]. Briefly, histograms were plotted using the regulon activity matrix , in which columns represent cells and rows represent the AUC regulon activity and the targetome of individual TFs, both which were queried using the regulon data frame .The average expression of microglia genes from each developmental stage was measured using the “AverageExpression” function in Seurat.
Slides were imaged using a Zeiss Axio Observer at 20× for representative images, and 40× for corrected total cell fluorescence quantification. In short, CTCF values were generated by manually segmenting microglia three separate times to obtain average total microglial area, mean intensity, and integrated density. Adjacent background selections of the same area as each combination of selected microglia were also taken simultaneously for comparison.
To study changes in the transcriptional profile of microglia over time, human brain samples were collected at different developmental stages. Pre-natal samples were obtained from second trimester pre-natal brain tissue (Fig.
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Analysis of N-linked Glycan Alterations in Tissue and Serum Reveals Promising Biomarkers for Intrahepatic CholangiocarcinomaThere is an urgent need for the identification of reliable prognostic biomarkers for patients with intrahepatic cholangiocarcinoma (iCCA) and alterations in N-glycosylation have demonstrated an immense potential to be used as diagnostic strategies for many cancers, including hepatocellular carcinoma (HCC). N-glycosylation is one of the most common post-translational modifications known to be altered based on the status of the cell. N-glycan structures on glycoproteins can be modified based on the addition or removal of specific N-glycan residues, some of which have been linked to liver diseases. However, little is known concerning the N-glycan alterations that are associated with iCCA. We characterized the N-glycan modifications quantitatively and qualitatively in three cohorts, consisting of two tissue cohorts: a discovery cohort (n=104 cases) and a validation cohort (n=75), and one independent serum cohort consisting of patients with iCCA, HCC, or benign chronic liver disease (n=67). N-glycan analysis in situ was correlated to tumor regions annotated on histopathology and revealed that bisected fucosylated N-glycan structures were specific to iCCA tumor regions. These same N-glycan modifications were significantly upregulated in iCCA tissue and serum relative to HCC and bile duct disease, including primary sclerosing cholangitis (PSC) (P | 0.0001). N-glycan modifications identified in iCCA tissue and serum were used to generate an algorithm that could be used as a biomarker of iCCA. We demonstrate that this biomarker algorithm quadrupled the sensitivity (at 90% specificity) of iCCA detection as compared with carbohydrate antigen 19-9, the current “gold standard” biomarker of CCA.Significance:. This work elucidates the N-glycan alterations that occur directly in iCCA tissue and utilizes this information to discover serum biomarkers that can be used for the noninvasive detection of iCCA.
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