DNA double-strand breaks cooperate with loss of Ink4 and Arf tumor suppressors to generate glioblastomas with frequent Met amplification

Glioblastomas (GBM) are highly radioresistant and lethal brain tumors. Ionizing radiation (IR)- induced DNA double-strand breaks (DSBs) are a risk factor for the development of GBM. In this study, we systematically examined the contribution of IR-induced DSBs to GBM development using transgenic mouse models harboring brain-targeted deletions of key tumor suppressors frequently lost in GBM, namely Ink4a, Ink4b, Arf, and/or PTEN. Using low linear energy transfer (LET) X-rays to generate simple breaks or high LET Fe ions to generate complex breaks, we found that DSBs induce high-grade gliomas in these mice which, otherwise, do not develop gliomas spontaneously. Loss of Ink4a and Arf was sufficient to trigger IR-induced glioma development but additional loss of Ink4b significantly increased tumor incidence. We analyzed IR-induced tumors for copy number alterations (CNAs) to identify oncogenic changes that were generated and selected for as a consequence of stochastic DSB events. We found Met amplification to be the most significant oncogenic event in these radiation-induced gliomas. Importantly, Met activation resulted in expression of Sox2, a GBM cancer stem cell (CSC) marker, and was obligatory for tumor formation. In sum, these results indicate that radiation-induced DSBs cooperate with loss of Ink4 and Arf tumor suppressors to generate high-grade gliomas that are commonly driven by Met amplification and activation

Separation of Sympathomimetic Amines of Abuse and Related Compounds by Micellar Electrokinetic Chromatography

Separation of twelve sympathomimetic amines and related compounds by micellar electrokinetic chromatography (MEKC) with UV absorbance detection is described. These amines were well separated within 25 min using 50 mM sodium tetraborate solution containing 15 mM sodium dodecylsulfate (SDS) of pH 9.3 as a running solution and detected at 210 nm. MEKC was performed with an applied voltage of 13 kV at 25 °C using a fused-silica capillary (50 cm×75 mm i.d.) with effective length of 37.5 cm. The detection limits of these compounds were in the range from 4 to 97 fmol/injection at a signal-to-noise ratio (S/N) of 3. The reproducibility of the method expressed as relative standard deviation (RSD) for within-day (n=6) and between-day (n=5) assays was less than 4.8 and 8.8%, respectively. The proposed method could be applied to the determination of an anorectic drug, phentermine, in Chinese tea with a detection limit of 99 μg/g (105 fmol/injection, S/N=3)

Mitochondria-Specific Accumulation of Amyloid β Induces Mitochondrial Dysfunction Leading to Apoptotic Cell Death

Mitochondria are best known as the essential intracellular organelles that host the homeostasis required for cellular survival, but they also have relevance in diverse disease-related conditions, including Alzheimer's disease (AD). Amyloid β (Aβ) peptide is the key molecule in AD pathogenesis, and has been highlighted in the implication of mitochondrial abnormality during the disease progress. Neuronal exposure to Aβ impairs mitochondrial dynamics and function. Furthermore, mitochondrial Aβ accumulation has been detected in the AD brain. However, the underlying mechanism of how Aβ affects mitochondrial function remains uncertain, and it is questionable whether mitochondrial Aβ accumulation followed by mitochondrial dysfunction leads directly to neuronal toxicity. This study demonstrated that an exogenous Aβ1–42 treatment, when applied to the hippocampal cell line of mice (specifically HT22 cells), caused a deleterious alteration in mitochondria in both morphology and function. A clathrin-mediated endocytosis blocker rescued the exogenous Aβ1–42-mediated mitochondrial dysfunction. Furthermore, the mitochondria-targeted accumulation of Aβ1–42 in HT22 cells using Aβ1–42 with a mitochondria-targeting sequence induced the identical morphological alteration of mitochondria as that observed in the APP/PS AD mouse model and exogenous Aβ1–42-treated HT22 cells. In addition, subsequent mitochondrial dysfunctions were demonstrated in the mitochondria-specific Aβ1–42 accumulation model, which proved indistinguishable from the mitochondrial impairment induced by exogenous Aβ1–42-treated HT22 cells. Finally, cellular toxicity was directly induced by mitochondria-targeted Aβ1–42 accumulation, which mimics the apoptosis process in exogenous Aβ1–42-treated HT22 cells. Taken together, these results indicate that mitochondria-targeted Aβ1–42 accumulation is the necessary and sufficient condition for Aβ-mediated mitochondria impairments, and leads directly to cellular death rather than along with other Aβ-mediated signaling alterations

MGMT promoter hypermethylation is a frequent, early, and consistent event in astrocytoma progression, and not correlated with TP53 mutation

Hypermethylation of the MGMT gene promoter and mutation of the TP53 tumor-suppressor gene are frequently present in diffuse astrocytomas. However, there is only anecdotal information about MGMT methylation status and TP53 mutations during progression of low-grade diffuse astrocytoma (AII) to anaplastic astrocytoma (AIII) and secondary glioblastoma (sGB). In this study biopsy specimens from 51 patients with astrocytic tumors with radiologically proved progression from low to high-grade malignancy were investigated for the presence and consistency of MGMT promoter hypermethylation and TP53 mutations. For 27 patients biopsy samples both of primary tumors and their recurrences were available. For the other 24 patients histology of either the low-grade lesion or the high-grade recurrence was available. It was found that MGMT promoter hypermethylation and TP53 mutations are both frequent and early events in the progression of astrocytomas and that their status is consistent over time. No correlation was found between MGMT methylation status and the presence of TP53 mutations. In addition, no correlation was found between MGMT promoter hypermethylation and the type of TP53 mutations. These results argue against the putative TP53 G:C>A:T transition mutations suggested to occur preferentially in MGMT hypermethylated tumors

Rodent Models of TDP-43 Proteinopathy: Investigating the Mechanisms of TDP-43-Mediated Neurodegeneration

Since the identification of phosphorylated and truncated transactive response DNA-binding protein 43 (TDP-43) as a primary component of ubiquitinated inclusions in amyotrophic lateral sclerosis and frontotemporal lobar degeneration with ubiquitin-positive inclusions, much effort has been directed towards ascertaining how TDP-43 contributes to the pathogenesis of disease. As with other protein misfolding disorders, TDP-43-mediated neuronal death is likely caused by both a toxic gain and loss of TDP-43 function. Indeed, the presence of cytoplasmic TDP-43 inclusions is associated with loss of nuclear TDP-43. Moreover, post-translational modifications of TDP-43, including phosphorylation, ubiquitination, and cleavage into C-terminal fragments, may bestow toxic properties upon TDP-43 and cause TDP-43 dysfunction. However, the exact neurotoxic TDP-43 species remain unclear, as do the mechanism(s) by which they cause neurotoxicity. Additionally, given our incomplete understanding of the roles of TDP-43, both in the nucleus and the cytoplasm, it is difficult to truly appreciate the detrimental consequences of aberrant TDP-43 function. The development of TDP-43 transgenic animal models is expected to narrow these gaps in our knowledge. The aim of this review is to highlight the key findings emerging from TDP-43 transgenic animal models and the insight they provide into the mechanisms driving TDP-43-mediated neurodegeneration

TMEM106B is a genetic modifier of frontotemporal lobar degeneration with C9orf72 hexanucleotide repeat expansions

Hexanucleotide repeat expansions in chromosome 9 open reading frame 72 (C9orf72) have recently been linked to frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis, and may be the most common genetic cause of both neurodegenerative diseases. Genetic variants at TMEM106B influence risk for the most common neuropathological subtype of FTLD, characterized by inclusions of TAR DNA-binding protein of 43 kDa (FTLD-TDP). Previous reports have shown that TMEM106B is a genetic modifier of FTLD-TDP caused by progranulin (GRN) mutations, with the major (risk) allele of rs1990622 associating with earlier age at onset of disease. Here, we report that rs1990622 genotype affects age at death in a single-site discovery cohort of FTLD patients with C9orf72 expansions (n = 14), with the major allele correlated with later age at death (p = 0.024). We replicate this modifier effect in a 30-site international neuropathological cohort of FTLD-TDP patients with C9orf72 expansions (n = 75), again finding that the major allele associates with later age at death (p = 0.016), as well as later age at onset (p = 0.019). In contrast, TMEM106B genotype does not affect age at onset or death in 241 FTLD-TDP cases negative for GRN mutations or C9orf72 expansions. Thus, TMEM106B is a genetic modifier of FTLD with C9orf72 expansions. Intriguingly, the genotype that confers increased risk for developing FTLD-TDP (major, or T, allele of rs1990622) is associated with later age at onset and death in C9orf72 expansion carriers, providing an example of sign epistasis in human neurodegenerative disease

Aging-related tau astrogliopathy (ARTAG):harmonized evaluation strategy

Pathological accumulation of abnormally phosphorylated tau protein in astrocytes is a frequent, but poorly characterized feature of the aging brain. Its etiology is uncertain, but its presence is sufficiently ubiquitous to merit further characterization and classification, which may stimulate clinicopathological studies and research into its pathobiology. This paper aims to harmonize evaluation and nomenclature of aging-related tau astrogliopathy (ARTAG), a term that refers to a morphological spectrum of astroglial pathology detected by tau immunohistochemistry, especially with phosphorylation-dependent and 4R isoform-specific antibodies. ARTAG occurs mainly, but not exclusively, in individuals over 60 years of age. Tau-immunoreactive astrocytes in ARTAG include thorn-shaped astrocytes at the glia limitans and in white matter, as well as solitary or clustered astrocytes with perinuclear cytoplasmic tau immunoreactivity that extends into the astroglial processes as fine fibrillar or granular immunopositivity, typically in gray matter. Various forms of ARTAG may coexist in the same brain and might reflect different pathogenic processes. Based on morphology and anatomical distribution, ARTAG can be distinguished from primary tauopathies, but may be concurrent with primary tauopathies or other disorders. We recommend four steps for evaluation of ARTAG: (1) identification of five types based on the location of either morphologies of tau astrogliopathy: subpial, subependymal, perivascular, white matter, gray matter; (2) documentation of the regional involvement: medial temporal lobe, lobar (frontal, parietal, occipital, lateral temporal), subcortical, brainstem; (3) documentation of the severity of tau astrogliopathy; and (4) description of subregional involvement. Some types of ARTAG may underlie neurological symptoms; however, the clinical significance of ARTAG is currently uncertain and awaits further studies. The goal of this proposal is to raise awareness of astroglial tau pathology in the aged brain, facilitating communication among neuropathologists and researchers, and informing interpretation of clinical biomarkers and imaging studies that focus on tau-related indicators