GSE158446
Critical Role of Synovial Tissue-resident Macrophage Niche in Joint Homeostasis and Suppression of Chronic Inflammation
During synovial tissue homeostasis, both monocyte-derived F4/80int, and self-renewing F4/80hi tissue-resident, macrophage populations were identified. In contrast, in HUPO mice, decreased synovial tissue-resident macrophages preceded chronic arthritis, opened a niche permitting the influx of activated monocytes, which differentiated into F4/80hi macrophages. Overall design: HUPO mice with established arthritis and littermate or age/gender matched control mice were sacrificed to harvest blood and ankle cells for RNA-seq. CM and NCM and 5 subsets of synovial tissue macrophages from ankles were stained as described, then sorted by flow cytometryacross 6 control and 7 HUPO populations of cells.   Quality control for the 49 samples was performed excluding5 low quality samples (alignment < 82%, duplicates > 66%, and mapped reads< 2.3x106) from groups with >3 replicates. To maintain consistent numbers of replicates (n=3) in each experimental group, we removed the least correlated sample from an additional 3 control mice under the assumption that the most similar samples better reflect steady-state conditions. We also removed 2 HUPO samples from F4/80hisubsets which appeared to have been contaminated based on the high expression of neutrophil-specific genes. Thus, 39 samples reflecting 3 replicates for each of the 13 groups remained. We defined 7,778 “expressed genes” with log2(FPKM+1)= 4 in at least 2 out of the39 individual samples.  K-means clustering for control mice and comparing HUPO and control mice.  The GENE-E software (http://softerware.broadinstitute.ord/GENE-E/) was employed for the pairwise Pearson's correlation and K-means clustering analyses, performed using the default settings. Principle component analysis (PCA) was performed using the prcomp functionin R with the FPKM matrix of the expressed genes. Gene ontology (GO)associations were determined by Gorilla(http://cbl-gorilla.cs.technion.ac.il).