or 8 weeks after stroke. Methods— Rats underwent permanent middle cerebral artery occlusion. Sensorimotor function was measured using limb-placing and body-swing symmetry tests, which normally show a partial recovery from initial deficits that plateaus ≈4 weeks after permanent middle cerebral artery occlusion. Dalfampridine was administered starting at 4 or 8 weeks after permanent middle cerebral artery occlusion in 2 blinded, vehicle-controlled studies. Plasma samples were collected and brain tissue was processed for histologic assessment. Results— Dalfampridine treatment (0.5–2.0 mg/kg) improved forelimb- and hindlimb-placing responses and body-swing symmetry in a reversible and dose-dependent manner. Plasma dalfampridine concentrations correlated with dose. Brain infarct volumes showed no differences between treatment groups. Conclusions— Dalfampridine improves sensorimotor function in the rat permanent middle cerebral artery occlusion model. Dalfampridine extended-release tablets (prolonged release fampridine outside the United States) are used to improve walking in patients with multiple sclerosis, and these preclinical data provide a strong rationale for examining the potential of dalfampridine to treat chronic stable deficits in stroke patients. Clinical Trial Registration— URL: http://www.clinicaltrials.gov. Unique identifier: NCT01605825

Use of Anisotropy, 3D Segmented Atlas, and Computational Analysis to Identify Gray Matter Subcortical Lesions Common to Concussive Injury from Different Sites on the Cortex.

Traumatic brain injury (TBI) can occur anywhere along the cortical mantel. While the cortical contusions may be random and disparate in their locations, the clinical outcomes are often similar and difficult to explain. Thus a question that arises is, do concussions at different sites on the cortex affect similar subcortical brain regions? To address this question we used a fluid percussion model to concuss the right caudal or rostral cortices in rats. Five days later, diffusion tensor MRI data were acquired for indices of anisotropy (IA) for use in a novel method of analysis to detect changes in gray matter microarchitecture. IA values from over 20,000 voxels were registered into a 3D segmented, annotated rat atlas covering 150 brain areas. Comparisons between left and right hemispheres revealed a small population of subcortical sites with altered IA values. Rostral and caudal concussions were of striking similarity in the impacted subcortical locations, particularly the central nucleus of the amygdala, laterodorsal thalamus, and hippocampal complex. Subsequent immunohistochemical analysis of these sites showed significant neuroinflammation. This study presents three significant findings that advance our understanding and evaluation of TBI: 1) the introduction of a new method to identify highly localized disturbances in discrete gray matter, subcortical brain nuclei without postmortem histology, 2) the use of this method to demonstrate that separate injuries to the rostral and caudal cortex produce the same subcortical, disturbances, and 3) the central nucleus of the amygdala, critical in the regulation of emotion, is vulnerable to concussion.