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S Gandy

Publications and source records attributed to S Gandy.

At least 19 recordsLinked to original sources
PubMed2004

Baculoviruses expressing the human familial Alzheimer's disease presenilin 1 mutation lacking exon 9 increase levels of an amyloid beta-like protein in Sf9 cells.

Presenilin 1 (PS1) plays a pivotal role in the production of the amyloid-beta protein (Abeta) that is central to the pathogenesis of Alzheimer's disease. PS1 regulates the intramembranous proteolysis of a 99-amino-acid C-terminal fragment of the amyloid precursor protein (APP-C99), a cleavage event that releases Abeta following a reaction catalyzed by an enzyme termed 'gamma-secretase'. The molecular mechanism of PS1-mediated, gamma-secretase cleavage remains largely unresolved. In particular, controversy surrounds whether PS1 includes the catalytic site of the gamma-secretase protease or whether instead PS1 mediates gamma-secretase activity indirectly, perhaps by regulating the trafficking or presentation of substrates to the 'authentic' protease, which may be a molecule distinct from PS1. To address this issue, the baculovirus expression system was used to co-express: (i) APP-C99; (ii) a pathogenic, constitutively active mutant form of PS1 lacking exon 9 (PS1DeltaE9); (iii) nicastrin and (iv) tropomyosin in Spodoptera frugiperda (Sf9) cells. Cells infected with APP-C99 alone produced an Abeta-like species, and levels of this species were enhanced by the addition of baculoviruses bearing the PS1DeltaE9 mutation. The addition to APP-C99-infected cells of baculoviruses bearing nicastrin, also a transmembrane protein, had a neutral or inhibitory effect on the reaction; tropomyosin viruses had the same effect as nicastrin viruses. These results suggest that PS1DeltaE9 molecules expressed in Sf9 cells retain the ability to modulate Abeta levels. Baculoviral-expressed PS1DeltaE9 provides a source of microgram quantities of bioactive molecules for use as starting material for purifying and reconstituting gamma-secretase activity from its individual purified component parts.

Alzheimer Disease↗
PubMed2003

The effects of GnRH administration postinsemination on serum concentrations of progesterone and pregnancy rates in dairy cattle exposed to mild summer heat stress.

The objective of this study was to evaluate whether administration of GnRH postinsemination would improve reproductive performance in heat-stressed dairy cattle. Estrous cycles of Holstein cows were synchronized using the OvSynch protocol and cows were artificially inseminated. Cows were then administered the following treatments: control (no GnRH; n=37), GnRH (100 microg) on Day 5 (GnRH-D5; n=34), or GnRH (100 microg) on Day 11 (GnRH-D11; n=34) postinsemination. Cows were provided access to both fans and sprinklers, and environmental data was collected hourly. Rectal temperatures and blood samples were obtained from cows on Days -9, -2, 0 (AI) and on alternate days from Day 5 to Day 19 postinsemination. Blood serum was collected for the analysis of progesterone (P(4)) by RIA. In a subset of cows (n=6/treatment) ultrasonography was performed on alternate days from Day 5 to Day 19 postinsemination to assess numbers of corpora lutea (CL) and CL cross-sectional areas. Pregnancy status of cows was confirmed at Day 30 postinsemination. Environmental data indicated that cows experienced mild heat stress during the trials (mean daily THI=73-77). Serum P(4) was greater (P<0.05) after Day 9 for GnRH-D5 cows and after Day 15 for GnRH-D11 cows through Day 19 postinsemination. The number of CLs present for GnRH-D5 cows was greater (P<0.05) on Day 17 than in either the control or GnRH-D11 treatment groups. On Day 17 postinsemination, both the GnRH-D5 and GnRH-D11 cows were observed to have greater (P<0.05) total CL tissue area than control cows. The interval from insemination to when serum P(4) returned to <1 ng/ml (i.e. luteolysis and return to estrus) did not differ (P>0.10) among treatment groups. Control cows (19%) tended to exhibit lower pregnancy rates (P<0.08) compared to the GnRH-D5 and GnRH-D11 treatment groups combined (35%). In summary, the treatment of heat-stressed dairy cows with GnRH postinsemination (Day 5 or 11) results in the appearance of more CL tissue, increased serum concentrations of P(4) and a tendency toward greater pregnancy rates.

Animals↗
PubMed2001

Alzheimer's disease. Molecular consequences of presenilin-1 mutation.

Alzheimer's disease is characterized by accumulation in the brain of a family of insoluble amyloid peptides (Abeta peptides), which are produced as a result of the normal processing of beta-amyloid precursor protein (beta-APP). Russo et al. claim that a truncated Abeta peptide that lacks the first ten amino acids accumulates in the brains of patients carrying a mutant form of pre-senilin 1 (PS1), a protein that is involved in cleavage of beta-APP. However, we have found that this same species is also overrepresented in Alzheimer's patients with mutations in beta-APP itself. Our findings do not support the conclusion of Russo et al. that pathogenic PS1 mutations may control cleavage of beta-APP by beta-secretase.

Aged↗
PubMed2000

Carboxyl-terminal fragments of Alzheimer beta-amyloid precursor protein accumulate in restricted and unpredicted intracellular compartments in presenilin 1-deficient cells.

Absence of functional presenilin 1 (PS1) protein leads to loss of gamma-secretase cleavage of the amyloid precursor protein (betaAPP), resulting in a dramatic reduction in amyloid beta peptide (Abeta) production and accumulation of alpha- or beta-secretase-cleaved COOH-terminal fragments of betaAPP (alpha- or beta-CTFs). The major COOH-terminal fragment (CTF) in brain was identified as betaAPP-CTF-(11-98), which is consistent with the observation that cultured neurons generate primarily Abeta-(11-40). In PS1(-/-) murine neurons and fibroblasts expressing the loss-of-function PS1(D385A) mutant, CTFs accumulated in the endoplasmic reticulum, Golgi, and lysosomes, but not late endosomes. There were some subtle differences in the subcellular distribution of CTFs in PS1(-/-) neurons as compared with PS1(D385A) mutant fibroblasts. However, there was no obvious redistribution of full-length betaAPP or of markers of other organelles in either mutant. Blockade of endoplasmic reticulum-to-Golgi trafficking indicated that in PS1(-/-) neurons (as in normal cells) trafficking of betaAPP to the Golgi compartment is necessary before alpha- and beta-secretase cleavages occur. Thus, although we cannot exclude a specific role for PS1 in trafficking of CTFs, these data argue against a major role in general protein trafficking. These results are more compatible with a role for PS1 either as the actual gamma-secretase catalytic activity or in other functions indirectly related to gamma-secretase catalysis (e.g. an activator of gamma-secretase, a substrate adaptor for gamma-secretase, or delivery of gamma-secretase to betaAPP-containing compartments).

Alzheimer Disease↗
PubMed2000

Regulation of Alzheimer beta-amyloid precursor trafficking and metabolism.

Alzheimer's disease (AD) is characterized by the intracranial accumulation of the 4 kDa amyloid-beta peptide (Abeta), following proteolysis of a approximately 700-amino acid, integral membrane precursor, the Alzheimer amyloid precursor protein (APP). The best evidence causally linking APP to AD has been provided by the discovery of mutations within the APP coding sequence that segregate with disease phenotypes in autosomal dominant forms of familial AD (FAD). Though FAD is rare ( < 10% of all AD), the hallmark features (amyloid plaques, neurofibrillary tangles, synaptic and neuronal loss, neurotransmitter deficits and dementia) are indistinguishable when FAD is compared with typical, common, 'non-familial', or sporadic, AD (SAD). Studies of some clinically relevant mutant APP molecules from FAD families have yielded evidence that APP mutations can lead to the enhanced generation or aggregability of Abeta, consistent with a pathogenic role in AD. Other genetic loci for FAD have been discovered which are distinct from the immediate regulatory and coding regions of the APP gene, indicating that defects in molecules other than APP can also specify cerebral amyloidogenesis and FAD. To date, all APP and non-APP FAD mutations can be demonstrated to have the common feature of promoting amyloidogenesis of Abeta. Epidemiological studies indicate that postmenopausal women on estrogen replacement therapy (ERT) have their relative risk of developing SAD diminished by about one third as compared with age-matched women not receiving ERT [M.X. Tang, D. Jacobs, Y. Stern, K. Marder, P. Schofield, B. Gurland, H. Andrews, R. Mayeux, Effect of estrogen during menopause on risk and age at onset of Alzheimer's disease, Lancet 348 (2000) 429432]. Because of the key role of cerebral Abeta accumulation in initiating AD pathology, it is most attractive that estradiol might modulate SAD risk or age-at-onset by inhibiting Abeta accumulation. A possible mechanistic basis for such a scenario is reviewed here.

Alzheimer Disease↗
PubMed2000

Ovariectomy and 17beta-estradiol modulate the levels of Alzheimer's amyloid beta peptides in brain.

OBJECTIVE: To test whether female gonadal hormone status and estrogen modulate the metabolism of Abeta peptides in vivo. BACKGROUND: AD is a neurodegenerative disorder characterized by accumulation of aggregated forms of the 40- and 42-amino acid Abeta peptides (Abeta40 and Abeta42). Estrogen replacement therapy in postmenopausal women is associated with decreased risk for AD or delay in disease onset or both. The mechanism by which estrogen exerts this neuroprotective effect is elusive. 17beta-estradiol (E2) was shown to reduce the release of Abeta peptides by primary neuronal cultures of murine and human origin. METHODS: For this purpose, four experimental sets of guinea pigs were used: intact animals, ovariectomized animals (ovx), and ovariectomized animals that received E2 at two different doses (ovx+low-dose E2 and ovx+high-dose E2). Brain Abeta40 and Abeta42 levels were assessed using Abeta40 and Abeta42-specific ELISA assays. RESULTS: Prolonged ovariectomy resulted in uterine atrophy and decreased serum E2 levels and was associated with a pronounced increase in brain Abeta levels. Total brain Abeta in the ovx animals was increased by 1. 5-fold on average as compared to intact controls. E2 treatment of ovariectomized animals led to uterine hypertrophy and a dose-dependent increase in serum E2 levels. In addition, both doses of E2 significantly reversed the ovariectomy-induced increase in brain Abeta levels. The high-dose E2 treatment did not lead to a further decrease in brain Abeta beyond that observed with the low-dose E2 treatment. CONCLUSIONS: Our results infer that cessation of ovarian estrogen production in postmenopausal women might facilitate Abeta deposition by increasing the local concentrations of Abeta40 and Abeta42 peptides in brain. In addition, our finding that E2 treatment is associated with diminution of brain Abeta levels suggests that modulation of Abeta metabolism may be one of the ways by which estrogen replacement therapy prevents or delays the onset of AD or both in postmenopausal women.

Amyloid beta-Peptides↗
PubMed2000

Diurnal variation in the femoral articular cartilage of the knee in young adult humans.

Our objective was to test the hypothesis that diurnal changes occur in thickness or volume of the femoral articular cartilage of the knee in asymptomatic young adults. Fat-suppressed three-dimensional (3D) spoiled gradient-echo magnetic resonance imaging (MRI) was employed. Six volunteers each were scanned early in the morning and at the end of a working day spent mainly standing. This protocol was repeated on 3 successive weeks. Femoral cartilage volumes were obtained via semiautomatic segmentation that employed a seeding algorithm. These segmentations then were regridded onto a 500-pixel template, and differences in the resulting thickness maps were assessed. Analysis of variance showed no significant diurnal variation in overall volume or thickness. The reproducibility for volume (test-retest coefficient of variation) was 1.6%. There were, however, statistically-significant diurnal changes in the thickness maps. Cartilage thickness decreased by up to 0.6 mm during the day in each of the following three specific locations: the patellofemoral compartment, the lateral tibiofemoral compartment, and the medial tibiofemoral compartment. Elsewhere, cartilage thickness was unchanged or increased by up to 0.5 mm. We conclude that, in asymptomatic young adults, cartilage volume does not change during the day; however, the cartilage does become thinner in locations that encounter the greatest biomechanical force.

Adult↗
PubMed2000

Neurohormonal signalling pathways and the regulation of Alzheimer beta-amyloid metabolism.

Alzheimer's disease (AD) is characterized by the intracranial accumulation of the 4 kDa amyloid beta peptide (A beta), following proteolysis of a approximately 700 amino acid, integral membrane precursor, the amyloid beta precursor protein (APP). The best evidence causally linking APP to AD has been provided by the discovery of mutations within the APP coding sequence that segregate with disease phenotypes in autosomal dominant forms of familial AD (FAD). Though FAD is rare (< 10% of all AD), the hallmark features--amyloid plaques, neurofibrillary tangles, synaptic and neuronal loss, neurotransmitter deficits, dementia--are indistinguishable when FAD is compared with typical, common, 'non-familial', or sporadic AD (SAD). Studies of some clinically relevant mutant APP molecules from FAD families have yielded evidence that APP mutations can lead to enhanced generation or aggregability of A beta, consistent with a pathogenic role in AD. Other genetic loci for FAD have been discovered which are distinct from the immediate regulatory and coding regions of the APP gene, indicating that defects in molecules other than APP can also specify cerebral amyloidogenesis and FAD. To date, all APP and non-APP FAD mutations can be demonstrated to have the common feature of promoting amyloidogenesis of A beta. Epidemiological studies indicate that postmenopausal women on oestrogen hormone replacement therapy (HRT) have their relative risk of developing SAD diminished by about one-third as compared with age-matched women not receiving HRT. Because of the key role of cerebral A beta accumulation in initiating AD pathology, it is most attractive that oestradiol might modulate SAD risk or age-at-onset by inhibiting A beta accumulation. A possible mechanistic basis for such a scenario is reviewed here.

Alzheimer Disease↗
PubMed2000

Post-menopausal estrogen deprivation and Alzheimer's disease.

Estrogen deprivation has been implicated as a risk factor in Alzheimer's disease (AD) as epidemiological data suggests that estrogen replacement therapy can protect against the onset, and progression of the disease. Biochemical data suggests that estrogen exerts its affect through the processing of the amyloid precursor protein to beta-amyloid which is deposited in the brains of patients with AD. The effects of estrogens may be more widespread, however, as it has been implicated in the maintenance of neuronal architecture and protection from free radicals. This review aims to discuss the various roles of estrogen in the development of AD.

Alzheimer Disease↗
PubMed2000

Ovariectomy and 17beta-estradiol modulate the levels of Alzheimer's amyloid beta peptides in brain.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by accumulation of aggregated forms of the 40- and 42-amino acid Abeta peptides (Abeta40 and Abeta42). Estrogen replacement therapy (ERT) in postmenopausal women is associated with decreased risk for AD and/or delay in disease onset. The mechanism by which estrogen exerts this neuroprotective effect is elusive. 17beta-estradiol (E2) was shown to reduce the release of Abeta peptides by primary neuronal cultures of murine and human origin. To test whether estrogen can modulate the metabolism of Abeta peptides in vivo, four experimental sets of guinea pigs were used: intact animals, ovariectomized animals, and ovariectomized animals that received E2 at two different doses. Ovariectomy was associated with a 1.5-fold average increase in total brain Abeta levels as compared to intact controls. E2 treatment significantly reversed the ovariectomy-induced increase in brain Abeta levels. The high-dose E2 treatment did not lead to further decrease in brain Abeta beyond the one observed with the low-dose E2 treatment. Our results infer that cessation of ovarian estrogen production in postmenopausal women might facilitate Abeta deposition by increasing the local concentrations of Abeta40 and Abeta42 peptides in brain and suggest that modulation of Abeta metabolism may be one of the ways by which ERT prevents and/or delays the onset of AD in postmenopausal women.

Amyloid beta-Peptides↗
PubMed2000

Mutant presenilin 1 increases the levels of Alzheimer amyloid beta-peptide Abeta42 in late compartments of the constitutive secretory pathway.

Mutations in the presenilin 1 (PS1) gene are associated with autosomal dominant, early-onset, familial Alzheimer's disease and result in increased release of the hyperaggregatable 42-amino acid form of the amyloid beta-peptide (A(beta)42). To determine which subcellular compartments are potential source(s) of released Abeta42, we compared the levels and spatial segregation of intracellular A(beta)40 and A(beta)42 peptides between N2a neuroblastoma cells doubly transfected with the "Swedish" familial Alzheimer's disease-linked amyloid precursor protein variant and either wild-type PS1 (PS1(wt)) or familial Alzheimer's disease-linked delta9 mutant PS1 (PS1delta9). As expected, PS1delta9-expressing cells had dramatically higher levels of intracellular Abeta42 than did cells expressing PS1wt. However, the highest levels of A(beta)42 colocalized not with endoplasmic reticulum or Golgi markers but with rab8, a marker for trans-Golgi network (TGN)-to-plasma membrane (PM) transport vesicles. We show that PS1 mutants are capable of causing accumulation of A(beta)42 in late compartments of the secretory pathway, generating there a readily releasable source of A(beta)42. Our findings indicate that PS1 "bioactivity" localizes to the vicinity of the TGN and/or PM and reconcile the apparent discrepancy between the preponderant concentration of PS1 protein in proximal compartments of the secretory pathway and the recent findings that PS1 "bioactivity" can control gamma-secretase-like processing of another transmembrane substrate, Notch, at or near the PM.

Alzheimer Disease↗
PubMed2000

Making research relevant to the primary health care team.

There is, as yet, no strong culture of research in primary care and much of the existing research is conceived and undertaken by people outside primary care. The poor implementation of research findings may, in part, be owing to the fact that those delivering the service are not involved in asking or answering questions that are relevant to their practice. This paper reports how three practices constructed a research agenda based on the unanswered questions of their primary care teams. The research questions prioritised by the teams tended towards patient behaviour and service organisation rather than clinical issues. This contrasts with national research priorities. The process has contributed towards the development of a culture of enquiry among team members. Other primary care teams may benefit from a similar approach. National research priority setting in primary care should take more account of the unanswered questions of primary care teams.

Family Practice↗
PubMed1999

Generation of the amyloid-beta peptide N terminus in Saccharomyces cerevisiae expressing human Alzheimer's amyloid-beta precursor protein.

The Alzheimer's amyloid-beta precursor protein (betaAPP) is a type 1 membrane-spanning protein from which the Alzheimer's disease amyloid-beta peptide (Abeta) is proteolytically derived. To date, attempts to identify the enzymes responsible for Abeta generation have failed. Here we report the accumulation of Abeta-immunoreactive peptides in yeast expressing human betaAPP. Characterization of these peptides by metabolic labeling, immunoprecipitation with Abeta-specific antibodies, and N-terminal radiosequencing indicates that these peptides include the Abeta peptide at their N termini. The Abeta-like peptides generated in yeast were recovered predominantly as 8- and 12-14-kDa species. A 4-kDa species was recovered either when a protease-deficient strain was used to prevent breakdown or when the 8- and 12-14-kDa species were treated with disaggregating agents. The likely existence in yeast of enzymes generating the Abeta N terminus indicates that the molecular identification of yeast beta-secretase-like enzymes may be accomplished using genetic screens or empirical approaches based upon the sequenced genome of Saccharomyces cerevisiae.

Alzheimer Disease↗
PubMed1999

Phylogenetically conserved CK-II phosphorylation site of the murine homeodomain protein Hoxb-6.

In an effort to characterize the signal transduction mechanisms that operate to regulate homeodomain protein function, we have analyzed the phosphorylation state of two homeodomain proteins, Hoxb-6 and Hoxc-8, in vitro and in vivo. The baculovirus expression system was employed to demonstrate that Hoxb-6 is phosphorylated in Sf9 cells while Hoxc-8 is not. Using two-dimensional tryptic phosphopeptide mapping and purified protein kinases, we demonstrate that Hoxb-6 is phosphorylated in vitro by casein kinase II and cAMP-dependent protein kinase. The casein kinase II phosphorylation site was mapped to serine-214. Two-dimensional tryptic phosphopeptide mapping of immunoprecipitated Hoxb-6 from mouse embryonic spinal cords demonstrates that the same peptide phosphorylated in vitro and in Sf9 cells by casein kinase II is also phosphorylated in vivo. The conservation of this site in several homeodomain proteins from various species is discussed.

Amino Acid Sequence↗
PubMed1999

Automatic realignment of time-separated MR images by genetic algorithm.

A simple yet highly efficient artificial intelligence technique utilizing a genetic algorithm is used to register time-separated pairs of MRI data sets. To encourage others to try the approach, the algorithm is presented by way of a simple example to a 2-D data set; it is equally applicable to 3-D data. The technique is reliably found to reduce mismatch in images of the distal-interphalangeal joint from the order of several mm to just 200 microm (one pixel). The method and transformation are general and would be suitable for locating physical changes between any image data sets. We believe the technique to be of use in functional imaging, measurement of disease progression with time (e.g., degradation of cartilage in arthritic disease) and pre/post-surgical studies.

Algorithms↗
PubMed1999

Neurohormonal Signaling Pathways and the Regulation of Alzheimer beta-Amyloid Precursor Metabolism.

Postmenopausal women on estrogen replacement appear to have their relative risk of developing Alzheimer's disease diminished by about one half. Because brain amyloid accumulation plays a key role in initiating Alzheimer's pathology, it is attractive to postulate that estrogen might modulate Alzheimer's risk by inhibiting amyloid accumulation. Data and cell biological models supporting such a scenario are reviewed here.

Journal Article↗