Characterisation of premature cell senescence in Alzheimer's disease using single nuclear transcriptomics.

Aging is associated with cell senescence and is the major risk factor for AD. We characterized premature cell senescence in postmortem brains from non-diseased controls (NDC) and donors with Alzheimer's disease (AD) using imaging mass cytometry (IMC) and single nuclear RNA (snRNA) sequencing (> 200,000 nuclei). We found increases in numbers of glia immunostaining for galactosidase beta (> fourfold) and p16INK4A (up to twofold) with AD relative to NDC. Increased glial expression of genes related to senescence was associated with greater ß-amyloid load. Prematurely senescent microglia downregulated phagocytic pathways suggesting reduced capacity for ß-amyloid clearance. Gene set enrichment and pseudo-time trajectories described extensive DNA double-strand breaks (DSBs), mitochondrial dysfunction and ER stress associated with increased ß-amyloid leading to premature senescence in microglia. We replicated these observations with independent AD snRNA-seq datasets. Our results describe a burden of senescent glia with AD that is sufficiently high to contribute to disease progression. These findings support the hypothesis that microglia are a primary target for senolytic treatments in AD.

A single nuclear transcriptomic characterisation of mechanisms responsible for impaired angiogenesis and blood-brain barrier function in Alzheimer's disease.

Brain perfusion and blood-brain barrier (BBB) integrity are reduced early in Alzheimer's disease (AD). We performed single nucleus RNA sequencing of vascular cells isolated from AD and non-diseased control brains to characterise pathological transcriptional signatures responsible for this. We show that endothelial cells (EC) are enriched for expression of genes associated with susceptibility to AD. Increased ß-amyloid is associated with BBB impairment and a dysfunctional angiogenic response related to a failure of increased pro-angiogenic HIF1A to increased VEGFA signalling to EC. This is associated with vascular inflammatory activation, EC senescence and apoptosis. Our genomic dissection of vascular cell risk gene enrichment provides evidence for a role of EC pathology in AD and suggests that reducing vascular inflammatory activation and restoring effective angiogenesis could reduce vascular dysfunction contributing to the genesis or progression of early AD.

Elevated Expression of Two Pore Potassium Channel THIK-1 in Alzheimer's Disease: An Inflammatory Mechanism.

INTRODUCTION: Tandem pore domain halothane-inhibited K+ channel 1 (THIK-1, coded by KCNK13) provides an upstream regulation of the activation of the NOD-like receptor pyrin domain containing 3 (NLRP3) inflammasome, which has been suggested as one of the key mechanisms of the pathological process in neurodegeneration mainly from in vitro and in vivo model systems studies. However, unequivocal evidence from neurodegenerative disorders has been lacking. OBJECTIVE: To investigate the involvement of the THIK-1/NLRP3 pathway in the pathological process of Alzheimer's disease (AD) and Parkinson's disease (PD). METHODS: This study investigated gene expression of markers in the THIK-1/NLRP3 pathway in an animal model representing AD as well as in human postmortem brains of AD and PD by quantitative real-time PCR. THIK-1 protein expression was determined using automated capillary electrophoresis immunoblotting. Furthermore, DNA methylation of KCNK13 was analysed in AD cohort by pyrosequencing. RESULTS: A substantial upregulation of KCNK13, glial activation markers, NLRP3 inflammasome components, and IL1B was observed in the animal study. Increased expression of KCNK13 support an inflammatory glial cell activation in both advanced AD and PD. The increase in KCNK13 expression was also supported by downregulation in DNA methylation of KCNK13 in AD. CONCLUSIONS: The association between THIK-1 K+ channels expression and pathology changes indicates a THIK-1-induced activation of this glial subtype in AD and PD. Therefore, specific blocks of the microglial THIK-1 K+ channels at the early stage of AD and PD may be beneficial for the patients.

Streptococcus suis in retail markets: how prevalent is it in raw pork?

Streptococcus suis present in raw pork meats sold in local retail markets was enumerated by Most Probable Number (MPN)-PCR method. This method combined the conventional MPN technique with a specifically designed PCR assay based on the amplification of a 294-bp S. suis species-specific 16S rRNA gene sequence. A total of 78 raw pork lean meat samples purchased at two different supermarkets (Site A and B) and a wet market (Site C) were tested. Results indicated that S. suis could be detected from the enriched MPN tubes of all, except one, sample homogenates. The concentration of S. suis ranged from <3 to 4600 MPN/g of pork meat, with a total bacterial count (TBC) varying from 3.6 log to 7.4 log CFU/g. Statistical analyses indicated that pork meats purchased from the supermarket at Site B in summer contained significantly higher concentration of S. suis organisms than those from other retailers in any season. A significant correlation existed between log S. suis concentration and log TBC of the samples. This study revealed that raw pork meats available in local supermarkets or wet markets could contain S. suis at concentrations that were usually difficult to detect with traditional culture method. Field application of this method may contribute to a measurable evaluation, and thus the effective control, of human S. suis infection due to raw pork or pig carcass handling.

Coronaviral hypothetical and structural proteins were found in the intestinal surface enterocytes and pneumocytes of severe acute respiratory syndrome (SARS).

Severe acute respiratory syndrome (SARS) is a newly emerging infectious disease that haunted the world from November 2002 to July 2003. Little is known about the biology and pathophysiology of the novel coronavirus that causes SARS. The tissue and cellular distributions of coronaviral hypothetical and structural proteins in SARS were investigated. Antibodies against the hypothetical (SARS 3a, 3b, 6, 7a and 9b) and structural proteins (envelope, membrane, nucleocapsid and spike) of the coronavirus were generated from predicted antigenic epitopes of each protein. The presence of these proteins were first verified in coronavirus-infected Vero E6 tissue culture model. Immunohistochemical studies on different human tissues, including a cohort of nine autopsies, two liver biopsies and intestinal biopsies of SARS patients, further confirmed the existence of coronaviral hypothetical and structural proteins in the cytoplasm of pneumocytes and small intestinal surface enterocytes in SARS patients. With this vast array of antibodies, no signal was observed in other cell types including those organs in which reverse transcriptase-polymerase chain reactions were reported to be positive. Structural proteins and the functionally undefined hypothetical proteins were expressed in coronavirus-infected cells with distinct expression pattern in different organs in SARS patients. These antipeptide antibodies can be useful for the diagnosis of SARS at the tissue level.