Hassan M Rostam
پوخته
Macrophages as antigen presenting cell (APC) play a vital role in orchestrating immune responses against foreign materials. The activation status of macrophages could be determined ...
زیاتر بخوێنەوە
Macrophages as antigen presenting cell (APC) play a vital role in orchestrating immune responses against foreign materials. The activation status of macrophages could be determined by tracking the expression of various cell markers that can be a signal for their immune activity behaviour following cellular stimulation either towards healing or inflammation. Previously numerous immunofluorescent cell markers have been used for distinguishing between pro-inflammatory macrophage phenotype (M1) and anti-inflammatory macrophage phenotype (M2) qualitatively, although most of those fluorescent cell markers express in both phenotypes. We have developed a new strategy to identify M1 and M2 phenotype quantitatively by using immunofluorescent cell markers. This approach enables the identification of different macrophage functional phenotypes quantitatively, and their degree of polarisation. Macrophages were polarised to M1 and M2 phenotypes by GM-CSF+IFN-γ and M-CSF+IL-4, respectively. Control cells were un-polarised (naïve) macrophages or monocytes were considered as macrophage progeny. For assessing cell polarisation all cell types were stained for nucleus. Also, their surface markers were stained with calprotectin for M1 cells and mannose receptor (MR) for M2 cells, followed by fluorescent microscopy examination. Cell images were analysed using CellProfiler software in order to measure the fluorescent signal intensity of the cell markers, and create a specific profile for each cell type. These profiles formed the basis for M1 and M2 phenotype identification. By using such fluorescent signal parameters we were able to identify M1 and M2 phenotypes effectively and distinguish them from naïve macrophages and monocytes.
