Carola Otth

Microbes and Alzheimer's Disease

We are researchers and clinicians working on Alzheimer’s disease (AD) or related topics, and we write to express our concern that one particular aspect of the disease has been neglected, even thoug ...

AβPP induces cdk5-dependent tau hyperphosphorylation in transgenic mice Tg2576

Previous studies of Abeta-induced neuronal damage of hippocampal cells in culture have provided strong evidence that deregulation of the Cdk5/p35 kinase system is involved in the neurodegeneration pathway. Cdk5 inhibitors and antisense probes neuroprotected hippocampal cells against the neurotoxic action of Abeta. To further investigate the mechanisms underlying the participation of Cdk5 in neuronal degeneration, the transgenic mouse containing the Swedish mutations, Tg2576, was used as an animal model. Immunocytochemical studies using anti-Abeta(1-17) antibody evidenced the presence of labeled small-clustered core plaques in the hippocampus and cortex of 18-month-old transgenic mice brains. The loss of granular cells without a compressed appearance was detected in the vicinity of the cores in the dentate gyrus of the hippocampus. Immunostaining of Tg2576 brain sections with antibodies AT8, PHF1 and GFAP labeled punctuate dystrophic neurites in and around the amyloid core. Reactive astrogliosis around the plaques in the hippocampus was also observed. Studies at the molecular level showed differences in the expression of the truncated Cdk5 activator p25 in the transgenic animal, as compared with wild type controls. However no differences in Cdk5 levels were detected, thus corroborating previous cellular findings. Interestingly, hyperphosphorylated tau epitopes were substantially increased as assessed with the AT8 and PHF1 antibodies, in agreement with the observation of a p25 increase in the transgenic animal. These observations strongly suggest that the increased exposure of Alzheimers type tau phosphoepitopes in the transgenic mice correlated with deregulation of Cdk5 leading to an increase in p25 levels. These studies also provide further evidence on the links between extraneuronal amyloid deposition and tau pathology.

Neuronal Cytoskeletal Dynamic Modification and Neurodegeneration Induced by Infection with Herpes Simplex Virus Type 1

Herpes simplex virus type 1 (HSV-1) and herpes simplex virus type 2 (HSV-2) belong to the family Herpesviridae, the subfamily Alphaherpesvirinae, and the genus Simplexvirus. They are ubiquitous, neurotropic, and the most common pathogenic cause of sporadic acute encephalitis in humans. Herpes simplex encephalitis (HSE) is associated with a high mortality rate and significant neurological, neuropsychological, and neurobehavioral sequelae, which afflict patients for life. HSV-1 has been suggested as an environmental risk factor for Alzheimers disease. However, the mechanisms involved in HSV-1 infection that may trigger the neurodegenerative process are still unknown. In general, HSV-1 induced cytoskeletal alterations reported to date involve the overall disruption of one or more elements of the cytoskeleton in cell lines. Axonal injury has recently attracted attention as a key predictor for the outcome of a number of brain disorders. Here we show that infection of mice neuronal cultures with HSV-1 result in marked neurite damage and neuronal death. Furthermore, in this in vitro model of infection, neurons manifested considerable alterations in microtubule dynamics and tau hyperphosphorylation. These results suggest a possible link between HSV-1 infection and neuronal cytoskeletal disruption.

Tau cleavage at D421 by caspase-3 is induced in neurons and astrocytes infected with herpes simplex virus type 1.

Herpes Simplex Virus Type 1 (HSV-1) is ubiquitous, neurotropic, and the most common pathogenic causes of sporadic acute encephalitis in humans. Herpes simplex encephalitis is associated with a high mortality rate and significant neurological, neuropsychological, and neurobehavioral sequelae, which afflict patients for life. HSV-1 infects limbic system structures in the central nervous system and has been suggested as an environmental risk factor for Alzheimers disease. However, the possible mechanisms that link HSV-1 infection with the neurodegenerative process are still largely unknown. In a previous study we demonstrated that HSV-1 triggers hyperphosphorylation of tau epitopes serine202/threonine205 and serine396/serine404 in neuronal cultures, resembling what occurs in neurodegenerative diseases. Therefore, the aim of the present study was to evaluate at the cellular level if another event associated with neurodegeneration, such as caspase-3 induced cleavage of tau, could also be triggered by HSV-1 infection in primary neuronal and astrocyte cultures. As expected, induction of caspase-3 activation and cleavage of tau protein at its specific site (aspartic acid 421) was observed by Western blot and immunofluorescence analyses in mice neuronal primary cultures infected with HSV-1. In agreement with our previous study on tau hyperphosphorylation, tau cleavage was also observed during the first 4 hours of infection, before neuronal death takes place. This tau processing has been previously demonstrated to increase the kinetics of tau aggregation in vitro and has also been observed in neurodegenerative pathologies. In conclusion, our findings support the idea that HSV-1 could contribute to induce neurodegenerative processes in age-associated pathologies such as Alzheimers disease.

Interleukin-3 prevents neuronal death induced by amyloid peptide

BackgroundInterleukin-3 (IL-3) is an important glycoprotein involved in regulating biological responses such as cell proliferation, survival and differentiation. Its effects are mediated via interaction with cell surface receptors. Several studies have demonstrated the expression of IL-3 in neurons and astrocytes of the hippocampus and cortices in normal mouse brain, suggesting a physiological role of IL-3 in the central nervous system. Although there is evidence indicating that IL-3 is expressed in some neuronal populations, its physiological role in these cells is poorly known.ResultsIn this study, we demonstrated the expression of IL-3 receptor in cortical neurons, and analyzed its influence on amyloid β (Aβ)-treated cells. In these cells, IL-3 can activate at least three classical signalling pathways, Jak/STAT, Ras/MAP kinase and the PI 3-kinase. Viability assays indicated that IL-3 might play a neuroprotective role in cells treated with Aβ fibrils. It is of interest to note that our results suggest that cell survival induced by IL-3 required PI 3-kinase and Jak/STAT pathway activation, but not MAP kinase. In addition, IL-3 induced an increase of the anti-apoptotic protein Bcl-2.ConclusionAltogether these data strongly suggest that IL-3 neuroprotects neuronal cells against neurodegenerative agents like Aβ.

Fibrillar amyloid-β1-42 modifies actin organization affecting the cofilin phosphorylation state: a role for Rac1/cdc42 effector proteins and the slingshot phosphatase.

The neuronal cytoskeleton regulates numerous processes that occur in normal homeostasis. Under pathological conditions such as those of Alzheimers disease (AD), major alterations in cytoskeleton organization have been observed and changes in both microtubules and actin filaments have been reported. Many neurodegenerative consequences of AD are linked to the production and accumulation of amyloid peptides (Aβ) and their oligomers, produced from the internal cleavage of the amyloid-β protein precursor. We previously reported that fibrillar Aβ1-42 (fAβ) treatment of hippocampal neurons induced an increase in Rac1 and Cdc42 activities linking fAβ effects with changes in actin dynamics. Here we show fAβ-induces increased activity of PAK1 and cyclin-dependent kinase 5, and that p21-activated kinase (PAK1) activation targets the LIMK1-cofilin signaling pathway. Increased cofilin dephosphorylation under conditions of enhanced LIM-Kinase 1 (LIMK1) activity suggests that fAβ co-stimulates bifurcating pathways impacting cofilin phosphorylation. Overexpression of slingshot (SSH) prevents the augment of F-actin induced by fAβ after 24 h, suggesting that fAβ-induced changes in actin assembly involve both LIMK1 and SSH. These results suggest that fAb may alter the PAK1/LIMK1/cofilin axis and therefore actin organization in AD.

Novel identification of peripheral dopaminergic D2 receptor in male germ cells

Dopamine is a recognized modulator in the central nervous system (CNS) and peripheral organ functions. The presence of peripheral dopamine receptors outside the CNS has suggested an intriguing interaction between the nervous system and other functional systems, such as the reproductive system. In the present study we analyzed the expression of D2R receptors in rat testis, rat spermatogenic cells and spermatozoa, in different mammals. The RT‐PCR analysis of rat testis mRNA showed specific bands corresponding to the two dopamine receptor D2R (L and S) isoforms previously described in the brain. Using Western blot analysis, we confirmed that the protein is present in rat testis, isolated spermatogenic cells and also in spermatozoa of a range of different mammals, such as rat, mouse, bull, and human. The immunohistochemistry analysis of rat adult testis showed that the receptor was expressed in all germ cells (pre‐ and post‐meiotic phase) of the tubule with staining predominant in spermatogonia. Confocal analysis by indirect immunofluorescence revealed that in non‐capacitated spermatozoa of rat, mouse, bull, and human, D2R is mainly localized in the flagellum, and is also observed in the acrosomal region of the sperm head (except in human spermatozoa). Our findings demonstrate that the two D2 receptor isoforms are expressed in rat testis and that the receptor protein is present in different mammalian spermatozoa. The presence of D2R receptors in male germ cells implies new and unsuspected roles for dopamine signaling in testicular and sperm physiology. J. Cell. Biochem. 100: 141–150, 2007.

Inflammatory and Neurodegeneration Markers during Asymptomatic HSV-1 Reactivation

BACKGROUND Currently, it is unclear whether asymptomatic recurrent reactivations of herpes simplex virus type 1 (HSV-1) occur in the central nervous systems of infected people, and if these events could lead to a progressive deterioration of neuronal function. In this context, HSV-1 constitutes an important candidate to be included among the risk factors for the development of neuropathies associated with chronic neuroinflammation. OBJECTIVE The aim of this study was to assess in vivo inflammatory and neurodegenerative markers in the brain during productive and latent HSV-1 infection using a mouse model of herpes simplex encephalitis. METHODS Neuroinflammation and neurodegeneration markers were evaluated in mice trigeminal ganglia and cerebral cortex during HSV-1 infection, by immunohistochemistry, western blot, and RT-PCR. RESULTS Neuronal ICP4 viral antigen expression indicative of a reactivation episode during asymptomatic latency of HSV-1 infection in mice was accompanied by upregulation of neuroinflammatory (toll-like receptor-4, interferon α/β, and p-IRF3) and early neurodegenerative markers (phospho-tau and TauC3). CONCLUSIONS HSV-1 reactivation from latency induced neuroinflammatory and neurodegenerative markers in the brain of asymptomatic mice suggesting that recurrent reactivations could be associated with cumulative neuronal dysfunctions.

Nutraceutical activators of AMPK/Sirt1 axis inhibit viral production and protect neurons from neurodegenerative events triggered during HSV-1 infection

Herpes simplex virus type-1 (HSV-1) is ubiquitous and is able to establish a lifelong persistent latent infection in neurons of infected individuals. It has been estimated that in approximately 70% of the population over 50 years old, the virus enters the brain and infects neurons, and possibly undergoes recurrent reactivation episodes during lifetime, especially in immunodepressed individuals. We previously showed that the sensors AMP-dependent kinase (AMPK) and Sirtuin 1 (Sirt1), involved in survival pathways and neuroprotection, were affected during the course of HSV-1 infection. To evaluate if natural activators of the AMPK/Sirt1 axis, such as Resveratrol and Quercetin could reduce viral propagation and/or counteract the effects of neuronal infection, we analyzed progeny virion production, neuronal viability and neurodegenerative events during HSV-1 infection. We found that the activators of AMPK/Sirt1 axis, increased the viability of infected neurons, significantly reduced the viral titer in the supernatant and the expression of viral genes. More importantly, pretreatment of neurons with Resveratrol or Quercetin significantly reduced the levels of caspase-3 cleaved- and hyperphosphorylated tau associated with HSV-1 infection. These results suggest that activators of the AMPK/Sirt1 axis could be potentially useful in reducing the risk of HSV-1 productive infection in neurons and the cellular damage associated with reactivation episodes.

Mechanisms associated with phagocytosis of Arcobacter butzleri by Acanthamoeba castellanii

Acanthamoeba castellanii is a free-living amoeba widely found in environmental matrices such as soil and water. Arcobacter butzleri is an emerging potential zoonotic pathogen that can be isolated from environmental water sources, where they can establish endosymbiotic relationships with amoebas. The aim of this study was to describe the implication of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoebic membrane, during the attachment of Arcobacter butzleri by blocking with different saccharides. Another objective was to describe the signaling pathways involved in phagocytosis of these bacteria using specific inhibitors and analyze the implication of phagolysosome formation on the survival of Arcobacter butzleri inside the amoeba. We infer that the attachment of Arcobacter butzleri to the amoeba is a process which involves the participation of mannose-binding proteins and membrane-associated receptors of glucose and galactose present in the amoeba. We also demonstrated an active role of protozoan actin polymerization in the phagocytosis of Arcobacter butzleri and a critical involvement of PI3K and RhoA pathways. Further, we demonstrated that the tyrosine kinase-induced actin polymerization signal is essential in Acanthamoeba-mediated bacterial uptake. Through phagolysosomal formation analysis, we conclude that the survival of Arcobacter butzleri inside the amoeba could be related with the ability to remain inside vacuoles not fused with lysosomes, or with the ability to retard the fusion between these structures. All these results help the understanding of the bacterial uptake mechanisms used by Acanthamoeba castellanii and contribute to evidence of the survival mechanisms of Arcobacter butzleri.