Search PubMed⌕ Search

Biomedical subjects

I R Griffiths

Publications and source records attributed to I R Griffiths.

At least 55 records · Page 3Linked to original sources
PubMed1996

Oligodendrocyte progenitors in the embryonic spinal cord express DM-20.

Oligodendrocyte progenitors, originating in the ventral ventricular zone of the embryonic rodent spinal cord, migrate and differentiate into the oligodendrocytes myelinating the future white matter. Transcripts for the dm-20 isoform of the proteolipid protein (plp) gene are detectable initially in cells of the ventral ventricular region of the embryonic central canal and subsequently throughout the white matter. The dm-20+ cells are present several days before oligodendrocytes or myelin sheaths are detectable. The purpose of the present study was to determine if DM-20 protein is present and whether DM-20+ cells can be linked to the oligodendrocyte lineage in the mouse spinal cord. Expression of plp and dm-20 transcripts and product was monitored using reverse transcription polymerase chain reaction (RT-PCR), and in situ hybridization and immunostaining of cryosections and associated cultures. Cell identification was performed using antigenic markers characterizing different stages of oligodendrocyte differentiation. We show a temporal and spatial progression of cells expressing dm-20 transcripts and product from the ventral ventricular zone at embryonic day 13 (E13.0), via the lateral borders of the floor plate to the ventral pia and white matter. The cells, initially devoid of myelin basic protein (MBP) and PLP, co-express these myelin proteins at approximately E16.5/17.0. Some DM-20+ cells co-label with definitive markers of the early oligodendrocyte lineage, are capable of mitosis and subsequently differentiate into oligodendrocytes. Other DM-20+ cells may represent earlier precursor cells. The expression of DM-20 in oligodendrocyte progenitors is consistent with a postulated role in glial cell development.

Animals↗
PubMed1995

Dominant-negative action of the jimpy mutation in mice complemented with an autosomal transgene for myelin proteolipid protein.

Mutations in genes encoding membrane proteins have been associated with cell death of unknown cause from invertebrate development to human degenerative diseases. A point mutation in the gene for myelin proteolipid protein (PLP) underlies oligodendrocyte death and dysmyelination in jimpy mice, an accurate model for Pelizaeus-Merzbacher disease. To distinguish the loss of PLP function from other effects of the misfolded protein, we took advantage of the X chromosomal linkage of the gene and have complemented jimpy with a wild-type PLP transgene. In this artificial heterozygous situation, the jimpy mutation emerged as genetically dominant. At the cellular level oligodendrocytes showed little increase in survival although endogenous PLP gene and autosomal transgene were truly coexpressed. In surviving oligodendrocytes, wild-type PLP was functional and immunodetectable in myelin. Moreover, compacted myelin sheaths regained their normal periodicity. This strongly suggests that, despite the presence of functional wild-type PLP, misfolded jimpy PLP is by itself the primary cause of abnormal oligodendrocyte death.

Animals↗
PubMed1995

O-2A progenitors of the mouse optic nerve exhibit a developmental pattern of antigen expression different from the rat.

In a previous study we demonstrated that differentiation and development of mouse oligodendrocytes is similar to that of the rat after the stage at which O4 is acquired. In this present study we compare directly the early differentiation of oligodendrocytes in the mouse and rat post natal optic nerve and show that the two species differ at the O-2A progenitor and proligodendroblast stages. Mouse progenitors show a variety of morphologies compared to the typical bipolar appearance in the rat. Many murine cells fail to immunolabel with A2B5, GD3, O4, and RmAb, classical markers for rat progenitors, proligodendroblasts, and immature oligodendrocytes. We find that these "unlabeled" cells stain for GAP-43 and that expression of GAP-43 overlaps A2B5 and GD3 in the earlier progenitors and O4, RmAb, and O1 in the later proligodendroblasts and immature oligodendrocytes. Our data suggest that in the development of the mouse O-2A progenitor cells there is a developmental discontinuity between the earlier markers such as A2B5 and GD3 and the later marker O4, which can be filled by GAP-43. We therefore consider that GAP-43 could be used in the mouse, in addition to the classical O-2A markers, for the study of the early oligodendrocyte lineage as it labels an otherwise undetectable O-2A population.

Animals↗
PubMed1995

The proteolipid protein gene.

Proteolipid protein (PLP) is the major myelin protein of the CNS and is believed to have a structural role in maintaining the intraperiod line of compact myelin. An isoform, DM-20, produced by alternative splicing of exon 3B is expressed earlier than PLP in the CNS and may be involved in glial cell development. DM-20 is also present in myelin-forming and non-myelin-forming Schwann cells, olfactory nerve ensheathing cells, some glial cell lines and cardiac myocytes. Molecular studies suggest the existence of a PLP gene family with sequence similarities between molecules of different species. Such studies also lend credence to the suggestion that PLP and/or DM-20 may function as a membrane pore. Mutations in the PLP gene occur in several animal species and cause severe pleiotropic effects on myelination. In man this presents as Pelizaeus-Merzbacher disease (PMD). The phenotype of such mutants is characterized by dysmyelination with myelin of abnormal periodicity, paucity of mature oligodendrocytes and astrocytosis. Duplication of the PLP gene in transgenic animals or in one form of PMD also results in dysmyelination. X-linked spastic paraplegia (SPG2) is allelic to PMD and is associated with PLP mutations in which the levels of the DM-20 isoform are probably relatively normal. The effects of PLP gene dosage on CNS myelination can be compared in many ways to the variety of phenotypes in the PNS in hereditary neuropathies of the Charcot-Marie-Tooth type in which the peripheral myelin-22 gene is mutated.

Animals↗
PubMed1995

Expression of the proteolipid protein gene in glial cells of the post-natal peripheral nervous system of rodents.

The proteolipid protein (PLP) gene encodes for two proteins, PLP and DM-20, which are produced by alternative splicing of exon 3B. PLP is the major CNS myelin protein and is postulated to play a structural role at the intraperiod line. Its developmental expression mirrors that of CNS myelination. DM-20 predominates in the embryo and prior to myelination of the CNS and may be involved in glial cell development. The PLP gene is expressed in the PNS in which DM-20 is the predominant isoform at all ages. In this study we describe the localization of the two isoforms in the post-natal rodent PNS using immunostaining and reverse transcriptase PCR. DM-20 is present in relatively high abundance in non-myelin-forming Schwann cells and within cytoplasmic regions of myelinated internodes, particularly the paranodes and Schmidt-Lanterman incisures and also the outer Schwann cytoplasm and perinuclear cytoplasm. DM-20 is also located in the perineuronal satellite cells of spinal, cranial and autonomic ganglia and in the ensheathing cells of the olfactory nerve layer of the olfactory bulb. PLP was detected by immunocytochemistry in the perinuclear region of myelinated internodes; PCR analysis indicated small amounts of PLP mRNA in the other locations but protein was not detected by immunostaining. Neither protein was identified in compact myelin of the PNS. DM-20 is the predominant product of the PLP gene expressed in a wide variety of peripheral glia. Its presence is not correlated to a myelin-forming state. Other studies have demonstrated early embryonic expression of the PLP gene throughout the PNS and all these features support the hypothesis that any putative role for DM-20 is unrelated to myelination but may involve glial cell development.

Animals↗
PubMed1995

Transgenic and natural mouse models of proteolipid protein (PLP)-related dysmyelination and demyelination.

The X chromosome-linked PLP/DM-20 gene is the CNS myelin gene most frequently associated with mutations, resulting in dysmyelination in several species including man (Pelizaeus-Merzbacher disease, X-linked Spastic Paraplegia). The pathology of most PLP gene mutations is characterized by hypomyelination, glial cell proliferation, increased numbers of microglia, and premature oligodendrocyte death. In most mutants, residual myelin structures have an abnormal ultrastructure and periodicity. Surprisingly, transgenic mice which carry extra copies of the wild type PLP gene show dysmyelination, demonstrating that the PLP gene is dosage sensitive. Pathological changes of transgenic mice vary from the phenotype of natural mutants. Specifically, many Golgi saccules of oligodendrocytes are vacuolated and the cytoplasm contains autophagic vacuoles hinting at a perturbation in protein trafficking. In fact, upon transgenic overexpression PLP becomes a prominent peripheral myelin protein, whereas in normal Schwann cells PLP is restricted from entering the myelin compartment. Surprisingly, transgenic animals which overexpress PLP/DM-20 at a low level appear normal during early development, but later spontaneously demyelinate. The mechanisms underlying this demyelination phenotype is unknown but an immune-mediated process has been suggested. All attempts to correct the phenotype of natural PLP mutants, such as jimpy mice, with a wild type transgene have had little effects, indicating a dominant-negative effect of the mutant gene product. On the other hand, mice with a targeted disruption of the PLP/DM-20 gene have surprisingly minor clinical signs. This suggests that the lethal phenotype associated with the majority of PLP gene mutations is a complex combination of loss and gain-of-function effects of a mutant myelin protein.

Animals↗
PubMed1994

Evidence that the agent of equine grass sickness may reach neurons by retrograde axonal transport.

Sera from acute and chronic cases of natural grass sickness or normal horses were injected into the parotid salivary gland of ponies. This gland receives its sympathetic innervation from the ipsilateral cranial cervical ganglion. None of the ponies showed any local or systemic signs of illness. After one week the cranial cervical ganglia, stellate and coeliaco-mesenteric ganglia were removed for histological study. Pathological changes were found only in the cranial cervical ganglion ipsilateral to a parotid salivary gland which had received an injection of grass sickness serum. Four out of five batches of test sera from cases of acute natural grass sickness were associated with chromatolytic changes in neurons; the remaining batch of serum produced no abnormalities. The most severe chromatolytic changes were induced by two samples obtained from horses whose signs of grass sickness had been present for less than 12 hours. A serum sample from a chronic case of grass sickness of three weeks duration did not produce chromatolysis but was associated with a moderately severe inflammatory infiltrate and neuronophagia in the ipsilateral cranial cervical ganglion. One batch of serum was size fractionated to separate components with molecular weights above or below 30 kDa. Only the fraction containing components above 30 kDa induced chromatolytic changes.

Acute Disease↗
PubMed1993

Leporine dysautonomia: further evidence that hares suffer from grass sickness.

The recently described dysautonomia of hares has many similarities to equine grass sickness, particularly when the autonomic ganglia of affected hares and horses are compared by light microscopy. This study shows that the ultrastructural findings are also similar, with a loss of ribosomes from the rough endoplasmic reticulum and distension of its cisternae; the Golgi apparatus is not recognisable in affected neurons. Membranous stacks were identified in autonomic neurons of affected hares, a feature not characteristic of equine grass sickness but often found in feline dysautonomia. Staining with wheat germ agglutinin, a lectin recognising Golgi membranes, showed a lack of reactivity in affected neurons again suggesting a lack of a normal Golgi apparatus.

Animals↗
PubMed1993

Oligodendrocyte development and differentiation in the rumpshaker mutation.

The jimpy rumpshaker (jprsh) mutation is an amino acid substitution in exon 4 (Ile186-->Thr) of the proteolipid protein (PLP) gene on the X chromosome. Affected mice show moderate hypomyelination of the central nervous system (CNS) with increased numbers of oligodendrocytes in the white matter of the spinal cord, a feature distinguishing them from other PLP mutations such as jp, in which premature cell death occurs with reduced numbers of oligodendrocytes. Myelin sheaths of jprsh immunostain for myelin basic protein (MBP) and DM-20, but very few contain PLP. This study examines the differentiation of oligodendrocytes cultured from the spinal cords of young mutant and wild type mice using various surface and cytoplasmic antigenic markers to define the stage of development. The majority of oligodendrocytes from mutant mice progress normally to express MBP; approximately 30%, relative to wild type, contain DM-20 at the in vivo age of 16 days, but very few immunostain for PLP or the O10 and O11 markers. The morphology of mutant cells in respect to membrane sheets and processes appears similar to normal. The jprsh oligodendrocyte is, therefore, characterized by a failure to express the markers indicative of the most mature cell; however, it is probably able to achieve a normal period of survival. These data, taken in conjunction with previous results, suggest that the PLP gene has at least two functions; one, probably involving PLP, is concerned with a structural role in normal myelin compaction; the other, perhaps related to DM-20 (or another lower molecular weight proteolipid), is essential for cell survival. The mutation in jprsh at residue 186 suggests that this region, which is common to PLP and DM-20, is not critical for this latter function.

Animals↗
PubMed1993

In vitro studies of axonally-regulated Schwann cell genes during Wallerian degeneration.

Wallerian degeneration in vivo is associated with marked downregulation of myelin protein genes such as P(o) and upregulation of other genes such as nerve growth factor receptor (NGF-R), glial fibrillary acidic protein (GFAP) and neural cell adhesion molecule (N-CAM). This study examines the expression of these genes during Wallerian degeneration in vitro and how manipulating Ca2+ affects this response. Small explants of sciatic nerve from normal young adult rats cultured for five days show similar reversal of the myelinating phenotype as found in vivo. If Ca++ is removed from the culture medium through the addition of EGTA, expression of the nerve growth factor receptor and glial fibrillary acidic protein genes is inhibited but downregulation of the P(o) gene still occurs. Explants cultured in medium containing EGTA are still capable of expressing nerve growth factor receptor if the medium is replaced by one containing Ca2+. Supplementation of normal medium with drugs modulating Ca2+, such as Bepridil which blocks the Na+Ca2+ exchanger or compound 48/80 which inhibits calmodulin, also prevent the expression of the nerve growth factor receptor gene during Wallerian degeneration in vitro. Treatment of the cervical sympathetic trunk with Bepridil leads to loss of the nerve growth factor receptor immunoreactivity which is normally present. The results indicate that Ca2+ may play a role in the expression of the nerve growth factor receptor gene during Wallerian degeneration and provide some indication that this effect may be directly on the Schwann cell rather than operating indirectly via the axon.

Animals↗
PubMed1993

Neurones in autonomic ganglia of normal horses contain phosphorylated neurofilaments.

Neurofilaments (NF) are composed of three polypeptides of differing molecular size, termed NF-L, NF-M and NF-H. The NF-H and, to a lesser degree, NF-M components are phosphorylated. In the majority of normal neurones, the location of phosphorylated NF is confined to neuronal processes, particularly the axon, and excluded from the perikaryon. Cell bodies of autonomic neurones of the rat do not contain phosphorylated NF. In many disease states, phosphorylated NF accumulate in the neuronal cell body and therefore in most circumstances their presence indicates abnormality. This paper reports that in at least two autonomic ganglia of normal horses (stellate and coeliomesenteric) the vast majority of neuronal perikarya immunostain strongly for phosphorylated NF. Pretreatment with alkaline phosphatase abolishes staining.

Animals↗
PubMed1993

Immunocytochemical and lectin histochemical study of neuronal lesions in autonomic ganglia of horses with grass sickness.

Equine grass sickness (EGS) is a primary dysautonomia characterised pathologically by lesions in autonomic ganglia, enteric plexi and specific nuclei in the CNS. Immunocytochemistry and lectin histochemistry of the autonomic ganglia were used to determine whether abnormalities can be detected in specific proteins or cellular organelles. EGS ganglia contained a mixture of morphologically normal and abnormal neurons, the former appearing identical to cells from control animals. Affected cells showed marked disturbances in neurofilament (NF) proteins and beta-tubulin, major components of the cytoskeleton; in most neurons immunoreactivity was reduced or absent while the distribution was altered in the remainder. Staining for neuron-specific enolase, a pan-neuronal marker, was severely reduced or absent, as was reactivity for the catecholaminergic enzyme tyrosine hydroxylase. However, affected neurons showed a marked increase in dopamine-beta-hydroxylase (D beta H), another enzyme associated with noradrenaline synthesis. Wheat germ agglutinin and Griffonia simplicifolia B4 lectin histochemistry was used to label membranes of the Golgi apparatus, which stained as discrete curvilinear perinuclear profiles. All affected neurons showed abnormalities with either complete loss of reaction or amorphous centrally located lectin staining. The results indicate perturbation in a wide variety of cytoplasmic and cytoskeletal proteins. In the majority of instances there is a decrease in stainable protein; the increase in D beta H may indicate a failure to be transported down the axon with resultant accumulation in the perikaryon. Loss of a recognisable Golgi structure appears to be an early event in the neuropathology of EGS.

Animals↗
PubMed1992

Rumpshaker: an X-linked mutation causing hypomyelination: developmental differences in myelination and glial cells between the optic nerve and spinal cord.

The X-linked mutation rumpshaker (rsh), which is probably an allele of jimpy (jp), causes hypomyelination in the CNS of mice. This study examines the developmental expression of the morphology, glial cells, and immunostaining of myelin proteins in the optic nerve and spinal cord. The optic nerve contains varying numbers of amyelinated and myelinated fibres. The majority of such sheaths are of normal thickness whereas in the spinal cord most axons are associated with a disproportionately thin sheath which changes little in thickness during development. In the optic nerve glial cell numbers are elevated in mutants during early and peak myelination but then fall slightly below normal in adults. In contrast, the number of glial cells is consistently elevated after 16 days of age in the spinal cord. The majority of the alterations to total glial cells are due to corresponding changes in the oligodendrocyte population. Immunostaining intensity is somewhat reduced for myelin basic protein (MBP) and the C-terminal common to proteolipid protein (PLP) and DM-20 and profoundly decreased for the PLP-specific peptide. Glial fibrillary acidic protein (GFAP) is increased in rsh. It is probable that some of the variation in myelination between optic nerve and cord in rsh is related to the difference in axon diameter in the two locations, as there are adequate numbers of oligodendrocytes at the time of myelination. However, the effect of the mutation on cell development in the brain and the spinal cord may be different. The immunostaining indicates a marked deficiency in PLP in myelin but suggests that DM-20 levels may be relatively normal. rsh shows several major differences from jp and other X-linked myelin mutants, particularly in relation to oligodendrocyte numbers, and will be useful to elucidate the role of the PLP gene in influencing oligodendrocyte differentiation and survival.

Animals↗
PubMed1992

Developmental expression of major myelin protein genes in the CNS of X-linked hypomyelinating mutant rumpshaker.

Rumpshaker (rsh) is an X-linked mutation causing hypomyelination of the CNS of mice and has recently been identified as an allele of jimpy (jp). The mutation (known as jprsh) differs in several respects from other X-linked myelin mutants, including jp, in that mice have normal longevity, oligodendrocyte numbers are not decreased, and cell death is not a feature. Myelin sheaths are deficient in immunostainable PLP protein. The present study examines the developmental expression of the major myelin protein genes and translatability of PLP and MBP mRNA. Differences between the spinal cord and brain of mutants are evident in that mRNA levels are more markedly decreased in the brain. Protein levels are severely reduced in both locations and to a proportionately greater extent than the mRNA, particularly in the spinal cord where PLP RNA and protein are approximately 80% and 10-20%, respectively, of age-matched wild type mice. DM-20 protein, the other major product of the PLP gene, is disproportionately expressed in rumpshaker as is a 10 kDa proteolipid. In vitro translation studies indicate a marked decrease in PLP translation products from mutant RNA. There is no deficiency in the number of PLP mRNA-expressing oligodendrocytes although the abundance per cell is reduced. The data suggest that the phenotypic effects of the mutation may be associated with reduced translation of major myelin proteins, in particular PLP and its incorporation into compact myelin. However, the mutation is compatible with survival of oligodendrocytes and their differentiation to the stage of expressing PLP/DM-20 mRNA.

Aging↗
PubMed1992

A review of the primary dysautonomias of domestic animals.

Primary dysautonomias appear to be the result of initial damage to the protein synthetic pathway of a specific neuronal population, but despite detailed morphological study of several species there is, as yet, no indication of the precise lesion or the nature of the causal agent. The very marked similarities between the species with regard to lesion type, distribution, the age group affected and the geographical restrictions of occurrence would suggest a very similar, if not common, aetiology. There is no explanation, however, for the 70 year gap between its appearance in horses and its subsequent occurrence in other species or why it is these species, with very different physiology, habits and habitats, that are affected. No reference could be found in the literature to any infectious agent or toxin causing a similar range of structural effects with a similar species specificity or lesion distribution. Many questions about dysautonomias remain. Why is the lesion distribution so specific? At what level of the synthetic pathway does the primary lesion occur? What are the unusual compounds demonstrated in "acute phase" serum from affected horses; are they a neurotoxic agent(s) and/or its metabolites, or the abnormal product of an affected animal? Why did the experimental ponies which developed autonomic lesions not become ill? When do the clinical signs appear in relation to the occurrence of the primary lesion? Why are adolescent and young adult animals most commonly affected? As the general understanding of neuronal function and the numerous factors which influence it improves, the many subtle distinctions and similarities amongst the myriad sub-populations of neurones will become clearer and common features may emerge which will link the seemingly disparate neuronal types involved in the primary dysautonomias.

Animals↗
PubMed1992

Primary demyelination induced by exposure to tellurium alters Schwann cell gene expression: a model for intracellular targeting of NGF receptor.

Exposure of developing rats to tellurium results in a highly synchronous segmental demyelination of peripheral nerves with sparing of axons; this demyelination is followed closely by a period of rapid remyelination. Demyelination occurs subsequent to a tellurium-induced block in the synthesis of cholesterol, the major myelin lipid. We utilized the techniques of Northern blotting, in situ hybridization, and immunocytochemistry to examine temporal alterations in Schwann cell gene expression related to demyelination and remyelination. Tellurium-induced demyelination is associated with downregulation of myelin protein expression and a corresponding upregulation of NGF receptor (NGF-R) and glial fibrillary acidic protein (GFAP) expression. Steady-state mRNA levels (expressed on a "per nerve" basis) for P0, the major myelin protein, were decreased by about 50% after 5 d of tellurium exposure, while levels of mRNA for NGF-R and GFAP were markedly increased (about 15-fold). In situ hybridization of teased fibers suggested that the increase in steady-state mRNA levels for NGF-R was primarily associated with demyelinated internodes and not with adjacent unaffected internodes. Although P0 message was almost totally absent from demyelinating internodes, it was also reduced in normal-appearing internodes as well. This suggests that limiting the supply of a required membrane component (cholesterol) may lead to partial downregulation of myelin gene expression in all myelinating Schwann cells. In partially demyelinated internodes, NGF-R and GFAP immunofluorescence appeared largely confined to the demyelinated regions. This suggests specific targeting of these proteins to local areas of the Schwann cell where there is myelin loss. These results demonstrate that demyelination is associated with reversion of the affected Schwann cells to a precursor cell phenotype. Because axons remain intact, our results suggest that these changes in Schwann cell gene expression do not require input from a degenerating axon, but instead may depend on whether concerted synthesis of myelin is occurring.

Animals↗