[Show abstract][Hide abstract] ABSTRACT: Neuromyelitis optica (NMO) is a devastating inflammatory disorder of the optic nerves and spinal cord characterized by frequently recurring exacerbations of humoral inflammation. NMO is associated with the highly specific NMO-IgG biomarker, an antibody that binds the aquaporin-4 water channel. Aquaporin-4 is present on glial endfeet in the central nervous system (CNS). In humans, the NMO-IgG portends more frequent exacerbations and a worse long-term clinical outcome.
We tested the longer-term outcome of mice with EAE injected with NMO-IgG and followed them for 60 days. Clinical exams and pathology of the spinal cord and optic nerves were compared to mice that received control human IgG.
Passively transferred human NMO-IgG leads to more severe neurology disability over two months after onset of disease. Clinical worsening is associated with an increased concentration of large demyelinating lesions primarily to subpial AQP4-rich regions of the spinal cord.
NMO-IgG is pathogenic in the context of EAE in mice.
[Show abstract][Hide abstract] ABSTRACT: Allografts continue to be used in clinical neurotransplantation studies, hence it is crucial to understand the mechanisms that govern allograft tolerance. We investigated the impact of transplantation site within the brain on graft survival. Mouse (FVB) glial precursors, transfected with luciferase have been injected (3x10⁵) into the forceps minor (FM) or striatum (STR). Immunodeficient rag2-/- and immunocompetent BALB/c mice were used as recipients. Magnetic resonance imaging confirmed that cells were precisely deposited at the selected coordinates. The graft viability was assessed non-invasively with bioluminescent imaging for a period of 16 days. Regardless of implantation site all grafts (n=10) deposited in immunodeficient animals revealed excellent survival. In contrast, immunocompetent animals accepted all grafts only at STR site (n=10), while all FM grafts were rejected (n=10). To investigate the factors that led to rejection of FM grafts, with acceptance of STR grafts, another group of animals (n=19) was sacrificed during pre-rejection period, on day 5. Nearinfrared fluorescence imaging with IRDye®800CW-PEG probe displayed similar blood-brain barrier disruption at both graft locations. The morphological distribution of FM grafts was cylindrical, parallel to the needle track, while cells transplanted into the STR accumulated along the border between the striatum and corpus callosum. There was a significantly less infiltration by both innate and adaptive immune cells in the STR grafts, especially along the calloso-striatal border. With allograft survival being dependent on the transplantation site, the anatomical coordinates of the graft target should always be taken into account as it may determine success or failure of therapy.
[Show abstract][Hide abstract] ABSTRACT: The development of cell-based therapies opens up new avenues for treating a myriad of diseases of the central nervous system (CNS). While significant effort is being directed toward development of patient-specific, autologous transplantable cells, at present, the majority of cell transplantation studies performed clinically utilize allografts. In this context, the issue of graft rejection and immunoprotection is of key importance. In this study, we transplanted mouse glial-restricted progenitors into immunodeficient, immunocompetent, and immunosuppressed mice and monitored their survival non-invasively using bioluminescence imaging (BLI). With the use of serial BLI, we evaluated both the prevalence and dynamics of cell rejection. We demonstrate that allografts in immunocompetent mice were rejected at a rate of 69.2% (n=13) indicating that graft tolerance is possible even without immunosuppression. Immunosuppression using a combination of rapamycin and FK506 or cyclosporin failed to fully protect the grafts. FK506 and Rapamycin treatment resulted in a slight improvement of immunoprotection (22.2% rejected, n=9) compared to Cyclosporin A (55.6% rejected, n=9), however the difference was not significant. Notably, immunohistochemistry revealed leukocytes infiltrating the graft area in both rejecting and non-rejecting immunocompetent animals, but not in immunodeficient animals. The induction of an inflammatory process, even in surviving allografts, has implications for their long-term survival and functionality.
[Show abstract][Hide abstract] ABSTRACT: PURPOSE
Efficient targeting of therapeutic cells for diffuse and multifocal neurological diseases has proven to be difficult using either intraparenchymal, intravenous, or intra-arterial administration. We hypothesized that transgene expression of the docking receptor VLA-4, that binds to VCAM-1 expressed on inflamed brain endothelium, can improve homing of human, fetal-derived glial-restricted precursor (GRP) cells following intra-arterial injection.
METHOD AND MATERIALS
GRPs were transfected with the α4 and β1 subunits of VLA-4, and then labeled with SPIO-Rhodamine. LPS (6 mg/kg) was injected i.p. in rats to induce global VCAM-1 expression. 1x106 GRPs were infused in the right carotid artery in 4 animals cohorts of n=3 each: VLA4-/LPS-, VLA4+/LPS-, VLA-/LPS+, and VLA4+/LPS+. MRI was performed at 9.4 T before and 1, 10, 20, and 30 min post injection.
Real-time, quantitative serial whole brain MR imaging revealed that, at first pass, VLA-4+ cells docked exclusively within the vascular bed of the ipsilateral carotid artery. Pixel-by-pixel analysis revealed that injection of VLA-4+ cells in LPS-treated animals resulted in 3,979±705 hypointense pixels as compared to 868±317 in VLA-4- controls. Twenty minutes after injection, VLA-4+ cells were retained at >95% immediate post-injection maximum values, while control values decreased up to 75%. Histology confirmed co-localization of GRPs with brain endothelial cells.
Targeted intra-arterial delivery of VLA-4 expressing hGRPs to inflamed endothelium is feasible and can be monitored in real time using MRI in a quantitative, dynamic manner.
Stem cell homing to inflammatory brain lesions is critical for successful therapy; improvements using cell engineering can be followed using MRI.
Radiological Society of North America 2011 Scientific Assembly and Annual Meeting; 11/2011
[Show abstract][Hide abstract] ABSTRACT: Transplantation of glial progenitor cells results in transplant-derived myelination and improved function in rodents with genetic dysmyelination or chemical demyelination. However, glial cell transplantation in adult CNS inflammatory demyelinating models has not been well studied. Here we transplanted human glial-restricted progenitor (hGRP) cells into the spinal cord of adult rats with inflammatory demyelination, and monitored cell fate in chemically immunosuppressed animals. We found that hGRPs migrate extensively, expand within inflammatory spinal cord lesions, do not form tumors, and adopt a mature glial phenotype, albeit at a low rate. Human GRP-transplanted rats, but not controls, exhibited preserved electrophysiological conduction across the spinal cord, though no differences in behavioral improvement were noted between the two groups. Although these hGRPs myelinated extensively after implantation into neonatal shiverer mouse brain, only marginal remyelination was observed in the inflammatory spinal cord demyelination model. The low rate of transplant-derived myelination in adult rat spinal cord may reflect host age, species, transplant environment/location, and/or immune suppression regime differences. We conclude that hGRPs have the capacity to myelinate dysmyelinated neonatal rodent brain and preserve conduction in the inflammatory demyelinated adult rodent spinal cord. The latter benefit is likely dependent on trophic support and suggests further exploration of potential of glial progenitors in animal models of chronic inflammatory demyelination.
[Show abstract][Hide abstract] ABSTRACT: In spinal cord injury (SCI) research there is a need for reliable measures to determine the extent of injury and assess progress due to natural recovery, drug therapy, surgical intervention or rehabilitation. Somatosensory evoked potentials (SEP) can be used to quantitatively examine the functionality of the ascending sensory pathways in the spinal cord. A reduction of more than 50% in peak amplitude or an increase of more than 10% in latency are threshold indicators of injury. However, in the context of injury, SEP peaks are often obscured by noise. We have developed a new technique to investigate the morphology of the SEP waveform, rather than focusing on a small number of peaks. In this study, we compare SEP signals before and after SCI using two rat models: a contusion injury model and a focal experimental autoimmune encephalomyelitis model. Based on mean slope changes over the signal, we were able to effectively differentiate pre-injury and post-injury SEP values with high levels of sensitivity (83.3%) and specificity (79.2%).
Journal of Clinical Neuroscience 09/2010; · 1.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Myelin oligodendrocyte glycoprotein (MOG) is commonly used as an immunogen to induce an immune response against endogenous myelin, thereby modeling multiple sclerosis in rodents. When MOG is combined with complete Freund's adjuvant (CFA), multifocal, multiphasic disease ensues; whereas when MOG is combined with incomplete Freund's adjuvant (IFA), clinical disease is usually absent. MOG-IFA immunized animals can be induced to have neurological disease after intraspinal injections of cytokines and ethidium bromide (EtBr). In this study, we investigated whether MOG-IFA immunized rats exhibited subclinical injury as defined by somatosensory evoked potential (SEP) recordings. The titration of anti-MOG-125 antibodies showed robust peripheral mounting of immune response against myelin in MOG-immunized rats. However the SEP measures showed no significant change over time. Upon injecting cytokine-EtBr in the spinal cord after MOG sensitization, the SEP recordings showed reduced amplitude and prolonged latency, suggestive of axonal injury and demyelination in the dorsal column, respectively. These findings were later confirmed using T2-weighted MRI and histological hematoxylin-eosin stain of the spinal cord. This report establishes that MOG-IFA immunization alone does not alter neuronal conduction in SEP-related neural-pathways and that longitudinal in-vivo SEP recordings provide a sensitive read-out for focal myelitis (MOG-IFA and intraspinal cytokine-EtBr) in rats.
Journal of the neurological sciences 06/2009; 284(1-2):81-9. · 2.32 Impact Factor