Aggregate-prone brain regions in Parkinson's disease are rich in unique N-terminus α-synuclein conformers with high proteolysis susceptibility.

In Parkinson's disease (PD), and other α-synucleinopathies, α-synuclein (α-Syn) aggregates form a myriad of conformational and truncational variants. Most antibodies used to detect and quantify α-Syn in the human brain target epitopes within the C-terminus (residues 96-140) of the 140 amino acid protein and may fail to capture the diversity of α-Syn variants present in PD. We sought to investigate the heterogeneity of α-Syn conformations and aggregation states in the PD human brain by labelling with multiple antibodies that detect epitopes along the entire length of α-Syn. We used multiplex immunohistochemistry to simultaneously immunolabel tissue sections with antibodies mapping the three structural domains of α-Syn. Discrete epitope-specific immunoreactivities were visualised and quantified in the olfactory bulb, medulla, substantia nigra, hippocampus, entorhinal cortex, middle temporal gyrus, and middle frontal gyrus of ten PD cases, and the middle temporal gyrus of 23 PD, and 24 neurologically normal cases. Distinct Lewy neurite and Lewy body aggregate morphologies were detected across all interrogated regions/cases. Lewy neurites were the most prominent in the olfactory bulb and hippocampus, while the substantia nigra, medulla and cortical regions showed a mixture of Lewy neurites and Lewy bodies. Importantly, unique N-terminus immunoreactivity revealed previously uncharacterised populations of (1) perinuclear, (2) glial (microglial and astrocytic), and (3) neuronal lysosomal α-Syn aggregates. These epitope-specific N-terminus immunoreactive aggregate populations were susceptible to proteolysis via time-dependent proteinase K digestion, suggesting a less stable oligomeric aggregation state. Our identification of unique N-terminus immunoreactive α-Syn aggregates adds to the emerging paradigm that α-Syn pathology is more abundant and complex in human brains with PD than previously realised. Our findings highlight that labelling multiple regions of the α-Syn protein is necessary to investigate the full spectrum of α-Syn pathology and prompt further investigation into the functional role of these N-terminus polymorphs.

PSA-NCAM Regulatory Gene Expression Changes in the Alzheimer's Disease Entorhinal Cortex Revealed with Multiplexed in situ Hybridization.

BACKGROUND: Alzheimer's disease (AD) is the most common form of dementia and is characterized by a substantial reduction of neuroplasticity. Our previous work demonstrated that neurons involved in memory function may lose plasticity because of decreased protein levels of polysialylated neural cell adhesion molecule (PSA-NCAM) in the entorhinal cortex (EC) of the human AD brain, but the cause of this decrease is unclear. OBJECTIVE: To investigate genes involved in PSA-NCAM regulation which may underlie its decrease in the AD EC. METHODS: We subjected neurologically normal and AD human EC sections to multiplexed fluorescent in situ hybridization and immunohistochemistry to investigate genes involved in PSA-NCAM regulation. Gene expression changes were sought to be validated in both human tissue and a mouse model of AD. RESULTS: In the AD EC, a cell population expressing a high level of CALB2 mRNA and a cell population expressing a high level of PST mRNA were both decreased. CALB2 mRNA and protein were not decreased globally, indicating that the decrease in CALB2 was specific to a sub-population of cells. A significant decrease in PST mRNA expression was observed with single-plex in situ hybridization in middle temporal gyrus tissue microarray cores from AD patients, which negatively correlated with tau pathology, hinting at global loss in PST expression across the AD brain. No significant differences in PSA-NCAM or PST protein expression were observed in the MAPT P301S mouse brain at 9 months of age. CONCLUSION: We conclude that PSA-NCAM dysregulation may cause subsequent loss of structural plasticity in AD, and this may result from a loss of PST mRNA expression. Due PSTs involvement in structural plasticity, intervention for AD may be possible by targeting this disrupted plasticity pathway.

N-terminal mutant huntingtin deposition correlates with CAG repeat length and symptom onset, but not neuronal loss in Huntington's disease.

Huntington's disease (HD) is caused by a CAG repeat expansion mutation in the gene encoding the huntingtin (Htt) protein, with mutant Htt protein subsequently forming aggregates within the brain. Mutant Htt is a current target for novel therapeutic strategies for HD, however, the lack of translation from preclinical research to disease-modifying treatments highlights the need to improve our understanding of the role of Htt protein in the human brain. This study aims to undertake an immunohistochemical screen of 12 candidate antibodies against various sequences along the Htt protein to characterize Htt distribution and expression in post-mortem human brain tissue microarrays (TMAs). Immunohistochemistry was performed on middle temporal gyrus TMAs comprising of up to 28 HD and 27 age-matched control cases, using 12 antibodies specific to various sequences along the Htt protein. From this study, six antibodies directed to the Htt N-terminus successfully immunolabeled human brain tissue. Htt aggregates and Htt protein expression levels for the six successful antibodies were subsequently quantified with a customized automated image analysis pipeline on the TMAs. A 2.5-12 fold increase in the number of Htt aggregates were detected in HD cases using antibodies MAB5374, MW1, and EPR5526, despite no change in overall Htt protein expression compared to control cases, suggesting a redistribution of Htt into aggregates in HD. MAB5374, MW1, and EPR5526 Htt aggregate numbers were positively correlated with CAG repeat length, and negatively correlated with the age of symptom onset in HD. However, the number of Htt aggregates did not correlate with the degree of striatal degeneration or the degree of cortical neuron loss. Together, these results suggest that longer CAG repeat lengths correlate with Htt aggregation in the HD human brain, and greater Htt cortical aggregate deposition is associated with an earlier age of symptom onset in HD. This study also reinforces that antibodies MAB5492, MW8, and 2B7 which have been utilized to characterize Htt in animal models of HD do not specifically immunolabel Htt aggregates in HD human brain tissue exclusively, thereby highlighting the need for validated means of Htt detection to support drug development for HD.

Tumour infiltrating lymphocyte density differs by meningioma type and is associated with prognosis in atypical meningioma.

Tumour infiltrating lymphocyte (TIL) density is prognostically significant in various tumours, but few studies have investigated its significance in meningioma. This study aimed to investigate how TIL density differs by meningioma histology and whether it is a predictor of meningioma recurrence. We studied CD3, CD8, CD4, FOXP3 and PD-1 positive (+) TIL density in a continuous cohort of 476 meningiomas resected at Auckland Hospital between 2002 and 2011 using tissue microarrays and computer assisted image analysis. TILs were identified in all meningiomas except one (median CD3+ TIL density across entire cohort 53.0 cells/mm2). Most TILs were CD8+ (median 33.6 cells/mm2) with smaller numbers of CD4+ TILs (median 2.9 cells/mm2). PD-1+ (median 0.32 cells/mm2) and FOXP3+ (median 0.0 cells/mm2) TILs were scarce. Reduced CD3+ (p=0.0066), CD8+ (p=0.0029) and PD-1+ (p=0.0375) TIL density was seen in WHO grade II/III meningioma compared with WHO grade I. Pairwise comparison confirmed statistically significant differences in TIL density existed between meningioma types (CD3, CD8, CD4, p<0.0001; FOXP3, p=0.0096; PD-1, p=0.0090) with chordoid meningioma having the lowest overall CD3+ TIL density (median 12.5 cells/mm2). Despite its low TIL density, chordoid meningioma had a higher FOXP3:CD8 ratio than several meningioma types. Atypical meningioma had a higher FOXP3:CD8 ratio than transitional meningioma (p=0.0045). No association between TIL density and recurrence was seen across the entire cohort or by WHO grade. However, CD3+ and CD8+ TIL density was associated with recurrence in atypical meningioma on multivariable analysis (CD3, p=0.0012; CD8, p=0.0071). A higher CD3+ and CD8+ TIL density was associated with improved recurrence free survival. Our findings suggest CD3+ and CD8+ TIL density is prognostically significant in atypical meningioma. Further investigation of this observation and its biological basis is warranted. The differences in TIL density by meningioma histology may be of relevance in studies of therapeutic immune checkpoint inhibition.

The epidemiology of patients undergoing meningioma resection in Auckland, New Zealand, 2002 to 2011.

The incidence of meningioma is known to vary by gender and ethnicity. This study aimed to describe the epidemiological characteristics of a 10-year cohort of patients undergoing meningioma resection at Auckland City Hospital, Auckland, New Zealand. Of particular interest was whether there was any difference in meningioma incidence and recurrence rates between New Zealand Maori and Pacific Island patients compared with other ethnic groups. The study was a retrospective analysis of 493 patients with pathologically confirmed meningioma over the period 1 January 2002 to 31 December 2011. Based on this neurosurgical cohort, the minimum incidence of meningioma in the Auckland region was 3.39 per 100,000 population per year (95% C.I. 3.02-3.80) for the study period. Meningioma was significantly more common in women than men by a ratio of 4.2:1. New Zealand Maori and Pacific Island patients had a significantly higher incidence of meningioma than other ethnic groups. New Zealand Maori had a meningioma incidence 2.74 times that of Europeans (95% C.I. 2.01-3.73, p < 0.001). Pacific Island patients had 2.03 times higher incidence of meningioma than Europeans (95% C.I. 1.42 - 2.89, p < 0.001). The overall meningioma recurrence rate was 21.6% with a mean follow-up of 77 months. Recurrence rates for meningioma among Pacific Island patients were significantly higher than for other ethnic groups (hazard ratio 1.73, p = 0.008). Multivariate analysis of clinical variables confirmed the significance of traditional prognostic factors such as WHO tumour grade and Simpson grade of surgical excision in predicting meningioma recurrence.

Isolated gluteal and paravertebral muscle weakness due to anti-3-hydroxy-3-methylglutaryl-coenzyme a reductase antibody-associated necrotizing autoimmune myopathy.

INTRODUCTION: A 56-year-old man with a distant history of statin use presented with progressive isolated very proximal lower limb and truncal weakness. Electromyogram (EMG) showed isolated gluteal and lumbar paraspinal muscle involvement. METHODS: Gluteus medius muscle biopsy was performed under general anesthesia. RESULTS: The biopsy showed a pauci-inflammatory necrotizing myopathy. Serum antibodies to 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGCR) were positive. He has since partially responded to corticosteroids and methotrexate. CONCLUSIONS: Anti-HMGCR-associated necrotizing autoimmune myopathy (NAM) can present in a restricted form after cessation of a statin. Biopsy of a symptomatic but uncommonly studied muscle is worthwhile. Muscle Nerve 54: 150-152, 2016.