Biomedical subjects

I D Duncan

Publications and source records attributed to I D Duncan.

At least 37 records · Page 2Linked to original sources

PubMed1999

Adult brain retains the potential to generate oligodendroglial progenitors with extensive myelination capacity.

Remyelination of focal areas of the central nervous system (CNS) in animals can be achieved by transplantation of glial cells, yet the source of these cells in humans to similarly treat myelin disorders is limited at present to fetal tissue. Multipotent precursor cells are present in the CNS of adult as well as embryonic and neonatal animals and can differentiate into lineage-restricted progenitors such as oligodendroglial progenitors (OPs). The OPs present in adults have a different phenotype from those seen in earlier life, and their potential role in CNS repair remains unknown. To gain insights into the potential to manipulate the myelinating capacity of these precursor and/or progenitor cells, we generated a homogenous culture of OPs from neural precursor cells isolated from adult rat subependymal tissues. Phenotypic characterization indicated that these OPs resembled neonatal rather than adult OPs and produced robust myelin after transplantation. The ability to generate such cells from the adult brain therefore opens an avenue to explore the potential of these cells for repairing myelin disorders in adulthood.

Aging↗

PubMed1999

Microtubule alterations in cultured taiep rat oligodendrocytes lead to deficits in myelin membrane formation.

The taiep rat is a myelin mutant in which hypomyelination and progressive demyelination of the CNS are accompanied by an accumulation of microtubules within oligodendrocytes. To investigate whether and how the myelin defects were caused by microtubule abnormalities, we have established a taiep oligodendrocyte culture system in which mutant cells produce abnormally high levels of tubulin and microtubule-associated proteins and exhibit myelin defects. The studies show that abnormal microtubule accumulation and tight microtubule bundles developed in the taiep oligodendrocytes, with a higher ratio of minus-end-distal to plus-end-distal microtubules in their processes. Initially, in culture, immature taiep oligodendrocytes which have higher levels of tubulin than controls extend roughly twice as much membrane sheet as controls. The membrane sheets of the mature taiep oligodendrocytes which display the microtubule accumulation, however, grew much less rapidly compared to controls. By the fifth day in culture, a majority of the taiep oligodendrocytes had ceased the expansion of their membrane sheets and in some cases the sheets retracted. The levels of the myelin proteins, proteolipid protein and myelin-associated glycoprotein, were also markedly diminished in the mature taiep oligodendrocytes. Treatment with the microtubule depolymerizing drug nocodazole prevented not only the accumulation of microtubules but also restored the normal distribution of proteolipid proteins within the taiep oligodendrocytes. These data demonstrate that myelin synthesis in the oligodendrocyte cultures relies on the formation of a normal microtubule array, and the microtubule abnormalities are directly responsible for the myelin deficit in the taiep oligodendrocytes.

Animals↗

PubMed1999

Should women with genital condyloma acuminata have routine diagnostic colposcopy in addition to cervical smear screening?

We aimed to evaluate the difference in the pick up rate of high grade cervical abnormalities using both primary colposcopy and cervical smear compared with cervical cytology screen only on women with genital warts. We also wished to establish whether this group was at high risk of sexually transmittable infections (STIs) and whether there is a correlation between those with high grade CIN and other STIs. This was a retrospective analysis based in the Department of Genito-Urinary Medicine, Dundee Teaching Hospitals Trust, Dundee, UK. We studied all women (n =117) attending the Department with genital warts between January 1995 and March 1997 who were screened by both cervical cytology and colposcopy High grade cervical abnormalities (CIN II-CIN III) were diagnosed in 10 patients (8.5.%) using cervical screening only and in 41 patients (35%) (CIN II-III) when primary colposcopy and histopathology were undertaken. Of 117 women screened for sexual transmittable infection there were 53 with a positive diagnosis (48%). Of the 53 positive there were 17 women (30%) with high grade CIN. We conclude that women with genital warts are a high risk group, both in terms of having high grade cervical abnormalities and sexually transmittable infection. The result suggests that, when practicable primary colposcopy should be performed as a routine screening tool, on this group of patients over and above cervical cytology screening. All patients with genital warts should be referred to genitourinary medicine for STI screening.

Journal Article↗

PubMed1999

Intraventricular transplantation of oligodendrocyte progenitors into a fetal myelin mutant results in widespread formation of myelin.

Transplantation of myelin-forming cells is a promising strategy for the treatment of myelin disorders. In this study, transplantation of glial cell progenitors into the cerebral ventricles of the embryonic myelin-deficient rat, a model of Pelizaeus-Merzbacher disease, was performed to assess the ability of these cells to incorporate into the developing brain and produce myelin. The donor cells migrated into the white and gray matter and produced myelin at widespread sites ranging from the corpus callosum and optic nerve to the cerebellum. These data suggest that myelin repair might be achieved by intraventricular delivery and transependymal incorporation of myelin-producing cells. Because these cells were genetically transduced to express a reporter gene, similar ex vivo manipulation with genes known to promote survival, migration, or proliferation of the transplanted cells could be used to enhance repair. Such a therapeutic strategy may be feasible in patients with inherited myelin disorders or in multiple sclerosis, particularly where the lesions are periventricular.

Animals↗

PubMed1998

Self-renewing canine oligodendroglial progenitor expanded as oligospheres.

We have previously shown that oligodendroglial progenitors (OP) can be generated from multipotent rat neural precursor cells. We now report the generation of a homogeneous culture of canine OP from neural precursor cells. In non-adherent cultures, homogeneous OP cultures were obtained in 6-8 weeks of treatment with B104 cell conditioned medium (B104CM). In adherent cultures where astrocytes grew as a layer of substrate, colonies of OP invariably appeared at 10-14 days in vitro (DIV) and the colonies were expanded as free-floating spheres (oligospheres), in the presence of B104CM, suggesting that astrocytes facilitate the generation of canine OP. The oligosphere cells were characterized by self-renewal in the presence of B104CM and by terminal differentiation into oligodendrocytes after withdrawal of B104CM. Transplantation studies indicated that the extensively expanded oligosphere cells retained myelination capacity. The oligospheres thus provide a valuable source for experimental cell therapy studies.

Animals↗

PubMed1998

Oligodendrocytes are not inherently programmed to myelinate a specific size of axon.

Current studies support the morphological classification of oligodendrocytes proposed by Del Rio Hortega ([1922] Bol. R. Soc. Esp. Hist. Nat. 10:25-29; [1924] C.R. Soc. Biol. 91:818-820), in which cells either myelinate multiple internodes that are associated with small axons, or they myelinate restricted/single internodes of large-diameter axons. The reasons why an oligodendrocyte myelinates a particular calibre of axon are unknown. Because progenitors are generated in restricted, subventricular zones, an intrinsic program would imply that germinal centres contain a mixture of cells, each committed to myelinate axons of a particular size. Conversely, each cell could have the potential ability to myelinate any size axon. We tested this latter hypothesis that oligodendrocyte progenitors are uncommitted in their ability to myelinate a particular axon size. We introduced oligodendrocyte lineage cells from the optic nerve, which normally encounter only small-diameter axons, to a myelin-deficient environment containing a large range of axon sizes. Dissociated, mixed glial cells from the optic nerve were characterised immunocytochemically and were grafted into the spinal cord ventral column of neonatal, myelin-deficient rat mutants. Examination of the patches of myelin produced by these cells at different times after transplantation revealed that optic nerve oligodendrocytes were capable of producing a widespread, nonselective myelination of axons that were destined to have both small or large calibres. Thus, an axonal or local signal, and not an intrinsic program, is probably responsible for the previously described oligodendrocyte diversity.

Animals↗

PubMed1998

The immune status of the myelin deficient rat and its immune responses to transplanted allogeneic glial cells.

This study examined the immunological responsiveness of the myelin deficient (md) rat, and its immune response to transplanted allogeneic glial cells, with and without immunosuppression therapy. Skin grafts from an ACI strain of rat were found to be acutely rejected by Wistar md rats. Anti-donor cytotoxic antibody was produced and alloreactive T helper cells were expanded in these mutants after skin sensitization. Equivalent high frequencies of precursor cytotoxic T lymphocytes (pCTLs) specific to the ACi MHC antigens were observed in both normal controls and mutants. An immune response was noted when allogeneic glial cells were transplanted into the spinal cords of md rats, which was effectively suppressed for 2 weeks post transplantation by treatment with either cyclosporin A (CsA) or a monoclonal antibody to the interleukin-2 receptor (IL-2R). These results demonstrate that md rats are immunocompetent, but that CNS allograft rejection can be prevented by immunosuppressive agents.

Animals↗

PubMed1998

Apoptotic glial cell death and kinetics in the spinal cord of the myelin-deficient rat.

This study examined the glial cell kinetics and death in the thoracic spinal cord of normal and myelin-deficient (md) rats between 1 and 21 days of age and determined whether the observed glial cell death primarily affected oligodendrocytes and had the morphologic and molecular features of apoptosis. In the md rat spinal cord there was an increase in cell division and death in a pattern that correlated with the onset of myelination. The dying cells were identified as oligodendrocytes ultrastructurally as many had the characteristic distention of the rough endoplasmic reticulum seen in the md rat glia. Double labeling using PLP in situ hybridization and a modified TUNEL method also suggested that the dying cells, in both mutant rats and control littermates, were oligodendrocytes. These findings were compared with previous studies on the md rat optic nerve and those in other PLP mutants.

Animals↗

PubMed1998

Myelin mosaicism and brain plasticity in heterozygous females of a canine X-linked trait.

The shaking (sh) pup, an animal model of Pelizaeus-Merzbacher disease, is characterized by severe central nervous system dysmyelination in affected males, and myelin mosaicism in some female heterozygotes as a result of X-linked inactivation. Heterozygous females develop a tremor of varying severity that usually disappears at 4 to 6 weeks, whereas male hemizygotes have severe, generalized tremor that persists throughout life. We have used these two myelin-deficient models to study the potential for recovery with time as reflected by brainstem auditory evoked responses (BAERs). At set time points, the state of myelination in the trapezoid body was studied microscopically. Sequential BAERs demonstrated consistently prolonged interpeak latencies during the period of gross tremor in heterozygotes, with the trend continuing to a lesser extent after tremor cessation. The random nature of X-linked inactivation resulted in variable myelin mosaicism that was reflected in variations in BAER changes within animals in the same litter. In most heterozygotes, the tremor resolved with time, the BAERs returned to near normal, and myelin mosaicism was lost. In contrast, in the affected males, the severity of tremor and lack of recovery was demonstrated by consistent abnormalities in BAER waves at all times studied, and severe and persistent myelin deficiency in the trapezoid body. These findings show that despite the normal tightly programmed temporal development of myelin in the brain in the heterozygous mosaic state, sufficient plasticity persists during the neonatal period for late-stage myelination to occur.

Animals↗

PubMed1998

Morphological and morphometric studies of the dysmyelinating mutant, the Long Evans shaker rat.

The Long Evans shaker (les) rat is a recently identified CNS myelin mutant with an autosomal recessive mode of inheritance. Although scattered myelin sheaths are present in some areas of the CNS, most notably the ventral spinal cord in the young neonatal rat, this myelin is gradually lost, and 8-12 weeks little myelin is present throughout the CNS. Despite this severe myelin deficiency, some mutants may live beyond one year of age. Rare, thin myelin sheaths that are present early in development lack myelin basic protein (MBP) and on ultrastructural examination are poorly compacted and lack a major dense line. Many oligodendrocytes develop an accumulation of vesicles and membranous bodies, but no abnormal cell death is observed. In the optic nerve, cell kinetic studies show an increase in proliferation at early time points in les, while total glial cell counts are also increased in les from 2 months of age. In situ hybridization studies demonstrate that the numbers of mature oligodendrocytes are similar to controls early in life and increase with time compared to controls. There is both a progressive astrocyte hypertrophy and microgliosis. While les has a mutation in the myelin basic protein (mbp) gene, it is dissimilar in both genotype and phenotype to the previously described mbp mouse mutants, shiverer (shi) and shiverer(mld). Unlike shi and its allele, where myelin increases with time and oligodendrocytes become ultrastructurally normal, les oligodendrocytes are permanently disabled, continue to demonstrate cytoplasmic abnormalities, and fail to produce myelin beyond the first weeks of life.

Animals↗

PubMed1998

Generation of oligodendroglial progenitors from neural stem cells.

To understand how the differentiation of stem cells to oligodendroglial progenitors is regulated, we established cultures of neural stem cells from neonatal rat striatum in the presence of epidermal growth factor (EGF) as free-floating neurospheres that were then exposed to an increasing amount of B104 cell-conditioned medium (B104CM). The resultant cells proliferated in response to B104CM but no longer to EGF. In vitro analysis and transplantation studies indicated that these cells were committed to the oligodendroglial lineage, and they were thus referred to as oligospheres. Further characterization of their expression of early markers, cell cycle, migration, and self-renewal suggests that they were pre-O2A progenitors. RT-PCR analysis indicated that the oligosphere cells expressed mRNAs of platelet-derived growth factor alpha receptor in addition to fibroblast growth factor receptor but not EGF receptor; the latter two receptor mRNAs were expressed by neurosphere cells. Thus, the progression of stem cells to oligodendroglial progenitors is likely induced by factors in B104CM.

Animals↗

PubMed1998

The value of cervical screening in women over 50 years of age--time for a multicentre audit.

This study reviews the cervical smear history of women developing CIN aged over 50 years to consider if they might be discharged sooner from the cervical screening programme in Tayside Region. From the OCCURS database all women over 50 years who developed CIN between 1 Jan 1993 and 30 June 1996 were identified and their smear history obtained. Results show that had women been discharged from the screening programme at age 50 following three consecutive negative smears and a negative exit smear then only two women with CIN 3 and one with microinvasive disease would have been missed in the subsequent three and a half years. A wider geographical survey of the incidence of CIN in this older age group is needed to determine whether it is cost beneficial and cost effective to continue cervical screening beyond the age of 50 years.

Adult↗

PubMed1997

Isolation and transplantation of multipotential populations of epidermal growth factor-responsive, neural progenitor cells from the canine brain.

Glial cell transplantation into myelin-deficient rodent models has resulted in myelination of axons and restoration of conduction velocity. The shaking (sh) pup canine myelin mutant is a useful model in which to test the ability to repair human myelin diseases, but as in humans, the canine donor supply for allografting is limited. A solution may be provided by self-renewing epidermal growth factor (EGF)-responsive multipotential neural progenitor cell populations ("neurospheres"). Nonadherent spherical clusters, similar in appearance to murine neurospheres, have been obtained from the brain of perinatal wildtype (wt) canine brain and expanded in vitro in the presence of EGF for at least 6 months. Most of the cells in these clusters express a nestin-related protein. Within 1-2 weeks after removal of EGF, cells from the clusters generate neurons, astrocytes, and both oligodendroglial progenitors and oligodendrocytes. Transplantation of lacZ-expressing wt neurospheres into the myelin-deficient (md) rat showed that a proportion of the cells differentiated into oligodendrocytes and produced myelin. In addition, cells from the neurosphere populations survived at least 6 weeks after grafting into a 14-day postnatal sh pup recipient and at least 2 weeks after grafting into an adult sh pup recipient. Thus, neurospheres provide a new source of allogeneic donor cells for transplantation studies in this mutant.

Animals↗

PubMed1997

Microtubule organization and stability in the oligodendrocyte.

The oligodendrocyte is the glial cell responsible for the formation and maintenance of CNS myelin. Because the development of neuronal morphology is known to depend on the presence of highly organized microtubule arrays, it may be hypothesized that the properties of microtubules influence the form and function of oligodendrocytes. The goals of the present study were to define the physical attributes of microtubules in oligodendrocytes maintained in vitro. The results of electron and confocal microscopy indicate that microtubules are present throughout the cell bodies and large and small processes of oligodendrocytes and are rarely associated with discrete microtubule-organizing centers. A modified "hooking" protocol demonstrated that the polarity orientation of microtubules is uniformly plus-end distal in small oligodendrocyte processes, compared with a nonuniform, predominantly plus-end distal orientation in large processes. Oligodendrocytes were exposed to the microtubule-depolymerizing drug nocodazole to examine microtubule stability in these cells. The results suggest that oligodendrocyte microtubules can be resolved into at least three distinct microtubule populations that differ in their kinetics of depolymerization in the presence of nocodazole. These findings suggest that the properties of the oligodendrocyte microtubule array reflect the functions of the different regions of this highly specialized cell.

Animals↗

PubMed1997

Induction of sodium channel clustering by oligodendrocytes.

As oligodendrocytes wrap axons of the central nervous system (CNS) with insulating myelin sheaths, sodium channels that are initially continuously distributed along axons become segregated into regularly spaced gaps in the myelin called nodes of Ranvier. It is not known whether the regular spacing of nodes results from regularly spaced glial contacts or is instead intrinsically specified by the axonal cytoskeleton. Contact with Schwann cells induces clustering of sodium channels along the axons of peripheral neurons in vitro and in vivo. Similarly, it has been suggested that astrocyte contact induces clustering of sodium channels along CNS axons. Here we show that oligodendrocytes are necessary for clustering of sodium channels in vitro and in vivo. The induction, but not the maintenance, of sodium-channel clustering along the axons of highly purified rat retinal ganglion cells in culture depends on a protein secreted by oligodendrocytes. Surprisingly, the oligodendrocyte-induced clusters are regularly spaced at the predicted interval in the absence of glial-axonal contact. Mutant rats that are deficient in oligodendrocytes develop few axonal sodium channel clusters in vivo. These results demonstrate a crucial role for oligodendrocytes in inducing clustering of sodium channels.

Animals↗

PubMed1997

Invasive cervical cancer after conservative therapy for cervical intraepithelial neoplasia.

BACKGROUND: Conservative outpatient therapy for cervical intraepithelial neoplasia (CIN) by ablative or excisional techniques is widely used. The main objective of this treatment is the prevention of invasive cervical cancer. We assessed the rate of invasive disease and the duration of the risk of developing invasive cervical cancer after such treatment. METHODS: Four UK centres have used life-table methods to analyse the long-term results of conservative treatment of CIN. We combined and updated data from these studies to investigate the rate of invasive disease after treatment and the duration of that risk. FINDINGS: The data comprised 44 699 woman-years of follow-up, with 2116 women under observation 8 years after treatment. 33 women developed invasive cancer, 14 of whom had microinvasion. The cumulative rate of invasion 8 years after treatment was 5.8 per 1000 women and the rate of invasive cancer during this period was 85 (95% CI 60-119) per 100,000 woman-years. The risk of developing cancer did not change throughout the 8 years of follow-up. INTERPRETATION: These data show that conservative outpatient therapy in women with CIN reduces the risk of invasive cancer of the cervix by 95% during the first 8 years after treatment. However, even with careful, long-term follow-up, the risk of invasive cervical cancer among these women is about five times greater than that among the general population of women throughout that period. Careful follow-up is essential for at least 10 years after conservative treatment of CIN.

Adult↗

PubMed1997

Normal temporal and spatial distribution of oligodendrocyte progenitors in the myelin-deficient (md) rat.

A point mutation in exon 3 of the proteolipid protein (PLP) gene of the myelin-deficient (md) rat leads to a failure of oligodendrocyte maturation and early death of oligodendrocytes, resulting in dysmyelination. It has been suggested that an alternative-splice isoform of PLP, known as DM-20, might be expressed in oligodendrocyte progenitors in the embryonic central nervous system (CNS), raising the possibility that early development of the oligodendrocyte lineage might also be affected in the md rat. To test this suggestion, we visualized oligodendrocyte progenitors in the embryonic md rat spinal cord and brain by in situ hybridization with a probe to the platelet-derived growth factor alpha receptor (PDGFR). We could detect no abnormalities in the time of first appearance of oligodendrocyte precursors, nor in their subsequent proliferation and dispersal throughout the CNS. These data strongly suggest that the PLP mutation in the md rat primarily or exclusively affects the later stages of oligodendrocyte lineage.

Animals↗

PubMed1997

Myelination following transplantation of EGF-responsive neural stem cells into a myelin-deficient environment.

Epidermal growth factor (EGF)-responsive stem cells have been identified in the murine central nervous system. These cells can be isolated from the brain and maintained in an undifferentiated state in vitro in the presence of EGF. After removing EGF, the cells cease mitosis and can be induced to differentiate into neurons, astrocytes, and oligodendrocytes. We demonstrate that when the undifferentiated stem cells (nestin-positive) are injected into the myelin-deficient rat spinal cord, they respond to cues within the mutant CNS and differentiate into myelinating oligodendrocytes, in contrast to their behavior in vitro, where they mainly form astrocytes. The cells provide a valuable model system for the study of the development of early oligodendrocytes from multipotent neural stem cells. Because these cells are influenced to divide using growth factors, rather than oncogenes, and because they appear to make appropriate lineage decisions when transplanted into a mutant environment, they may provide an excellent source of cells for a variety of future therapies using cellular transplantation.

Animals↗