Many rules in biology exist, and even more exceptions to these rules. For years, immunologists have tried to classify macrophages into two subtypes, classically activated M1, and alternatively activated M2 macrophages. However, with the growing number of activated macrophage subtypes needing description, established nomenclature falls short. Researchers are seeking an alternative to the traditional black-and-white principle.


Optimal culture conditions for macrophages:
Dr. Hagen Wieland from the development department at PromoCell works on standardizing solutions for cell generation and differentiation.
Macrophage polarization is a dynamic process
Researchers have tried to classify macrophages based either on their belonging to the mononuclear phagocytic system, their origin, or their functional phenotype. In the early 1990s, Michael Stein and colleagues described the different effects of interleukin-4 (IL-4) compared to interferon gamma (IFN-γ) and/or lipopolysaccharide (LPS) on the activation phenotype of macrophages. The term “alternative activation” was created to describe IL-4 -stimulated macrophages, and to distinguish them from “classical activation” of IFN-γ /LPS -stimulated macrophages. Several years later, the M1 (classically activated) - M2 (alternatively activated) model was introduced to describe the two different macrophage responses, reflecting the T-helper cells nomenclature (Mills et al., 2000). M1 macrophages were described as having inflammatory functions, and were crucial for the resistance to pathogens and their elimination. M2 macrophages had anti-inflammatory functions, and were important for maintaining tissue integrity. “The limitations of the M1/M2 model for defining macrophage polarization are becoming more and more clear, as these two terms identify extremes of a continuum that do not occur in vivo,” remarks Wieland. Macrophages exist usually between M1 and M2 as the polarization process is dynamic, and cells often display characteristics of both states at the same time.
A spectrum of macrophage activation stages:
The traditional M1/M2 model is not satisfactory to reflect the heterogeneity of macrophages. See full reference list in our Application Note about cryopreserved macrophages.
Recommendations for a uniform nomenclature of macrophages
Various attempts to account for the cells’ plasticity and to classify the macrophages into different subpopulations have resulted in an unclear nomenclature and confusing categorization. This diversity of terminology makes it difficult to describe scientific results clearly, and impedes scientific progress and translational approaches. In the search for a solution to the problem, a group of researchers met at the International Congress of Immunology in Milan in August 2013. There they drafted a macrophage-activation nomenclature and reporting standards for in vitro experiments. The common framework proposal was then published one year later (Murray et al., 2014), and included the following recommendations:- Reproducible in vitro experimental standards: Peripheral blood monocytes in humans and bone marrow-derived monocytes in mice should be used as references for generating macrophages that can be then stimulated with IFN-γ or IL-4 to give rise to defined subpopulations. In addition, scientists should note whether macrophages were polarized using M-CSF or GM-CSF.
- Minimal reporting standards: Precise descriptions of how macrophages are isolated, polarized, activated, and analyzed are crucial, and allow a direct comparison among different laboratories.
- Definition of the activator: Researchers should describe how they activate the macrophages, and adopt a nomenclature linked to their activation standards, for example, M(IL-4), M(Ig), M(IL-10), M(GC), M(IFN-γ), M(LPS) and so forth, and avoid the complexity of M2a, M2b, etc.
- Avoid certain terms: Terms to avoid include “regulatory,” as all macrophages are regulatory in some capacity.
- Markers of activation: Using combinations of markers helps describe activation outcomes.

Macrophage plasticity and polarization in different types of pathologies:
Macrophages exhibit a high plasticity and can switch between polarization/activation states. This enables them to perform different functions. For example, M1-like macrophages are more predominant in early phases of inflammatory responses, whereas M2-like macrophages are more commonly associated with chronic inflammation processes. (RA = rheumatoid arthritis) (Liu et al 2014, Murray and Wynn 2011, Sica and Mantovani 2012)
A better standardization is crucial for new therapeutic approaches
Different subsets of macrophages are essential to the pathogenesis of many diseases, where they often play opposing roles (Murray et al., 2011). M2-like macrophages regulate important metabolic functions in adipose tissue, maintain insulin sensitivity, and prevent the onset of type 2 diabetes, whereas disease progression is often associated with a switch from M2 to M1 phenotype (Lumeng et al., 2007). Tissue inflammation is an important feature of cancer biology, and is often linked to a polarization of M1 into M2 macrophages. These promote tumor growth, angiogenesis and metastasis (Joshi et al., 2014). Moreover, M1 macrophage-derived cytokines are important mediators in chronic inflammation, autoimmune diseases, and atherosclerosis. A more detailed understanding of the mechanisms that regulate the activation of human macrophages is likely to lead to the development of more effective strategies for treating various important inflammatory diseases. “The standardization and reproducibility of macrophage generation in vitro is the starting point for the development of new therapies,” concludes Wieland. “Only when different laboratories are able to generate the same cells in the same way will it be possible to succeed in developing robust strategies to address the multitude of macrophage-related disorders.” This could enable macrophage researchers to succeed at building their tower of Babel, and prevent the failure experienced by the biblical people.Macrophage Polarization: Plasticity in Health and Disease
Macrophages are known as the “big eaters” in the human body, and are present in almost all tissues.
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Related resources
Related pages & videos
MURRAY ET AL., (2014): MACROPHAGE ACTIVATION AND POLARIZATION: NOMENCLATURE AND EXPERIMENTAL GUIDELINES
The lack of consensus on how to define macrophage activation in experiments in vitro and in vivo impedes progress in multiple ways, including the fact that many researchers still consider there to be only the two types of activated macrophages often termed M1 and M2.
VIDEO: MACROPHAGES SUBTYPES: POLARIZATION, ACTIVATION AND PLASTICITY
Macrophages are multitasking cells at the first line of defense against pathogens invasion. But they can also play a major role in destroying tissues and developing cancer, autoimmune disorder or allergies. Learn more about the complexity of macrophage subtypes.