Project description:Vascular cell types are under-represented in standard single-nucleus RNA-Seq studies of human frozen post-mortem brain tissue. The dataset represents the pilot data obtained with the first version of a microvessel- enrichment protocol to obtain a higher percentage of endothelial cells as the key cellular component of the blood-brain barrier.
Project description:Proteomic characterization of human brain tissue is increasingly utilized to identify potential novel biomarker and drug targets for a variety of neurological diseases. In whole tissue studies, results may be driven by changes in the proportion of the largest and most abundant organelles or tissue cell-type composition. Spatial proteomics approaches enhance our knowledge of disease mechanisms and changing signaling pathways at the subcellular level by taking into account the importance of cellular localization, which critically influences protein function. Density gradient-based ultracentrifugation methods allow for subcellular fractionation and have been utilized in cell lines and fresh mouse brain tissue to predict the subcellular location of thousands of proteins, but to our knowledge have not been used in frozen human brain tissue. The use of frozen human tissue for tissue fractionation faces two major challenges, the post-mortem interval, during which proteins may leach from their usual location into the cytosol, and freezing, which results in membrane breakdown. Despite these challenges, in this proof-of-concept study, we show that the majority of proteins segregate reproducibly into crude density-based centrifugation fractions, that the fractions are enriched for the appropriate organellar markers, and that significant differences in protein localization can be observed between tissue from individuals with Alzheimer’s Disease and control individuals.
Project description:We set up a pilot study using Affymetrix Gene Chip® Porcine Genome Arrays to evaluate the impact of time lags from death on gene expression profiling of porcine skeletal muscle at four post mortem time points (up to 24 hrs) during the routine processing of fresh tights Post mortem skeletal muscle samples were obtained from three commercial hybrid pigs of female sex (of about 160 kg each) raised in the same commercial farm and slaughtered under normal conditions at the same abattoir within 1 minute of each other after stunning by CO2 (concentration 87%) using a dip lift system (Butina, Denmark). A portion of semimembranosus muscle (3-5 g) was sampled from the left legs at 20 minutes (T0) after death following the normal operation of the abattoir. Other samples were collected from the same muscle at the same position after 2 (T1), 6 (T2) and 24 (T3) hrs post mortem in the same abattoir following the cold chain at the abattoir until 4 °C. After sampling, tissues were snap frozen in liquid nitrogen and stored at -80 °C till RNA extraction.
Project description:We set up a pilot study using Affymetrix Gene Chip® Porcine Genome Arrays to evaluate the impact of time lags from death on gene expression profiling of porcine skeletal muscle at four post mortem time points (up to 24 hrs) during the routine processing of fresh tights
Project description:We performed high-throughput snRNA-seq using the 10X Genomics Chromium platform on archived post-mortem dorsolateral prefrontal cortex (BA9) tissue in MDD subjects who died by suicide and in control subjects to identify cell-type specific differentially expressed genes.
