Age-related changes to tumor necrosis factor receptors affect neuron survival in the presence of beta-amyloid

Inflammation including local accumulations of tumor necrosis factor alpha (TNF-alpha) is a part of Alzheimer's disease pathology and may exacerbate age-related neurodegeneration. Most studies on TNF-alpha and TNF neuronal receptors are conducted by using embryonic neurons. Few studies consider age-related deficits that may occur in neurons. Age-related changes in susceptibility to TNF-alpha through TNF receptor 1 (TNFR1) and receptor 2 (TNFR2) expression could increase susceptibility to beta-amyloid (1-42, Abeta42). Evidence is conflicting about which receptor mediates survival and/or apoptosis. We determined how aging affects receptor expression in cultured adult rat cortical neurons. Old neurons were more susceptible to Abeta42 toxicity than middle-aged neurons, and the addition of TNF-alpha was neuroprotective in middle-aged neurons, but exacerbated the toxicity from Abeta42 in old neurons. These pathologic and protective responses in old and middle-aged neurons, respectively, correlated with higher starting TNFR1 and TNFR2 mRNA levels in old vs. middle-aged neurons. Middle-aged neurons treated with TNF-alpha plus Abeta42 did not show an increase in either TNFR1 or TNFR2 mRNA, but old neurons showed an up-regulation in TNFR2 mRNA and not TNFR1 mRNA. Despite these mRNA changes, surface immunoreactivity of both TNFR1 and TNFR2 increased with the dose of TNF-alpha in middle-aged neurons. However, middle-aged neurons treated with TNF-alpha plus Abeta42 showed an up-regulation in both TNFR1 and TNFR2 surface expression, whereas old neurons failed to up-regulate surface expression of either receptor. These findings support the hypothesis that age-related changes in TNF-alpha surface receptor expression contribute to the neuronal loss associated with inflammation in Alzheimer's disease.     

Comparative study of microglia activation induced by amyloid-beta and prion peptides: role in neurodegeneration

The inflammatory responses in Alzheimer's disease (AD) and prion-related encephalopathies (PRE) are dominated by microglia activation. Several studies have reported that the amyloid-beta (Abeta) peptides, which are associated with AD, and the pathogenic isoform of prion protein (PrPSc) have a crucial role in neuronal death and gliosis that occur in both of these disorders. In this study, we investigate whether Abeta and PrPSc cause microglia activation per se and whether these amyloidogenic peptides differentially affect these immunoeffector cells. In addition, we also determined whether substances released by Abeta- and PrP-activated microglia induce neuronal death. Cultures of rat brain microglia cells were treated with the synthetic peptides Abeta1-40, Abeta1-42 and PrP106-126 for different time periods. The lipopolysaccharide was used as a positive control of microglia activation. Our results show that Abeta1-40 and PrP106-126 caused similar morphological changes in microglia and increased the production of nitric oxide and hydroperoxides. An increase on inducible nitric oxide synthase expression was also observed in microglia treated with Abeta1-40 or PrP106. However, these peptides affected in a different manner the secretion of interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) secretion. In cocultures of microglia-neurons, it was observed that microglia treated with Abeta1-40 or PrP106-126 induced a comparable extent of neuronal death. The neutralizing antibody for IL-6 significantly reduced the neuronal death induced by Abeta- or PrP-activated microglia. Taken together, the data indicate that Abeta and PrP peptides caused microglia activation and differentially affected cytokine secretion. The IL-6 released by reactive microglia caused neuronal injury.     

Involvement of microglial receptor for advanced glycation endproducts (RAGE) in Alzheimer's disease: identification of a cellular activation mechanism

Receptor-mediated interactions with amyloid beta-peptide (Abeta) could be important in the evolution of the inflammatory processes and cellular dysfunction that are prominent in Alzheimer's disease (AD) pathology. One candidate receptor is the receptor for advanced glycation endproducts (RAGE), which can bind Abeta and transduce signals leading to cellular activation. Data are presented showing a potential mechanism for Abeta activation of microglia that could be mediated by RAGE and macrophage colony-stimulating factor (M-CSF). Using brain tissue from AD and nondemented (ND) individuals, RAGE expression was shown to be present on microglia and neurons of the hippocampus, entorhinal cortex, and superior frontal gyrus. The presence of increased numbers of RAGE-immunoreactive microglia in AD led us to further analyze RAGE-related properties of these cells cultured from AD and ND brains. Direct addition of Abeta(1-42) to the microglia increased their expression of M-CSF. This effect was significantly greater in microglia derived from AD brains compared to those from ND brains. Increased M-CSF secretion was also demonstrated using a cell culture model of plaques whereby microglia were cultured in wells containing focal deposits of immobilized Abeta(1-42). In each case, the Abeta stimulation of M-CSF secretion was significantly blocked by treatment of cultures with anti-RAGE F(ab')2. Treatment of microglia with anti-RAGE F(ab')2 also inhibited the chemotactic response of microglia toward Abeta(1-42). Finally, incubation of microglia with M-CSF and Abeta increased expression of RAGE mRNA. These microglia also expressed M-CSF receptor mRNA. These data suggest a positive feedback loop in which Abeta-RAGE-mediated microglial activation enhances expression of M-CSF and RAGE, possibly initiating an ascending spiral of cellular activation.     

Effects of statins on microglia

High serum cholesterol level has been shown as one of the risk factors for Alzheimer's disease (AD), and epidemiological studies indicate that treatment with cholesterol-lowering substances, statins, may provide protection against AD. An acute-phase reaction and inflammation, with increased levels of proinflammatory cytokines, are well known in the AD brain. Notably, there is evidence for antiinflammatory activities of statins, such as reduction in proinflammatory cytokines. Consequently, it is of interest to analyze the effects of statins on microglia, the main source of inflammatory factors in the brain, such as in AD. The aims of this study were to determine the effects of statins (atorvastatin and simvastatin) on microglial cells with regard to the secretion of the inflammatory cytokine interleukin-6 (IL-6) and cell viability after activation of the cells with bacterial lipopolysaccharides (LPS) or beta-amyloid1-40 (Abeta1-40) and in unstimulated cells. Cells of the human microglial cell line CHME-3 and primary cultures of rat neonatal cortical microglia were used. Incubation with LPS or Abeta1-40 induced secretion of IL-6, and Abeta1-40, but not LPS, reduced cell viability. Both atorvastatin and simvastatin reduced the basal secretion of IL-6 and the cell viability of the microglia, but only atorvastatin reduced LPS- and Abeta1-40-induced IL-6 secretion. Both statins potentiated the Abeta1-40-induced reduction in cell viability. The data indicate the importance of also considering the microglial responses to statins in evaluation of their effects in AD and other neurodegenerative disorders with an inflammatory component.     

High mobility group box protein-1 inhibits microglial Abeta clearance and enhances Abeta neurotoxicity

One pathogenic characteristic of Alzheimer's disease (AD) is the formation of extracellular senile plaques with accumulated microglia. According to the amyloid hypothesis, the increase or accumulation of amyloid-beta (Abeta) peptides in the brain parenchyma is the primary event that influences AD pathology. Although the role of microglia in AD pathology has not been clarified, their involvement in Abeta clearance has been noted. High mobility group box protein-1 (HMGB1) is an abundant nonhistone chromosomal protein. We reported recently that HMGB1 was associated with senile plaques and the total protein level significantly increased in AD brain. In this study, diffuse HMGB1 immunoreactivity was observed around dying neurons in the kainic acid- and Abeta1-42 (Abeta42)-injected rat hippocampi. HMGB1 also colocalized with Abeta in the Abeta42-injected rats but not in transgenic mice, which show massive Abeta production without neuronal loss in their brains. Furthermore, coinjection of HMGB1 delayed the clearance of Abeta42 and accelerated neurodegeneration in Abeta42-injected rats. These results suggest that HMGB1 released from dying neurons may inhibit microglial Abeta42 clearance and enhance the neurotoxicity of Abeta42. HMGB1 may thus be another target in the investigation of a therapeutic strategy for AD.     

Validation of ELISA methods for quantification of total tau and phosporylated-tau181 in human cerebrospinal fluid with measurement in specimens from two Alzheimer's disease studies

Pyrazole derivatives are well documented to possess anti-inflammatory activity but their effects on microglial activation are unknown. We determined the efficacy of the novel pyrazole compound 2-MBAPA (R/S-(±)-2-Methylbenzylamino 2-oxo-N-[4-cyano-1-phenyl-1H-pyrazol-5-yl] acetamide) on activated microglia under conditions relevant to inflammation in Alzheimer's disease (AD) brain. The compound at a non-toxic concentration inhibited secretion of tumor necrosis factor (TNF)-α by activated human microglia and attenuated toxicity of conditioned medium from activated human microglia towards human SH-SY5Y neuroblastoma cells in vitro. The 2-MBAPA neuroprotection was further demonstrated in vivo using an animal model of AD. The compound inhibited microgliosis, but not astrogliosis, in amyloid-β peptide (Aβ)(1-42)-injected rat brain. 2-MBAPA also diminished neuronal loss in the dentate gyrus caused by Aβ(1-42) injection. These results indicate that this novel pyrazole compound confers neuroprotection by inhibiting microglial activation. Therefore, further studies with 2-MBAPA and novel analogues based on this lead compound are warranted in an effort to develop new pharmacological agents that may be useful for slowing down progression of AD and other neuroinflammatory disorders associated with activated microglia.     

Red mold rice ameliorates impairment of memory and learning ability in intracerebroventricular amyloid beta-infused rat by repressing amyloid beta accumulation

Amyloid beta (Abeta) peptide related to the onset of Alzheimer's disease (AD) damaged neurons and further resulted in dementia. Monascus-fermented red mold rice (RMR), a traditional Chinese medicine as well as health food, includes monacolins (with the same function as statins) and multifunctional metabolites. In this study, ethanol extract of RMR (RE) was used to evaluate neuroprotection against Abeta40 neurotoxicity in PC12 cells. Furthermore, the effects of dietary administration of RMR on memory and learning abilities are confirmed in an animal model of AD rats infused with Abeta40 into the cerebral ventricle. During continuous Abeta40 infusion for 28 days, the rats of test groups were administered RMR or lovastatin. Memory and learning abilities were evaluated in the water maze and passive avoidance tasks. After sacrifice, cerebral cortex and hippocampus were collected for the examination of AD risk factors. The in vitro results clearly indicate that RE provides stronger neuroprotection in rescuing cell viability as well as repressing inflammatory response and oxidative stress. RMR administration potently reverses the memory deficit in the memory task. Abeta40 infusion increases acetylcholinesterase activity, reactive oxygen species, and lipid peroxidation and decreases total antioxidant status and superoxide dismutase activity in brain, but these damages were potently reversed by RMR administration, and the protection was more significant than that with lovastatin administration. The protection provided by RMR is able to prevent Abeta fibrils from being formed and deposited in hippocampus and further decrease Abeta40 accumulation, even though Abeta40 solution was infused into brain continuously.     

Decreased release of the angiogenic peptide vascular endothelial growth factor in Alzheimer's disease: recovering effect with insulin and DHEA sulfate

Changes of vascular endothelial growth factor (VEGF) secretion have recently been demonstrated in patients with Alzheimer's disease (AD). Since VEGF has been involved in brain angiogenesis, neuroprotection and cerebromicrovascular exchange of substrates and nutrients, the study of VEGF could have important relapses into the pathogenesis and treatment of AD. Within this context, 35 healthy subjects (16 of young and 19 of old age), 18 patients with dementia of the vascular type (VAD) and 22 with dementia of the Alzheimer's type (AD) were included in the study. VEGF levels were determined in the supernates of circulating natural killer (NK) immune cells isolated by immunomagnetic separation (pure CD16 + CD56 + NK cells at a final density of 7.75 x 10(6) cells/ml). VEGF was measured in spontaneous conditions (without modulation) and after exposure of NK cells with IL-2, lipopolysaccharide (LPS), dehydroepiandrosterone sulfate (DHEAS), LPS + insulin, amyloid-beta (Abeta) fragment 1-42, the inactive sequence Abeta(40-1) and Abeta(1-42) + insulin. A significant decrease in VEGF released by NK cells was demonstrated in AD subjects compared to the other groups. No differences of VEGF levels were found between healthy subjects of old age and the VAD group. The incubation with LPS and DHEAS significantly increased, in a dose-dependent manner, VEGF levels in AD as well as in healthy subjects of young and old age and in VAD patients. The incubation of NK cells with Abeta(1-42) completely suppressed VEGF generation in AD subjects, also reducing VEGF release in the other groups. The co-incubation of NK with LPS + insulin, at different molar concentrations, significantly restored (4- and 6-fold increase from LPS alone) VEGF in AD, also enhancing VEGF secretion in healthy subjects and the VAD group, while the co-incubation of NK with Abeta(1-42) + insulin promptly abolished the negative effects of Abeta(1-42) on VEGF release. These data might suggest that the decreased VEGF secretion by peripheral immune cells of AD subjects could have a negative role for brain angiogenesis, neuroprotection and for brain microvascular permeability to nutrients, increasing brain frailty towards hypoxic injuries. On the contrary, insulin and DHEAS could have beneficial effects in AD, as well as in VAD and in physiological aging, by increasing, in a dose-dependent fashion, VEGF availability by peripheral and resident immune and endothelial cells, so contributing to increase its circulating pool.     

Age-related changes in neuronal glucose uptake in response to glutamate and beta-amyloid

Energy supplies that may decline with age are crucial for cells to maintain ionic homeostasis and prevent neuron death. We examined baseline glucose transporter expression and rate of glucose uptake in cultured hippocampal neurons from embryonic, middle-age (12-month-old), and old (24-month-old) rats and exposed the neurons to glutamate, beta-amyloid, and mitochondrial inhibitors. Without stress, the rate of glucose uptake was similar in middle-age and old neurons, and the rate of glucose uptake in embryonic neurons was threefold greater than that in middle-age and old neurons. Glucose uptake increased in the presence of mitochondrial inhibitors (FCCP and oligomycin) for embryonic and middle-age neurons. The old neurons failed to increase glucose uptake. In the presence of glutamate, FCCP, and oligomycin, embryonic neurons showed a decrease in glucose uptake and the middle-age and old neurons showed no change in glucose uptake. Middle-age neurons took up significantly more glucose than old neurons when under mitochondrial and glutamate stress. In the presence of beta-amyloid, only embryonic neurons increased glucose uptake; middle-age and old neurons did not. Fluorescence imaging of immunoreactive glut3 in response to beta-amyloid demonstrated a 16-49% increase in glut3 immunoreactivity at the plasma membrane for the three ages. The results suggest that old neurons were not able to upregulate glucose uptake to ensure cell survival. Neuron aging does not indicate a defect in normal glut3 function; rather, our results suggest that mechanisms regulating glucose uptake under stress fail to react in time to ensure cell survival.     

P2X7 receptor modulation of beta-amyloid- and LPS-induced cytokine secretion from human macrophages and microglia

To test whether extracellular ATP can play a role in the neuroimmunopathology of Alzheimer's disease (AD), we evaluated the capacity of the ATP-binding purinoreceptor, P2X7, to modulate cytokine secretion on cultured human macrophages and microglia pre-activated 24 h with the 42 amino acid beta-amyloid peptide (Abeta(1-42)) or lipopolysaccharide. Thirty minutes of exposure to the selective P2X7 agonist 2'-3'-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP) resulted in the secretion of IL-1beta after either Abeta(1-42) or LPS stimulation of human macrophages that was dependent on the concentration of the stimulus used to pre-activate the cells. Further tests on human microglia treated with BzATP (300 microM) resulted in a 1.5- and 3.5-fold enhancement of IL-1alpha and IL-1beta secretion, respectively, from cells pre-activated by 10 microM Abeta(1-42) and a 1.6- and 3.9-fold enhancement of IL-1alpha and IL-1beta secretion, respectively, from cells pre-activated by 1 microg/ml LPS. BzATP induction of IL-1alpha and IL-1beta secretion from microglia was completely reversed by pre-incubation of the cells with the P2X7 antagonist, adenosine 5'-triphosphate 2',3'-acyclic dialcohol (oxidized ATP). In contrast to its effects on IL-1alpha and IL-1beta secretion, BzATP induced TNF-alpha after LPS stimulation, but not after stimulation with Abeta(1-42), induced IL-18 secretion regardless of whether microglia were pre-activated and attenuated IL-6 secretion after either LPS or Abeta(1-42) pre-activation. These results demonstrate that extracellular ATP can modulate Abeta-induced cytokine secretion from human macrophages and microglia and thus may play a role in the neuroimmunopathology of AD.     

RAGE-Abeta interactions in the pathophysiology of Alzheimer's disease

RAGE is a cell surface molecule primarily identified for its capacity to bind advanced glycation end-products and amphoterin. Immunocytochemical studies demonstrated that in Alzheimer's Disease (AD) the expression of RAGE is elevated in neurons close to neuritic plaque beta-amyloid (Abeta) deposits and in the cells of Abeta containing vessels. Cross-linking of surface bound Abeta 1-40 to endothelial cells, yielded a band of 50 kDa identified as RAGE. Using the soluble extracellular domain of recombinant human RAGE, we found that Abeta binds to RAGE with a Kd = 57 +/- 14 nM, a value close to those found for mouse brain endothelial cells and rat cortical neurons. The interaction of Abeta with RAGE in neuronal, endothelial, and RAGE-transfected COS-1 cells induced oxidative stress, as assessed by the TBARS and MTT assays. ELISA demonstrated a 2.5 times increase of RAGE in AD over control brains. Activated microglia also showed elevated expression of RAGE. In the BV-2 microglial cell line, RAGE bound Abeta in dose dependent manner with a Kd of 25 +/- 9 nM. Soluble Abeta induced the migration of microglia along a concentration gradient, while immobilized Abeta arrested this migration. Abeta-RAGE interaction also activated NF-kappaB, resulting in neuronal up-regulation of macrophage-colony stimulating factor (M-CSF) which also induced microglial migration. Taken together, our data suggest that RAGE-Abeta interactions play an important role in the pathophysiology of Alzheimer's Disease.     

CD40 promotion of amyloid beta production occurs via the NF-kappaB pathway

The CD40 receptor is a member of the tumor necrosis factor (TNF) super-family of trans-membrane receptors. Interaction of CD40 with its ligand CD40L mediates a broad range of immune and inflammatory responses in the periphery and in the central nervous system. Recently it has been suggested that CD40/CD40L interaction is involved in amyloid precursor protein (APP) processing and Alzheimer's disease (AD)-like pathology in transgenic mouse models of AD. We have previously shown that pharmacologically inhibiting CD40/CD40L interaction improves memory deficits in the PSAPP AD mouse model. We have also recently shown that CD40 deficiency mitigates amyloid deposition in APPsw and PSAPP mouse models. In the present report, using human embryonic kidney cells (HEK293) over-expressing both the APPsw mutation and CD40, we demonstrate that CD40/CD40L interaction directly increases the production of APP metabolites (Abeta 1-40, Abeta 1-42, CTFs, sAPPbeta and sAPPalpha). The results also show that CD40/CD40L interaction affects APP processing via the NF-kappaB pathway. Using NFkappaB inhibitors and SiRNAs to silence diverse elements of the NFkappaB pathway, we observe a reduction in levels of both Abeta 1-40 and Abeta 1-42. Taken together, our results further suggest that CD40L stimulation may be a key component in AD pathology and that elements of the NF-kappaB pathway may be suitable targets for therapeutic approaches against AD.     

Disruption of neurogenesis in the subventricular zone of adult mice,and in human cortical neuronal precursor cells in culture,by amyloid beta-peptide: implications for the pathogenesis of Alzheimer's disease

The adult mammalian brain contains populations of stem cells that can proliferate and then differentiate into neurons or glia. The highest concentration of such neural progenitor cells (NPC) is located in the subventricular zone (SVZ) and these cells can produce new olfactory bulb and cerebral cortical neurons. NPC may provide a cellular reservoir for replacement of cells lost during normal cell turnover and after brain injury. However, neurogenesis does not compensate for neuronal loss in age-related neurodegenerative disorders such as Alzheimer's disease (AD), suggesting the possibility that impaired neurogenesis contributes to the pathogenesis of such disorders. We now report that amyloid beta-peptide (Abeta), a self-aggregating neurotoxic protein thought to cause AD, can impair neurogenesis in the SVZ/cerebral cortex of adult mice and in human cortical NPC in culture. The proliferation and migration of NPC in the SVZ of amyloid precursor protein (APP) mutant mice, and in mice receiving an intraventricular infusion of Abeta, were greatly decreased compared to control mice. Studies of NPC neurosphere cultures derived from human embryonic cerebral cortex showed that Abeta can suppress NPC proliferation and differentiation, and can induce apoptosis. The adverse effects of Abeta on neurogenesis were associated with a disruption of calcium regulation in the NPC. Our data show that Abeta can impair cortical neurogenesis, and suggest that this adverse effect of Abeta contributes to the depletion of neurons and the resulting olfactory and cognitive deficits in AD.     

Interleukin-6, beta-amyloid peptide and NMDA interactions in rat cortical neurons

Neuronal damage in Alzheimer's disease (AD) is thought to involve direct toxicity of beta-amyloid peptide (Abeta) and excitotoxicity involving NMDA receptors (NMDARs) and altered Ca(2+) dynamics. Inflammation agents produced by microglia or astrocytes and associated with senile plaques such as the cytokine interleukin-6 (IL-6) could also contribute. To investigate this possibility, neuronal damage (lactate dehydrogenase assay, LDH, assay) was measured in cultures of rodent cortical neurons chronically treated with IL-6, Abeta or Abeta plus IL-6 and acutely treated with NMDA. Both Abeta and NMDA produced neuronal damage and this effect was larger with combined treatment. IL-6 did not produce significant neuronal damage but the largest neuronal damage was observed in cultures exposed to all three factors. IL-6 and Abeta enhanced Ca(2+) responses to NMDA and combined treatment produced the largest effect. These results are consistent with a role for interactions between Abeta, NMDA and IL-6 in the neuronal loss in AD.     

Modulation of calcium/calmodulin kinase-II provides partial neuroprotection against beta-amyloid peptide toxicity

Beta-amyloid (Abeta) peptide-induced neurotoxicity has been implicated in the pathogenesis of Alzheimer's disease (AD). The exact mechanism by which Abeta peptides trigger neuronal death is not well defined and may be related to an abrupt increase in intracellular calcium, leading to the activation of many pro-apoptotic pathways. While modulation of intracellular calcium increase receives much attention for pharmaceutical intervention, Ca2+-mediated pro-apoptotic signalling pathways have not been systematically studied. We have reported our study on the roles of calcium/calmodulin-dependent protein kinase II (CaMKII) in Abeta peptide neurotoxicity. By treating the primary cortical neurons exposed to Abeta peptides (Abeta(25-35) and Abeta(1-42)) with two selective CaMKII inhibitors, autocamtide-related inhibitory peptide (AIP) and KN93, Abeta peptide neurotoxicity was significantly reduced. Release of LDH and DNA fragmentation/condensation (by DAPI staining) in neurons exposed to Abeta peptides were significantly decreased in the presence of AIP and KN93. While these inhibitors significantly attenuated Abeta peptide-triggered activation of caspase-2 and caspase-3, and AIP significantly decreased the degree of tau phosphorylation of the Abeta peptide-treated neurons at early time, they could elicit partial neuroprotection only. Pharmacological inhibitor targeting calmodulin, W7, did not provide neuroprotection. Morphine, which activates CaMKII via micro receptors, augments Abeta-induced LDH release, caspase-2 and caspase-3 activities and neuronal apoptosis. Taken together, although CaMKII plays a role in Abeta peptide neurotoxicity, pharmacological inhibition cannot afford complete neuroprotection.     

β-amyloid protein structure determines the nature of cytokine release from rat microglia

Activated microglia represent a major source of inflammatory factors in Alzheimer's disease and a possible source of cytotoxic factors. beta-Amyloid (Abeta) peptide, the predominant component in amyloid plaques, has been shown to activate microglia and stimulate their production of inflammatory factors. The present study was performed to analyze the responses of microglia to different forms of Abeta, with regard to release of the proinflammatory cytokines interleukin-1alpha (IL-1alpha), IL-1beta, tumor necrosis factor-alpha (TNF-alpha), IL-6, and interferon-gamma (IFN-gamma), as well as the IL-1 receptor antagonist (IL-1ra). Primary cultures of microglia from rat neonatal cerebral cortex were incubated with freshly dissolved Abeta1-40 or Abeta1-42, Abeta1-40 fibrils, Abeta1-40 betaamy balls, or vehicle. Abeta1-40 fibrils did not significantly stimulate any of these cytokines. Freshly dissolved Abeta1-40 resulted in a marked increase in the release of IL-1beta, and freshly dissolved Abeta1-42 significantly stimulated both IL-1alpha and IFN-gamma secretion. The Abeta1-40 betaamy balls stimulated the secretion of IL-1alpha and IL-1beta. Incubation with Abeta peptides did not affect the secretion of IL-1ra, IL-6, or TNF-alpha. In the case of IL-1beta, the response is correlated with the presence of Abeta peptide as monomers and oligomers.     

Stimulation of PGE receptors EP2 and EP4 protects cultured neurons against oxidative stress and cell death following beta-amyloid exposure

Alzheimer's disease (AD) is associated with gliosis, neuroinflammation and higher levels of prostaglandins. Conflicting roles for cyclooxygenases and prostaglandins in the etiopathology of AD have been reported. We hypothesized that PGE2 signaling through EP2 and EP4 G-protein-coupled receptors could protect against amyloid beta-peptide (Abeta) neurotoxicity by increasing the cAMP signaling cascade. Using primary neuronal cultures, we investigated the presence of EP receptors (EP1-4) and the action of PGE2 and EP receptor agonists on neuronal susceptibility to Abeta1-42 toxicity. Low concentrations (1 microm) of PGE2, butaprost (EP2 agonist), and 1-hydroxy-PGE1 (EP4/EP3 agonist) were neuroprotective against Abeta1-42 toxicity, while sulprostone (EP3/EP1 agonist) at similar doses had no detectable effects. EP2 and EP4 receptor-mediated neuroprotection would involve changes in cAMP levels, as both EP2 and EP4 agonists increased intracellular cAMP concentration by approximately doubling basal levels, and both exhibited neuroprotective actions against Abeta-induced toxicity. The protein kinase A (PKA) inhibitor RpcAMPS significantly attenuated the neuroprotection by butaprost, but not that by 1-hydroxy-PGE1, implying differences between EP2 and EP4 receptor protective mechanisms. Additionally, the increase in reactive oxygen species generated following exposure to Abeta was reduced by stimulation of both EP2 and EP4 receptors. Together, these results indicate that PGE2 can protect neurons against Abeta toxicity by acting on given receptors and stimulating a cascade of intracellular events, including the cAMP-PKA pathway. We propose that development and testing of specific PGE2 receptor agonists downstream of cyclooxygenase could lead to therapeutic applications.     

Alzheimer's disease cybrids replicate beta-amyloid abnormalities through cell death pathways

Alzheimer's disease (AD) is characterized by the deposition in brain of beta-amyloid (Abeta) peptides, elevated brain caspase-3, and systemic deficiency of cytochrome c oxidase. Although increased Abeta deposition can result from mutations in amyloid precursor protein or presenilin genes, the cause of increased Abeta deposition in sporadic AD is unknown. Cytoplasmic hybrid ("cybrid") cells made from mitochondrial DNA of nonfamilial AD subjects show antioxidant-reversible lowering of mitochondrial membrane potential (delta(gYm), secrete twice as much Abeta(1-40) and Abeta(1-42), have increased intracellular Abeta(1-40) (1.7-fold), and develop Congo red-positive Abeta deposits. Also elevated are cytoplasmic cytochrome c (threefold) and caspase-3 activity (twofold). Increased AD cybrid Abeta(1-40) secretion was normalized by inhibition of caspase-3 or secretase and reduced by treatment with the antioxidant S(-)pramipexole. Expression of AD mitochondrial genes in cybrid cells depresses cytochrome c oxidase activity and increases oxidative stress, which, in turn, lowers delta(psi)m. Under stress, cells with AD mitochondrial genes are more likely to activate cell death pathways, which drive caspase 3-mediated Abeta peptide secretion and may account for increased Abeta deposition in the AD brain. Therapeutic strategies for reducing neurodegeneration in sporadic AD can address restoration of delta(psi)m and reduction of elevated Abeta secretion.