Highlights d Rapid establishment of glioblastoma organoids (GBOs) in a defined medium with biobank d GBOs maintain parental tumor cellular heterogeneity, gene expression, and mutations d GBO transplantation exhibits efficient engraftment and aggressive infiltration d Tumor-specific treatment responses in GBOs to drugs and CART cells

Molecular probes for selective identification of protein aggregates are important to advance our understanding of the molecular pathogenesis underlying cerebral amyloidoses. Here we report the chemical design of pentameric thiophene derivatives, denoted luminescent conjugated oligothiophenes (LCOs), which could be used for real-time visualization of cerebral protein aggregates in transgenic mouse models of neurodegenerative diseases by multiphoton microscopy. One of the LCOs, p-FTAA, showed conformation-dependent optical properties and could be utilized for ex vivo spectral assignment of distinct prion deposits from two mouse-adapted prion strains. p-FTAA also revealed staining of transient soluble pre-fibrillar non-thioflavinophilic Aβ-assemblies during in vitro fibrillation of Aβ peptides. In brain tissue samples, Aβ deposits and neurofibrillary tangles (NFTs) were readily identified by a strong fluorescence from p-FTAA and the LCO staining showed complete co-localization with conventional antibodies (6E10 and AT8), indicating that p-FTAA detects all the immuno-positive aggregated proteinaceous species in Alzheimer disease, but with significantly shorter imaging time (100 fold) compared to immunofluorescence. In addition, a patchy islet-like staining of individual Aβ plaque was unveiled by the anti-oligomer A11 antibody during co-staining with p-FTAA, suggesting that pre-fibrillar species are likely an intrinsic component of Aβ plaques in human brain. The major hallmarks of Alzheimer's disease, namely Aβ aggregates versus NFTs could also be distinguished due to distinct emission spectra from p-FTAA. Overall, we demonstrate that LCOs can be utilized as powerful practical research tools for studying protein aggregation diseases and facilitate the study of amyloid origin, evolution and maturation, Aβ−tau interactions and pathogenesis both ex vivo and in vivo. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptThe formation of highly ordered aggregates of intra-or extracellular proteins underlies a wide range of neurodegenerative conditions including prion, Parkinson's, Huntington's and Alzheimer's (AD) diseases. Hence, molecular probes that specifically target protein aggregates and allow in vitro or in vivo imaging of these pathological hallmarks, are of great importance. Small hydrophobic probes that cross the blood-brain barrier (BBB) can be monitored in vivo with positron emission tomography (PET), single-photon emission computerized tomography (SPECT) or multiphoton microscopy (1-7). The latter is especially applicable in transgenic mouse models where mechanistic insights regarding the pathological events involved in the formation of protein deposits can be obtained. Additionally, molecular imaging probes may also help in early diagnosis of neurodegenerative diseases and in monitoring the effect of therapeutic interventions. However, a major drawback of these conventional probes is that only a subset of aggregates that roughly corresponds to histologically identifiable amyloid ...

Microglial cells closely interact with senile plaques in Alzheimer’s disease and acquire the morphological appearance of an activated phenotype. The significance of this microglial phenotype and the impact of microglia for disease progression have remained controversial. To uncover and characterize putative changes in the functionality of microglia during Alzheimer’s disease, we directly assessed microglial behavior in two mouse models of Alzheimer’s disease. Using in vivo two-photon microscopy and acute brain slice preparations, we found that important microglial functions - directed process motility and phagocytic activity - were strongly impaired in mice with Alzheimer’s disease-like pathology compared to age-matched non-transgenic animals. Notably, impairment of microglial function temporally and spatially correlated with Aβ plaque deposition, and phagocytic capacity of microglia could be restored by interventionally decreasing amyloid burden by Aβ vaccination. These data suggest that major microglial functions progressively decline in Alzheimer’s disease with the appearance of Aβ plaques, and that this functional impairment is reversible by lowering Aβ burden, e.g. by means of Aβ vaccination.

The Alzheimer disease-associated presenilin (PS) proteins apparently provide the active site of ␥-secretase, an unusual intramembrane-cleaving aspartyl protease. PSs principally occur as high molecular weight protein complexes that contain nicastrin (Nct) and additional so far unidentified components. Recently, PEN-2 has been implicated in ␥-secretase function. Here we identify PEN-2 as a critical component of PS1/␥-secretase and PS2/␥-secretase complexes. Strikingly, in the absence of PS1 and PS1/PS2, PEN-2 levels are strongly reduced. Similarly, PEN-2 levels are reduced upon RNA interference-mediated down-regulation of Nct. On the other side, down-regulation of PEN-2 by RNA interference is associated with reduced PS levels, impaired Nct maturation, and deficient ␥-secretase complex formation. We conclude that PEN-2 is an integral ␥-secretase complex component and that ␥-secretase complex components are expressed in a coordinated manner.The Alzheimer disease-associated polytopic membrane proteins presenilin 1 (PS1) 1 and presenilin 2 (PS2) are required for the intramembranous ␥-secretase cleavage of the ␤-amyloid precursor protein (APP) (1). Following an initial cleavage by ␤-secretase within the APP ectodomain, ␥-secretase cleavage releases the 40 -42-amino acid amyloid ␤-peptide (A␤) from the membrane (1). The majority of familial Alzheimer disease cases are associated with mutations in the PS1 gene (1). Apparently all PS1 mutations investigated cause an increased generation of the highly amyloidogenic A␤42 (1). Absence of PS1 reduces ␥-secretase activity (2, 3) and absence of PS1/PS2 eliminates ␥-secretase function completely (4, 5). Mounting evidence suggests that PSs are unusual aspartyl proteases with ␥-secretase activity (6). All PSs contain two aspartates within transmembrane domains 6 and 7 that are critically required for ␥-secretase activity (7,8). Moreover, ␥-secretase inhibitors designed to mimic the transition state of the catalytic mechanism of aspartyl proteases can be covalently cross-linked to PSs (9, 10). Finally, PSs are apparently members of a group of polytopic membrane-bound aspartyl proteases that are all characterized by a GXGD (X ϭ variable amino acid) signature motif in which the C-terminal active site aspartate is embedded (11). Besides the PSs, the bacterial type 4 prepilin peptidases (11, 12) and signal peptide peptidase and its related proteins carry this signature motif (13,14).The PSs reside in high molecular weight complexes (15)(16)(17)(18)). An integral component of these high molecular weight complexes is the membrane glycoprotein nicastrin (Nct) (18 -20). Down-regulation of Nct in cultured mammalian or Drosophila cells inhibits ␥-secretase cleavage of APP (18, 21) and site 3 (S3) cleavage of Notch (21) and reduces PS levels (18,21,22). On the other side, absence of PS1 and PS1/PS2 causes a strong inhibition of Nct maturation (18, 23).PSs are not only required for the ␥-secretase-mediated processing of APP but also for intramembrane proteolysis of several other type I transm...

In Alzheimer’s disease, microglia cluster around β-amyloid deposits, suggesting that these cells are important for amyloid plaque formation, maintenance and/or clearance. We crossed two distinct APP transgenic mouse strains with CD11b-HSVTK mice, in which nearly complete ablation of microglia was achieved for up to 4 weeks after ganciclovir application. Neither amyloid plaque formation and maintenance nor amyloid-associated neuritic dystrophy depended on the presence of microglia.