Understanding microglial responses in the frontal cortex of alzheimer´s disease patients

Microglial cells, the immune cells of the brain, and the neuroinflammatory process associated, have been postulated as a critical factor in AD pathogenesis, since the identification of genetic risk factors related to microglial function. However, the microglial role in the development/progression of AD has not been determined yet. In this sense, we have previously reported a limited activation and microglial degeneration in the hippocampus of AD patients in contrast to the proinflammatory view based on findings in amyloidogenic models. Here, we have further analyzed the functional/phenotypic profile displayed by microglial cells in other vulnerable brain region of AD patients, the frontal cortex. Immunohistochemistry and image analysis approaches were performed in the frontal cortex of post mortem samples from controls (Braak 0-II) and AD patients (Braak V-VI) including familial cases. Microglia of Braak V-VI individuals were observed forming clusters and showed, both plaque (Iba1+/TMEM119+/P2ry12-/CD45high/Trem2+) and inter-plaque (Iba1+/ TMEM119+/P2ry12-/CD45high/Trem2-) microglial activation, similar that observed in amyloidogenic mice. By contrast, homeostatic and ramified microglial cells of non-demented Braak II cases presented Iba1+/P2ry12+/TMEM119+/CD45low/Trem2- profile. Furthermore, different microglial responses were observed between sporadic and familial AD cases. These different microglial phenotypes associated with AD pathology show the heterogeneity and complexity of the microglial phenotypes and suggest different functional states of these glial cells in a region-specific manner. These data need to be considered for better understand the immunological mechanisms underlying AD progression. Modulating brain inflammatory responses might be a promising avenue to prevent cognitive dysfunction in AD patients. ISCiii:PI18/01557(AG)-PI18/01556(JV);Junta Andalucia:UMA18-FEDERJA211(AG). All cofinanced by FEDER funds (European-Union).Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Microglial response differences between amyloidogenic transgenic models and Alzheimer’s disease patients

Aims: The continuing failure to develop an effective treatment for Alzheimer’s disease (AD) reveals the complexity for AD pathology. Increasing evidence indicates that neuroinflammation involving particularly microglial cells contributes to disease pathogenesis. Here we analyze the differences in the microglial response between APP/PS1 model and human brains. Methods: RT-PCR, western blots, and immunostaining were performed in the hippocampus of human post mortem samples (from Braak II to Braak V-VI) and APP751SL/PS1M146L mice. In vitro studies to check the effect of S1 fractions on microglial cells were assayed. Results: In APP based models the high Abeta accumulation triggers a prominent microglial response. On the contrary, the microglial response detected in human samples is, at least, partial or really mild. This patent difference could simple reflect the lower and probably slower Abeta production observed in human hippocampal samples, in comparison with models or could reflect the consequence of a chronic long-standing microglial activation. However, beside this differential response, we also observed a prominent microglial degenerative process in Braak V-VI samples that, indeed, could compromise their normal role of surveying the brain environment and respond to the damage. This microglial degeneration, particularly relevant at the dentate gyrus of the hippocampal formation, might be mediated by the accumulation of toxic soluble phospho-tau species. Conclusions: These differences need to be considered when delineating animal models that better integrate the complexity of AD pathology and, therefore, guarantee clinical translation. Correcting dysregulated brain inflammatory responses might be a promising avenue to restore cognitive function. Supported by grants FIS PI15/00796 and FIS PI15/00957 co-financed by FEDER funds from European Union, and by Junta de Andalucia Proyecto de Excelencia CTS385 2035.Financiado por FIS PI15/00796 y FIS PI15/0095, cofinanciado por los fondos FEDER de la Unión Europea, y por Junta de Andalucia Proyecto de Excelencia CTS385 2035. Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Lithium, as a neuroprotective therapy for Alzheimer’s disease pathology, modifies abeta plaque toxicity

BACKGROUND: Despite the relatively large information about the Alzheimer’s disease (AD) pathology, no effective disease-modifying treatment has been yet developed. Lithium, a primary drug to treat bipolar disorder, has been suggested as a potential treatment against AD. In this work we have evaluated whether lithium treatment could ameliorate the neuropathology progression of the transgenic PS1M146L/APPSwe-London mice. Unlike most transgenic animal models, which do not exhibit the neurodegenerative spectrum of disease observed in the patient population, this AD model exhibits a prominent amyloid pathology along with a selective and significant neuronal loss in the hippocampus and entorhinal cortex. Therefore, this model is highly valuable for evaluating the effectiveness of potential neuroprotective therapies for AD. METHODS: For lithium treatment, PS1/APP mice (3 month old at the beginning of treatment) were fed, ad libitum, with diet supplemented with lithium carbonate (1.2g/kg, Harlan, Spain). The treatment lasts 6 months. After behavioural studies, mice were anesthetized and brains dissected out (hippocampus and cortex). Hemibrains were processed for immunohistochemistry, stereological and image analysis quantification, and the other hemibrains for RT-PCR and Western blot studies. RESULTS: Our data demonstrate that chronic oral administration of lithium, before the pathology onset, resulted in less toxic plaque formation that significantly ameliorated the degenerative processes and behavioural/memory deficits occurring during disease progression in our PS1/APP model. Specifically, and of great relevance for AD prevention, early lithium intervention was able to arrest neuronal loss in hippocampus and entorhinal cortex of highly vulnerable populations. Besides, lithium reduced the axonal dystrophic pathology, associated to amyloid plaques, by increasing the Abeta compaction. Moreover, a significant lower accumulation of phospho-tau, LC3-II and ubiquitinated proteins was detected. Our study highlights that the switch of plaque quality by lithium could be mediated by astrocyte activation and the release of heat shock proteins, which concentrated in the core of the plaques.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Disentangling the contribution of tau and abeta pathologies in transgenic models of Alzheimer's disease

AIMS: Amyloid-beta (Abeta) deposits and intraneuronal hyperphosphorylated tau are major pathological hallmarks of Alzheimer’s disease (AD). The coexistence of these aggregates in AD brains leads to synaptic dysfunction, neuronal loss and cognitive decline. Failures in protein homeostasis, along with defective glial responses, have been identified as pathological mechanisms linked to this disorder. Thus, our main objective is to better understand the differential impact of Abeta- and tau-aggregates to these processes in the hippocampus of AD models. METHODS: We analyzed APP- (APPSL/PS1M146L) and Tau- (ThyTau22 and hP301S) based models from 2 to 18 months of age. Tau and Abeta pathologies were assessed by western blotting and immunohistochemistry. Confocal microscopy was used to study microglia/aggregates relationship. Levels of synaptic proteins, autophagy and inflammatory markers were determined by quantitative PCR, WB and immunohistochemistry. RESULTS: Tau and Abeta pathologies initiated as early as 2 months of age and increased progressively with aging. Even though only APP/PS1 hippocampus showed dystrophic neurites positive to proteostatic and presynaptic markers, their protein levels were altered in both types of models from 6-9 months compared to age-matched WT mice. Inflammatory markers and microglial reactivity were barely increased in the hippocampus of ThyTau mice in contrast to P301S and APP/PS1 mice which displayed a prominent microglial response. CONCLUSIONS: Clarifying the effects of Abeta and tau separately would indeed enable the development of novel therapeutic strategies and drugs targeting pathways related to these proteinopathies. Supported by grants FIS PI15/00796 and PI15/00957 co-financed by FEDER funds from European Union, by Junta de Andalucia Proyecto de Excelencia CTS385 2035 and by grant PPIT.UMA.B1/2017.26Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tec

Analyzing hippocampal synaptic damage and glial response in a mouse model of tauopathy

Tau pathology is highly related to synaptic and neuronal loss, leading to cognitive decline and dementia in Alzheimer’s disease (AD) and other tauopathies. Tau transgenic mice are widely used to investigate the specific contribution of this protein to AD since they reproduce the synaptic and cognitive dysfunction in parallel to an age-dependent accumulation of hyperphosphorylated forms of tau (phospho-tau). The aim of this study was to investigate the progression of tau aggregation and analyze its relationship with microglial activation and synaptic damage within the hippocampus of a transgenic tau model. 2, 6, 9, 12 and 18 month-old THY-Tau22 transgenic and WT mice were analyzed. Tau pathology was assessed by western blotting and immunohistochemistry (AT8, AT100). Confocal microscopy was used to study microglial/phospho-tau relationship, and Thioflavin-S staining to evidence fibrillar aggregates. Levels of general (Synaptophysin) and subtype-specific (ChAT, VGAT, VGLUT-1) synaptic proteins were determined by WB and immunohistochemistry. Inflammatory markers were assessed by quantitative PCR (CD45, CD68, TREM2), immunohistochemistry (Iba-1) and image analysis. Tau pathology was detectable in the hippocampus from 2 months of age and increased progressively during aging. Presynaptic protein levels were significantly decreased from 9-12 months compared to age-matched WT mice. Even though some inflammatory markers were slightly increased in the hippocampus, microglial reactivity was found to be generally attenuated and some cells even exhibited reduction in their prolongations and a clear degenerative phenotype at advanced ages similar to that seen in the hippocampus of AD patients. Finally, this model could be a relevant tool to further understand the specific role of tau in both microglial response and synaptic pathology in AD.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Abeta from APP/PS1 Alzheimer mice hippocampus induced synaptic damage in vivo and in vitro

We aim to investigate the effects of Abeta from young APP/PS1 mouse model of Alzheimer`s disease (AD) on the synaptic integrity, as the loss of synapses strongly correlates with cognitive deficits in patients. Plaque-associated abnormal swellings of neuronal processes represent the first indicator of disease development and might compromise neuronal integrity and synaptic function. Here, we examined the synaptic nature of dystrophic neurites, and the reduction of both synapses and vesicles density in presynaptic terminals along with the progressive accumulation of autophagic structures and Abeta within hippocampal synaptosomes during the aging. We analysed both the direct synaptotoxic effect of plaques in the hippocampus of this model and also the repercussion of the soluble (S1) fraction in neuronal cultures. Hippocampal synapses were observed under both optic and electron microscopy. Synapses and vesicle density were quantified in periplaque and control (plaque-free) areas by electron microscopy. Primary neuronal cultures were incubated for 48 hours with 6-month-old APP/PS1 and wild-type S1 fractions. In addition, Abeta immunodepletion was carried out with different anti-Abeta antibodies and the levels of synaptic proteins were measured by Western-blot (WB). Both synapse number and synaptic-vesicles density were significantly decreased in young APP/PS1 mice, close to the Abeta deposits, in several hippocampal layers. Importantly, there was a correlation between the synaptic deficiencies and the distance to plaques, which presented oligomeric forms in their periphery. Some presynaptic elements were abnormally swollen, containing autophagic vesicles. In addition, we found by WB a decrease in several hippocampal synaptic markers as early as 4 months of age in this model, and also in neuronal cultures incubated with S1 fractions. Significantly, the neuronal reduction in VGLUT was reversed after Abeta immunodepletion. Plaque-associated oligomeric Abeta induced an early deleterious effect on synapses that correlates with memory deficits in young APP/PS1 mice. Moreover, soluble Abeta derived from these transgenic mice reduced synaptic protein content in vitro, which was restored after immunodepletion of Abeta species. Therefore, this model produced synaptotoxic Abeta and may represent a valuable tool to test novel treatments to protect synapses as an early therapeutic approach for AD.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Microtubule stabilization reduces amyloid pathology and improves synaptic/memory deficits in APP/PS1 mice

Aims: Cognitive decline in Alzheimer's disease (AD) is highly related to synaptic/neuronal loss. Tau hyperphosphorylation destabilizes microtubules leading to axonal transport failure and generation of dystrophic neurites, thus contributing to synaptic dysfunction. The effect of microtubule stabilization on amyloid-beta pathology has not been assessed in vivo yet. This study evaluated the effect of the microtubule-stabilizing agent, Epothilone D (EpoD) in the pathology of an amyloidogenic mouse model. Methods: APP751SL/PS1M146L mice (3-month-old) were treated weekly with intraperitoneal injections of EpoD (2 mg/kg) or vehicle for 3 months. For memory performance, animals were tested on the objectrecognition, Y-maze and Morris water maze. Hippocampal proteinopathies were quantified by image analysis after immunostaining. Somatostatin (SOM)-numerical density was calculated by stereology. APPswe-N2a cells were treated with EpoD 100nM for 12/24 hours. Protein levels were analysed by Western/dot-blot. Results: EpoD-treated mice improved their performance of cognitive tests, while hippocampal phospho-tau and Ab (especially oligomers) accumulation decreased, together with synaptic/neuritic pathology. Remarkably, EpoD exerted a neuroprotective effect on SOM-interneurons, a highly AD-vulnerable GABAergic subpopulation. Conclusions: EpoD improved microtubule dynamics and axonal transport in an AD-like context, reducing tau and Ab accumulation and promoting neuronal and cognitive protection, underlining the cross-talk between cytoskeleton pathology and proteinopathy. Therefore, microtubule-stabilizing drugs could be candidates for slowing AD at both tau and Ab pathologies.Supported by PI18/01557 (to AG) and PI18/01556 (to JV) grants from ISCiii of Spain, co-financed by FEDER funds (European Union), CIBERNED collaborative grant (to AG and JV), and by PPIT.UMA.B1.2017/26 grant (to RSV). Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

IFNL4 ss469415590 polymorphism is associated with unfavourable clinical and immunological status in HIV-infected individuals

AbstractThe IFNL4 ss469415590 polymorphism, in high linkage disequilibrium with the IL28B rs12979860 variant, has been associated with hepatitis C virus clearance. We evaluated whether ss469415590 is associated with clinical and immunovirological parameters in human immunodeficiency virus-infected subjects. We found an independent association of the IFNL4 ss469415590 polymorphism with higher prevalence of AIDS-defining illnesses and lower CD4 T cell numbers. These results suggest the existence of common host defence mechanisms against different viral infections

Nosocomial Outbreaks Caused by Leuconostoc mesenteroides subsp. mesenteroides

From July 2003 through October 2004, 42 patients became infected by strains of Leuconostoc mesenteroides subsp. mesenteroides (genotype 1) in different departments of Juan Canalejo Hospital in northwest Spain. During 2006, 6 inpatients, also in different departments of the hospital, became infected (genotypes 2–4). Parenteral nutrition was the likely source