ABSTRACT
As mentioned earlier, macrophages polarize into different phenotypes (inflammatory and anti-inflammatory) by responding to their local microenvironment. In response to injury, macrophages are recruited to sites of injury by responding to chemo-attractant signals produced during inflammation. While macrophages exhibit considerable heterogeneity in their activation profiles, two major activation schemes, “classical activation” and “alternative activation” have been characterized in detail in recent years. The classical pathway involves recognition of PAMPs such as LPS by PRRs and stimulation by IFNg produced by T helper 1 (TH1) or CD8+ cells or natural killer (NK) cells leading to a TH1-type inflammatory response (Ma et al. 2003; Taylor et al. 2005; Rubartelli and Lotze 2007; Classen et al. 2009; Martinez et al. 2009). The alternatively activated macrophage pathway involves stimulation of macrophages by IL4 and/or IL-13, inducing cells such as eosinophils, basophils and naive T lymphocytes to generate a TH2-type anti-inflammatory response (Ma et al. 2003; Taylor et al. 2005; Mantovani 2006; Rubartelli and Lotze 2007; Martinez et al. 2009). The nomenclature of classically activated macrophages “M1” and alternatively activated macrophages “M2” was first designated by analogy to the TH1 and TH2 immune responses (Rubartelli and Lotze 2007). In addition to signaling through IFNg, antigen presenting cells are stimulated through TLRs to secrete TNFa, a pro-inflammatory cytokine which synergizes with IFNg to sustain inflammation (Mosser and Edwards 2008). This M1 inflammatory macrophage is geared towards phagocytosis, recognition and destruction of foreign organisms, pathogens and tumor cells. M1 macrophages in turn secrete pro-inflammatory cytokines including TNFa, IL-1, IL-6 and IL-12, up-regulate expression of reactive oxygen radicals, iNOS, MMP9 and MHC class II receptors on the cell surface, and release nitric oxide (NO) which aids in their microbicidal and tumoricidal capability. Agonists of several TLRs, such as LPS (TLR4 agonist) and lipotechoic acid (TLR2 agonist), bind to TLRs on the cell surface in conjunction with other cell surface co-receptors. These include CD14, CD11b/CD18, and MD2 for TLR4 and CD36 for TLR2. In conjunction with these co-receptors, TLRs trigger intracellular signaling pathways that lead to gene induction through the action of several transcription factors, including NF-kB, STATs and interferon-regulatory factors (IRFs) that are involved in M1 macrophage polarization. Alternatively activated macrophages (M2) differ from classically activated macrophages by their activation patterns, expression of cytokines and growth factors and their involvement in anti-inflammatory, wound healing and TH2 responses (Gordon 2003; Gordon and Taylor 2005). M2 macrophages generally express high levels of IL-10, TGF-b, arginase-1 (Arg1), receptors such as the SR-A/scavenger receptor (CD204), MRCI/mannose receptor (CD206), b-glucan receptor (dectin-1) and cell surface markers such as Found in Inflammatory Zone-1 (Fizz1) and chitinase 3-like 3/chitinase 3-like
4 (Ym1/Ym2) (Enfield and Leibovich 2011). They also express low levels of TNFa, IL-12, IL-1b, IL-6, IL-8, IL-23 and iNOS (Enfield and Leibovich 2011). Exceptions exist, however, where distinct forms of M2 cells have been described to express low IL-12 and high IL-10 phenotype with the capacity to produce TNF, IL-1 and IL-6 (Mantovani et al. 2004). This highlights the complex ability of macrophages to change their phenotype to serve their required functions in different microenvironments. As mentioned previously, recent studies have categorized alternatively activated M2 macrophages into subsets (M2a, M2b, M2c and M2d) depending on their responses to different cytokines and growth factors (Martinez et al. 2009). M2a macrophages are prototypically induced by IL-4 and IL-13 through the common IL-4Ra subunit of the receptors for these cytokines (Martinez et al. 2009). However, in mouse wounds lacking IL-4 and IL-13, or lacking the IL-4Ra subunit, macrophages having M1 and M2-like characteristics were still found, clearly indicating that IL-4 and IL-13 are not essential for the induction of macrophages with an M2-like phenotype (Daley et al. 2010). M2b macrophages are induced by immune complex and IL-1b or TLR agonists. M2c macrophages are induced by glucocorticoids, IL-10 and TGFb (Martinez et al. 2009). M2d macrophages are induced by TLR agonists and adenosine in an IL-4Ra-independent manner (Enfield and Leibovich 2011) (Fig. 1). A more detailed description of the M2d macrophage activation program in the context of wound healing and angiogenesis will be presented in the next section. Several reports indicate that TAMs have diverse phenotypes, having both M1 and M2 markers, but are generally associated with M2 functions that support tumor progression and angiogenesis (Van Ginderachter et al. 2006). Mantovani and colleagues present TAMs as polarized M2 mononuclear phagocytes producing high IL-10 and low IL-12 (Mantovani et al. 2004; Mantovani and Sica 2010). Biswas et al report that TAMs from murine fibrosarcoma express a mixed phenotype with M1 and M2 properties (high dectin-1, high IL-10 and low IL-12), coexisting with M1 expression of IFN-inducible cytokines such as CXCL10. They further demonstrated that upon treatment with LPS, NF-kB activation was inhibited, but IFNginducible chemokines were expressed through enhanced activation of IRF-3 and STAT1 (Biswas et al. 2006). Additionally, they reported that arginase-1, which is considered to be an M2 marker, is regulated in a complex manner in TAMs from murine fibrosarcoma. They suggest that both arginase-1 and iNOS could be induced by the same IRF-3/STAT1 pathway, which could explain the mixed M1/M2 gene expression profile in TAMs. Thus, macrophages can take on different phenotypes (M1, M2, M2-like) to achieve the goal of tumor promotion in the tumor microenviroment.
