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Biomedical subjects

Todd E Golde

Publications and source records attributed to Todd E Golde.

At least 19 recordsLinked to original sources
PubMed2003

Amyloid-beta immunization effectively reduces amyloid deposition in FcRgamma-/- knock-out mice.

Direct immunization with amyloid beta protein (Abeta) and passive transfer of anti-Abeta antibodies reduce Abeta accumulation and attenuate cognitive deficits in transgenic models of Alzheimer's disease (AD). The reduction in Abeta deposition has been proposed to involve microglial phagocytosis of Abeta immune complexes via Fc receptors (FcRs). We have examined the efficacy of Abeta immunization in amyloid precursor protein (APP) transgenic mice crossed into FcR-gamma chain knock-out mice (FcRgamma-/-). As might be expected from previous studies on macrophages, phagocytosis of Abeta immune complexes via FcR was completely impaired in microglia cells isolated from FcRgamma-/- mice. Thus, we immunized APP Tg2576 transgenic mice that were crossed in the FcRgamma-/- background with Abeta1-42 and then analyzed the effect on Abeta accumulation. In APP Tg2576 transgenic mice crossed to FcRgamma-/-, Abeta1-42 immunization significantly attenuated Abeta deposition, as assessed by both biochemical and immunohistological methods. The reduction in Abeta accumulation was equivalent to the reduction in deposition seen in Abeta1-42 immunized, age-matched, FcR-sufficient Tg2576 mice. We conclude that after Abeta immunization, the effects of anti-Abeta antibodies on Abeta deposition in APP Tg2576 transgenic mice are not dependent on FcR-mediated phagocytic events.

Age Factors↗
PubMed2003

TCR-mediated Notch signaling regulates proliferation and IFN-gamma production in peripheral T cells.

Notch genes encode membrane receptors that regulate cell fate decisions in metazoa. Notch receptors and ligands are expressed in developing lymphoid tissue and mature lymphocytes and the role of Notch signaling in early T and B cell development has been studied extensively. However, its contribution to mature T cell function is unknown. TCR-mediated T cell activation is a fundamental process of the adaptive immune system that has been studied for decades; however, the details of this process are incompletely understood. In this study, we present evidence that Notch is required for TCR-mediated activation of peripheral T cells. Inhibition of Notch activation dramatically decreases T cell proliferation in both CD4 and CD8 cells and blocks both NF-kappaB activity and IFN-gamma production in peripheral T cells. Our data reveal a new, nondevelopmental function of Notch as a previously unknown key link in peripheral T cell activation and cytokine secretion.

Amyloid Precursor Protein Secretases↗
PubMed2003

Intramembrane-cleaving aspartic proteases and disease: presenilins, signal peptide peptidase and their homologs.

Recent studies demonstrate that presenilins (PSs) and signal peptide peptidase (SPP) are members of a novel protease family of integral membrane proteins that may utilize a catalytic mechanism similar to classic aspartic proteases such as pepsin, renin and cathepsin D. The defining features of the PSs and SPP are their ability to cleave substrate polypeptides within a transmembrane region, the presence of two active site aspartate residues in adjacent membrane-spanning regions and a conserved PAL motif near their COOH-terminus. PSs appear to be the catalytic subunit of multiprotein complexes that possess gamma-secretase activity. Because this activity generates the amyloid beta peptide (Abeta) deposited in the brain of patients with Alzheimer's disease (AD), PSs are considered therapeutic targets in AD. In contrast to PSs that are not active unless part of a larger complex, SPP does not appear to require protein co-factors. Because of its requirement for hepatitis C virus maturation and a possible immune modulatory role, SPP is also considered a potential therapeutic target. Four additional PS/SPP homologs have been identified in humans; yet, their functions have not been elucidated. Herein, we will review the recent advances in our understanding of the PS/SPP family of proteases as well as discuss aspects of intramembrane cleavage that are not well understood.

Amino Acid Sequence↗
PubMed2003

Triple-transgenic model of Alzheimer's disease with plaques and tangles: intracellular Abeta and synaptic dysfunction.

The neuropathological correlates of Alzheimer's disease (AD) include amyloid-beta (Abeta) plaques and neurofibrillary tangles. To study the interaction between Abeta and tau and their effect on synaptic function, we derived a triple-transgenic model (3xTg-AD) harboring PS1(M146V), APP(Swe), and tau(P301L) transgenes. Rather than crossing independent lines, we microinjected two transgenes into single-cell embryos from homozygous PS1(M146V) knockin mice, generating mice with the same genetic background. 3xTg-AD mice progressively develop plaques and tangles. Synaptic dysfunction, including LTP deficits, manifests in an age-related manner, but before plaque and tangle pathology. Deficits in long-term synaptic plasticity correlate with the accumulation of intraneuronal Abeta. These studies suggest a novel pathogenic role for intraneuronal Abeta with regards to synaptic plasticity. The recapitulation of salient features of AD in these mice clarifies the relationships between Abeta, synaptic dysfunction, and tangles and provides a valuable model for evaluating potential AD therapeutics as the impact on both lesions can be assessed.

Alzheimer Disease↗
PubMed2003

The non-cyclooxygenase targets of non-steroidal anti-inflammatory drugs, lipoxygenases, peroxisome proliferator-activated receptor, inhibitor of kappa B kinase, and NF kappa B, do not reduce amyloid beta 42 production.

Epidemiological evidence suggests that chronic use of non-steroidal anti-inflammatory drugs (NSAIDs) reduces the risk of Alzheimer's disease. Recently, NSAIDs have been shown to decrease amyloid pathology in a transgenic mouse model of Alzheimer's disease. This benefit may be partially attributable to the ability of NSAIDs to selectively reduce production of the amyloidogenic A beta 42 peptide in both cultured cells and transgenic mice. Although this activity does not appear to require the action of cyclooxygenases in cultured cells, it is not known whether other NSAID-sensitive targets contribute to this A beta 42 effect. In this study, we have used both pharmacological and genetic means to determine if other known cellular targets of NSAIDs could mediate the reduction in A beta 42 secretion from cultured cells. We find that altered arachidonic acid metabolism via NSAID action on cyclooxygenases and lipoxygenases does not alter A beta 42 production. Furthermore, we demonstrate that alterations in activity of peroxisome proliferator-activated receptors, I kappa B kinase beta or nuclear factor kappa B do not affect A beta 42 production. Thus, NSAIDs do not appear to alter A beta 42 production indirectly through previously identified cellular targets and may interact directly with the gamma-secretase complex itself to affect amyloid production.

Amyloid beta-Peptides↗
PubMed2003

Evidence that nonsteroidal anti-inflammatory drugs decrease amyloid beta 42 production by direct modulation of gamma-secretase activity.

Chronic use of nonsteroidal anti-inflammatory drugs (NSAIDs) is associated with a lower risk of developing Alzheimer's disease. Recent evidence indicates that some NSAIDs specifically inhibit secretion of the amyloidogenic A beta 42 peptide in cultured cells and mouse models of Alzheimer's disease. The reduction of A beta 42 peptides is not mediated by inhibition of cyclooxygenases (COX) but the molecular mechanism underlying this novel activity of NSAIDs has not been further defined. We now demonstrate that NSAIDs efficiently reduce the intracellular pool of A beta 42 in cell-based studies and selectively decrease A beta 42 production in a cell-free assay of gamma-secretase activity. Moreover, we find that presenilin-1 (PS1) mutations, which affect gamma-secretase activity, differentially modulate the cellular A beta 42 response to NSAID treatment. Overexpression of the PS1-M146L mutation enhances the cellular drug response to A beta 42 lowering NSAIDs as compared with cells expressing wild-type PS1. In contrast, expression of the PS1-Delta Exon9 mutation strongly diminishes the A beta 42 response, showing that PS1 mutations can modulate the cellular drug response to NSAID treatment both positively and negatively. Enhancement of the NSAID drug response was also observed with overexpression of the APP V717F mutation but not with Swedish mutant APP, which affects beta-secretase cleavage. In sum, these results strongly suggest that NSAIDs represent a founding group of compounds that lower A beta 42 production by direct modulation of gamma-secretase activity or its substrate.

Amyloid Precursor Protein Secretases↗
PubMed2003

Peptide-based, irreversible inhibitors of gamma-secretase activity.

The characterization of the enzymes responsible for amyloid beta-peptide (Abeta) production is considered to be a primary goal towards the development of future therapeutics for the treatment of Alzheimer's disease. Inhibitors of gamma-secretase activity were critical in demonstrating that the presenilins (PSs) likely comprised at least part of the active site of the gamma-secretase enzyme complex, with two highly conserved membrane aspartates presumably acting as catalytic residues. However, whether or not these aspartates are actually the catalytic residues of the enzyme complex or are merely essential for normal PS function and/or maturation is still unknown. In this paper, we report the development of reactive inhibitors of gamma-secretase activity that are functionally irreversible. Since such inhibitors have been shown to bind catalytic residues in other aspartyl proteases (e.g., HIV protease), they might be used to determine if the transmembrane aspartates of PSs are involved directly in substrate cleavage.

Amyloid Precursor Protein Secretases↗
PubMed2003

Abeta42-lowering nonsteroidal anti-inflammatory drugs preserve intramembrane cleavage of the amyloid precursor protein (APP) and ErbB-4 receptor and signaling through the APP intracellular domain.

Epidemiological studies indicate that long term use of nonsteroidal anti-inflammatory drugs (NSAIDs) confers protection from Alzheimer's disease, and some NSAIDs were shown to specifically decrease production of the amyloidogenic Abeta42 peptide, most likely by direct modulation of gamma-secretase activity. In contrast to gamma-secretase inhibitors, Abeta42-lowering NSAIDs do not impair S3 cleavage in the NOTCH receptor and release of the NOTCH intracellular domain, a finding with conceptual implications for the development of safer drugs targeting Abeta production through gamma-secretase modulation. Intramembrane cleavage and release of an intracellular signaling domain has recently been demonstrated in a number of additional gamma-secretase substrates. We now show in cell-based assays that intramembrane cleavage of APP and ErbB-4 receptor is not impaired by the Abeta42-lowering NSAIDs, sulindac sulfide and ibuprofen. Generation of the APP intracellular domain (AICD) was further not inhibited in a cell-free assay at concentrations far exceeding those effective in reducing Abeta42 production. Closer inspection of AICD signaling showed that stabilization of the AICD peptide by FE65 and AICD-mediated transcription were also retained at Abeta42-lowering concentrations. These results demonstrate that S3-like/intramembrane cleavage is preserved by Abeta42-lowering NSAIDs in at least three substrates of gamma-secretase APP, ErbB-4, and NOTCH and underline the striking specificity by which these drugs target Abeta42 production.

Amino Acid Sequence↗
PubMed2003

Overexpression of nicastrin increases Abeta production.

Gamma-secretase cleavage is the final proteolytic step that releases the amyloid beta-peptide (Abeta) from the amyloid beta-protein precursor (APP). Significant evidence indicates that the presenilins (PS) are catalytic components of a high molecular weight gamma-secretase complex. The glycoprotein nicastrin was recently identified as a functional unit of this complex based on 1) binding to PS and 2) the ability to modulate Abeta production following mutation of a conserved DYIGS region. In contrast to the initial report, we find that overexpression of wild-type (WT) nicastrin increases Abeta production, whereas DYIGS mutations (MT) have little or no effect. The increase in Abeta production is associated with an increase in gamma-secretase activity but not with a detectable increase in PS1 levels. Subcellular fractionation studies show that WT but not MT nicastrin matures into buoyant membrane fractions enriched in gamma-secretase activity. These data support the hypothesis that nicastrin is an essential component of the gamma-secretase complex. The finding that WT nicastrin overexpression can increase gamma-secretase activity without altering levels of the presumed catalytic component (PS) of the enzyme may point to a role for nicastrin in facilitating cleavage by regulating substrate interactions with the gamma-secretase complex.

Amyloid Precursor Protein Secretases↗
PubMed2003

Physiologic and pathologic events mediated by intramembranous and juxtamembranous proteolysis.

Intramembranous proteolysis (IP) is a recently recognized mechanism for transmembrane signal transduction that involves proteolysis of transmembrane proteins within their membrane-spanning domains. Juxtamembranous proteolysis (JP) is similar, but proteolytic cleavage of a transmembrane protein occurs at a site close to, but not within, the transmembrane domain of the target protein. In both IP and JP, a soluble domain of a transmembrane protein is released from its membrane tether. This domain can then transmit a signal either locally or at some distance from the site of cleavage. In certain signaling pathways, JP and IP are linked. JP on one side of the membrane results in secondary IP, which then releases a signaling domain from the membrane. Whereas well-characterized proteases such as caspases, the proteasome, and metalloprotease disintegrins, have been implicated in JP, three families of multipass membrane proteases (MpMPs) have now been shown to carry out IP. Recent studies of events mediated by IP and JP indicate that they regulate key cellular signaling events including pathways involved in sterol regulation, cell fate selection, and growth regulation. Moreover, IP and JP have important roles in certain diseases such as Alzheimer's disease. Because some of the proteases mediating IP and JP can be selectivity inhibited, inhibitors targeting these proteases are likely to alter both physiologic and pathologic events triggered by IP and JP.

Alzheimer Disease↗
PubMed2003

Immune responses against Abeta1-42 in HLA class II transgenic mice: implications for Abeta1-42 immune-mediated therapies.

We have investigated whether polymorphic differences in the major histocompatibility complex (MHC) class II molecules influence humoral and cellular immune responses against Abeta1-42. To analyze the effects of mouse MHC class II and tolerance effects of overexpression of human APP in mice, we immunized Tg2576 and non-transgenic littermates bred into two different MHC backgrounds with Abeta1-42 and compared both B and T cell responses. We found that in the presence of the mouse C57BL/6 background, both B and T cell responses against Abeta1-42 were significantly suppressed. To directly test the contribution of human MHC class II, we immunized various human HLA class II transgenic (TG) mice with Abeta1-42 and analyzed anti-Abeta immune responses. HLA-DR3 and HLA-DQ8 TG mice generated modest B and T cell responses against Abeta1-42. The presence of HLA-DR3/DQ8 in double TG mice enhanced the overall immune response against Abeta1-42. In contrast, HLA-DR4 TG mice mounted strong T cell responses but failed to generate high titer antibody responses against Abeta1-42, whereas, the HLA-DQ6 TG mice were not able to mount significant B or T cell responses against Abeta1-42. These studies in mice suggest that the presence of certain MHC class II molecules or combinations of class II molecules can potentially influence the overall immune response against Abeta1-42.

Amyloid beta-Peptides↗
PubMed2003

NSAIDs and enantiomers of flurbiprofen target gamma-secretase and lower Abeta 42 in vivo.

Epidemiologic studies demonstrate that long-term use of NSAIDs is associated with a reduced risk for the development of Alzheimer disease (AD). In this study, 20 commonly used NSAIDs, dapsone, and enantiomers of flurbiprofen were analyzed for their ability to lower the level of the 42-amino-acid form of amyloid beta protein (Abeta42) in a human H4 cell line. Thirteen of the NSAIDs and the enantiomers of flurbiprofen were then tested in acute dosing studies in amyloid beta protein precursor (APP) transgenic mice, and plasma and brain levels of Abeta and the drug were evaluated. These studies show that (a). eight FDA-approved NSAIDs lower Abeta42 in vivo, (b). the ability of an NSAID to lower Abeta42 levels in cell culture is highly predicative of its in vivo activity, (c). in vivo Abeta42 lowering in mice occurs at drug levels achievable in humans, and (d). there is a significant correlation between Abeta42 lowering and levels of ibuprofen. Importantly, flurbiprofen and its enantiomers selectively lower Abeta42 levels in broken cell gamma-secretase assays, indicating that these compounds directly target the gamma-secretase complex that generates Abeta from APP. Of the compounds tested, meclofenamic acid, racemic flurbiprofen, and the purified R and S enantiomers of flurbiprofen lowered Abeta42 levels to the greatest extent. Because R-flurbiprofen reduces Abeta42 levels by targeting gamma-secretase and has reduced side effects related to inhibition of cyclooxygenase (COX), it is an excellent candidate for clinical testing as an Abeta42 lowering agent.

Alzheimer Disease↗
PubMed2002

Identification of a novel family of presenilin homologues.

Presenilin 1 and presenilin 2 are polytopic membrane proteins, whose genes are mutated in some individuals with Alzheimer's disease. Presenilins have been shown to influence limited proteolysis of amyloid beta protein precursor (APP), Notch and ErbB4, and have been proposed to be gamma-secretases that perform the terminal cleavage of APP. In this model, two conserved and apparently intramembranous aspartic acids participate in catalysis. Highly sequence-similar presenilin homologues are known in plants, invertebrates and vertebrates. In this work, we have used a combination of different sequence database search methods to identify a new family of proteins homologous to presenilins. Members of this family, which we term presenilin homologues (PSH), have significant sequence similarities to presenilins and also possess two conserved aspartic acid residues within adjacent predicted transmembrane segments. The PSH family is found throughout the eukaryotes, in fungi as well as plants and animals, and in archaea. Five PSHs are detectable in the human genome, of which three possess "protease-associated" domains that are consistent with the proposed protease function of PSs. Based on these findings, we propose that PSs and PSHs represent different sub-branches of a larger family of polytopic membrane-associated aspartyl proteases.

Alzheimer Disease↗
PubMed2002

Inclusion body myositis-like phenotype induced by transgenic overexpression of beta APP in skeletal muscle.

Inclusion body myositis (IBM), the most common age-related muscle disease in the elderly population, is an incurable disorder leading to severe disability. Sporadic IBM has an unknown etiology, although affected muscle fibers are characterized by many of the pathobiochemical alterations traditionally associated with neurodegenerative brain disorders such as Alzheimer's disease. Accumulation of the amyloid-beta peptide, which is derived from proteolysis of the larger amyloid-beta precursor protein (betaAPP), seems to be an early pathological event in Alzheimer's disease and also in IBM, where in the latter, it predominantly occurs intracellularly within affected myofibers. To elucidate the possible role of betaAPP mismetabolism in the pathogenesis of IBM, transgenic mice were derived in which we selectively targeted betaAPP overexpression to skeletal muscle by using the muscle creatine kinase promoter. Here we report that older (>10 months) transgenic mice exhibit intracellular immunoreactivity to betaAPP and its proteolytic derivatives in skeletal muscle. In this transgenic model, selective overexpression of betaAPP leads to the development of a subset of other histopathological and clinical features characteristic of IBM, including centric nuclei, inflammation, and deficiencies in motor performance. These results are consistent with a pathogenic role for betaAPP mismetabolism in human IBM.

Amyloid beta-Protein Precursor↗
PubMed2002

A physiologic signaling role for the gamma -secretase-derived intracellular fragment of APP.

Presenilins mediate an unusual intramembranous proteolytic activity known as gamma-secretase, two substrates of which are the Notch receptor (Notch) and the beta-amyloid precursor protein (APP). Gamma-secretase-mediated cleavage of APP, like that of Notch, yields an intracellular fragment [APP intracellular domain (AICD)] that forms a transcriptively active complex. We now demonstrate a functional role for AICD in regulating phosphoinositide-mediated calcium signaling. Genetic ablation of the presenilins or pharmacological inhibition of gamma-secretase activity (and thereby AICD production) attenuated calcium signaling in a dose-dependent and reversible manner through a mechanism involving the modulation of endoplasmic reticulum calcium stores. Cells lacking APP (and hence AICD) exhibited similar calcium signaling deficits, and-notably-these disturbances could be reversed by transfection with APP constructs containing an intact AICD, but not by constructs lacking this domain. Our findings indicate that the AICD regulates phosphoinositide-mediated calcium signaling through a gamma-secretase-dependent signaling pathway, suggesting that the intramembranous proteolysis of APP may play a signaling role analogous to that of Notch.

Amyloid Precursor Protein Secretases↗
PubMed2002

FAD-linked mutations in presenilin 1 alter the length of Abeta peptides derived from betaAPP transmembrane domain mutants.

gamma-Secretase is an enzymatic activity responsible for the final cleavage of the amyloid precursor protein leading to the production of the amyloid beta-peptide (Abeta). gamma-Secretase is likely an aspartyl protease, since its activity can be inhibited by both pepstatin and active-site directed aspartyl protease inhibitors. Recent work has indicated that presenilins 1 and 2 may actually be the gamma-secretase enzymes. Presenilin (PS) mutations, which lead to an increase in the production of a longer form of Abeta, are also the most common cause of familial Alzheimer's disease (FAD). Therefore, in an attempt to better characterize the substrate preferences of gamma-secretase, we performed experiments to determine how FAD-linked mutations in PS1 would affect the generation of Abeta peptides from full length precursor substrates that we have previously demonstrated to be proteolytically cleaved at alternative sites and/or by enzymatic activities that are pharmacologically distinct. Presenilin mutations increased the production of Abeta peptides from sites distal to the primary cleavage site ('longer' peptides) and in several cases also decreased production of 'shorter' peptides. These results support a model in which the FAD-linked mutants subtly alter the conformation of the gamma-secretase complex to favor the production of long Abeta.

Amino Acid Sequence↗
PubMed2002

Cholesterol-dependent gamma-secretase activity in buoyant cholesterol-rich membrane microdomains.

Buoyant membrane fractions containing presenilin 1 (PS1), an essential component of the gamma-secretase complex, and APP CTFbeta, a gamma-secretase substrate, can be isolated from cultured cells and brain by several different fractionation procedures that are compatible with in vitro gamma-secretase assays. Analysis of these gradients for amyloid beta protein (Abeta) and CTFgamma production indicated that gamma-secretase activity is predominantly localized in these buoyant membrane microdomains. Consistent with this localization, we find that gamma-secretase activity is cholesterol dependent. Depletion of membrane cholesterol completely inhibits gamma-secretase cleavage, which can be restored by cholesterol replacement. Thus, altering cholesterol levels may influence the development of Alzheimer's disease (AD) by influencing production and deposition of Abeta within cholesterol rich membrane microdomains.

Alzheimer Disease↗