Focal and diffuse cortical degenerative changes in a marmoset model of multiple sclerosis.

BACKGROUND: Degenerative features, such as neuronal, glial, synaptic and axonal loss, have been identified in neocortical and other grey matter structures in patients with multiple sclerosis, but mechanisms for neurodegeneration are unclear. Cortical demyelinating lesions are a potential cause of this degeneration, but the pathological and clinical significance of these lesions is uncertain as they remain difficult to identify and study in vivo. In this study we aimed to describe and quantify cellular and subcellular pathology in the cortex of myelin oligodendrocyte glycoprotein-induced marmoset experimental autoimmune encephalomyelitis using quantitative immunohistochemical methods. RESULTS: We found evidence of diffuse axonal damage occurring throughout cortical grey matter with evidence for synaptic loss and gliosis and a 13.6% decrease in neuronal size and occurring in deep cortical layers. Evidence of additional axonal damage and a 29.6-36.5% loss of oligodendrocytes was found in demyelinated cortical lesions. Leucocortical lesions also showed neuronal loss of 22.2% and a 15.8% increase in oligodendrocyte size. CONCLUSIONS: The marmoset experimental autoimmune encephalomyelitis model, therefore, shows both focal and generalized neurodegeneration. The generalized changes cannot be directly related to focal lesions, suggesting that they are either a consequence of diffusible inflammatory factors or secondary to remote lesions acting through trans-synaptic or retrograde degeneration.

Methods for magnetically labeling stem and other cells for detection by in vivo magnetic resonance imaging.

Superparamagnetic iron oxide (SPIO) nanoparticles are being used for intracellular magnetic labeling of stem cells and other cells in order to monitor cell trafficking by magnetic resonance imaging (MRI) as part of cellular-based repair, replacement and treatment strategies. This review focuses on the various methods for magnetic labeling of stem cells and other mammalian cells and on how to translate experimental results from bench to bedside.

Effective antigen-specific immunotherapy in the marmoset model of multiple sclerosis.

Mature T cells initially respond to Ag by activation and expansion, but high and repeated doses of Ag cause programmed cell death and can suppress T cell-mediated diseases in rodents. We evaluated repeated systemic Ag administration in a marmoset model of experimental allergic encephalomyelitis that closely resembles the human disease multiple sclerosis. We found that treatment with MP4, a chimeric, recombinant polypeptide containing human myelin basic protein and human proteolipid protein epitopes, prevented clinical symptoms and did not exacerbate disease. CNS lesions were also reduced as assessed in vivo by magnetic resonance imaging. Thus, specific Ag-directed therapy can be effective and nontoxic in primates.

Neurotransplantation of magnetically labeled oligodendrocyte progenitors: magnetic resonance tracking of cell migration and myelination.

Demyelination is a common pathological finding in human neurological diseases and frequently persists as a result of failure of endogenous repair. Transplanted oligodendrocytes and their precursor cells can (re)myelinate axons, raising the possibility of therapeutic intervention. The migratory capacity of transplanted cells is of key importance in determining the extent of (re)myelination and can, at present, be evaluated only by using invasive and irreversible procedures. We have exploited the transferrin receptor as an efficient intracellular delivery device for magnetic nanoparticles, and transplanted tagged oligodendrocyte progenitor cells into the spinal cord of myelin-deficient rats. Cell migration could be easily detected by using three-dimensional magnetic resonance microscopy, with a close correlation between the areas of contrast enhancement and the achieved extent of myelination. The present results demonstrate that magnetic resonance tracking of transplanted oligodendrocyte progenitors is feasible; this technique has the potential to be easily extended to other neurotransplantation studies involving different precursor cell types.

Serial MR imaging of experimental autoimmune encephalomyelitis induced by human white matter or by chimeric myelin-basic and proteolipid protein in the common marmoset.

BACKGROUND AND PURPOSE: Experimental autoimmune encephalomyelitis (EAE) in the marmoset was monitored by serial MR imaging to determine correlates to the natural-history MR studies in multiple sclerosis (MS). The relationships of MR-revealed lesions to clinical status and histopathologic findings were also explored. METHODS: We induced EAE by subcutaneous inoculation in two marmosets by human white matter (HWM) and in seven marmosets by MP4 (a chimeric recombinant fusion protein of myelin-basic and proteolipid protein) in adjuvant along with intravenous inactivated pertussis vaccine to facilitate the disease process. The HWM-inoculated animals were induced with Freund's adjuvant as the established model of marmoset EAE. The MP4-inoculated animals were induced with either Freund's incomplete adjuvant or TiterMax as part of a preclinical treatment trial. MR imaging was performed at 1.5 T at baseline, and repeated at 1- to 2-week intervals for a period of up to 16 weeks in six EAE-induced marmosets, and intermittently for up to 70 weeks in three EAE-induced and two control marmosets. Proton density- (PD-) and T2-weighted, pre- and postgadopentetate dimeglumine enhancement, T1-weighted, and magnetization transfer (MT) images were obtained. The brains were prepared for histologic evaluation of lesion distribution and counts, characterization of lesions as demyelinating or inflammatory, and histopathologic scoring. The clinical, MR, and pathologic scoring were done on grading systems, and correlated for evaluation. RESULTS: White matter (WM) changes after EAE induction were observed first at 9 days in the HWM-induced animals and at 2.5 weeks in the MP4-induced animals, with subsequent week-to-week fluctuations on PD- and T2-weighted images. Contrast-enhancing lesions were not observed in all animals. MR-revealed WM lesions correlated to histopathologic analysis of EAE lesions, measuring from 0.5 mm to 1.5 mm. The lesion count and extent of demyelination was greater in the HWM-induced animals than in the MP4-induced animals. Some MR-revealed lesions correlated directly to clinical symptoms, but the majority of lesions were clinically silent. CONCLUSION: On MR images, lesions in the EAE marmoset model were confined to the WM, and their development, resolution, distribution, and enhancing characteristics fluctuated over the duration of the study. The dynamic presentation of MR-revealed lesions confirms the parallels between EAE in the marmoset and relapsing-remitting MS. Clinical symptoms alone were not representative of ongoing pathologic brain lesions. Therefore, serial MR imaging serves as a very important adjunct to clinical and histologic surveillance of the development of new and the persistence of existing brain lesions in this animal model of MS.

Study of relapsing remitting experimental allergic encephalomyelitis SJL mouse model using MION-46L enhanced in vivo MRI: early histopathological correlation.

MION-46L, a superparamagnetic iron oxide contrast agent, was investigated for its ability to increase the sensitivity of in vivo 3D MRI in the detection of brain lesions in a chronic experimental allergic encephalomyelitis (crEAE) mouse model. Lesion conspicuity on postcontrast 3D MRI was dramatically enhanced as compared to precontrast images corresponding to areas of inflammatory and demyelinating lesions. MION-46L could be detected on Prussian blue iron stain in the vascular endothelium, the perivascular space, and in macrophages within perivascular cuffs and areas of inflammation and demyelination. By taking advantage of the MION-46L induced macroscopic susceptibility effect, acute early lesions measuring only 100 microm in diameter could be detected. MION-46L enhanced MRI may be used to 1) provide a unique sensitivity in EAE lesion detection and correlate imaging to histopathology; 2) help to understand EAE lesion development and its underlying pathophysiology; and 3) eventually assist in preclinical screening of new experimental therapies directed at patients with multiple sclerosis (MS).

In vivo three-dimensional MR microscopy of mice with chronic relapsing experimental autoimmune encephalomyelitis after treatment with insulin-like growth factor-I.

PURPOSE: The purpose of this study was to determine the ability of three-dimensional in vivo MR microscopy to depict the treatment effects of insulin-like growth factor-I (IGF-I) in SJL mice with chronic relapsing experimental autoimmune encephalomyelitis (crEAE). METHODS: The experiments were performed at 4.7-T on 10 crEAE mice and on one set of control animals. Five crEAE mice were treated with IGF-I and five were treated with a placebo. RESULTS: In the crEAE mice treated with the placebo, in vivo MR microscopy showed areas of abnormal signal throughout the cerebrum, brain stem, and cerebellum. These findings were not present in either the IGF-I-treated mice or the normal control animals. The diffuse alterations in signal intensity in the placebo-treated crEAE mice were not identified on histologic sections of the same areas. CONCLUSION: Differences between the IGF-I- and placebo-treated groups may reflect changes in stabilization or permeability of cell membranes and/or of the blood-brain barrier, although other alternative contrast mechanisms could be playing a role. In vivo MR microscopy depicted changes resulting from treatment of crEAE with IGF-I.

Fetal damage caused by parvoviral infections.

The single-stranded DNA parvoviruses occur in humans and many species of animals. In general, they are species-specific and capable of producing disease at any stage of life. Parvoviruses have a requirement to replicate in cells in a permissive S-phase of DNA mitosis. The infections may be cytolytic to select cell groups resulting in specific developmental defects or may produce more generalized effects such as anemia, pancytopenia, or hemorrhage. The fetus is at particular risk for damage because of the vast number of cells in active mitosis. The teratogenic effects may be severe, often resulting in fetal death. Infections in childhood and adulthood are more frequently mild to subclinical. Some of the teratogenic effects recognized in animal species have been identified in humans. With increased knowledge of parvovirus effects in animals, more pathogenic effects may be related to human parvoviral disease. The need for vaccination, currently used annually in many domestic animal species, continues to be evaluated for humans.

Virus isolation and quinolinic acid in primary and chronic simian immunodeficiency virus infection.

OBJECTIVE: In this 2.5-year study of simian immunodeficiency virus (SIVsm) infection in rhesus monkeys, quinolinic acid (QUIN) levels and virus isolation determinations were made in serial cerebrospinal fluid (CSF) and blood samples to evaluate the relationship between these parameters over the course of infection. METHODS: Eight rhesus monkeys were inoculated in the saphenous vein with SIVsm. Four animals were maintained as uninoculated controls. CSF and blood samples were obtained every 1-4 weeks over the course of study. SIV isolation was determined in H9 cells for the CSF and in primary rhesus lymphocyte co-cultures for peripheral blood mononuclear cells (PBMC). QUIN was quantitated in CSF and serum by electron-capture negative chemical ionization gas chromatography mass spectrometry. RESULTS: All SIV-inoculated animals became CSF and PBMC isolation-positive by 1-3 weeks post-inoculation. Control animals remained SIV-negative. One SIV-positive animal was humanely euthanized at 2 weeks post-inoculation. The three SIV-inoculated animals that were CSF isolation-negative after the fifth week post-inoculation maintained CSF QUIN values < 100 nM, remained CSF and PBMC isolation-negative, and clinically healthy in the chronic course of disease. In contrast, the four SIV-inoculated animals that were CSF isolation-positive 6-8 weeks post-inoculation had CSF QUIN levels as high as 153-565 nM during the second month post-inoculation and remained CSF virus isolation-negative, persistently PBMC isolation-positive, and experienced clinical symptoms of SIV in the chronic course of disease. Three of these four animals have succumbed to SIV infection. DISCUSSION: Initial QUIN responses and viral isolation status in the first month post-inoculation were consistent among SIV-inoculated animals with CSF and serum QUIN values significantly higher than those of controls. A divergence within the SIV-inoculated group of animals became apparent within the second month of primary SIV infection and was maintained throughout the course of infection. Persistent PBMC viral isolation and marked elevations of QUIN were linked to symptomatic disease and a poor prognosis for survival. Predominantly negative PBMC viral isolation and slight, but significant, elevations of QUIN were linked to asymptomatic disease with a favorable prognosis for survival.

Relationship of neurologic status in macaques infected with the simian immunodeficiency virus to cerebrospinal fluid quinolinic acid and kynurenic acid.

Increased concentrations of the excitotoxin quinolinic acid (QUIN) have been implicated in the neurologic deficits and brain atrophy that may accompany infection with the human immunodeficiency virus type-1. Key neuropathologic features of the AIDS encephalitis are replicated in some macaques following infection with the simian immunodeficiency virus (SIV). In the present studies, cerebrospinal fluid (CSF) QUIN concentrations increased within 2 weeks following infection of 11 rhesus macaques (Macaca mulatta) with a neurotropic sooty mangabey isolate of the simian immunodeficiency virus (SIVsm) and were sustained to greater than 2 standard deviations above uninfected control macaques. Highest CSF QUIN concentrations (up to 400-fold above pre-inoculation levels) were observed in 6 SIVsm-infected macaques with motor and behavioral abnormalities during life, brain atrophy on MRI scan and inflammatory lesions within the brain and meninges. Four of the 6 neurologic macaques deteriorated rapidly within 12 weeks after inoculation and had substantially larger increases in CSF QUIN levels than 2 other neurologic macaques and 5 macaques without neurologic signs which survived for longer than 37 weeks. Increases in serum QUIN and CSF kynurenic acid also occurred but generally to a lesser degree than the increases in CSF QUIN. In some animals, increases in serum L-kynurenine concentrations and reductions in CSF and serum L-tryptophan occurred and were consistent with activation of indoleamine-2, 3-dioxygenase, the first enzyme of the kynurenine pathway in extrahepatic tissues. CSF QUIN exceeded serum QUIN in 8.8% of samples from macaques with neurologic signs, supporting increased QUIN synthesis within the central nervous system. Production of [13C6]QUIN was demonstrated in one SIVsm-infected macaque and one uninfected control macaque following an intracisternal injection of [13C6]L-tryptophan and suggests that L-tryptophan is a substrate for QUIN synthesis within the nervous system or meninges, although the cellular localization of QUIN synthesis remain to be determined. We conclude that increases in kynurenine pathway metabolism occur in SIV-infected macaques and are most prominent in macaques with neurologic signs. Macaques infected with SIV offer a model to investigate the relationship between the metabolism of neuroactive kynurenines and neurologic disturbances associated with retroviral infection.

HLA-DR expression in macaque neuroendothelial cells in vitro and during SIV encephalitis.

HLA-DR expression in neuroendothelial cells (NEC) was studied during the course of SIV encephalitis in rhesus monkeys. HLA-DR determinants were detected on NEC in monkeys with SIV encephalitis, but not in control animals. In situ hybridization with an SIV probe indicated that HLA-DR expression was not a consequence of SIV replication within NEC. Cultured rhesus NEC stimulated with gamma interferon expressed HLA-DR to a higher degree than cultured brain fibroblasts or astrocytes. These data support the contention that NEC participate in retrovirus-induced inflammation and autoimmunity within the central nervous system.