Session Title: Pediatric Rheumatology – Pathogenesis and Genetics - Poster
Session Type: ACR Poster Session C
Session Time: 9:00AM-11:00AM
Background/Purpose: CD163 is involved in the regulation and resolution of innate inflammation and the removal of free hemoglobin from the blood via internalization of the hemoglobin-haptoglobin complex. It is also a cell surface marker of alternatively activated macrophages overexpressed in inflammatory disorders such as systemic juvenile idiopathic arthritis. To better understand the activation of myeloid cells in rheumatic disorders, a flow cytometry panel has been developed to define polarized monocyte and macrophage phenotypes. Along with detecting surface protein markers, the novel technique called PrimeFlow™ allows specific mRNA sequences to be detected at single cell resolution. This panel was utilized to define the expression of CD163 in polarized monocyte and macrophage populations.
Results: PrimeFlow™ staining of CD163 mRNA could reliably detect increased CD163 expression in THP-1 monocytic cells, primary CD14+ blood monocytes, and monocyte-derived macrophages. The PrimeFlow™ CD163 mRNA probe was also used simultaneously to detect protein surface markers, and this flow cytometry panel provided the same degree of positively stained cells as the results of staining for individual cell surface markers without mRNA labeling. This combined panel demonstrated that some alternatively activated populations, such as M2a (IL-4 conditions), show an increase in CD163 surface protein expression with no change in CD163 mRNA levels. In marked contrast and in agreement with qPCR data, increase in both CD163 mRNA and protein levels is limited to the IL-10 stimulated M2c conditions. The PrimeFlow™ technique was determined to be highly sensitive for increased mRNA expression, with a strong signal over a broad range of IL-10 concentrations in contrast to qPCR data and surface protein detection. The kinetics of CD163 induction with IL-10 stimulation were shown to be similar for both mRNA and protein. There is minimal increase in both mRNA and protein expression after 5 hours of stimulation, but significant increase after 24 hours.
To cite this abstract in AMA style:Tan R, Thornton S, Sproles A, Do T, Schick J, DeLay M, Schulert G. Single-Cell Analysis of CD163 mRNA and Protein Expression By Primeflow™ in Polarized Monocyte and Macrophage Populations [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/single-cell-analysis-of-cd163-mrna-and-protein-expression-by-primeflow-in-polarized-monocyte-and-macrophage-populations/. Accessed December 2, 2020.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/single-cell-analysis-of-cd163-mrna-and-protein-expression-by-primeflow-in-polarized-monocyte-and-macrophage-populations/