Peter Koson

Complement activating antibodies to myelin oligodendrocyte glycoprotein in neuromyelitis optica and related disorders

BackgroundSerum autoantibodies against the water channel aquaporin-4 (AQP4) are important diagnostic biomarkers and pathogenic factors for neuromyelitis optica (NMO). However, AQP4-IgG are absent in 5-40% of all NMO patients and the target of the autoimmune response in these patients is unknown. Since recent studies indicate that autoimmune responses to myelin oligodendrocyte glycoprotein (MOG) can induce an NMO-like disease in experimental animal models, we speculate that MOG might be an autoantigen in AQP4-IgG seronegative NMO. Although high-titer autoantibodies to human native MOG were mainly detected in a subgroup of pediatric acute disseminated encephalomyelitis (ADEM) and multiple sclerosis (MS) patients, their role in NMO and High-risk NMO (HR-NMO; recurrent optic neuritis-rON or longitudinally extensive transverse myelitis-LETM) remains unresolved.ResultsWe analyzed patients with definite NMO (n = 45), HR-NMO (n = 53), ADEM (n = 33), clinically isolated syndromes presenting with myelitis or optic neuritis (CIS, n = 32), MS (n = 71) and controls (n = 101; 24 other neurological diseases-OND, 27 systemic lupus erythematosus-SLE and 50 healthy subjects) for serum IgG to MOG and AQP4. Furthermore, we investigated whether these antibodies can mediate complement dependent cytotoxicity (CDC). AQP4-IgG was found in patients with NMO (n = 43, 96%), HR-NMO (n = 32, 60%) and in one CIS patient (3%), but was absent in ADEM, MS and controls. High-titer MOG-IgG was found in patients with ADEM (n = 14, 42%), NMO (n = 3, 7%), HR-NMO (n = 7, 13%, 5 rON and 2 LETM), CIS (n = 2, 6%), MS (n = 2, 3%) and controls (n = 3, 3%, two SLE and one OND). Two of the three MOG-IgG positive NMO patients and all seven MOG-IgG positive HR-NMO patients were negative for AQP4-IgG. Thus, MOG-IgG were found in both AQP4-IgG seronegative NMO patients and seven of 21 (33%) AQP4-IgG negative HR-NMO patients. Antibodies to MOG and AQP4 were predominantly of the IgG1 subtype, and were able to mediate CDC at high-titer levels.ConclusionsWe could show for the first time that a subset of AQP4-IgG seronegative patients with NMO and HR-NMO exhibit a MOG-IgG mediated immune response, whereas MOG is not a target antigen in cases with an AQP4-directed humoral immune response.

Truncated tau from sporadic Alzheimer's disease suffices to drive neurofibrillary degeneration in vivo

Truncated tau protein is the characteristic feature of human sporadic Alzheimers disease. We have identified truncated tau proteins conformationally different from normal healthy tau. Subpopulations of these structurally different tau species promoted abnormal microtubule assembly in vitro suggesting toxic gain of function. To validate pathological activity in vivo we expressed active form of human truncated tau protein as transgene, in the rat brain. Its neuronal expression led to the development of the neurofibrillary degeneration of Alzheimers type. Furthermore, biochemical analysis of neurofibrillary changes revealed that massive sarcosyl insoluble tau complexes consisted of human Alzheimers tau and endogenous rat tau in ratio 1:1 including characteristic Alzheimers disease (AD)‐specific proteins (A68). This work represents first insight into the possible causative role of truncated tau in AD neurofibrillary degeneration in vivo.

Neurodegeneration caused by expression of human truncated tau leads to progressive neurobehavioural impairment in transgenic rats

Human truncated tau protein is an active constituent of the neurofibrillary degeneration in sporadic Alzheimers disease. We have shown that modified tau protein, when expressed as a transgene in rats, induced AD characteristic tau cascade consisting of tau hyperphosphorylation, formation of argyrophilic tangles and sarcosyl-insoluble tau complexes. These pathological changes led to the functional impairment characterized by a variety of neurobehavioural symptoms. In the present study we have focused on the behavioural alterations induced by transgenic expression of human truncated tau. Transgenic rats underwent a battery of behavioural tests involving cognitive- and sensorimotor-dependent tasks accompanied with neurological assessment at the age of 4.5, 6 and 9 months. Behavioural examination of these rats showed altered spatial navigation in Morris water maze resulting in less time spent in target quadrant (p<0.05) and fewer crossings over previous platform position (p<0.05) during probe trial. Spontaneous locomotor activity and anxiety in open field was not influenced by transgene expression. However beam walking test revealed that transgenic rats developed progressive sensorimotor disturbances related to the age of tested animals. The disturbances were most pronounced at the age of 9 months (p<0.01). Neurological alterations indicating impaired reflex responses were other added features of behavioural phenotype of this novel transgenic rat. These results allow us to suggest that neurodegeneration, caused by the non-mutated human truncated tau derived from sporadic human AD, result in the neuronal dysfunction consequently leading to the progressive neurobehavioural impairment.

Truncated tau expression levels determine life span of a rat model of tauopathy without causing neuronal loss or correlating with terminal neurofibrillary tangle load

We have previously demonstrated in a transgenic rat model of tauopathy that human misfolded truncated tau derived from Alzheimer’s disease suffices to drive neurofibrillary degeneration in vivo. We employed this model to investigate the impact of truncated tau expression levels on life span, neuronal loss and the final load of neurofibrillary tangles (NFTs) in transgenic rats. Two independent transgenic lines (SHR72, SHR318), that display different expression levels of truncated tau, were utilized in this study. We found that transgene expression levels in the brain of SHR72 rats were 44% higher than in SHR318 rats and that truncated tau protein levels determined the survival rate of transgenic rats. The line with higher expression levels of truncated tau (SHR72) showed decreased median survival (222.5 days) when compared with the line with lower expression (SHR318; 294.5 days). Interestingly, NFT loads (total NFT/total neurons) were very similar in terminal stages of disease in both transgenic lines (SHR72 – 10.9%; SHR318 – 11.6%), despite significantly different expression levels of truncated tau. Moreover, mean neuron numbers in the hippocampus (CA1–3) and brain stem (gigantocellular reticular nucleus) in the two transgenic rat strains in the terminal stages of disease were similar, and did not differ significantly from those observed in age‐matched non‐transgenic controls. These findings suggest that the expression levels of misfolded truncated tau determine the life span in a transgenic rat model of tauopathy without causing neuronal loss or correlating with terminal NFT load.

First transgenic rat model developing progressive cortical neurofibrillary tangles.

Neurofibrillary degeneration induced by misfolded protein tau is considered to be one of the key pathological hallmarks of Alzheimers disease (AD). In the present study, we have introduced a novel transgenic rat model expressing a human truncated tau that encompasses 3 microtubule binding domains (3R) and a proline-rich region (3R tau151-391). The transgenic rats developed progressive age-dependent neurofibrillary degeneration in the cortical brain areas. Neurofibrillary tangles (NFTs) satisfied several key histological criteria used to identify neurofibrillary degeneration in human Alzheimers disease including argyrophilia, Congo red birefringence, and Thioflavin S reactivity. Neurofibrillary tangles were also identified with antibodies used to detect pathologic tau in the human brain, including DC11, recognizing an abnormal tau conformation and antibodies that are specific for hyperphosphorylated forms of tau protein. Moreover, neurofibrillary degeneration was characterized by extensive formation of sarkosyl insoluble tau protein complexes consisting of rat endogenous and truncated tau species. Interestingly, the transgenic rats did not show neuronal loss either in the cortex or in the hippocampus. We suggest that novel transgenic rat model for human tauopathy represents a valuable tool in preclinical drug discovery targeting neurofibrillary degeneration of Alzheimers type.

The central biobank and virtual biobank of BiOMarKaPD: a resource for studies on neurodegenerative diseases

Biobanks are important resources for biomarker discovery and assay development. Biomarkers for Alzheimer’s and Parkinson’s disease (BIOMARKAPD) is a European multicenter study, funded by the EU Joint Programme-Neurodegenerative Disease Research, which aims to improve the clinical use of body fluid markers for the diagnosis and prognosis of Alzheimer’s disease (AD) and Parkinson’s disease (PD). The objective was to standardize the assessment of existing assays and to validate novel fluid biomarkers for AD and PD. To support the validation of novel biomarkers and assays, a central and a virtual biobank for body fluids and associated data from subjects with neurodegenerative diseases have been established. In the central biobank, cerebrospinal fluid (CSF) and blood samples were collected according to the BIOMARKAPD standardized pre-analytical procedures and stored at Integrated BioBank of Luxembourg. The virtual biobank provides an overview of available CSF, plasma, serum, and DNA samples at each site. Currently, at the central biobank of BIOMARKAPD samples are available from over 400 subjects with normal cognition, mild cognitive impairment (MCI), AD, frontotemporal dementia (FTD), vascular dementia, multiple system atrophy, progressive supranuclear palsy, PD, PD with dementia, and dementia with Lewy bodies. The virtual biobank contains information on over 8,600 subjects with varying diagnoses from 21 local biobanks. A website has been launched to enable sample requests from the central biobank and virtual biobank.

Progressive neurofibrillary degeneration does not induce loss of pyramidal neurons in the cortex of the novel transgenic rat model for human tauopathy

paramagnetic resonance we observed significantly increased accumulation of ascorbyl free radicals in brains of transgenic animals (up to 1.5-fold increase; P < 0.01). Moreover, dysregulation of apoptotic and oxidative stress response genes was determined in these animals. We found significant up-regulation of anti-apoptotic gene from Bcl-2 family (up to 2.7fold increase) that is involved in mitochondrial membrane potential regulation and control of the production of reactive oxygen species. In line with the data transgene-expressing primary neurons harboured significantly increased levels of depolarized mitochondria (w1.2-fold above control) suggesting protective role of Bcl-x up-regulation in neurodegeneration. Conclusions: These data demonstrate impaired redox balance, increased oxidative stress and anti-apoptotic markers elevation induced by misfolded truncated tau expression. Furthermore, dysregulation of genes involved in apoptotic and redox pathways correlates to mitochondrial dysfunction and free radicals elevation in transgenic rat model. Moreover, our results indicate that toxic gain of function of human misfolded tau is associated with activation of anti-apoptotic defense mechanisms. This work was supported by research funding agencies of Slovak republic: APVV No. 0634-07, LPP-0043-09 and international agency: ICGEB grant No. CRP/SVK08-01.

P1-100: Neuronal loss is age-related and is not linked to massive neurofibrillary degeneration induced by expression of human truncated Tau protein in transgenic animals

GLT-1 dysfunction contributes to AD-related cognitive decline. Therefore, the objective of this study was to determine the consequences of reducing GLT-1 expression in a novel animal model of AD. Methods: To investigate this we crossed mice lacking one allele for GLT-1( /-) with APPswe/ PS1deltaE9 transgenic mice generating four experimental groups: APPswe/PS1deltaE9/GLT-1( / ); APPswe/PS1deltaE9/GLT-1( /-); nontransgenic GLT-1( / ); and non-transgenic GLT-1( /-). Using the Morris water maze we tested male mice aged to 6 months. Animals trained three days with a visible escape platform (training phase) followed with five testing days with a submerged (hidden) platform (testing phase). On the final test day a probe trial was performed (platform removed). Results: Across the three consecutive days of training all groups showed a significant reduction (p 0.001) in the latency to find the exposed platform that was statistically indistinguishable between groups. In the testing phase there was a statistically significant difference between groups (p 0.01) that was accounted for by the APPswe/PS1deltaE9/GLT-1( /-) mice which took significantly longer to find the hidden platform in comparison to the other groups (p 0.002). Additionally, the probe trial on the fifth day of testing showed that the APPswe/PS1deltaE9/GLT-1( /-) mice displayed no preference for the target quadrant and had significantly fewer entries into the target quadrant when compared to the other groups (p 0.01). Conclusions: Our findings show that partial loss of GLT-1 exacerbates the behavioral disturbances observed in transgenic mice expressing mutant APP and PS1, suggesting that GLT-1 dysfunction may contribute to cognitive decline in AD.

P1-096: Progressive neurobehavioral impairment in tau transgenic rat model correlates with axonopathy and neurofibrillary degeneration, but not with neuronal loss

Background: Transgenic animal models of Alzheimer s disease expressing mutated or nonmutated proteins involved in the etiology of the disease provide reliable tool for closer investigation of pathological processes in in vivo conditions. Moreover transgenic animal models enable us to study individual steps of pathological cascade, or to correlate pathological findings with functional neurobehavioral response of examined animals. Methods: Histopathological (Gallyas silver method, Thioflavin S), immunohistochemical (AT8) characterization of neuropathological lesions in brain stem and spinal cord tissues and quantitative neuromorphological analysis of neuron numbers was done with the use of a computer-based stereological system. Transgenic rat neurobehavioral examination was performed with use of validated testing protocols focused on measuring sensorimotor (beam walking test), neuromuscular (prehensile traction test, grip strength test) and neurological functions (placing, righting, postural reflexes) at different age periods (3.5, 5, 6 and 7 months old). Correlation of axonal degeneration with neurobehavioral response and plasma/CSF levels of phosphorylated form of neurofilament H (using ELISA kit) was also performed. Results: First signs of axonal degeneration were detected at 3-4 months old transgenic rats. Similarly on functional neurobehavioural level, transgenic rats developed hindlimb clasping phenotype and alterations of sensorimotor functions at that age. During aging, observed neurobehavioural changes became more profound due to progressive maturation of degenerative changes in axons. At 6 months old transgenic rats also neurofibrillary tangles were detected, what on functional level contributed to a rapid deterioration of sensorimotor, neuromuscular, neurological functions and premature death of animals. Moreover, quantitative assessment of total neuron numbers of transgenic rats at the terminal stage did not reveal the difference between transgenic and nontransgenic rats. Conclusions: In our study we found that axonal damage and neurofibrillary degeneration, but not neuronal loss faithfully mirror the progressive neurobehavioural impairment of transgenic rats expressing human truncated tau protein.