Active glycogen synthase kinase-3 and tau pathology-related tyrosine phosphorylation in pR5 human tau transgenic mice

We studied underlying pathomechanisms in tauopathies using pR5 mice that express the P301L tau mutation found in familial forms of frontotemporal dementia. In a longitudinal study we investigated the functional status of glycogen synthase kinase-3 and correlated it with the appearance of distinct tau phospho-epitopes. Neurons displaying increases in activating phosphorylation of glycogen synthase kinase-3α/β at tyrosine 279/216 also showed an intense rather than moderate AT8 (phospho-Ser202/Thr205 tau) immunoreactivity, and immunoreactivity for AT100 (phospho-Ser212/Thr214 tau) and phosphorylated Ser422, phospho-epitopes associated with fibrillar tau pathology. These neurons were rare in 8.5-month-old, but numerous in 18.5- and 28-month-old pR5 mice. Two antibodies that detect phosphotyrosine residues more generally only stained these neurons. In contrast, we did not find increased phosphotyrosine in neuronal perikarya of mice with an amyloid-β plaque pathology. Our results suggest a link between increased tyrosine phosphorylation and tau aggregation. They also reveal for the mouse models studied, that tau- rather than an amyloid-β peptide-induced pathology is associated with increased neuronal tyrosine phosphorylation

Analysis of ribosomal protein S6 baseline phosphorylation and effect of tau pathology in the murine brain and human hippocampus

We examined the distribution pattern of the phosphorylated 40S ribosomal subunit protein S6, a downstream target of the mTOR pathway, in the brains of 24-months-old human tau transgenic pR5 mice, non-transgenic littermates and in human hippocampi. We studied baseline levels of phosphorylated S6 and a possible effect of tau pathology. S6 phosphorylated at Ser235/236 (pS6Ser235/236) or Ser240/244 (pS6Ser240/244) has been used as a read-out of mTOR activity in several studies. The mTOR pathway regulates a wide variety of cellular functions including cell growth, ribosome biosynthesis, translational control and autophagy. Its dysregulation might underlie the neurodegenerative pathology of Alzheimer's disease and other tauopathies. pS6Ser235/236 and pS6Ser240/244 immunoreactivity in the mouse brain were widespread and similar distributed, but intensive pS6Ser235/236 immunoreactivity was more selective, especially highlighting certain brainstem regions. In the human hippocampus mainly granulovacuolar inclusions in neurons displayed pS6Ser235/236 immunoreactivity. In contrast, a considerable number of neurons displayed pS6Ser240/244 immunoreactivity in the cytoplasm without labeling of granulovacuolar inclusions. Except for a tendency of lower numbers of intensely phosphorylated S6 positive neurons in pR5 mice, the pattern of distribution of pS6Ser235/236 and pS6Ser240/244 immunoreactivity was largely unchanged when compared with non-transgenic mice and also when human hippocampi from AD cases and controls were compared. Similar to pR5 mice most neurons with hyper-phosphorylated tau in human hippocampi displayed no or only weak labeling for phosphorylated S6, suggesting that phosphorylated S6 is not especially associated with pathological tau, but is rather a feature of unaffected neurons. (C) 2017 Elsevier B.V. All rights reserved

Granulovacuolar degeneration and unfolded protein response in mouse models of tauopathy and A beta amyloidosis

Histopathological studies on the brains of tauopathy cases including cases with Alzheimer's disease (AD) demonstrate that neurons with hyperphosphorylated protein tau display granulovacuolar degeneration (GVD), as evidenced by vacuolar lesions harboring a central granule, together with markers of the activated unfolded protein response (UPR). In order to examine whether this hallmark is reproduced in animal models we studied the presence of GVD and the activated UPR in two complementary mouse models, pR5 mice with a tau pathology and APPSLxPS1mut mice with an amyloid plaque pathology. Neither GVD nor a significant activation of the UPR was found in both APPSLxPS1mut mice and in those regions in the pR5 brain where only neurons with an early stage of tau hyperphosphorylation were present In contrast, those neurons that displayed a tau phospho-epitope signature that only appeared in old pR5 mice and also correlated with Gallyas-positive tangle staining harbored granulovacuolar lesions that were labeled with the GVD markers casein kinases 1 delta and 1 epsilon. Granulovacuolar lesions in pR5 mice were also labeled with the UPR markers phosphorylated PKR-like endoplasmic reticulum kinase, phosphorylated inositol-requiring enzyme 1 alpha and phosphorylated eukaryotic initiation factor 2 alpha. However, GVD was rarely observed in neurons bearing mature neurofibrillary tangles as evidenced by Congo red staining. Our results suggest that NFT-formation activates the UPR in pR5 mice and that it is the early stages of neurofibrillary tangle formation that are accompanied by GVD, in line with observations from studies on human autopsy cases. (C) 2014 Elsevier Inc. All rights reserved

Distribution of spleen tyrosine kinase and tau phosphorylated at tyrosine 18 in a mouse model of tauopathy and in the human hippocampus

Purpose: Spleen tyrosine kinase (Syk) has been shown to phosphorylate tyrosine 18 of tau in vitro. It has been proposed that increased immunoreactivity for double-phosphorylated Syk in hippocampal neurons of Alzheimer's disease cases indicates a not yet defined neurodegenerative process. To investigate this possibility we have studied Syk and tau phosphorylated at tyrosine 18 (pTyr18) in transgenic mice and human hippocampi. Methods: We performed immunohistochemistry, immunofluorescence labeling and Western blotting and compared the distribution of Syk double-phosphorylated at tyrosines 525 and 526 and pTyr18 in human tau transgenic pR5 mice and human hippocampi with low and high Braak stages for neurofibrillary tangle pathology. Results: pTyr18 appeared early during the course of neurodegeneration in pR5 mice and was widely distributed in the pR5 brain, including neuronal somata and fiber tracts. In contrast, only strongly pTyr18- and AT100-(tau phosphorylated at Thr212 and Ser214) positive neurons with a fibrillary tau pathology in old pR5 mice and microglia displayed immunoreactivity for double-phosphorylated Syk. In human hippocampi, phosphorylated Syk was mainly present in granulovacuolar inclusions in hippocampal pyramidal neurons and did not co-locate with pTyr18 in these neurons. We observed pTyr18-positive neurons and neurons with granular pSyk immunoreactivity already at early Braak stages and their number was markedly increased in Braak stage VI. Conclusion: Syk appears unlikely to be the major kinase that phosphorylates tyrosine 18 of tau in tauopathy. It possibly phosphorylates tyrosine 18 of tau and regulates other tau kinases in neurons with a fibrillary tau pathology. (C) 2017 Elsevier B.V. All rights reserved

Variants of arterial supply to the inferior (diaphragmatic) surface of the ventricles of the heart and the influence on age at death

Coronary artery systems of the inferior wall of the ventricles vary considerably. Schlesinger's concept distinguishes dominance of the right or left coronary artery (LCA) or balanced type. LCA dominance has been reported to be associated with increased mortality. Early angiography studies have shown that the anterior interventricular artery (AIVA), a branch of the LCA, often continues on the inferior surface of the heart and may replace the inferior interventricular artery. Others considered an AIVA on the inferior surface of the heart a rare variant. A long AIVA is a strong predictor of death in acute anterior wall myocardial infarction. We determined coronary artery variance at the inferior surface of the ventricles in 134 dissected human hearts and analyzed a possible association between coronary artery variance and age at death. The AIVA extended to the inferior side in 64.9% of the hearts, but rarely reached the basal half of the inferior interventricular groove. Most frequently (53%), it extended into the apical two-fifths of the length of the inferior ventricular walls. An AIVA extending to the apical 40% of the length of the inferior ventricular walls may therefore be considered a common variant. In 20.1% of the hearts, a right inferior diagonal artery was also found. Statistical analysis neither revealed an association between mean AIVA length at the inferior ventricular surface and type of coronary artery dominance nor an association between AIVA length at the inferior ventricular surface or coronary artery dominance type and age at death

The relation between tau pathology and granulovacuolar degeneration of neurons

Neurofibrillary tangles arising from aggregated microtubule-associated protein tau occur in aged brains and are hallmarks of neurodegenerative diseases. A subset of neurons containing aggregated tau displays granulovacuolar degeneration (GVD) that is characterized by membrane-bound cytoplasmic vacuoles, each containing an electron dense granule (GVB). Tau pathology induces GVBs in experimental models, but GVD does not generally follow tau pathology in the human brain. The entorhinal cortex, DRN, and LC are among the regions that display pathological changes of tau earliest, whereas neurons with GVBs occur first in the hippocampus and have been found in oral raphe nuclei only at the most advanced GVD stage. To date, there is no detailed report about neurons with GVD in aminergic nuclei. We studied the relation between tau pathology and GVD in field CA1 of the hippocampus, entorhinal cortex, dorsal (DRN) and median (MRN) raphe nucleus, and locus coeruleus from elderly subjects with Braak & Braak stages of tau pathology ranging from 0 to VI. Tau pathology and GVBs were visualized by means of immunolabeling and quantified. Percentages of neurons containing GVBs were significantly related to percentages of AT8-positive neurons in the regions examined. GVD and tau pathology were found together in neurons to a different extent in regions of the brain. 53.2% of AT8-immunoreactive neurons in CA1, 19.8% in layer II of the entorhinal cortex, 29.6% in the DRN, and 31.4% in the locus coeruleus contained GVBs. Age-related factors, the percentage of neurons with pretangles in a region of the brain, and the metabolism of a neuron possibly influence the prevalence of neurons with GVBs

Effect of tau-pathology on charged multivesicular body protein 2b (CHMP2B)

Charged multivesicular body protein 2b (CHMP2B) is a subunit of the endosomal sorting complex required for transport (ESCRT)-III that mediates scission of budded membranes. Neurons with CHMP2B-positive granulovacuolar inclusions in the cytoplasm are much more frequent in hippocampi of cases with Alzheimer's disease when compared with controls. We analyzed immunolabeled brain sections from tau-transgenic mice, APP-transgenic mice, non-transgenic mice, and human hippocampi to investigate the relation between CHMP2B and tau and plaque pathology that are major histopathological features of Alzheimer's disease. Neurons undergoing granulovacuolar degeneration (GVD) were found in human hippocampi and old tau-trangenic mice but not in the APP-transgenic strains. 57% of neurons with GVD displayed GVD-granules double-labeled for CHMP2B and the GVD-marker casein kinase 1 delta in 24 months-old tau-transgenic mice and 5.7% of neurons with tau hyperphosphorylated at Thr212 and Ser214 (immunoreactive with antibody AT100) displayed CHMP2B-positive GVD-granules, in human hippocampi it was 100% and 46% respectively. The number of neurons with GVD-inclusions increased in tau-transgenic mice with the number of AT100-positive neurons, suggesting a link between tau-pathology and GVD. GVD-granules in human hippocampi also displayed immunoreactivity for Vps4a, another protein component of ESCRT-III. In cases with aging-related tau astrogliopathy (ARTAG), astrocytes containing hyper-phosphorylated tau immunoreactive with antibody AT8 displayed strong CHMP2B immunoreactivity. The results suggest dysregulation of CHMP2B together with tau-pathology and possibly a disturbance of the regulation of vesicular compartments. The absence of combined A beta- and tau-associated pathology in the transgenic mice may account for the difference in CHMP2B-immunoreactivity between the transgenic mice and human hippocampus