Izaskun Rodal

The microtubule-associated protein tau plays a critical role in the pathogenesis of Alzheimer’s disease (AD) and several related disorders collectively known as tauopathies. Development of tau pathology is associated with progressive neuronal loss and cognitive decline. In the brains of AD patients, tau pathology spreads following a predictable, anatomically defined progression pattern that can be followed by immunohistochemistry looking at brain post-mortem samples from Alzheimer patients at different stages of the disease. Furthermore, since it has been proposed that AD may be a synaptopathy and dendritic spines of pyramidal neurons are the major targets of cortical synapses, the analysis of dendritic spines is a useful tool to study the correlation between tau phosphorylation at specific sites, synaptopathy and cognitive impairment. Finally, characterization of phosphorylated tau in detergent-insoluble protein aggregates could also be an indication of the neuropathological staging in AD. Here, we describe these three complementary protocols to follow the development of tau pathology in Alzheimer’s disease.

The possibility that Alzheimer’s disease (AD) has a microbial aetiology has been proposed by several researchers. Here, we provide evidence that tissue from the central nervous system (CNS) of AD patients contain fungal cells and hyphae. Fungal material can be detected both intra- and extracellularly using specific antibodies against several fungi. Different brain regions including external frontal cortex, cerebellar hemisphere, entorhinal cortex/hippocampus and choroid plexus contain fungal material, which is absent in brain tissue from control individuals. Analysis of brain sections from ten additional AD patients reveals that all are infected with fungi. Fungal infection is also observed in blood vessels, which may explain the vascular pathology frequently detected in AD patients. Sequencing of fungal DNA extracted from frozen CNS samples identifies several fungal species. Collectively, our findings provide compelling evidence for the existence of fungal infection in the CNS from AD patients, but not in control individuals.

The identification of biomarkers for Alzheimer’s disease is important for patient management and to assess the effectiveness of clinical intervention. Cerebrospinal fluid (CSF) biomarkers constitute a powerful tool for diagnosis and monitoring disease progression. We have analyzed the presence of fungal proteins and DNA in CSF from AD patients. Our findings reveal that fungal proteins can be detected in CSF with different anti-fungal antibodies using a slot-blot assay. Additionally, amplification of fungal DNA by PCR followed by sequencing distinguished several fungal species. The possibility that these fungal macromolecules could represent AD biomarkers is discussed.

Among neurogenerative diseases, amyotrophic lateral sclerosis (ALS) is a fatal illness characterized by a progressive motor neuron dysfunction in the motor cortex, brainstem and spinal cord. ALS is the most common form of motor neuron disease; yet, to date, the exact etiology of ALS remains unknown. In the present work, we have explored the possibility of fungal infection in cerebrospinal fluid (CSF) and in brain tissue from ALS patients. Fungal antigens, as well as DNA from several fungi, were detected in CSF from ALS patients. Additionally, examination of brain sections from the frontal cortex of ALS patients revealed the existence of immunopositive fungal antigens comprising punctate bodies in the cytoplasm of some neurons. Fungal DNA was also detected in brain tissue using PCR analysis, uncovering the presence of several fungal species. Finally, proteomic analyses of brain tissue demonstrated the occurrence of several fungal peptides. Collectively, our observations provide compelling evidence of fungal infection in the ALS patients analyzed, suggesting that this infection may play a part in the etiology of the disease or may constitute a risk factor for these patients.

Argyrophilic grain disease (AGD) is a sporadic 4 R tauopathy that usually presents in combination with other sporadic tauopathies or with Alzheimer's disease (AD) pathology, and may contribute to dementia in older age patients. In previous studies, a

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a genetic arteriopathy related to Notch3 mutations, is difficult to diagnosis. The goal of this study was to determine the value of clinical, immuno

Parkin mutations produce Parkinson's disease (PD) in humans and nigrostriatal dopamine lesions related to increased free radicals in mice. We examined the effects of NP7, a synthetic, marine derived, free radical scavenger which enters the brain, on H(2)O(2) toxicity in cultured neurons and glia from wild-type (WT) and parkin null mice (PK-KO). NP7, 5-10 microM, prevented the H(2)O(2) induced apoptosis and necrosis of midbrain neuronal and glial cultures from WT and PK-KO mice. NP7 suppressed microglial activation and the H(2)O(2) induced drop-out of dopamine neurons(.) Furthermore, NP7 prevented the increased phosphorylation of ERK and AKT induced by H(2)O(2). NP7 may be a promising neuroprotector against oxidative stress in PD.

Deposition of proteins leading to amyloid takes place in some neurodegenerative diseases such as Alzheimer’s disease and Huntington’s disease. Mutations of tau and parkin proteins produce neurofibrillary abnormalities without deposition of amyloid. Here we report that mature, parkin null, over-expressing human mutated tau (PK^−/−/Tau^VLW) mice have altered behaviour and dopamine neurotransmission, tau pathology in brain and amyloid deposition in brain and peripheral organs. PK^−/−/Tau^VLW mice have abnormal behaviour and severe drop out of dopamine neurons in the ventral midbrain, up to 70%, at 12 months and abundant phosphorylated tau positive neuritic plaques, neuro-fibrillary tangles, astrogliosis, microgliosis and plaques of murine β-amyloid in the hippocampus. PK^−/−/Tau^VLW mice have organomegaly of the liver, spleen and kidneys. The electron microscopy of the liver confirmed the presence of a fibrillary protein deposits with amyloid characteristics. There is also accumulation of mouse tau in hepatocytes. These mice have lower levels of CHIP-HSP70, involved in the proteosomal degradation of tau, increased oxidative stress, measured as depletion of glutathione which, added to lack of parkin, could trigger tau accumulation and amyloidogenesis. This model is the first that demonstrates β-amyloid deposits caused by over-expression of tau and without modification of the amyloid precursor protein, presenilins or secretases. PK^−/−/Tau^VLW mice provide a link between the two proteins more important for the pathogenesis of Alzheimer disease.