Klaus Fassbender

Role of the toll-like receptor 4 in neuroinflammation in Alzheimer's disease.

Microglial activation is a key feature in Alzheimer’s disease and is considered to contribute to progressive neuronal injury by release of neurotoxic products. The innate immune receptor Toll-like-receptor 4 (TLR4), localized on the surface of microglia, is a first-line host defense receptor against invading microorganisms. Here, we show that a spontaneous loss-of-function mutation in the Tlr4 gene strongly inhibits microglial and monocytic activation by aggregated Alzheimer amyloid peptide resulting in a significantly lower release of the inflammatory products IL-6, TNFα and nitric oxide. Treatment of primary murine neuronal cells with supernatant of amyloid peptide-stimulated microglia demonstrates that Tlr4 contributes to amyloid peptide-induced microglial neurotoxicity. In addition, stimulation experiments in transfected HEK293 cells allowed to define a tri-molecular receptor complex consisting of TLR4, MD-2 and CD14 necessary for full cellular activation by aggregated amyloid peptide. A clinical relevance of these findings is supported by a marked upregulation of Tlr4 mRNA in APP transgenic mice and by an increased expression of TLR4 in Alzheimer’s disease brain tissue associated with amyloid plaque deposition. Together, these observations provide the first evidence for a role of the key innate immune receptor, TLR4, in neuroinflammation in Alzheimer’s disease.

Diagnosis and treatment of patients with stroke in a mobile stroke unit versus in hospital: a randomised controlled trial.

BACKGROUND Only 2-5% of patients who have a stroke receive thrombolytic treatment, mainly because of delay in reaching the hospital. We aimed to assess the efficacy of a new approach of diagnosis and treatment starting at the emergency site, rather than after hospital arrival, in reducing delay in stroke therapy. METHODS We did a randomised single-centre controlled trial to compare the time from alarm (emergency call) to therapy decision between mobile stroke unit (MSU) and hospital intervention. For inclusion in our study patients needed to be aged 18-80 years and have one or more stroke symptoms that started within the previous 2·5 h. In accordance with our week-wise randomisation plan, patients received either prehospital stroke treatment in a specialised ambulance (equipped with a CT scanner, point-of-care laboratory, and telemedicine connection) or optimised conventional hospital-based stroke treatment (control group) with a 7 day follow-up. Allocation was not masked from patients and investigators. Our primary endpoint was time from alarm to therapy decision, which was analysed with the Mann-Whitney U test. Our secondary endpoints included times from alarm to end of CT and to end of laboratory analysis, number of patients receiving intravenous thrombolysis, time from alarm to intravenous thrombolysis, and neurological outcome. We also assessed safety endpoints. This study is registered with ClinicalTrials.gov, number NCT00153036. FINDINGS We stopped the trial after our planned interim analysis at 100 of 200 planned patients (53 in the prehospital stroke treatment group, 47 in the control group), because we had met our prespecified criteria for study termination. Prehospital stroke treatment reduced the median time from alarm to therapy decision substantially: 35 min (IQR 31-39) versus 76 min (63-94), p<0·0001; median difference 41 min (95% CI 36-48 min). We also detected similar gains regarding times from alarm to end of CT, and alarm to end of laboratory analysis, and to intravenous thrombolysis for eligible ischaemic stroke patients, although there was no substantial difference in number of patients who received intravenous thrombolysis or in neurological outcome. Safety endpoints seemed similar across the groups. INTERPRETATION For patients with suspected stroke, treatment by the MSU substantially reduced median time from alarm to therapy decision. The MSU strategy offers a potential solution to the medical problem of the arrival of most stroke patients at the hospital too late for treatment. FUNDING Ministry of Health of the Saarland, Germany, the Werner-Jackstädt Foundation, the Else-Kröner-Fresenius Foundation, and the Rettungsstiftung Saar.

The LPS receptor (CD14) links innate immunity with Alzheimer's disease

To rapidly respond to invading microorganisms, humans call on their innate immune system. This occurs by microbe‐detecting receptors, such as CD14, that activate immune cells to eliminate the pathogens. Here, we link the lipopolysaccharide receptor CD14 with Alzheimers disease, a severe neurodegenerative disease resulting in dementia. We demonstrate that this key innate immunity receptor interacts with fibrils of Alzheimer amyloid peptide. Neutralization with antibodies against CD14 and genetic deficiency for this receptor significantly reduced amyloid peptide induced microglial activation and microglial toxicity. The observation of strongly enhanced microglial expression of the LPS receptor in brains of animal models of Alzheimers disease indicates a clinical relevance of these findings. These data suggest that CD14 may significantly contribute to the overall neuroinflammatory response to amyloid peptide, highlighting the possibility that the enormous progress currently being made in the field of innate immunity could be extended to research on Alzheimers disease.

Stent-Assisted Mechanical Recanalization for Treatment of Acute Intracerebral Artery Occlusions

Background and Purpose— The purpose of this study was to demonstrate a new approach to the use of a self-expanding stent in the treatment of acute ischemic stroke. Methods— Twenty-two consecutive patients with acute intracerebral artery occlusions were treated with a self-expandable intracranial stent, which was withdrawn in its unfolded state. For this technique, we used the Solitaire AB/FR, which is the only intracranial stent that is fully recoverable. Eight patients had an occlusion of the basilar artery, 12 had a middle cerebral artery occlusion, and 2 had terminal carotid artery occlusions; 6 of these had to be treated first for an acute occlusion originating in the internal carotid artery. Recanalization results were assessed by follow-up angiography immediately after the procedure. Neurologic status was evaluated before and after treatment (90-day follow-up) according to the National Institutes of Health Stroke Scale and modified Rankin scale. Results— Successful revascularization was achieved in 20 of 22 (90.9%) patients (thrombolysis in cerebral infarction [TICI] 2a/b and 3), a TICI 3 state was accomplished in 12 patients, and partial recanalization or slow distal branch filling with filling of more than two-thirds of the vessel territory was achieved in 8 patients (TICI 2b). There was immediate flow restoration in 21 of 22 (95.4%) cases after deployment of the device. The stent was removed in its unfolded state in all patients. The mean time from stroke symptom onset to recanalization was 277 minutes, with a standard deviation of 118 minutes. Mean National Institutes of Health Stroke Scale score on admission was 19.4, with a standard deviation of 5.7. Almost two-thirds of the patients (63.6%) improved by >10 points on the National Institutes of Health Stroke Scale at discharge, and 50% showed a modified Rankin scale score of ≤2 at 90 days (59% with a modified Rankin scale ≤3). Mortality was 18.1%. In 1 case, an asymptomatic intracranial hemorrhage was detected on control computed tomography, and 2 patients had a symptomatic intracranial hemorrhage. Conclusion— Withdrawal of an unfolded, fully recoverable, intracranial stent yielded very promising angiographic and clinical results. It combines the advantages of prompt flow restoration and mechanical thrombectomy.

TLR2 Is a Primary Receptor for Alzheimer’s Amyloid β Peptide To Trigger Neuroinflammatory Activation

Microglia activated by extracellularly deposited amyloid β peptide (Aβ) act as a two-edged sword in Alzheimer’s disease pathogenesis: on the one hand, they damage neurons by releasing neurotoxic proinflammatory mediators (M1 activation); on the other hand, they protect neurons by triggering anti-inflammatory/neurotrophic M2 activation and by clearing Aβ via phagocytosis. TLRs are associated with Aβ-induced microglial inflammatory activation and Aβ internalization, but the mechanisms remain unclear. In this study, we used real-time surface plasmon resonance spectroscopy and conventional biochemical pull-down assays to demonstrate a direct interaction between TLR2 and the aggregated 42-aa form of human Aβ (Aβ42). TLR2 deficiency reduced Aβ42-triggered inflammatory activation but enhanced Aβ phagocytosis in cultured microglia and macrophages. By expressing TLR2 in HEK293 cells that do not endogenously express TLR2, we observed that TLR2 expression enabled HEK293 cells to respond to Aβ42. Through site-directed mutagenesis of tlr2 gene, we identified the amino acids EKKA (741–744) as a critical cytoplasmic domain for transduction of inflammatory signals. By coexpressing TLR1 or TLR6 in TLR2-transgenic HEK293 cells or silencing tlrs genes in RAW264.7 macrophages, we observed that TLR2-mediated Aβ42-triggered inflammatory activation was enhanced by TLR1 and suppressed by TLR6. Using bone marrow chimeric Alzheimer’s amyloid precursor transgenic mice, we observed that TLR2 deficiency in microglia shifts M1- to M2-inflammatory activation in vivo, which was associated with improved neuronal function. Our study demonstrated that TLR2 is a primary receptor for Aβ to trigger neuroinflammatory activation and suggested that inhibition of TLR2 in microglia could be beneficial in Alzheimer’s disease pathogenesis.

Screening of innate immune receptors in neurodegenerative diseases: A similar pattern

In Alzheimers disease (AD), Parkinsons disease (PD), dementia with Lewy bodies (DLB) and amyotrophic lateral sclerosis (ALS), neuroinflammatory responses are considered to contribute to neuronal injury. Recently, the innate immune receptors, toll-like receptors (TLRs) and the LPS receptor (CD14) have been related to neurodegeneration. In this study, we systematically assessed the expression of most TLRs and CD14 in AD, PD/DLB and ALS using murine models of these diseases and human post-mortem brain tissues. A common upregulation of TLR2 and CD14 was found in all three animal models. While these two receptors could also be detected in AD patient tissues, they were absent from DLB and ALS tissues. This uniform pattern of innate immune response in animal models of neurodegenerative diseases clearly indicates that this response is part of a non-specific neuroinflammatory effector phase rather than a disease-specific event. The less dynamic disease progression in humans and the location (extracellular versus intracellular) of the aggregated proteins deposits might explain the divergent results seen between animal models and human tissues.

Streamlining of prehospital stroke management: the golden hour.

Thrombolysis with alteplase administered within a narrow therapeutic window provides an effective therapy for acute ischaemic stroke. However, mainly because of prehospital delay, patients often arrive too late for treatment, and no more than 1-8% of patients with stroke obtain this treatment. We recommend that all links in the prehospital stroke rescue chain must be optimised so that in the future more than a small minority of patients can profit from time-sensitive acute stroke therapy. Measures for improvement include continuous public awareness campaigns, education of emergency medical service personnel, the use of standardised, validated scales for recognition of stroke symptoms and for triaging to the appropriate institution, and advance notification to the receiving hospital. In the future, use of telemedicine technologies for interaction between the emergency site and hospital, and the strategy of treatment directly at the emergency site (mobile stroke unit concept), could contribute to more efficient use of resources and reduce the time taken to instigate treatment to within 60 min--the golden hour--of the onset of the symptoms of stroke.

Innate immune receptor expression in normal brain aging

Brain aging often results in cognitive impairment and is considered to be a major risk factor for neurodegenerative diseases. Earlier studies reported inflammatory responses in aged brain that could contribute to age-related neurodegeneration. Recently, innate immune receptors such as toll-like receptors (TLRs), so far implicated in defense against microorganisms, have been linked to pathogenesis of Alzheimers disease. Therefore, we asked whether the transcription of TLRs (1-9) and CD14, could also be altered in physiological brain aging. Using real-time polymerase chain reaction (PCR), we indeed observed that TLR1, TLR2, TLR4, TLR5, TLR7 and CD14 expression was up-regulated in mouse brain in correlation with age. In contrast, transcriptions of TLR3, TLR6 and TLR8 were unchanged and the one of TLR9 was down-regulated. In situ hybridization further confirmed these results and identified the cellular source of TLR2 and TLR7 as mononuclear phagocytes. Together, this first systematic analysis demonstrates altered regulation of those innate immune receptors even in normal brain aging, which might be of relevance for understanding susceptibility to neurodegenerative processes associated with aging.

High doses of simvastatin, pravastatin, and cholesterol reduce brain cholesterol synthesis in guinea pigs.

Recent epidemiological studies suggest that inhibitors of 3-hydroxy-3-methyl-glutaryl CoA reductase, so-called statins, are effective in lowering the prevalence of Alzheimers disease. Whether the effect of statins is due to a local inhibition of cholesterol synthesis in the brain or whether it is mediated by the reduced levels of cholesterol in the circulation is not known. In the present work, we tested the possibility that high doses of lipophilic and hydrophilic statins, simvastatin and pravastatin, respectively, or a diet high in cholesterol could affect cholesterol homeostasis in the brain of guinea pigs. The total brain cholesterol levels were not affected by high-dose simvastatin or pravastatin treatment. Significantly lower levels of the cholesterol precursor lathosterol and its ratio to cholesterol were found in the brains of simvastatin and pravastatin-treated animals. 24S-Hydroxycholesterol, the transportable form of cholesterol across the blood-brain barrier, was significantly lower in the brain of pravastatin-treated animals. Excessive cholesterol feeding resulted in higher serum cholesterol levels but did not affect total brain cholesterol level. However, de novo cholesterol synthesis in the brain seemed to be down-regulated, as indicated by lower absolute levels and cholesterol-related ratios of lathosterol compared with controls. The passage of deuterium-labeled cholesterol across the blood-brain barrier in one animal was found to be approximately 1%. Our results suggest that brain cholesterol synthesis in guinea pigs can be slightly, but significantly, influenced by high doses of lipophilic and hydrophilic statins as well as by high dietary cholesterol intake, while total brain cholesterol content and thus, cholesterol homeostasis is maintained.

Early prediction of neurological outcome after cardiopulmonary resuscitation: a multimodal approach combining neurobiochemical and electrophysiological investigations may provide high prognostic certainty in patients after cardiac arrest.

A reliable and reproducible method for precisely predicting the neurological outcome of patients with hypoxic-ischemic encephalopathy after cardiac arrest is urgently needed in neurological intensive care units. We prospectively investigated the predictive power of serum concentrations of neuron-specific enolase (NSE) and protein S-100B (S-100B) measured on days 1, 2, 3 and 7 as well as somatosensory-evoked potentials (SEPs) recorded within 48 h and on day 7 after cardiopulmonary resuscitation (CPR) in 27 patients (14 females, 13 males; mean age 61.3 ± 17.3 years) with hypoxic-ischemic encephalopathy. During the first 7 days after CPR, median values of NSE and S-100B were increased in patients who remained unconscious after CPR compared to those patients who regained consciousness (significance up to ≤0.001). The best predictor of negative outcome was an NSE cutoff point ≧43 µg/l on day 2; this had a sensitivity of 90.9% and a specificity of 100%. Additional use of S-100B on day 2 did not increase sensitivity, but this could be markedly increased by combining NSE and S-100B on days 1, 3 and 7. SEPs showing bilateral loss of cortical responses identified patients who did not regain consciousness with a specificity of 100%.