Acute and chronic tissue injury results in the generation of a myriad of environmental cues that macrophages respond to by changing their phenotype and function. This phenotypic regulation is critical for controlling tissue inflammation and resolution. Here, we have identified the adaptor protein disabled homolog 2 (DAB2) as a regulator of phenotypic switching in macrophages.
Samantha E. Adamson, Rachael Griffiths, Radim Moravec, Subramanian Senthivinayagam, Garren Montgomery, Wenshu Chen, Jenny Han, Poonam R. Sharma, Garrett R. Mullins, Stacey A. Gorski, Jonathan A. Cooper, Alexandra Kadl, Kyle Enfield, Thomas J. Braciale, Thurl E. Harris, Norbert Leitinger
Submitter: Muhammad Ashfaq-Khan | firstname.lastname@example.org
Authors: Muhammad Ashfaq-Khan and Detlef Schuppan
Institute of Translational Immunology and Research Center for Immunotherapy, University Medical Center, Mainz, Germany
Published May 3, 2016
Disabled homology-2 (DAB2) is mainly expressed in brain, kidney, intestine and ovary, and induced in (mouse) mammary glands during pregnancy and lactation (1-6). It has many functions, such as the modulation of wnt signalling (7), modulates immunity (6) and plays an important role during embroygenesis (8). Notably, DAB2 has been dubbed a tumour suppresser gene, since it is downregulated in several cancers such as urothelial, bladder, nasopharyngeal, gastric and breast cancer. However, DAB2 has also been implicated in potentiation of TGF-b as a tumor promoter and in driving epithelial-mesenchymal transition which is linked to tumorigenesis (9-12). Most cancers harbour a unique subset of macrophages: freshly recruited, monocyte derived, and tissue resident, resulting in tumour associated macrophages (TAM) (13). These TAM exhibit features of alternative (M2) macrophages that by suppressing tumour-directed T cell responses and by driving angiogenesis promote tumour growth and metastasis (14).
In a recent paper in the Journal of Clinical Investigation (15) the authors demonstrated that DAB2 downregulated M1 macrophage and upregulated M2 macrophage poolarization in mouse bone marrow derived macrophages treated either with LPS and INF-g or IL-4 (to produce M1 and M2 macrophages, respectively). Similarly, murine RAW 246.7 macrophages were exposed to various stumuli such as IL-4, IL-13 and IL-10 to produce M2 like macrophages, or to retinoic acid and TGF-β that are known inducers of DAB2. Intraperitoneal injection of LPS in myeloid cell deleted LysM-Cre Dab2fl/fl mice resulted in increased TNF-α and IL-6 expression in lung and liver after 1 and 4 hours. Moreover induction of fatty liver disease in these mice leads to adipose tissue inflammation. Finally, DAB2 was shown to be a negative regulator of the NF-κB–signaling pathway via interaction with TRAF6. The authors concluded that DAB2 anatgonizes macrophage activation and an increased M1 polarization.
We believe that the term “macrophage activation” or “macrophage polarization “should be used with caution, since in a multistage immune response the M1 and M2 bimodal polarization scheme may too simplistic (16, 17). Based on their data, the authors suggested DAB2 not only as a novel regulator of macrophage phenotypic but also of functional polarization in inflammatory disorders and cancers, while they did not showed the role of DAB2 expression e.g. in tumour associated macrophages. This puts a question mark on the possible role of DAB2 in other settings such as CNS inflammation and cancers where TAM are prototypic M2 (like) macrophages. Thus upregulation of DAB2 has been reported in lesional macrophages (= in murine experimental autoimmune encephalitis, a model of multiple sclerosis, where M1 macrophages (microglia) and a Th1 (Th17) T cell response predominate and where disease severity was reduced in DAB2 deficient mice (18).
Therefore, the regulatory role of the DAB2 gene in macrophage (functional) polarization may influence different diseases, including cancers, as well as in different organs in different ways. In our opinion, this complex and obviously context-dependent role of DAB2 in monocytes/macrophages has not been addressed thoroughly enough to make a clear statement about its function in macrophages and inflammation in general. In summary, the authors’ claim that “DAB2 controls macrophage phenotype polarization”, qualifying it as an attractive pharmacological target to treat chronic inflammatory conditions or cancers appears premature.
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2. Huang CH, Cheng JC, Chen JC, and Tseng CP. Evaluation of the role of Disabled-2 in nerve growth factor-mediated neurite outgrowth and cellular signalling. Cell Signal. 2007;19(6):1339-47.
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15. Adamson SE, Griffiths R, Moravec R, Senthivinayagam S, Montgomery G, Chen W, Han J, Sharma PR, Mullins GR, Gorski SA, et al. Disabled homolog 2 controls macrophage phenotypic polarization and adipose tissue inflammation. J Clin Invest. 2016;126(4):1311-22.
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18. Jokubaitis VG, Gresle MM, Kemper DA, Doherty W, Perreau VM, Cipriani TL, Jonas A, Shaw G, Kuhlmann T, Kilpatrick TJ, et al. Endogenously regulated Dab2 worsens inflammatory injury in experimental autoimmune encephalomyelitis. Acta Neuropathol Commun. 2013;1(32).
Submitter: Norbert Leitinger | email@example.com
Authors: Subramanian Senthivinayagam, Samantha E. Adamson, Thurl E. Harris, and Norbert Leitinger
University of Virginia, P.O. Box 800735, Charlottesville, Virginia 22908
Published May 3, 2016
We thank Drs. Ashfaq-Khan and Schuppan for critically reading our manuscript and raising the important question whether Dab2 will also control the phenotypic polarization status of tumor-associated macrophages (TAMs). We certainly agree that macrophages are highly plastic and that their phenotypic polarization spectrum is more complex than can be described by the oversimplifying M1/M2 paradigm. In fact, macrophage phenotypic polarization critically depends on the tissue microenvironment and in that context, our lab recently identified a distinct macrophage phenotype Mox, which develops in a microenvironment enriched in oxidized phospholipids, such as seen in atherosclerotic lesions (1).
Interaction of tumor cells with immune cells including macrophages is a critical determinant of tumor cell survival and growth. Indeed, evidence suggests that macrophages in the tumor microenvironment (TAMs) are involved in complex chemical cross talk with tumor and non-tumor cells. In spite of their highly plastic nature, TAMs in most cases exhibit an M2-like phenotype (2). M2-like TAMs support tumor growth via multiple mechanisms including but not limited to evading cell death, angiogenesis and metastasis, by secreting immunosuppressive cytokines such as IL-10, CCL-17 and CCL-22 as well as proangiogenic and tissue remodeling factors like VEGF or MMP9 (3, 4). Several experimental studies have shown that targeting TAMs may have beneficial effects including reducing tumor growth (5). Recently, treatment approaches using a CSF1-R inhibitor or histidine-rich glycoprotein (HRG) were shown to reduce M2 expression markers in TAMs and to skew their phenotype from tumor promoting M2-like into a tumor-toxic M1-like phenotype (6, 7). Hence, regulators of macrophage phenotypic polarization may be attractive targets for developing anti-tumor therapies.
In our recent manuscript published in the Journal of Clinical Investigation (8) we have shown that manipulation of endogenous Dab2 levels controls phenotyping switching of macrophages in vitro and in vivo. While over-expression of Dab2 inhibited LPS- and IFN-induced M1 marker gene expression, siRNA-mediated knock down facilitated expression of M1 markers while reducing IL-4 induced alternative activation in both bone marrow-derived macrophages and macrophage cell lines. Moreover, using a mouse model where Dab2 is specifically deleted in myeloid cells we have shown Dab2 knock out facilitated M1 macrophage polarization both in conditions under acute high grade inflammation and chronic low grade inflammation. We also identify a mechanism, by which Dab2 inhibits NFkB-dependent gene expression, namely by binding to TRAF6. Hence, we concluded that Dab2 is a novel regulator of macrophage phenotypic polarization. However, as the authors pointed out, our study has not addressed the role of Dab2 in tumor macrophages. Nevertheless, based on our observations, it is tempting to speculate that manipulating Dab2 levels in M2-like TAMs will phenotypially repolarize TAMs to become tumor-toxic. However, future studies will be required to shed more light on the role of Dab2 in TAMs.
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