Role of Delayed Neuroglial Activation in Impaired Cerebral Blood Flow Restoration Following Comorbid Injury.

Besides other causes, ischemia and Alzheimer's disease pathology is also linked to decreased cerebral blood flow (CBF). There is little or no consensus about the role of neuroglial cells in maintaining CBF in various neuropathologies. This consensus becomes scarcer when it comes to clinical and experimental cases of comorbid Abeta-amyloid (Aß) toxicity and ischemia. Here, a comorbid rat model of Aß toxicity and endothelin-1 induced ischemia (ET1) not only demonstrated the appearance of axotomized phagocytosed pyknotic neurons (NeuN) immediately after the injury, but also showed a diversity of continuously changing neuroglia (MHC Class II/OX6, Iba1) and macrophage (Iba1/CD68) phenotypes with round, stout somas, and retracted processes. This is indicative of a response to a concomitant increase in large fluid-filled spaces due to the vascular leakage. Ironically 4 weeks after the injury despite a conclusive reduction in neurons, CBF restoration in ET1 rats was associated with a massive increase in neuroglial cell numbers, hypertrophy, ramification, and soma sizes bordering the continuously reducing lesion core and inflamed vasculature, possibly to shield their leaky phenotype. Astrocytes were also found to be releasing matrix metalloproteinase9 (MMP9), which stabilized matrix ligand ß-dystroglycan (ß-DG) in repaired or functional vessels. Changing neuroglia phenotypes, responses, motility, astrocytic recruitment of MMP9, and ß-DG stabilization implies the role of communication between neuroglia and endothelium in recovering CBF, in the absence of neurons, in ET1 rats compared to Aß+ET1 rats, which showed characteristics delayed neuroglial activation. Stimulation of timely neuroglial reactivity may serve as a viable strategy to compensate for the neuronal loss in restoring CBF in comorbid cases of ischemia and Aß toxicity.

Correction to: Role of Delayed Neuroglial Activation in Impaired Cerebral Blood Flow Restoration Following Comorbid Injury.

The original version of this article contained a random order of part labels for Fig. 4. The correct caption of Fig. 4 with correct order of part labels is given below.

Developing Trojan horses to induce, diagnose and suppress Alzheimer's pathology.

There are many obstacles impeding the Alzheimer's disease (AD) research. For instance, its early diagnosis to identify individuals at risk has not been successful so far. AD animal models cannot be created without genetic pre-disposition or surgical manipulation. Single gene/protein delivery has so far failed to achieve significant clinical improvements in multifactorial AD. We hypothesize that the blood-brain barrier (BBB) penetration issues are the major obstacle in the development of current Alzheimer's causative, diagnostic, and multi-targeted therapeutic probes, and partly the cause of the failure of more than 99% of intervention trials. To overcome this problem, shuttle peptides or monoclonal antibodies for receptors on BBB can act as molecular Trojan horses to transport the fused novel classes of re-engineered AD causative agents, diagnostic probes, or multiple function neurovascular medicines across the BBB via receptor-mediated transport to cause, diagnose, or improve the AD phenotype, respectively. Here, we propose the design of such Trojan horses, comprising three essential components that could (i) reverse Aß amyloidosis, (ii) clear liberated Aß, and (iii) improve angiogenesis or endothelial metabolic dysfunction, besides alleviating the inflammation, to eventually enhance neuronal health, cerebral blood flow, and cognitive function. Such Trojan horses can aid in AD research by diagnosing Aß-oligomers at earlier stages, creating improved animal models by exposing transgenic animals to amyloid-inducing agents, and allowing treatment by novel neurovascular medicines.

Dipyridamole plus Triflusal versus Triflusal Alone in Infarct Reduction after Middle Cerebral Artery Occlusion.

BACKGROUND AND PURPOSE: The objective of this work is to study the dose-dependent effect of combination therapy with dipyridamole and triflusal over that of triflusal alone on infarct size after middle cerebral artery occlusion (MCAO) ischemia. MATERIALS AND METHODS: Male Wistar rats were subjected to a permanent MCAO in the right hemisphere. Rats received triflusal alone and with dipyridamole via oral route. Three days after surgery, infarct volumes were measured. RESULTS: The lower dose regime of triflusal (10 mg/kg) and dipyridamole (200 mg/kg) caused the greatest decrease in infarct size compared with higher dose regime of triflusal (30 mg/kg) and dipyridamole (200 mg/kg) (P <.01), triflusal (30 mg/kg) alone (P <.07), and vehicle-treated controls. CONCLUSIONS: The lower dose combination of dipyridamole and triflusal appears to be more effective than triflusal alone after MCAO-induced cerebral ischemia. Therefore, there is a strong rationale to continue to examine the protective effects of triflusal and dipyridamole after cerebral ischemia.

Neural plasticity and memory: molecular mechanism.

Deciphering the cellular and molecular mechanisms of memory has been an important topic encompassing the learning and memory domain besides the neurodegenerative disorders. Synapses accumulate cognitive information from life-lasting alterations of their molecular and structural composition. Current memory storage models identify posttranslational modification imperative for short-term information storage and mRNA translation for long-term information storage. However, the precise account of these modifications has not been summarized at the individual synapse level. Therefore, herein we describe the spatiotemporal reorganization of synaptic plasticity at the dendritic spine level to elucidate the mechanism through which synaptic substructures are remodeled; though at the molecular level, such mechanisms are still quite unclear. It has thus been concluded that the existing mechanisms do not entirely elaborate memory storage processes. Further efforts are therefore encouraged to delineate the mechanism of neuronal connectivity at the chemical level as well, including inter- or intramolecular bonding patterns at the synaptic level, which may be a permissive and vital step of memory storage.

Protein markers of cerebrovascular disruption of neurovascular unit: immunohistochemical and imaging approaches.

Currently, there is great interest in the assembly and function of cerebral endothelial, glial and neuronal cells that form the anatomical basis of the neurovascular unit (NVU) to maintain blood-brain barrier (BBB) and cerebral blood flow. Recent studies have provided considerable insight into the assembly of the components of the NVU. However, there is still paucity of data regarding the identification and expression pattern of these components in various pathologies of the central nervous system. Here, we provide a brief overview of the histological and imaging methods to study BBB disruption in various experimental settings especially the ischemia models associated with necrosis. Emphasis is on the immunohistochemistry of various protein markers of NVU. An understanding of the alterations in the expression pattern of these markers in various neurodegenerative disorders could lead to a better understanding of the conditions that cause BBB disruption as well as to the development of new restorative and protective treatments.

A randomized controlled longitudinal naturalistic trial testing the effects of automatic self transcending meditation on heart rate variability in late life depression: study protocol.

BACKGROUND: The prevalence and socioeconomic cost of late life depression (LLD) is on the rise, while the response rate to antidepressant trials remains poor. Various mind-body therapies are being embraced by patients as they are considered safe and potentially effective, yet little is known regarding the effectiveness of such therapies to improve LLD symptoms. Among the mind-body therapies currently in practice, the results of our pilot study have shown that a particular meditation technique called Sahaj Samadhi Meditation, which belongs to the category of meditation termed automatic self-transcending meditation (ASTM) may have some promise in improving cardiovascular autonomic disturbances associated with LLD as well as ameliorating symptoms of depression and anxiety. METHODS/DESIGN: Patients between the ages of 60 and 85 with LLD will be randomized either to ASTM plus treatment as usual (TAU) or TAU alone to assess changes in cardiovascular autonomic parameters, neuropsychological symptoms of depression and anxiety as well as quality of life. The instructional phase of the intervention consists of 4 consecutive days of meditation training, after which participants are encouraged to meditate twice daily for twenty minutes each time at home. The intervention also includes once weekly follow up sessions for the subsequent 11 weeks. The planned study has one and a half year recruitment period. Participants will be assessed at baseline and at 4, 8, 12 and 24 weeks post intervention. DISCUSSION: This study should provide a unique data source from a randomized, controlled, longitudinal trial to investigate the effects of a form of ASTM on cardiovascular autonomic and neuropsychological health in LLD. TRIAL REGISTRATION: Clinicaltrials.gov NCT02149810, date registered: 05/28/2014.

A modular inquiry-based semester theme that integrates data science education and bioinformatics in protein structure function courses.

BACKGROUND: With an exponential growth in biological data and computing power, familiarity with bioinformatics has become a demanding and popular skill set both in academia and industry. There is a need to increase students' competencies to be able to take on bioinformatic careers, to get them familiarized with scientific professions in data science and the academic training required to pursue them, in a field where demand outweighs the supply. METHODS: Here we implemented a set of bioinformatic activities into a protein structure and function course of a graduate program. Concisely, students were given hands-on opportunities to explore the bioinformatics-based analyses of biomolecular data and structural biology via a semester-long case study structured as inquiry-based bioinformatics exercises. Towards the end of the term, the students also designed and presented an assignment project that allowed them to document the unknown protein that they identified using bioinformatic knowledge during the term. RESULTS: The post-module survey responses and students' performances in the lab module imply that it furthered an in-depth knowledge of bioinformatics. Despite having not much prior knowledge of bioinformatics prior to taking this module students indicated positive feedback. CONCLUSION: The students got familiar with cross-indexed databases that interlink important data about proteins, enzymes as well as genes. The essential skillsets honed by this research-based bioinformatic pedagogical approach will empower students to be able to leverage this knowledge for their future endeavours in the bioinformatics field.

Structural insight into the differential effects of omega-3 and omega-6 fatty acids on the production of Abeta peptides and amyloid plaques.

Several studies have shown the protective effects of dietary enrichment of various lipids in several late-onset animal models of Alzheimer Disease (AD); however, none of the studies has determined which structure within a lipid determines its detrimental or beneficial effects on AD. High-sensitivity enzyme-linked immunosorbent assay (ELISA) shows that saturated fatty acids (SFAs), upstream omega-3 FAs, and arachidonic acid (AA) resulted in significantly higher secretion of both Aß 40 and 42 peptides compared with long chain downstream omega-3 and monounsaturated FAs (MUFA). Their distinct detrimental action is believed to be due to a structural template found in their fatty acyl chains that lack SFAs, upstream omega-3 FAs, and AA. Immunoblotting experiments and use of APP-C99-transfected COS-7 cells suggest that FA-driven altered production of Aß is mediated through γ-secretase cleavage of APP. An early-onset AD transgenic mouse model expressing the double-mutant form of human amyloid precursor protein (APP); Swedish (K670N/M671L) and Indiana (V717F), corroborated in vitro findings by showing lower levels of Aß and amyloid plaques in the brain, when they were fed a low fat diet enriched in DHA. Our work contributes to the clarification of aspects of structure-activity relationships.

Additive effects of fatty acid mixtures on the levels and ratio of amyloid ß40/42 peptides differ from the effects of individual fatty acids.

Several studies have shown the protective and/or deleterious effects of dietary enrichment of single fatty acids (FAs) in several animal and cell-culture models of Alzheimer's disease (AD). However, potential interactions among dietary fatty acids are traditionally ignored. None of these studies has examined and compared the differential effects of FAs in combination, as well as alone, for their effects on amyloid ß production or AD. Here we investigated the effects of omega-9 (oleic acid) and omega-6 (linoleic and arachidonic acids) fatty acids, either alone or combined, on Aß production by APP-695 and SP-C99 transfected COS-7 cells. Overall, our results are the first to demonstrate that mixtures of FAs alter the production of Aß40 and Aß42 peptides and consequently the Aß40:42 ratio differently from individual FAs. Here we show that the effects of a single lipid on Aß production are not attributed to that single FA alone. Rather, the overall lipid composition influences the specificity and level of the regulated intramembranous proteolysis of APP by the γ-secretase complex. Our results reinforce the importance of studying composite lipids/nutrients rather than single lipids or nutrients.

Differential temporal and spatial post-injury alterations in cerebral cell morphology and viability.

Combination of ischemia and ß-amyloid (Aß) toxicity has been shown to simultaneously increase neuro-inflammation, endogenous Aß deposition, and neurodegeneration. However, studies on the evolution of infarct and panorama of cellular degeneration as a synergistic or overlapping mechanism between ischemia and Aß toxicity are lacking. Here, we compared fluorojade B (FJB) and hematoxylin and eosin (H&E) stains primarily to examine the chronology of infarct, and the viability and morphological changes in neuroglia and neurons located in different brain regions on d1, d7, and d28 post Aß toxicity and endothelin-1 induced ischemia (ET1) in rats. We demonstrated a regional difference in cellular degeneration between cortex, corpus callosum, striatum, globus pallidus, and thalamus after cerebral injury. Glial cells in the cortex and corpus callosum underwent delayed FJB staining from d7 to d28, but neurons in cortex disappeared within the first week of cerebral injury. Striatal lesion core and globus pallidus of Aß + ET1 rats showed extensive degeneration of neuronal cells compared with ET1 rats alone starting from d1. Differential and exacerbated expressions of cyclooxygenase-2 might be the cause of excessive neuronal demise in the striatum of Aß + ET1 rats. Such an investigation may improve our understanding to identify and manipulate a critical therapeutic window post comorbid injury.

Engineering of fluorescent or photoactive Trojan probes for detection and eradication of ß-Amyloids.

Trojan horse technology institutes a potentially promising strategy to bring together a diagnostic or cell-based drug design and a delivery platform. It provides the opportunity to re-engineer a novel multimodal, neurovascular detection probe, or medicine to fuse with blood-brain barrier (BBB) molecular Trojan horse. In Alzheimer's disease (AD) this could allow the targeted delivery of detection or therapeutic probes across the BBB to the sites of plaques and tangles development to image or decrease amyloid load, enhance perivascular Aß clearance, and improve cerebral blood flow, owing principally to the significantly improved cerebral permeation. A Trojan horse can also be equipped with photosensitizers, nanoparticles, quantum dots, or fluorescent molecules to function as multiple targeting theranostic compounds that could be activated following changes in disease-specific processes of the diseased tissue such as pH and protease activity, or exogenous stimuli such as, light. This concept review theorizes the use of receptor-mediated transport-based platforms to transform such novel ideas to engineer systemic and smart Trojan detection or therapeutic probes to advance the neurodegenerative field.

The spatial cerebral damage caused by larger infarct and ß-amyloid toxicity is driven by the anatomical/functional connectivity.

Large cerebral infarctions are major predictors of death and severe disability from stroke. Conversely, data concerning these types of infarctions and the affected adjacent brain circuits are scarce. It remains to be determined if the co-morbid concurrence of large infarct and ß-amyloid (Aß) toxicity can precipitate the early development of dementia. Here, we described a dose-dependent effect of a unilateral striatal injection of vasoconstrictive endothelin-1 (ET-1) along with Aß toxicity on CNS pathogenesis; driven by the anatomical and functional networks within a brain circuit. After 21 days of treatment, a high dose (60 pmol) of ET-1 (E60) alone caused the greatest increase in neuroinflammation, mainly in the ipsilateral striatum and distant regions with synaptic links to the striatal lesion such as white matter (subcortical white matter, corpus callosum, internal capsule, anterior commissure), gray matter (globus pallidus, thalamus), and cortices (cingulate, motor, somatosensory, entorhinal). The combined E60 + Aß treatment also extended perturbation in the contralateral hemisphere of these rats, such as increased deposition of amyloid precursor protein fragments associated with the appearance of degenerating cells and the leakage of laminin from the basement membrane across a compromised blood-brain barrier. However, the cerebral damage induced by the 6 pmol ET-1 (E6), Aß and E6 + Aß rats was not detrimental enough to injure the complete network. The appreciation of the causal interactions among distinct anatomical units in the brain after ischemia and Aß toxicity will help in the design of effective and alternative therapeutics that may disassociate the synergistic or additive association between the infarcts and Aß toxicity.

Pathological Changes in Microvascular Morphology, Density, Size and Responses Following Comorbid Cerebral Injury.

Aberrations in brain microcirculation and the associated increase in blood-brain-barrier (BBB) permeability in addition to neuroinflammation and Aß deposition observed in Alzheimer's disease (AD) and ischemia have gained considerable attention recently. However, the role of microvascular homeostasis as a pathogenic substrate to disturbed microperfusion as well as an overlapping etiologic mechanism between AD and ischemia has not been thoroughly explored. In this study, we employ temporal histopathology of cerebral vasculature in a rat model of ß-amyloid (Aß) toxicity and endothelin-1 induced-ischemia (ET1) to investigate the panorama of cerebral pathology and the protein expression on d1, d7, and d28 post-injury. The combination of Aß and ET1 pathological states leads to an alteration in microvascular anatomy, texture, diameter, density, and protein expression, in addition to disturbed vessel-matrix-connections, inter-compartmental water exchange and basement membrane profile within the lesion epicenter localized in the striatum of Aß+ET1 brains compared to Aß and ET1 rats. We conclude that the neural microvascular network, in addition to the neural tissue, is not only sensitive to structural deterioration but also serves as an underlying vascular etiology between ischemia and AD pathologies. Such investigation can provide prospects to appreciate the interrelationships between structure and responses of cerebral microvasculature and to provide a venue for vascular remodeling as a new treatment strategy.

Spatial Dynamics of Vascular and Biochemical Injury in Rat Hippocampus Following Striatal Injury and Aß Toxicity.

The hippocampus, a brain region vital for memory and learning, is sensitive to the damage caused by ischemic/hypoxic stroke and is one of the main regions affected by Alzheimer's disease. The pathological changes that might occur in the hippocampus and its connections, because of cerebral injury in a distant brain region, such as the striatum, have not been examined. Therefore, in the present study, we evaluated the combined effects of endothelin-1-induced ischemia (ET1) in the striatum and ß-amyloid (Aß) toxicity on hippocampal pathogenesis, dictated by the anatomical and functional intra- and inter-regional hippocampal connections to the striatum. The hippocampal pathogenesis induced by Aß or ET1 alone was not severe enough to significantly affect the entire circuit of the hippocampal network. However, the combination of the two pathological states (ET1 + Aß) led to an exacerbated increase in neuroinflammation, deposition of the amyloid precursor protein (APP) fragments with the associated appearance of degenerating cells, and blood-brain-barrier disruption. This was observed mainly in the hippocampal formation (CA2 and CA3 regions), the dentate gyrus as well as distinct regions with synaptic links to the hippocampus such as entorhinal cortex, thalamus, and basal forebrain. In addition, ET1 + Aß-treated rats also demonstrated protracted loss of AQP4 depolarization, dissolution of ß-dystroglycan, and basement membrane laminin with associated IgG and dysferlin leakage. Spatial dynamics of hippocampal injury in ET1 + Aß rats may provide a valuable model to study new targets for clinical therapeutic applications, specifically when areas remotely connected to hippocampus are damaged.

Altered Insulin/Insulin-Like Growth Factor Signaling in a Comorbid Rat model of Ischemia and ß-Amyloid Toxicity.

Ischemic stroke and diabetes are vascular risk factors for the development of impaired memory such as dementia and/or Alzheimer's disease. Clinical studies have demonstrated that minor striatal ischemic lesions in combination with ß-amyloid (Aß) load are critical in generating cognitive deficits. These cognitive deficits are likely to be associated with impaired insulin signaling. In this study, we examined the histological presence of insulin-like growth factor-I (IGF-1) and insulin receptor substrate (IRS-1) in anatomically distinct brain circuits compared with morphological brain damage in a co-morbid rat model of striatal ischemia (ET1) and Aß toxicity. The results demonstrated a rapid increase in the presence of IGF-1 and IRS-1 immunoreactive cells in Aß + ET1 rats, mainly in the ipsilateral striatum and distant regions with synaptic links to the striatal lesion. These regions included subcortical white matter, motor cortex, thalamus, dentate gyrus, septohippocampal nucleus, periventricular region and horizontal diagonal band of Broca in the basal forebrain. The alteration in IGF-1 and IRS-1 presence induced by ET1 or Aß rats alone was not severe enough to affect the entire brain circuit. Understanding the causal or etiologic interaction between insulin and IGF signaling and co-morbidity after ischemia and Aß toxicity will help design more effective therapeutics.

The Dynamics of Impaired Blood-Brain Barrier Restoration in a Rat Model of Co-morbid Injury.

Defect in brain microperfusion is increasingly recognized as an antecedent event to Alzheimer's disease (AD) and ischemia. Nevertheless, studies on the role of impaired microperfusion as a pathological trigger to neuroinflammation, Aß deposition as well as blood-brain barrier (BBB) disruption, and the etiological link between AD and ischemia are lacking. In this study, we employ in vivo sequential magnetic resonance imaging (MRI) and computed tomography (CT) imaging in a co-morbid rat model of ß-amyloid toxicity (Aß) and ischemia (ET1) with subsequent histopathology of striatal lesion core and penumbra at 1, 7, and 28 days post injury. Within 24 h, cerebral injury resulted in increased BBB permeability due to the dissolution of ß-dystroglycan (ß-DG) and basement membrane laminin by active matrix metalloproteinase9 (MMP9). As a result, net flow of circulating IgG down a hydrostatic gradient into the parenchyma led to vasogenic edema and impaired perfusion, thus increasing the apparent hyperintensity in true fast imaging with steady-state free precession (true FISP) imaging and acute hypoperfusion in CT. This was followed by a slow recruitment of reactive astroglia to the affected brain and depolarization of aquaporin4 (AQP4) expression resulting in cytotoxic edema-in an attempt to resolve vasogenic edema. On d28, functional BBB was restored in ET1 rats as observed by astrocytic MMP9 release, ß-DG stabilization, and new vessel formation. This was confirmed by reduced hyperintensity on true FISP imaging and normalized cerebral blood flow in CT. While, Aß toxicity alone was not detrimental enough, Aß+ET1 rats showed delayed differential expression of MMP9, late recruitment of astroglial cells, protracted loss of AQP4 depolarization, and thus delayed BBB restoration and cerebral perfusion.

Microbial Proteins as Novel Industrial Biotechnology Hosts to Treat Epilepsy.

Epilepsy is characterized by the hyperexcitability of various neuronal circuits that results due to the imbalance between glutamate-mediated excitation of voltage-gated cation channels and γ-amino butyric acid (GABA)-mediated inhibition of anion channels leading to aberrant, sporadic oscillations or fluctuations in neuronal electrical activity. Epilepsy with a risk of mortality and around 65 million sufferers of all ages all over the world is limited therapeutically with high rates of adverse reactions, lack of complete seizure control, and over 30% patients with refractory epilepsy. The only alternative to medicines is to identify and surgically remove the seizure foci in the brain or to abort the seizures just as they begin using an implanted cerebral electrode. However, these alternatives are unable to precisely aim aberrant neuronal circuits while leaving others unaltered. Epilepsy animal models also constitute the identical constraint. Thus, a better target-specific approach is needed to study and treat epilepsy. Unicellular green algae Chlamydomonas reinhardtii expresses a channelrhodopsin-2 (ChR2) sodium ion channel protein that controls the phototaxis movement of algae in response to blue light. Similarly, archaeon Natronomonas pharaonis (NpHR) expresses a monovalent Cl- channel protein halorhodopsin that responds to yellow light. These features of ChR2 and NpHR proteins can be used in optogenetic techniques to manipulate the bi-directional firing pattern of neuronal circuits in an attempt to better understand the pathophysiology of epileptic seizures as well as to discover novel potential drugs to treat epilepsy.

Correction: DHA Supplemented in Peptamen Diet Offers No Advantage in Pathways to Amyloidosis: Is It Time to Evaluate Composite Lipid Diet?

[This corrects the article DOI: 10.1371/journal.pone.0024094.].

Why therapies for Alzheimer's disease do not work: Do we have consensus over the path to follow?

Alzheimer's disease (AD) represents a personal tragedy of enormous magnitude, which imposes a daunting worldwide challenge for health-care providers and society as well. In last five decades, global research in clinics and laboratories has illuminated many features of this sinister and eventually fatal disease. Notwithstanding this development, the Alzheimer's research apparently has come across a phase of disappointment and a little reservation about the direction to follow. Persistently distressing controversies and a significant number of missing facts shed further uncertainty about the path forward. A detailed description of some of the main controversies in AD research may assist the field towards finding a resolution. Here I reviewed some alarming concerns or controversies related to these primary issues and emphasized on a possible mechanism to settle them.