Guangliang Ding

Effects of administration route on migration and distribution of neural progenitor cells transplanted into rats with focal cerebral ischemia, an MRI study

We tested the hypotheses that administration routes affect the migration and distribution of grafted neural progenitor cells (NPCs) in the ischemic brain and that the ischemic lesion plays a role in mediating the grafting process. Male Wistar rats (n=41) were subjected to 2-h middle cerebral artery occlusion (MCAo), followed 1 day later by administration of magnetically labeled NPCs. Rats with MCAo were assigned to one of three treatment groups targeted for cell transplantation intra-arterially (IA), intracisternally (IC), or intravenously (IV). MRI measurements consisting of T2-weighted imaging and three-dimensional (3D) gradient echo imaging were performed 24 h after MCAo, 4 h after cell injection, and once a day for 4 days. Prussian blue staining was used to identify the labeled cells, 3D MRI to detect cell migration and distribution, and T2 map to assess lesion volumes. Intra-arterial (IA) administration showed significantly increased migration, a far more diffuse distribution pattern, and a larger number of transplanted NPCs in the target brain than IC or IV administration. However, high mortality with IA delivery (IA: 41%; IC: 17%; IV: 8%) poses a serious concern for using this route of administration. Animals with smaller lesions at the time of transplantation have fewer grafted cells in the parenchyma.

Magnetic resonance imaging investigation of axonal remodeling and angiogenesis after embolic stroke in sildenafil-treated rats

Interaction between angiogenesis and axonal remodeling after stroke was dynamically investigated by MRI in rats with or without sildenafil treatments. Male Wistar rats were subjected to embolic stroke and treated daily with saline (n = 10) or with sildenafil (n = 11) initiated at 24 h and subsequently for 7 days after onset of ischemia. T*2-weighted imaging, cerebral blood flow (CBF), and diffusion tensor imaging (DTI) measurements were performed from 24 h to 6 weeks after embolization. T*2 and fractional anisotropy (FA) maps detected angiogenesis and axonal remodeling after stroke, respectively, starting from 1 week in sildenafil-treated rats. Areas demarcated by MRI with enhanced angiogenesis, elevated local CBF, and augmented axonal remodeling were spatially and temporally matched, and FA values were significantly correlated with the corresponding CBF values (R = 0.66, P < 4 × 10−5) at 6 weeks after stroke. Axonal projections were reorganized along the ischemic boundary after stroke. These MRI measurements, confirmed by histology, showed that sildenafil treatment simultaneously enhanced angiogenesis and axonal remodeling, which were MRI detectable starting from 1 week after stroke in rats. The spatial and temporal consistency of MRI metrics and the correlation between FA and local CBF suggest that angiogenesis, by elevating local CBF, promotes axonal remodeling after stroke.

Angiogenesis and improved cerebral blood flow in the ischemic boundary area detected by MRI after administration of sildenafil to rats with embolic stroke.

To dynamically investigate the long-term response of an ischemic lesion in rat brain to the administration of sildenafil, male Wistar rats subjected to embolic stroke were treated with sildenafil (n=11) or saline (n=10) at a dose of 10 mg/kg administered subcutaneously 24-h after stroke and daily for an additional 6 days. Magnetic resonance images were acquired and functional performance was measured in all animals at 1 day, 2 days and weekly for 6 weeks post-stroke. All rats were sacrificed 6 weeks after stroke and endothelial barrier antigen immunostaining was employed for morphological analysis and quantification of cerebral vessels. Map-ISODATA was computed from T(1), T(2) and T(1sat) maps. ISODATA derived tissue signatures characterize the degree of ischemic injury. Based on the map-ISODATA calculated at 6 weeks, the ischemic lesion for each animal was divided into two specific regions, the ischemic boundary and ischemic core. The temporal profiles of cerebral blood flow (CBF) and tissue signature were retrospectively tracked in these two regions and were compared with histological evaluation and functional outcome. After 1 week of sildenafil treatment, the ischemic lesion exhibited two significantly different regions, with higher CBF level and correspondingly, lower tissue signature value in the boundary region than in the core region. Sildenafil treatment did not significantly reduce the lesion size, but did enhance angiogenesis. Functional performance was significantly increased after sildenafil treatment compared with the control group. Administration of sildenafil to rats with embolic stroke enhances angiogenesis and selectively increases the CBF level in the ischemic boundary, and improves neurological functional recovery compared to saline-treated rats.

MRI identification of white matter reorganization enhanced by erythropoietin treatment in a rat model of focal ischemia

Background and Purpose— The objectives of the present study were to: (1) noninvasively identify white matter reorganization and monitor its progress within 6 weeks after the onset of stroke; and (2) quantitatively investigate the effect of recombinant human erythropoietin treatment on this structural change using in vivo measurement of diffusion anisotropy. Methods— Male Wistar rats were subjected to middle cerebral artery occlusion and treated with recombinant human erythropoietin intraperitoneally at a dose of 5000 U/kg of body weight (n=11) or the same volume of saline (n=7) daily for 7 days starting 24 hours after middle cerebral artery occlusion. MRI measurements of T2- and diffusion-weighted images and cerebral blood flow were performed and neurological severity score was assessed at 1 day and weekly for 6 weeks after middle cerebral artery occlusion. Luxol fast blue and Bielschowsky staining were used to demonstrate myelin and axons, respectively. Results— White matter reorganization occurred along the ischemic lesion boundary after stroke. The region of white matter reorganization seen on the tissue slice coincided with the elevated area on the fractional anisotropy map, which can be accurately identified. The increase in elevated fractional anisotropy pixels corresponded with progress of white matter reorganization and was associated with improvement of neurological function. Treatment with recombinant human erythropoietin after stroke significantly enhanced white matter reorganization, restored local cerebral blood flow, and expedited functional recovery. Conclusions— White matter reorganization can be detected by fractional anisotropy. Elevated fractional anisotropy pixels may be a good MRI index to stage white matter remodeling and predict functional outcome.

Angiogenesis Detected After Embolic Stroke in Rat Brain Using Magnetic Resonance T2*WI

Background and Purpose— This study uses T2* weighted imaging (T2*WI) to measure the temporal evolution of cerebral angiogenesis in rats subjected to embolic stroke up to 6 weeks after stroke onset with or without sildenafil treatment. Method— Male Wistar rats were subjected to embolic stroke and treated with saline (n=10) or with sildenafil (n=11), with treatment initiated at 24 hours and continued daily for 7 days after onset of ischemia. T2*WI measurements were performed at 24 hours after embolization and weekly up to 6 weeks using a 7-Tesla system. Histological measurements were obtained at 6 weeks after MRI scans. Results— Using T2*WI, cerebral angiogenesis was detected starting from 4 weeks and from 2 weeks after onset of embolic stroke in saline and sildenafil treated rats, respectively. Significant differences in the temporal and spatial features of angiogenesis after embolic stroke up to 6 weeks after onset of stroke were found between saline and sildenafil treated rats and were identified with T2*WI. MRI permeability parameter, Ki, complementarily detected angiogenesis after ischemia in embolic stroke rats. Sildenafil treatment of stroke rats significantly enhanced the angiogenesis, as confirmed histologically. Conclusions— T2*WI can quantitatively measure the temporal evolution of angiogenesis in rats subjected to embolic stroke. Compared to control rats, sildenafil treatment significantly increased angiogenesis in treated animals up to 6 weeks after stroke.

Impairment of the glymphatic system after diabetes

The glymphatic system has recently been shown to clear brain extracellular solutes and abnormalities in glymphatic clearance system may contribute to both initiation and progression of neurological diseases. Despite that diabetes is known as a risk factor for vascular diseases, little is known how diabetes affects the glymphatic system. The current study is the first investigation of the effect of diabetes on the glymphatic system and the link between alteration of glymphatic clearance and cognitive impairment in Type-2 diabetes mellitus rats. MRI analysis revealed that clearance of cerebrospinal fluid contrast agent Gd-DTPA from the interstitial space was slowed by a factor of three in the hippocampus of Type-2 diabetes mellitus rats compared to the non-DM rats and confirmed by florescence imaging analysis. Cognitive deficits detected by behavioral tests were highly and inversely correlated to the retention of Gd-DTPA contrast and fluorescent tracer in the hippocampus of Type-2 diabetes mellitus rats. Type-2 diabetes mellitus suppresses clearance of interstitial fluid in the hippocampus and hypothalamus, suggesting that an impairment of the glymphatic system contributes to Type-2 diabetes mellitus-induced cognitive deficits. Whole brain MRI provides a sensitive, non-invasive tool to quantitatively evaluate cerebrospinal fluid and interstitial fluid exchange in Type-2 diabetes mellitus and possibly in other neurological disorders, with potential clinical application.

Ischemic cerebral tissue response to subventricular zone cell transplantation measured by iterative self-organizing data analysis technique algorithm

To investigate the changes of the ischemic lesion in rat brain after subventricular zone (SVZ) cell transplantation and the influence of the grafted cells on the appearance of angiogenesis, Svz cells, superparamagnetically labeled, were intracisternally transplanted into the rat brain 48 h after onset of embolic stroke. A complete set of magnetic resonance (MR) images was acquired for all animals with (n = 8) and without (n = 3) cell grafting at approximately 24 h, 72 h, and weekly for 6 weeks after stroke. Transplanted cells were tracked by high-resolution three-dimensional gradient-echo images and the interaction between the cells and ischemic lesion was detected by ISODATA (Iterative Self-Organizing Data Analysis Technique Algorithm) calculated from T1, T2 and T1sat maps. Tissue status from ISODATA was characterized by a specific signature, which represents the deviation from normal tissue in the feature space. Transplanted SVZ cells selectively migrated towards the ischemic side of the rat brain and approached the lesion boundary within 1-week after grafting. Cell treated rats exhibited a significant reduction of average lesion size compared with control rats (P < 0.05). A significant reduction of tissue signature (P < 0.001) induced by cell transplantation was localized to the position of grafted cells, and these sites exhibited stably restored cerebral blood flow (CBF) (approximately 85% of normal CBF). Angiogenesis was present in sites either immediately adjacent to or surrounded by the grafted cells. Our data indicate that map-ISODATA accurately and dynamically characterizes the ischemic lesion and its response to cell therapy.

A Model of Mini-Embolic Stroke Offers Measurements of the Neurovascular Unit Response in the Living Mouse

Background and Purpose— To measure cerebral vascular and neuronal responses after stroke in the living mouse, we generated a mouse model of embolic stroke localized to the parietal cortex. Methods— Male C57/6J or male transgenic mice (2 to 3 months old) expressing yellow fluorescent protein (YFP) were used in the present study. A single fibrin-rich clot (8 mm in length) was injected into a branch of the right middle cerebral artery (MCA). MRI measurements were performed to measure ischemic lesion. Using confocal and 2-photon microscopy, changes in the embolus, dendrites, and dendritic spines were measured in the living mouse. Results— Eight of 11 mice (73%) had the embolus localized to a branch of the right MCA in the parietal cortex. Expansion of the embolus within the artery was observed 24 hours after stroke. The presence of ischemic lesion in the parietal cortex was verified by MRI measurements, and histopathological analysis revealed that these mice (n=8) had a cortical infarct volume of 4.9±3.6% of the contralateral hemisphere. In the living mouse, substantial loss of YFP-labeled axonal and dendritic structures as well as the formation of abnormal dendritic bulbs were detected in the ischemic boundary regions 24 hours after stroke compared with that 1 hour after stroke. Conclusion— This model offers a novel approach to study the neurovascular unit in cerebral cortex after stroke in the living mouse.

Analysis of Combined Treatment of Embolic Stroke in Rat with r-tPA and a GPIIb/IIIa Inhibitor

Suppression of platelet activation improves the efficacy of thrombolytic therapy for stroke. Thus, combination treatment with recombinant tissue plasminogen activator (r-tPA) and 7E3 F(ab′)2, a GPIIb/IIIa inhibitor that binds the platelet to fibrin, may improve the efficacy of thrombolytic therapy in embolic stroke. Magnetic resonance imaging (MRI) was used to monitor treatment response in rats subjected to embolic middle cerebral artery (MCA) occlusion (MCAo). Animals were randomized into treated (n = 12) and control (n = 10) groups and received intravenous combination therapy or saline, respectively, 4 hours after MCAo. Magnetic resonance imaging (MRI) measurements performed 1 hour after MCAo showed no difference between groups. However, an increased incidence (50%) of MCA recanalization was found in the treated group at 24 hours compared with 20% in the control group. The area of low cerebral blood flow at 24 and 48 hours was significantly smaller in the combination treatment group, and the lesion size, as indicated from the T2 and T1 maps, differed significantly between groups. Fluorescence microscopy measurements of cerebral microvessels perfused with fluorescein isothiocyanate-dextran and measurements of infarct volume revealed that the combination treatment significantly increased microvascular patency and reduced infarct volume, respectively, compared with the control rats. The efficacy of combination treatment 4 hours after ischemia is reflected by MRI indices of tissue perfusion, MCA recanalization, and reduction of lesion volume. The treatment also reduced secondary microvascular perfusion deficits.

Multiparametric ISODATA analysis of embolic stroke and rt-PA intervention in rat

To increase the sensitivity of MRI parameters to detect tissue damage of ischemic stroke, an unsupervised analysis method, Iterative Self-Organizing Data Analysis Technique Algorithm (ISODATA), was applied to analyze the temporal evolution of ischemic damage in a focal embolic cerebral ischemia model in rat with and without recombinant tissue plasminogen activator (rt-PA) treatment. Male Wistar rats subjected to embolic stroke were investigated using a 7-T MRI system. Rats were randomized into control (n=9) and treated (n=9) groups. The treated rats received rt-PA via a femoral vein at 4 h after onset of embolic ischemia. ISODATA analysis employed parametric maps or weighted images (T1, T2, and diffusion). ISODATA results with parametric maps are superior to ISODATA with weighted images, and both of them were highly correlated with the infarction size measured from the corresponding histological section. At 24 h after embolic stroke, the average map ISODATA lesion sizes were 37.7+/-7.0 and 39.2+/-5.6 mm2 for the treated and the control group, respectively. Average histological infarction areas were 37.9+/-7.4 mm2 for treated rats and 39.4+/-6.1 mm2 for controls. The R2 values of the linear correlation between map ISODATA and histological data were 0.98 and 0.96 for treated and control rats, respectively. Both histological and map ISODATA data suggest that there is no significant difference in infarction area between non-treated and rt-PA-treated rats when treatment was administered 4 h after the onset of embolic stroke. The ISODATA lesion size analysis was also sensitive to changes of lesion size during acute and subacute stages of stroke. Our data demonstrate that the multiparameter map ISODATA approach provides a more sensitive quantitation of the ischemic lesion at all time points than image ISODATA and single MRI parametric analysis using T1, T2 or ADCw.