Phenotypic analysis of the Plp1 gene overexpressing mouse model #72 : implications for demyelination and remyelination failure

Duplication of the proteolipid protein (PLP1) gene, which encodes the most abundant protein of central nervous system (CNS) myelin, is the most common cause of Pelizaeus Merzbacher disease (PMD). Various animal models have been generated to study the effect of Plp1 gene overexpression on oligodendrocyte and myelin sheath integrity. The #72 line harbours 3 additional copies of the murine Plp1 gene per haploidic chromosomal set. Homozygous #72 mice appear phenotypically normal until three months of age, after which they develop seizures leading to premature death at around 4 months of age. An earlier study examining the optic nerve showed a progressive demyelination accompanied by marked microglial and astrocytic responses. Using electron microscopy and immunohistochemistry, I demonstrated that initial myelination of the #72 corpus callosum was followed by a progressive demyelination, probably mediated by a distal “dying back” phenomenon of the myelin sheath. No evidence of effective remyelination was observed despite the presence and proliferation of oligodendrocyte progenitor cells (OPCs). A marked increase in density and reactivity of microglia/macrophages and astrocytes, and the occurrence of axonal swellings, accompanied the demyelination. In situ and in vitro evaluation of adult #72 OPCs provided evidence of impaired OPC differentiation. Transplantation of neurospheres (NS) into adult #72 mouse corpus callosum confirmed that axons were capable of undergoing remyelination. Furthermore, NS transplanted into neonatal CNS integrated into the parenchyma and survived up to 120 days, demonstrating the potential of early cell replacement therapy. Taking advantage of the spatially distinct pathologies between the retinal and chiasmal region of the #72 optic nerve, I evaluated the capability of diffusion weighted MRI to identify lesion type. I found significant differences between #72 and wild type optic nerves, as well as between the two distinct pathological regions within the #72 optic nerve. These results confirm the potential of the #72 mouse to serve as a model to study chronic demyelination. The study also demonstrates the utility of the #72 mouse to evaluate cell transplant strategies for the treatment of chronic CNS white matter lesions and PMD. Additionally, DW MRI has potential as a modality capable of diagnosing myelin-related white matter changes, and may be applicable to the clinical setting

Comparative magnetic resonance imaging and histopathological correlates in two SOD1 transgenic mouse models of amyotrophic lateral sclerosis

Amyotrophic Lateral Sclerosis (ALS) is a progressive and fatal disease due to motoneuron degeneration. Magnetic resonance imaging (MRI) is becoming a promising non-invasive approach to monitor the disease course but a direct correlation with neuropathology is not feasible in human. Therefore in this study we aimed to examine MRI changes in relation to histopathology in two mouse models of ALS (C57BL6/J and 129S2/SvHsd SOD1G93A mice) with different disease onset and progression. A longitudinal in vivo analysis of T2 maps, compared to ex vivo histological changes, was performed on cranial motor nuclei. An increased T2 value was associated with a significant tissue vacuolization that occurred prior to motoneuron loss in the cranial nuclei of C57 SOD1G93A mice. Conversely, in 129Sv SOD1G93A mice, which exhibit a more severe phenotype, MRI detected a milder increase of T2 value, associated with a milder vacuolization. This suggests that alteration within brainstem nuclei is not predictive of a more severe phenotype in the SOD1G93A mouse model. Using an ex vivo paradigm, Diffusion Tensor Imaging was also applied to study white matter spinal cord degeneration. In contrast to degeneration of cranial nuclei, alterations in white matter and axons loss reflected the different disease phenotype of SOD1G93A mice. The correspondence between MRI and histology further highlights the potential of MRI to monitor progressive motoneuron and axonal degeneration non-invasively in vivo. The identification of prognostic markers of the disease nevertheless requires validation in multiple models of ALS to ensure that these are not merely model-specific. Eventually this approach has the potential to lead to the development of robust and validated non-invasive imaging biomarkers in ALS patients, which may help to monitor the efficacy of therapies

Characterisation of spinal cord in a mouse model of spastic paraplegia related to abnormal axono-myelin interactions by in vivo quantitative MRI.

International audienceIn inherited neurodegenerative disorders the engineering of genetically modified mice for the causative genes have provided new insights in the understanding of axono-glial interactions. Patients lacking the major proteins of the central nervous system myelin, the proteolipoproteins (PLP1) exhibit an ascending axonopathy, named spastic paraplegia type 2. Our objective was to examine the interest of using quantitative MRI for non invasive detection of spinal cord (SC) consequences of the PLP1 defect in a mouse model of SPG2 (PLP1-/Y). For this purpose an MRI acquisition and retrospective correction chain was set up to map apparent diffusion coefficients (ADC) and T2 in the mouse cervical SC which improve the intra- and inter-animal homogeneity. This reliable imaging processing protocol allowed to detect significant changes between PLP1-/Y and wild type 15-month old SC, mainly no longer detected ex vivo after SC fixation. On the basis of ADC(//) and ADC( perpendicular) variations, white matter (WM) damages were characterised on both the myelin and axonal components. The microstructural changes observed in the Plp1 deficient grey matter (GM) were concomitantly related to the isotropic increase of GM ADC. The T2 reduction measured in the WM as well as the GM of the mutant SC seems to be also an interesting marker of the SC axono-glial dysfunction. The present study demonstrated the interest of quantitative MRI for phenotyping in vivo the WM and GM changes in SC neurodegenerative disorders related to myelin and impaired glia-axonal interaction

Removal of antagonistic spindle forces can rescue metaphase spindle length and reduce chromosome segregation defects

Regular Abstracts - Tuesday Poster Presentations: no. 1925Metaphase describes a phase of mitosis where chromosomes are attached and oriented on the bipolar spindle for subsequent segregation at anaphase. In diverse cell types, the metaphase spindle is maintained at a relatively constant length. Metaphase spindle length is proposed to be regulated by a balance of pushing and pulling forces generated by distinct sets of spindle microtubules and their interactions with motors and microtubule-associated proteins (MAPs). Spindle length appears important for chromosome segregation fidelity, as cells with shorter or longer than normal metaphase spindles, generated through deletion or inhibition of individual mitotic motors or MAPs, showed chromosome segregation defects. To test the force balance model of spindle length control and its effect on chromosome segregation, we applied fast microfluidic temperature-control with live-cell imaging to monitor the effect of switching off different combinations of antagonistic forces in the fission yeast metaphase spindle. We show that spindle midzone proteins kinesin-5 cut7p and microtubule bundler ase1p contribute to outward pushing forces, and spindle kinetochore proteins kinesin-8 klp5/6p and dam1p contribute to inward pulling forces. Removing these proteins individually led to aberrant metaphase spindle length and chromosome segregation defects. Removing these proteins in antagonistic combination rescued the defective spindle length and, in some combinations, also partially rescued chromosome segregation defects. Our results stress the importance of proper chromosome-to-microtubule attachment over spindle length regulation for proper chromosome segregation.postprin

Psr1p interacts with SUN/sad1p and EB1/mal3p to establish the bipolar spindle

Regular Abstracts - Sunday Poster Presentations: no. 382During mitosis, interpolar microtubules from two spindle pole bodies (SPBs) interdigitate to create an antiparallel microtubule array for accommodating numerous regulatory proteins. Among these proteins, the kinesin-5 cut7p/Eg5 is the key player responsible for sliding apart antiparallel microtubules and thus helps in establishing the bipolar spindle. At the onset of mitosis, two SPBs are adjacent to one another with most microtubules running nearly parallel toward the nuclear envelope, creating an unfavorable microtubule configuration for the kinesin-5 kinesins. Therefore, how the cell organizes the antiparallel microtubule array in the first place at mitotic onset remains enigmatic. Here, we show that a novel protein psrp1p localizes to the SPB and plays a key role in organizing the antiparallel microtubule array. The absence of psr1+ leads to a transient monopolar spindle and massive chromosome loss. Further functional characterization demonstrates that psr1p is recruited to the SPB through interaction with the conserved SUN protein sad1p and that psr1p physically interacts with the conserved microtubule plus tip protein mal3p/EB1. These results suggest a model that psr1p serves as a linking protein between sad1p/SUN and mal3p/EB1 to allow microtubule plus ends to be coupled to the SPBs for organization of an antiparallel microtubule array. Thus, we conclude that psr1p is involved in organizing the antiparallel microtubule array in the first place at mitosis onset by interaction with SUN/sad1p and EB1/mal3p, thereby establishing the bipolar spindle.postprin