Soluble amyloid precursor protein alpha inhibits tau phosphorylation through modulation of GSK3ß signaling pathway.

We recently found that sAPPα decreases amyloid-beta generation by directly associating with ß-site amyloid precursor protein (APP)-converting enzyme 1 (BACE1), thereby modulating APP processing. Because inhibition of BACE1 decreases glycogen synthase kinase 3 beta (GSK3ß)-mediated Alzheimer's disease (AD)-like tau phosphorylation in AD patient-derived neurons, we determined whether sAPPα also reduces GSK3ß-mediated tau phosphorylation. We initially found increased levels of inhibitory phosphorylation of GSK3ß (Ser9) in primary neurons from sAPPα over-expressing mice. Further, recombinant human sAPPα evoked the same phenomenon in SH-SY5Y cells. Further, in SH-SY5Y cells over-expressing BACE1, and HeLa cells over-expressing human tau, sAPPα reduced GSK3ß activity and tau phosphorylation. Importantly, the reductions in GSK3ß activity and tau phosphorylation elicited by sAPPα were prevented by BACE1 but not γ-secretase inhibition. In accord, AD mice over-expressing human sAPPα had less GSK3ß activity and tau phosphorylation compared with controls. These results implicate a direct relationship between APP ß-processing and GSK3ß-mediated tau phosphorylation and further define the central role of sAPPα in APP autoregulation and AD pathogenesis.

Potential Autoepitope within the Extracellular Region of Contactin-Associated Protein-like 2 in Mice.

AIMS: Implicated in autoimmune encephalitis, neuromyotonia and genetic forms of autism, here we report that contactin-associated protein-like 2 (CNTNAP2) contains a potential autoepitope within the extracellular region. METHODOLOGY: CNTNAP2 sequence-similar regions (CSSRs) from human pathogens were identified. Sera from autistic and control children were obtained and analyzed for the presence of antibodies able to bind CSSRs. One such candidate CSSR was evaluated for evidence of autoimmune responses to CNTNAP2 in a mouse model of acute infection. RESULTS: Autistic and control children sera contained antibodies able to discrete regions of CNTNAP2. In a murine model of acute infection, a CSSR derived from the N-terminal extracellular region of CNTNAP2 resulted in anti-CNTNAP2 antibody production, proinflammatory cytokine elevation, cerebellar and cortical white matter T-cell infiltration as well as motor dysfunction. CONCLUSION: Taken together, these data suggest that CNTNAP2 contains a potential autoepitope within the extracellular region.

GFAP expression and social deficits in transgenic mice overexpressing human sAPPα.

Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed ß-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment.

Aberrant T-lymphocyte development and function in mice overexpressing human soluble amyloid precursor protein-α: implications for autism.

Abnormalities in T-lymphocyte populations and function are observed in autism. Soluble amyloid precursor protein α (sAPP-α) is elevated in some patients with autism and is known to be produced by immune cells. In light of the well-established role of sAPP-α in proliferation, growth, and survival of neurons, we hypothesized an analogous role in the immune system. Thus, we explored whether sAPP-α could modulate immune development and function, especially aspects of the pinnacle cell of the adaptive arm of the immune system: the T cell. To do this, we generated mice overexpressing human sAPP-α and characterized elements of T-cell development, signal transduction, cytokine production, and innate/adaptive immune functions. Here, we report that transgenic sAPP-α-overexpressing (TgsAPP-α) mice displayed increased proportions of CD8(+) T cells, while effector memory T cells were decreased in the thymus. Overall apoptotic signal transduction was decreased in the thymus, an effect that correlated with dramatic elevations in Notch1 activation; while active-caspase-3/total-caspase-3 and Bax/Bcl-2 ratios were decreased. Greater levels of IFN-γ, IL-2, and IL-4 were observed after ex vivo challenge of TgsAPP-α mouse splenocytes with T-cell mitogen. Finally, after immunization, splenocytes from TgsAPP-α mice displayed decreased levels IFN-γ, IL-2, and IL-4, as well as suppressed ZAP70 activation, after recall antigen stimulation. Given elevated levels of circulating sAPP-α in some patients with autism, sAPP-α could potentially drive aspects of immune dysfunction observed in these patients, including dysregulated T-cell apoptosis, aberrant PI3K/AKT signaling, cytokine alterations, and impaired T-cell recall stimulation.

Memantine and catatonia: a case report and literature review.

Catatonia is a movement disorder with various possible etiologies. The majority of cases are associated with an underlying mood or psychotic disorder, while others are caused by medical conditions. Currently, benzodiazepines are the first-line psychopharmacologic agents in the treatment of catatonia. However, several cases have been reported in which treatment with memantine proved to be effective. We present the case of a 92-year-old female with major depressive disorder and associated catatonic symptoms. In this case, the patient's symptoms remitted quickly after the initiation of memantine. We review the possible causes of catatonia and pharmacologic treatments for the condition and highlight the possible benefits of N-methylD-aspartic acid receptor antagonists such as memantine in the treatment of catatonia.

Antiretroviral medications disrupt microglial phagocytosis of ß-amyloid and increase its production by neurons: implications for HIV-associated neurocognitive disorders.

Up to 50% of long-term HIV infected patients, including those with systemically well-controlled infection, commonly experience memory problems and slowness, difficulties in concentration, planning, and multitasking. Deposition of Aß plaques is also a common pathological feature of HIV infection. However, it is not clear whether this accumulation is due to AD-like processes, HIV-associated immunosuppression, Tat protein-induced Aß elevations, and/or the effects of single highly active antiretroviral therapy (ART). Here we evaluated the effects of several ART medications (Zidovudine, Lamivudine, Indinavir, and Abacavir) alone and in combination on: 1) Aß1-40, 42 generation in murine N2a cells transfected with the human "Swedish" mutant form of APP; 2) microglial phagocytosis of FITC-Aß1-42 peptides in cultured murine N9 microglia. We report for the first time that these antiretroviral compounds (10 µM) generally increase Aß generation (~50-200%) in SweAPP N2a cells and markedly inhibit microglial phagocytosis of FITC-Aß1-42 peptides in murine microglia. The most significant amyloidogenic effects were observed with combined ART (p < 0.05); suggesting certain ART medications may have additive amyloidogenic effects when combined. As these antiretroviral compounds are capable of penetrating the blood brain barrier and reaching the concentrations employed in the in vitro studies, these findings raise the possibility that ART may play a casual role in the elevated Aß found in the brains of those infected with HIV. Therefore these compounds may consequently contribute to cognitive decline observed in HIV associated neurocognitive disorders (HAND).

Flipping the switches: CD40 and CD45 modulation of microglial activation states in HIV associated dementia (HAD).

Microglial dysfunction is associated with the pathogenesis and progression of a number of neurodegenerative disorders including HIV associated dementia (HAD). HIV promotion of an M1 antigen presenting cell (APC) - like microglial phenotype, through the promotion of CD40 activity, may impair endogenous mechanisms important for amyloid- beta (Aß) protein clearance. Further, a chronic pro-inflammatory cycle is established in this manner. CD45 is a protein tyrosine phosphatase receptor which negatively regulates CD40L-CD40-induced microglial M1 activation; an effect leading to the promotion of an M2 phenotype better suited to phagocytose and clear Aß. Moreover, this CD45 mediated activation state appears to dampen harmful cytokine production. As such, this property of microglial CD45 as a regulatory "off switch" for a CD40-promoted M1, APC-type microglia activation phenotype may represent a critical therapeutic target for the prevention and treatment of neurodegeneration, as well as microglial dysfunction, found in patients with HAD.

ADAM10 activation is required for green tea (-)-epigallocatechin-3-gallate-induced alpha-secretase cleavage of amyloid precursor protein.

Recently, we have shown that green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) exerts a beneficial role on reducing brain Abeta levels, resulting in mitigation of cerebral amyloidosis in a mouse model of Alzheimer disease. EGCG seems to accomplish this by modulating amyloid precursor protein (APP) processing, resulting in enhanced cleavage of the alpha-COOH-terminal fragment (alpha-CTF) of APP and corresponding elevation of the NH(2)-terminal APP product, soluble APP-alpha (sAPP-alpha). These beneficial effects were associated with increased alpha-secretase cleavage activity, but no significant alteration in beta-or gamma-secretase activities. To gain insight into the molecular mechanism whereby EGCG modulates APP processing, we evaluated the involvement of three candidate alpha-secretase enzymes, a-disintegrin and metalloprotease (ADAM) 9, 10, or 17, in EGCG-induced non-amyloidogenic APP metabolism. Results show that EGCG treatment of N2a cells stably transfected with "Swedish" mutant human APP (SweAPP N2a cells) leads to markedly elevated active ( approximately 60 kDa mature form) ADAM10 protein. Elevation of active ADAM10 correlates with increased alpha-CTF cleavage, and elevated sAPP-alpha. To specifically test the contribution of ADAM10 to non-amyloidogenic APP metabolism, small interfering RNA knockdown of ADAM9, -10, or -17 mRNA was employed. Results show that ADAM10 (but not ADAM9 or -17) is critical for EGCG-mediated alpha-secretase cleavage activity. In summary, ADAM10 activation is necessary for EGCG promotion of non-amyloidogenic (alpha-secretase cleavage) APP processing. Thus, ADAM10 represents an important pharmacotherapeutic target for the treatment of cerebral amyloidosis in Alzheimer disease.

Rosiglitazone reduces serum homocysteine levels, smooth muscle proliferation, and intimal hyperplasia in Sprague-Dawley rats fed a high methionine diet.

Homocysteine (Hcy) is a metabolite of the essential amino acid methionine. Hyperhomocysteinemia is associated with vascular disease, particularly carotid stenosis. Rosiglitazone, a ligand of the peroxisome proliferator-activated receptor gamma , attenuates balloon catheter-induced carotid intimal hyperplasia in type 2 diabetic rats. We studied 4 groups (n = 7 per group) of adult female Sprague-Dawley rats fed (a) powdered laboratory chow (control), (b) control diet with rosiglitazone (3.0 mg/kg/d), (c) diet containing 1.0% l -methionine, and (d) diet containing methionine and rosiglitazone. After 1 week on high methionine diet, the rats were administered an aqueous preparation of rosiglitazone by oral gavage. One week after initiation of rosiglitazone, balloon catheter injury of the carotid artery was carried out using established methods, and the animals continued on their respective dietary and drug regimens for another 21 days. At the end of the experimental period, blood samples were collected, and carotid arteries and liver were harvested. Serum Hcy increased significantly on methionine diet compared with controls (28.9 +/- 3.2 vs 6.3 +/- 0.04 micromol/L). Development of intimal hyperplasia was 4-fold higher in methionine-fed rats; this augmentation was significantly reduced ( P < .018) in rosiglitazone-treated animals. Rosiglitazone treatment significantly ( P < .001) suppressed Hcy levels and increased the activity of the Hcy metabolizing enzyme, cystathionine-beta-synthase in the liver samples. Hcy (100 micromol/L) produced a 3-fold increase in proliferation of rat aortic vascular smooth muscle cells; this augmentation was inhibited by incorporating rosiglitazone (10 micromol/L). After balloon catheter injury to the carotid artery of animals on a high methionine diet, there was an increase in the rate of development of intimal hyperplasia consistent with the known effects of Hcy. It is demonstrated for the first time that the peroxisome proliferator-activated receptor gamma agonist rosiglitazone can attenuate the Hcy-stimulated increase in the rate of development of intimal hyperplasia indirectly by increasing the rate of catabolism of Hcy by cystathionine-beta-synthase and directly by inhibiting vascular smooth muscle cell proliferation. These findings may have important implications for the prevention of cardiovascular disease and events in patients with hyperhomocysteinemia (HHcy).

Inhibition of angiogenesis by Abeta peptides.

Abeta peptides are naturally occurring peptides forming beta-sheet aggregates that constitute an integral component of senile plaques and vascular deposits in Alzheimer's disease. Since several peptides adopting a beta-sheet conformation have been shown to be anti-angiogenic, we investigated the effect of Abeta on angiogenesis. We show that in vitro, Abeta dose-dependently inhibits the formation of capillaries by human brain endothelial cells plated on Matrigel and stimulates capillary degeneration at high doses. Preparations of Abeta peptides containing a higher content of beta-sheet structures are more potently anti-angiogenic in vitro. Ex vivo, Abeta dose-dependently opposes angiogenesis in rat aortae and in human middle cerebral arteries. In vivo, Abeta dose dependently inhibits angiogenesis in the chick chorioallantoic membrane assay and suppresses bFGF-induced blood vessel formation in the corneal micropocket and Matrigel plug assays. Since angiogenesis is required for tumor growth, we explored the effect of Abeta on human glioblastoma (U87MG) and human lung adenocarcinoma (A549) tumors. We show that intra-tumoral injection of Abeta potently inhibits the growth and vascularization of human glioblastoma and human lung adenocarcinoma tumor xenografts in nude mice. Similarly to the intra-tumoral injection regimen, Abeta delivered intraperitoneally also suppressed the growth of human lung adenocarcinoma tumor xenografts. Altogether our data show that Abeta is an angiogenesis inhibitor.

Pro-inflammatory effect of freshly solubilized beta-amyloid peptides in the brain.

It has recently been shown that the level of soluble beta-amyloid (Abeta) peptides correlates well with the severity of synaptic loss and the density of neurofibrillary tangles observed in Alzheimer's disease (AD) brain. However, the biological activity of soluble forms of Abeta peptides in the brain remains to be determined. We have investigated ex vivo the effect of freshly solubilized Abeta1-40 peptides (fsAbeta) on prostaglandin E2 (PGE2) production in rat brain slices. PGE2 levels increased rapidly following treatment with fsAbeta, an effect that was prevented by SB202190, a selective inhibitor of p38 mitogen-activated protein kinase (p38 MAPK), and by NS-398, which preferentially inhibits cyclooxygenase-2 (COX-2) compared to COX-1. In an attempt to determine the cellular systems of the brain responsible for prostaglandin production in response to fsAbeta, the effect of fsAbeta was tested on isolated brain microvessels, primary cultures of brain smooth muscle cells/pericytes and endothelial cells, and a human neuron-like cell line (IMR32). Our data show that fsAbeta ex vivo can stimulate prostaglandin accumulation in incubates of isolated rat brain microvessels. In addition, fsAbeta appears to cause a concentration-dependent enhancement of prostaglandin accumulation in primary cultures of brain microvessel-derived smooth muscle cells/pericytes but not of brain endothelial cells. Finally, fsAbeta also stimulated PGF2alpha accumulation in cultures of differentiated IMR32 cells, but to a lesser extent than in brain smooth muscle cell/pericyte cultures. Deposition of aggregated forms of Abeta in the brain has been thought to trigger an inflammatory response which accompanies the neuropathologic events of AD. Our data provide evidence that fsAbeta triggers a pro-inflammatory reaction in rat brain, and suggest that the cerebrovasculature may constitute an important source of pro-inflammatory eicosanoids.

Statins inhibit A beta-neurotoxicity in vitro and A beta-induced vasoconstriction and inflammation in rat aortae.

Freshly solubilized A beta peptides synergistically increase the magnitude of the constriction induced by endothelin-1 (ET-1), via the activation of a pro-inflammatory pathway. We report that mevinolin and mevastatin, two inhibitors of the 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase are able to completely abolish the vasoactive properties of A beta in rat aortae. Mevinolin also appears to oppose the increased vascular reactivity to ET-1 induced by interleukin 1-beta and phospholipase A(2) suggesting that statins display some anti-inflammatory properties. We show that freshly solubilized A beta stimulates prostaglandin E(2) and F(2 alpha) production (by 6 and 3.6 times, respectively) in isolated rat aortae and that mevinolin completely antagonizes this effect confirming the anti-inflammatory action of mevinolin ex vivo in rat aortae. In addition, we observed that A beta vasoactivity is not mediated nor modulated by mevalonic acid suggesting that the anti-inflammatory action of the statins are not related to an inhibition of HMG-CoA reductase activity. Differentiated human neuroblastoma cells (IMR32) were used to assess the neurotoxic effect of pre-aggregated A beta by quantifying the release of lactate dehydrogenase (LDH) in the cell culture medium. A beta appears to enhance LDH release by 30% in IMR32 cells, an effect that can be completely opposed by mevastatin. Taken together these data show that statins can antagonize the effect of A beta in different assays and provide new clues to understand the prophylactic action of the statins against Alzheimer's disease.