Biomedical subjects

Nicholas Marsh-Armstrong

Publications and source records attributed to Nicholas Marsh-Armstrong.

6 recordsLinked to original sources

PubMed2026

Cerebrospinal delivery of a bidirectional AAV9 vector improves optic nerve and retinal pathology in a sheep model of Tay-Sachs disease.

Tay-Sachs disease (TSD) is a fatal neurodegenerative lysosomal storage disease. The Jacob sheep is the only large-animal model of TSD, yet ocular pathology and the therapeutic potential of gene therapy remain poorly defined. Sheep cohorts included normal controls (n = 3); untreated TSD-affected (n = 4); intravenous AAV9-Bic_HexA/HexB-treated (n = 3); and intracerebroventricular, cisterna magna, and lumbar intrathecal AAV9- Bic_HexA/HexB-treated sheep (cerebrospinal fluid [CSF] therapy; n = 7). Retinal histopathology and immunohistochemistry, retinal whole-mount analyses for retinal ganglion cell (RGC) morphology and density, optic nerve evaluation with p-phenylenediamine (PPD )semi-thin sections, qPCR assessment for vector genomes, and RNAscope probes for transgene expression were performed. Untreated TSD sheep exhibited RGCs with abundant microvesicular cytoplasmic expansion and optic nerve spheroids, with storage material variably staining with periodic acid-Schiff. Marked astrocytosis, microgliosis, and GM2 accumulation within RGCs were present. Optic nerve axon counts and RGC density were significantly reduced, and optic nerve damage scores increased, in untreated and IV-treated sheep but were rescued with short-term CSF therapy. GM2 volume and signal intensity per RGC were significantly reduced following short-term CSF therapy. Minimal but detectable retinal vector genomes and transgene expression were observed. These findings demonstrate retinal and optic nerve pathology in Jacob sheep with TSD and AAV9 therapy.

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PubMed2006

Use of a ROSA26:GFP transgenic line for long-term Xenopus fate-mapping studies.

Widespread and persistent marker expression is a prerequisite for many transgenic applications, including chimeric transplantation studies. Although existing transgenic tools for the clawed frog, Xenopus laevis, offer a number of promoters that drive widespread expression during embryonic stages, obtaining transgene expression through metamorphosis and into differentiated adult tissues has been difficult to achieve with this species. Here we report the application of the murine ROSA26 promoter in Xenopus. GFP is expressed in every transgenic tissue and cell type examined at post-metamorphic stages. Furthermore, transgenic ROSA26:GFP frogs develop normally, with no apparent differences in growth or morphology relative to wild-type frogs. ROSA26 transgenes may be used as a reliable marker for embryonic fate-mapping of adult structures in Xenopus laevis. Utility of this transgenic line is illustrated by its use in a chimeric grafting study that demonstrates the derivation of the adult bony jaw from embryonic cranial neural crest.

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PubMed2005

Thyroid hormone controls multiple independent programs required for limb development in Xenopus laevis metamorphosis.

Thyroid hormone (TH) is required for limb development in Xenopus laevis. Specific cell types in the growing limb were targeted for expression of a dominant negative form of the TH receptor by sperm-mediated transgenesis. Limb muscle development, the innervation of muscle from the spinal cord, and cartilage growth can be inhibited without affecting patterning of the limb or differentiation of other cell types. Remodeling of the skin occurs late in metamorphosis after the limb has formed. The coordination of these independent programs is affected in part by the control that TH exerts over DNA replication in all cell types of the limb.

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PubMed2004

An outer segment localization signal at the C terminus of the photoreceptor-specific retinol dehydrogenase.

Photoreceptor retinol dehydrogenase (prRDH) is a membrane-associated cytosolic protein that localizes to the outer segments (OS) of rods and cones. Here, we demonstrate that the C-terminal 16 amino acids of prRDH confer membrane association as well as cone and rod OS targeting on a linked green fluorescent protein. Membrane association in transfected 293 cells and in transgenic Xenopus photoreceptors is mediated by fatty acylation at one or more evolutionarily conserved cysteines within the prRDH C-terminal tail. In bovine OS, native prRDH is similarly acylated, and hydrolysis of this linkage releases the protein from the membrane. Efficient OS localization requires both membrane association and the prRDH sequence. (V/I)XPX at the extreme C terminus, which closely resembles the C-terminal sequence that targets opsin/rhodopsin to the OS. Taken together, these data imply that the C-terminal. (V/I)XPX sequence is a general OS localization signal that can function in the context of both integral and peripheral membrane proteins. This strategy for OS localization resembles those used for protein localization to mitochondria, peroxisomes, endosomes, and endoplasmic reticulum; in each case, a short N- or C-terminal sequence is shared among structurally diverse proteins that are targeted to the same subcellular destination.

Alcohol Oxidoreductases↗

PubMed2003

Thyroid hormone controls the development of connections between the spinal cord and limbs during Xenopus laevis metamorphosis.

During premetamorphic stages, Xenopus laevis tadpoles expressing either a dominant-negative thyroid hormone (TH) receptor or a type-III iodothyronine deiodinase transgene in the nervous system have reduced TH-induced proliferation in the spinal cord and produce fewer hindlimb-innervating motorneurons. During prometamorphic stages, innervation of the hindlimbs is reduced, and few functional neuromuscular connections are formed. By metamorphic climax, limb movement is impaired, ranging from uncoordinated leg swimming to complete quadriplegia. This phenotype is due to transgene action in the tadpole spinal cord. The requirement of TH for neurogenesis during premetamorphosis is the earliest TH-regulated process reported to date in the sequence of metamorphic changes in anurans. The muscle formed during limb growth was previously shown to be a direct target of TH control. Here, we show that the same is true of the development of spinal cord cells that innervate the limbs.

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PubMed2003

Role of visual pigment properties in rod and cone phototransduction.

Retinal rods and cones share a phototransduction pathway involving cyclic GMP. Cones are typically 100 times less photosensitive than rods and their response kinetics are several times faster, but the underlying mechanisms remain largely unknown. Almost all proteins involved in phototransduction have distinct rod and cone variants. Differences in properties between rod and cone pigments have been described, such as a 10-fold shorter lifetime of the meta-II state (active conformation) of cone pigment and its higher rate of spontaneous isomerization, but their contributions to the functional differences between rods and cones remain speculative. We have addressed this question by expressing human or salamander red cone pigment in Xenopus rods, and human rod pigment in Xenopus cones. Here we show that rod and cone pigments when present in the same cell produce light responses with identical amplification and kinetics, thereby ruling out any difference in their signalling properties. However, red cone pigment isomerizes spontaneously 10,000 times more frequently than rod pigment. This high spontaneous activity adapts the native cones even in darkness, making them less sensitive and kinetically faster than rods. Nevertheless, additional factors are probably involved in these differences.

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