Classically (M1) and alternatively activated (M2) macrophages exhibit distinct functions. In vivo, there is a spectrum of macrophage phenotypes exists, and in vitro, there is low or non-selective M2 marker protein expression. Therefore, by leveraging the wealth of knowledge that we have on macrophage marker development, Creative Biolabs performs the most comprehensive analysis of the transcriptional signature of M1 and M2 macrophages in humans and mice. Our strategy can identify M1-exclusive or M2-exclusive markers, which can be used to distinguish between M1 and M2 macrophages. Our skilled scientists provide efficiencies with shorter timelines to accelerate our clients' M1/M2 macrophage marker development projects.
There is a spectrum of macrophage phenotypes and functions, due to the diversity and overlap of cues in the microenvironment. Macrophages help maintain homeostasis. They are extremely plastic cells that are able to respond and adapt to external stimuli. The pro-inflammatory M1 and anti-inflammatory M2 are at the extremes. When there are lipopolysaccharide (LPS), tumor necrosis factor (TNF)-α, or interferon (IFN)-γ, macrophages adopt M1 phenotype and produce cytokines, such as IFN-γ, TNF-α, interleukin (IL)-6, IL-12, and reactive oxygen species (ROS). On the other hand, M2 macrophages are induced by IL-4, IL-10, IL-13, or glucocorticoids, which produce anti-inflammatory cytokines, such as transforming growth factor (TGF)-β and IL-10. They are characterized by their scavenger, angiogenic, and pro-invasive properties. As a consequence of the immunosuppressive tumor microenvironment, tumor-associated macrophages (TAMs) are reported to be M2-like macrophages, playing a critical role in tumor growth, invasion, and metastasis.
Historically, cDNA subtraction was used for M1/M2 marker identification started in mouse macrophages. To distinguish M1from M2 macrophages efficiently, a comprehensive transcriptome-base approach for M1/M2 macrophage marker identification and an effective validation method is highly needed.
Fig.1 Macrophage signature in classically activated M1 and alternatively activated M2 macrophages. (Jablonski, 2015)
Creative Biolabs has extensive experience in employing RNAseq and microarrays to evaluate how gene expression changes throughout the process of M1/M2 macrophage activation. These transcriptome-based approaches to characterize polarized macrophages offer a rich resource for identifying novel markers for M1/M2 macrophages. The markers we identified are M1-exclusive or M2-exclusive, and they are further confirmed by flow cytometry. After that, RT-PCR is usually used to examine a panel of transcripts to verify the reproducibility of the gene expression changes. This assay is also used to determine the expression kinetics of previously established markers as well as novel markers identified by this strategy. The markers identified could be used to distinguish between M1 and M2 macrophage, and better characterize TAM as well as other macrophage populations collected from humans and mice.
Creative Biolabs is well equipped and versed in M1/M2 macrophage marker discovery and development. Our scientists will work together closely to address the scientific and technical challenges to facilitate our clients' projects. For more detailed information, please feel free to contact us or send us your inquiry or question.