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Biomedical subjects

K A Nave

Publications and source records attributed to K A Nave.

At least 19 recordsLinked to original sources
PubMed2006

Alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid-mediated excitotoxic axonal damage is attenuated in the absence of myelin proteolipid protein.

In vivo and in vitro studies have shown that alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-receptor-mediated excitotoxicity causes cytoskeletal damage to axons. AMPA/kainate receptors are present on oligodendrocytes and myelin, but currently there is no evidence to suggest that axon cylinders contain AMPA receptors. Proteolipid protein (PLP) and DM20 are integral membrane proteins expressed by CNS oligodendrocytes and located in compact myelin. Humans and mice lacking normal PLP/DM20 develop axonal swellings and degeneration, suggesting that local interactions between axons and the oligodendrocyte/myelin unit are important for the normal functioning of axons and that PLP/DM20 is involved in this process. To determine whether perturbed glial-axonal interaction affects AMPA-receptor-mediated axonal damage, AMPA (1.5 nmol) was injected into the caudate nucleus of anesthetized Plp knockout and wild-type male mice (n = 13). Twenty-four hours later, axonal damage was detected by using neurofilament 200 (NF 200) immunohistochemistry and neuronal damage detected via histology. AMPA-induced axonal damage, assessed with NF 200 immunohistochemistry, was significantly reduced in Plp knockout mice compared with wild-type mice (P = 0.015). There was no significant difference in the levels of neuronal perikaryal damage between the Plp knockout and wild-type mice. In addition, there was no significant difference in the levels of glutamate receptor subunits GluR1-4 or KA2 in Plp knockout compared with wild-type littermates. The present study suggests that PLP-mediated interactions among oligodendrocytes, myelin, and axons may be involved in AMPA-mediated axonal damage.

Animals↗
PubMed2003

Differential expression of L- and S-MAG upon cAMP stimulated differentiation in oligodendroglial cells.

Myelin-associated glycoprotein (MAG), an immunoglobulin-like cell signaling protein involved in axon-glial interactions, displays two intracellular C-termini as a result of alternative mRNA splicing. During brain development, the two MAG mRNAs that encode L-MAG and S-MAG differ in their relative abundance. We have investigated the differential expression of L- and S-MAG upon cAMP treatment in the oligodendroglial cell line Oli-neu, a cell line able to differentiate in vitro. We have engineered GFP and VSVG fusions by small insertions into the alternatively spliced exons of the cloned MAG gene and reintroduced them into Oli-neu cells. The individually tagged MAG isoforms were expressed under the control of the MAG promoter and regulatory region. In this system, L-MAG was the predominant isoform before the stimulation of cells with cAMP, whereas upon cAMP treatment the S-MAG isoform was predominantly expressed in cells with a high degree of morphological differentiation. We suggest that the regulation of the MAG alternative splicing and the morphological differentiation in oligodendrocytes are controlled both by the same cAMP-responsive differentiation step.

Animals↗
PubMed2001

Membrane traffic in myelinating oligodendrocytes.

In the central nervous system (CNS), the myelin sheath is synthesised by oligodendrocytes as a specialised subdomain of an extended plasma membrane, reminiscent of the segregated membrane domains of polarised cells. Myelination takes place within a relatively short period of time and oligodendrocytes must have adapted membrane sorting and transport mechanisms to achieve such a high rate of myelin synthesis and to maintain the unique organisation of the myelin membrane. In adult life, maintenance of the functional myelin sheath requires a carefully orchestrated balance of myelin synthesis and turnover. Imbalance in these processes may cause dys- or demyelination and disease. This review summarises what is currently known about myelin protein trafficking and mistrafficking in oligodendrocytes. We also present data demonstrating distinct transport pathways for myelin structural proteins and the expression of SNARE proteins in differentiating oligodendrocytes. Myelinating glial cells may well serve as a model system for studying general aspects of membrane trafficking and organisation of membrane domains.

Animals↗
PubMed2001

Myelin proteolipid proteins promote the interaction of oligodendrocytes and axons.

Although proteolipid protein (PLP) and its DM20 isoform are the major membrane proteins of CNS myelin, their absence causes surprisingly few developmental defects. In comparison, missense mutations of the X-linked Plp gene cause severe dysmyelination. Previous studies have established roles for PLP/DM20 in the formation of the intraperiod line and in maintaining axonal integrity. We now show that a normal number of oligodendrocytes are present in mice lacking PLP/DM20. However, in heterozygous females, which are natural chimeras for X-linked genes, oligodendrocytes lacking PLP/DM20 are in direct competition with wild-type oligodendrocytes that have a distinct advantage. PLP+ oligodendrocytes and PLP+ myelin sheaths make up the greater majority, and this feature is generalised in the CNS throughout life. Moreover, in the absence of PLP/DM20, a proportion of small-diameter axons fails to myelinate, remaining ensheathed but lacking a compact sheath, or show delayed myelination. These findings suggest that PLP/DM20 is also involved in the early stages of axon-oligodendrocyte interaction and wrapping of the axon.

Animals↗
PubMed2001

The transcription factor Sox10 is a key regulator of peripheral glial development.

The molecular mechanisms that determine glial cell fate in the vertebrate nervous system have not been elucidated. Peripheral glial cells differentiate from pluripotent neural crest cells. We show here that the transcription factor Sox10 is a key regulator in differentiation of peripheral glial cells. In mice that carry a spontaneous or a targeted mutation of Sox10, neuronal cells form in dorsal root ganglia, but Schwann cells or satellite cells are not generated. At later developmental stages, this lack of peripheral glial cells results in a severe degeneration of sensory and motor neurons. Moreover, we show that Sox10 controls expression of ErbB3 in neural crest cells. ErbB3 encodes a Neuregulin receptor, and down-regulation of ErbB3 accounts for many changes in development of neural crest cells observed in Sox10 mutant mice. Sox10 also has functions not mediated by ErbB3, for instance in the melanocyte lineage. Phenotypes observed in heterozygous mice that carry a targeted Sox10 null allele reproduce those observed in heterozygous Sox10(Dom) mice. Haploinsufficiency of Sox10 can thus cause pigmentation and megacolon defects, which are also observed in Sox10(Dom)/+ mice and in patients with Waardenburg-Hirschsprung disease caused by heterozygous SOX10 mutations.

Animals↗
PubMed2001

PMP22 transgenic dorsal root ganglia cultures show myelin abnormalities similar to those of human CMT1A.

Charcot-Marie-Tooth 1A (CMT1A) neuropathy is caused by duplication of the peripheral myelin protein 22 (PMP22) gene, leading to protein overexpression. Although this protein has a role in regulating Schwann cell growth and peripheral myelin compaction, how altered concentrations of PMP22 impair myelination is unknown. We established dorsal root ganglia (DRG) cultures from a transgenic rat overexpressing PMP22 (PMP22tg) to study the behavior of PMP22tg Schwann cells in early stages of development and myelination. We used reverse transcriptase-polymerase chain reaction and light and electron microscopy to study PMP22 expression and myelin formation. Myelin ultrastructure was evaluated in sural nerves from CMT1A patients to compare experimental and human findings. PMP22tg DRG cultures contained a greater number of internodes devoid of myelin, in the absence of remyelination, and increased periodicity of myelin lamellae compared with normal cultures. Widening of myelin lamellae was also observed in CMT1A biopsy specimens. Our results suggest that both functions of PMP22, in regulating Schwann cell differentiation and contributing to peripheral myelin compaction, are affected by its overexpression. The presence of similar myelin abnormalities in PMP22tg cultures and human nerves emphasizes the importance of developing in vitro models of hereditary neuropathies to study their underlying pathomechanisms.

Animals↗
PubMed2001

Multiple splice isoforms of proteolipid M6B in neurons and oligodendrocytes.

Proteolipids are abundant integral membrane proteins, initially described as structural proteins of CNS myelin. More recently, two neuronal proteins related to proteolipid protein (PLP), termed M6A and M6B, were identified, suggesting a common function of proteolipids in oligodendrocytes and neurons. We have analyzed the X-linked M6B gene and discovered an unexpected complexity of protein isoforms. Two promoters and alternative exons yield at least eight M6B proteins and polypeptides, differentially expressed in neurons and oligodendrocytes. Six isoforms are tetraspan membrane proteins that differ by highly conserved amino- and carboxy-terminal domains, termed alpha, beta, psi, and omega. In MDCK cells, the beta-domain of M6B stabilizes tetraspan proteolipids at the cell surface, whereas non-beta isoforms are more abundant in intracellular compartments. Cotransfection experiments suggest a physical interaction of M6B and mutant PLP, when retained in the endoplasmic reticulum, that may also contribute to oligodendrocyte dysfunction in Pelizaeus-Merzbacher disease.

Alternative Splicing↗
PubMed2001

Mutant Plp/DM20 cannot be processed to secrete PLP-related oligodendrocyte differentiation/survival factor.

Most of the mutations within the PLP gene result in degeneration of oligodendrocytes and this is believed to be caused by intracellular trafficking defects. Previous studies have demonstrated that cells expressing the wild type PLP gene release a factor promoting differentiation/survival of oligodendrocyte and that this factor is the C-terminal portion of the protein itself. In this study we asked how the naturally occurring mutations of the PLP gene (jimpy, jimpy msd, and rumpshaker) affect this activity. We developed a transient expression system for retroviral production and transduction that enabled the expression of mutant PLP/DM20 cDNAs in NIH3T3 cells. None of the NIH3T3 cells producing mutant PLP/DM20s secreted the PLP-related factor that increases the number of oligodendrocytes. Since it has been shown that rumpshaker DM20 can be transported to the cell surface, but its folding is incorrect, absence of secretion of this factor is more heavily attributable to incorrect protein folding than to the defect in the PLP/DM20 trafficking.

3T3 Cells↗
PubMed2000

Uncoupling of myelin assembly and schwann cell differentiation by transgenic overexpression of peripheral myelin protein 22.

We have generated previously transgenic rats that overexpress peripheral myelin protein 22 (PMP22) in Schwann cells. In the nerves of these animals, Schwann cells have segregated with axons to the normal 1:1 ratio but remain arrested at the promyelinating stage, apparently unable to elaborate myelin sheaths. We have examined gene expression of these dysmyelinating Schwann cells using semiquantitative reverse transcription-PCR and immunofluorescence analysis. Unexpectedly, Schwann cell differentiation appears to proceed normally at the molecular level when monitored by the expression of mRNAs encoding major structural proteins of myelin. Furthermore, an aberrant coexpression of early and late Schwann cell markers was observed. PMP22 itself acquires complex glycosylation, suggesting that trafficking of the myelin protein through the endoplasmic reticulum is not significantly impaired. We suggest that PMP22, when overexpressed, accumulates in a late Golgi-cell membrane compartment and uncouples myelin assembly from the underlying program of Schwann cell differentiation.

Animals↗
PubMed2000

Neuronal basic helix-loop-helix proteins (NEX and BETA2/Neuro D) regulate terminal granule cell differentiation in the hippocampus.

The transcription factors neuronal helix-loop-helix protein (NEX)/mammalian atonal homolog 2 (Math-2), BETA2/neuronal determination factor (NeuroD), and NeuroD-related factor (NDRF)/NeuroD2 comprise a family of Drosophila atonal-related basic helix-loop-helix (bHLH) proteins with highly overlapping expression in the developing forebrain. The ability of BETA2/NeuroD and NDRF to convert ectodermal cells into neurons after mRNA injection into Xenopus oocytes suggested a role in specifying neuronal cell fate. However, neuronal bHLH genes are largely transcribed in CNS neurons, which are fully committed. Here we analyze a defect in mice lacking BETA2/NeuroD, and in NEX*BETA2/NeuroD double mutants, demonstrating that bHLH proteins are required in vivo for terminal neuronal differentiation. Most strikingly, presumptive granule cells of the dentate gyrus are generated but fail to mature, lack normal sodium currents, and show little dendritic arborization. Long-term hippocampal slice cultures demonstrate secondary alterations of entorhinal and commissural/associational projections. The primary developmental arrest appears to be restricted to granule cells in which an autoregulatory system involving all three neuronal bHLH genes has failed.

Action Potentials↗
PubMed2000

Shaping of the autoreactive T-cell repertoire by a splice variant of self protein expressed in thymic epithelial cells.

Intrathymic expression of tissue-specific self antigens may be involved in immunological tolerance and protection from autoimmune disease. We have analyzed the role of T-cell tolerance to proteolipid protein (PLP), the main protein of the myelin sheath, in susceptibility to experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis. Intrathymic expression of PLP was largely restricted to the shorter splice variant, DM20. Expression of DM20 by thymic epithelium was sufficient to confer T-cell tolerance to all epitopes of PLP in EAE-resistant C57BL/6 mice. In contrast, the major T-cell epitope in SJL/J mice was only encoded by the central nervous system-specific exon of PLP, but not by thymic DM20. Thus, lack of tolerance to this epitope offers an explanation for the exquisite susceptibility of SJL/J mice to EAE. As PLP expression in the human thymus is also restricted to the DM20 isoform, these findings have implications for selection of the autoimmune T-cell repertoire in multiple sclerosis.

Alternative Splicing↗
PubMed1999

Distinct phenotypes associated with increasing dosage of the PLP gene: implications for CMT1A due to PMP22 gene duplication.

Increased dosage of the proteolipid protein (Plp) gene causes CNS disease (Pelizaeus-Merzbacher disease [PMD]), which has many similarities to disorders of the PNS associated with duplication of the peripheral myelin protein-22 (PMP22) gene locus. Transgenic mice carrying extra copies of the wild-type Plp gene provide a valid model of PMD. Variations in gene dosage can cause a wide range of phenotypes from severe, lethal dysmyelination through late-onset demyelination. A predilection for different fiber diameters may occur within the various phenotypes with dysmyelination being more obvious in large fibers and late-onset degeneration predominantly affecting small fibers. Although the frequency of apoptotic oligodendrocytes is increased with high gene dosage, the number of mature oligodendrocytes appears adequate. Oligodendrocytes in the dysmyelinated CNS express a range of genes typical of mature cells, yet are unable to assemble sufficient myelin. Oligodendrocytes contain abnormal vacuoles and stain intensely for PLP and other proteins such as MAG. The findings suggest that with high gene dosage much of the PLP, and possibly other proteins, is missorted and degraded in the lysosomal system.

Animals↗
PubMed1999

Transgenic mouse models of CMT1A and HNPP.

We have generated several PMP22 animal mutants with altered PMP22 gene dosage. A moderate increase in the number of PMP22 genes led to hypomyelination comparable to CMT1A, whereas high copy numbers of transgenic PMP22 resulted in phenotypes resembling more severe forms of hereditary motor and sensory neuropathies. In contrast, eliminating one of the two normal PMP22 genes by gene targeting caused unstable focal hypermyelination (tomacula) similar to the pathology in HNPP. A related but more severe phenotype was observed in mice that lack PMP22 completely. Detailed analysis of the different PMP22 mutants revealed, in addition to the obvious myelinopathy, distal axonopathy as a characteristic feature. We conclude that the maintenance of axons might be a promising target for therapeutic interventions in these demyelinating hereditary neuropathies. Furthermore, our results strongly support the concept that PMP22-related neuropathies (and most likely also other forms of inherited motor and sensory neuropathies) should be viewed as the consequence of impaired neuron-Schwann cell interactions that are likely already to be operative during development. Such considerations should be taken into account in the design of potential novel treatment strategies.

Animals↗
PubMed1999

The "CMT rat": peripheral neuropathy and dysmyelination caused by transgenic overexpression of PMP22.

We have generated a transgenic rat model of Charcot-Marie-Tooth disease type 1A (CMT1A) providing formal proof that this neuropathy can be caused by increased expression of peripheral myelin protein-22 (PMP22). Heterozygous PMP22-transgenic rats develop muscle weakness and gait abnormalities as well as reduced nerve conduction velocities and EMG abnormalities, which closely resemble recordings in patients with CMT1A. Dys- and demyelination, Schwann cell hypertrophy, and "onion bulb" formation are also similar to findings in humans. When bred to homozygosity, transgenic rats completely fail to elaborate myelin, but all myelin-forming Schwann cells segregate with axons in the normal one-to-one ratio. Although arrested at this "promyelin" stage, differentiation proceeds in homozygous rats at the molecular level, as demonstrated by high-level expression of myelin structural genes. Intracellular trafficking of the wild-type protein is not visibly impaired, even when strongly overexpressed, suggesting that PMP22 blocks myelin assembly in a late Golgi/cell membrane compartment of the affected Schwann cell.

Animals↗
PubMed1999

Myelination in the absence of galactolipids and proteolipid proteins.

The galactolipids galactocerebroside and sulfatide and the proteolipid protein (PLP) and its splice variant DM20 are the most abundant lipid and protein components of central nervous system myelin. Recent studies have found that mice lacking either the galactolipids or PLP are able to form myelin sheaths with apparently normal periodicity and near normal compaction. Here, we have generated galactolipid/proteolipid double mutants to examine the possibility that these molecules have overlapping functions. We show that the absence of the galactolipids and PLP has pleotropic effects on myelin formation. While oligodendrocytes in the postnatal day 20 galactolipid/proteolipid-deficient mouse are able to elaborate myelin with close to normal intraperiod lines, there is an increased frequency of uncompacted myelin sheaths as well as unmyelinated axons. Moreover, the double mutants display extensive white matter vacuolization of the cerebellum that initiates around postnatal day 16, which correlates with the onset of a severe ataxic phenotype and an increased percentage of apoptotic nuclei in the cerebellar internal granule cell layer. These data indicate that the galactolipids and PLP/DM20 are not required for intraperiod line formation, but they suggest a role for these molecules in mediating myelin compaction and in maintaining the integrity of the cerebellum.

Aging↗
PubMed1999

Transgenic Lewis rats overexpressing the proteolipid protein gene: myelin degeneration and its effect on T cell-mediated experimental autoimmune encephalomyelitis.

Transgenic Lewis rats overexpressing proteolipid protein (PLP) genes in peripheral and central nervous myelin were produced by microinjecting murine genomic PLP sequences into fertilized eggs. The mouse PLP gene shares 98.7% homology in the nucleotide sequence with its rat counterpart, but both are fully identical on protein level. Homozygous rats show tremors early in postnatal life, eventually develop seizures, and die before they reach weaning age, while hemizygous animals are phenotypically normal and have a normal life expectancy. Transgene expression in the central nervous system (CNS) has profound consequences for myelin formation and maintenance: approximately twofold overexpression of PLP/DM-20, as seen in homozygotes, results in apoptosis of mature, and a developmental arrest of the remaining immature oligodendrocytes. Severe dysmyelination ensues, associated with reactive astrogliosis and microglia activation/proliferation. Activation of microglia is also prominent in hemizygous rats with low levels of transgene overexpression. In these animals, myelin sheaths remain intact, but there is low-grade myelin degeneration throughout life witnessed by myelin uptake and activation of microglia and astrocytes, in the absence of the expression of major histocompatibility complex class II gene products. There were no spontaneous lymphocytic infiltrates in areas of myelin degeneration. However, hemizygous LEW.PLP rats were more sensitive to experimental autoimmune encephalomyelitis mediated by T cells specific for PLP, but not another encephalitogenic myelin protein, MBP.

Animals↗
PubMed1998

Axonal swellings and degeneration in mice lacking the major proteolipid of myelin.

Glial cells produce myelin and contribute to axonal morphology in the nervous system. Two myelin membrane proteolipids, PLP and DM20, were shown to be essential for the integrity of myelinated axons. In the absence of PLP-DM20, mice assembled compact myelin sheaths but subsequently developed widespread axonal swellings and degeneration, associated predominantly with small-caliber nerve fibers. Similar swellings were absent in dysmyelinated shiverer mice, which lack myelin basic protein (MBP), but recurred in MBP*PLP double mutants. Thus, fiber degeneration, which was probably secondary to impaired axonal transport, could indicate that myelinated axons require local oligodendroglial support.

Animals↗