Original ArticlesInduction of breast cancer stem cells by M1 macrophages through Lin-28B-let-7-HMGA2 axis
Introduction
Macrophages have been demonstrated to be the most abundant immune cells in solid tumors, comprising 5–40% of tumor mass. In early tumors, macrophages appear to possess “classically activated” (M1) phenotype to play proinflammatory tumoricidal roles [1]. As tumors are established, macrophages tend to be educated by tumor cells towards immunosuppressive “alternatively activated” phenotype (M2) to aid tumor progression [2]. Mounting evidence suggests that an elevated number of TAMs with M2 profile is correlated with therapy failure and poor prognosis in patients [[3], [4], [5], [6]], indicating TAM an important target in anti-tumor therapy [7,8]. Attempts have been made to increase M1/M2 ratio in tumors by inducing TAMs to switch from M2 to M1 or injecting polarized M1 into tumors [[9], [10], [11]], showing that increased M1 macrophages efficiently reduced tumor burden and malignancy in mouse models. Furthermore, modulation of TAMs is also required by other immunotherapies to succeed. Change of TAM phenotype to M1 augmented the anti-tumor efficacy of CD8+ T cells in lung and melanoma cancers in vivo [12,13]. Monoclonal antibodies targeting programmed cell death protein-1 (α-PD-1) have shown notable clinical efficacy in patients with various cancer. However, PD-1- TAMs could capture α-PD-1 from T cell surface and dramatically reduce the efficacy of α-PD-1 in activating T cells [14]. These data suggest that modulating TAMs would be a promising cancer therapy and also beneficial for other established immunotherapies. Though M2 phenotype has been considered to be the major phenotype of TAM, TAMs with a mixed M1/M2 phenotype are also present in patients of pancreatic cancer, T cell/histiocyte rich large B cell lymphoma and ovarian cancer [[15], [16], [17], [18]]. In pancreatic ductal adenocarcinoma (PDAC) patients, TAMs exhibiting M1 phenotype were detected and both pro- and anti-inflammatory macrophages contributed equally to EMT in PDAC initiation and development [15]. Very recently, it has also been revealed that polarized M1 macrophages enhances the metastasis potential of ovarian cancer cell lines via NF-κB pathway [19]. These evidences indicate that the pro-inflammatory signals of tumor-associated macrophages might function in tumor progression, so strategies using M1 macrophages in cancer treatment should be regarded with some caution and more studies should be focused on the effect of M1 macrophages on cancer cells.
Cancer stem cells (CSCs) are a minority subpopulation of highly tumorigenic cells in tumors, which have been identified in a wide variety of cancers [20]. It has been suggested that CSCs may arise from non-stem cancer cells (NSCCs) upon microenvironment signals [21,22]. However, the mechanisms of CSC formation still remain elusive. Ablation of macrophages in mouse mammary fat pad severely impaired the function of stem/progenitor cells in developing mammary gland and almost completely blocked tumor initiation in mouse models, suggesting a critical correlation between macrophages and CSCs [23]. M2 TAMs have been indicated to support CSC signatures [24,25]. Both M1 and M2 TAMs are observed in tumor tissues and the transient activation status is essential for macrophages to acquire M2 phenotype. Also, pro-inflammatory cytokines such as interleukin (IL)-1β, IL-6, and IL-8 contributed to the establishment of CSC niche. Since CSCs are a major cause for tumor metastasis, drug resistance and relapse and M1 macrophages are being introduced into cancer treatments, it is worth investigating any potential correlation between macrophages and CSCs.
Accumulating evidence indicates that stem cell-like properties can be induced in cancer cells via epithelial-mesenchymal transition (EMT), during which epithelial cells lose their cell polarity and cell-cell adhesion and gain migratory and invasive properties. Previously, we reported that in breast cancer cells, Stat3-coordinated Lin-28B-let-7-HMGA2 axis effectively initiated EMT in response to inflammatory cytokine Oncostatin M (OSM) and inflammatory pathways IL-6/Stat3/Jagged-1/Notch could increase drug resistance of gastric cancer cells via EMT/CSC process [26,27].
In this study, we found that M1 macrophages contribute to the initiation of CSC phenotypic transformation in breast cancer cells. M1-associated inflammatory cytokine network triggers the expansion and self-renewal of CSCs through Lin-28B-let-7-HMGA2 axis. Our data also suggest that upon interplay with tumor cells, M1 transdifferentiate into M2 phenotype, which function in supporting CSC maintenance. This study not only sheds a new light on the potential mechanism of CSC generation, but more importantly, indicates that when M1 macrophage is utilized as a powerful tool to kill cancer cells, it is of great importance to suppress potential CSC formation and maintenance by blocking key pathways and preventing M1 from switching to M2 phenotype.
Section snippets
Cell culture
Human breast cancer cell lines MCF-7, T47D, BT474, and MDA231 were cultured in Dulbecco's Modified Eagle's Medium (DMEM, Invitrogen) supplemented with 10% fetal bovine serum (FBS) in a 5% CO2 atmosphere at 37 °C. Human monocytic cell line THP-1 was cultured in RPMI 1640 medium supplemented with 10% FBS. Stable depletion of HMGA2 in MDA-MB-231 cells was achieved by transfection of the HMGA2 shRNA expression vector and the transfected cells were maintained in DMEM containing 10% FBS and 400 ng/ml
M1 macrophage induces CSC subpopulation in breast cancer cells
Human monocytic cell line THP-1 is widely used as a model for monocyte/macrophage differentiation. We produced polarized M1 and M2 macrophage from THP-1 and human monocytes according to established methods [28] (Supplemental Fig. 1A), which have also been used in studying the effect of polarized M1 on tumor cells both in vivo and in vitro [10,11]. THP-1 cells treated with IFN-γ/LPS firmly attached to the plates and most of the cells extended multiple elongated processes. IL-4/IL-13-induced
Discussion
Nowadays, it has been widely accepted that tumorigenesis is orchestrated by innate immunity [42]. As a major component in innate immunity and the most abundant immune cells infiltrating in solid tumor, macrophages regulate tumor initiation and progression. The phenotype of macrophage is considered to switch from proinflammatory M1 in tumorigenesis to immunosuppressive M2 since tumors are established, with its role changing from tumoricidal to pro-malignant in tumor. An increased M1/M2 ratio
Compliance with ethics guide lines
All authors declare they have no conflict of interest. This study was approved by the ethics committee of Institute of Basic Medical Sciences. All animal studies were performed in accordance with the institutional guidelines and the experiments were approved by the Animal Care and Use Committee of the institute.
Conflicts of interest
All authors read and approved the final version of the manuscript, and the authors declare no conflict interest.
Acknowledgements
Grant Support: National Key Technologies R&D Program for New Drugs (2013ZX09102056), the National High-Tech Research and Development Plan (863 Program, No. 2014AA020604), National Natural Science Foundation of China (No. 31500702, 31370825, 81272232, 81402562, 81572845, and 81401311), Beijing Natural Science Foundation (No. 7162144, 7122124, and 7132163), and China Postdoctoral Science Foundation (No. 2015T81095). The authors are grateful to all staffs who contributed to this study.
References (67)
- et al.
Alternatively activated (M2) macrophages promote tumour growth and invasiveness in hepatocellular carcinoma
J. Hepatol.
(2015) - et al.
Reprogramming tumor-associated macrophages by antibody targeting inhibits cancer progression and metastasis
Cell Rep.
(2016) - et al.
Stochastic state transitions give rise to phenotypic equilibrium in populations of cancer cells
Cell
(2011) - et al.
Tumor-associated macrophages promote cancer stem cell-like properties via transforming growth factor-beta1-induced epithelial-mesenchymal transition in hepatocellular carcinoma
Cancer Lett.
(2014) - et al.
Tumor-associated macrophage-induced invasion and angiogenesis of human basal cell carcinoma cells by cyclooxygenase-2 induction
J. Investig. Dermatol.
(2009) - et al.
ALDH1 is a marker of normal and malignant human mammary stem cells and a predictor of poor clinical outcome
Cell Stem Cell
(2007) - et al.
The epithelial-mesenchymal transition generates cells with properties of stem cells
Cell
(2008) - et al.
Tumor-derived hyaluronan induces formation of immunosuppressive macrophages through transient early activation of monocytes
Blood
(2007) - et al.
Characteristic alteration of monocytes with increased intracellular IL-10 and IL-12 in patients with advanced-stage gastric cancer
J. Surg. Res.
(2004) - et al.
Melanoma exosomes promote mixed M1 and M2 macrophage polarization
Cytokine
(2018)
Slug and Sox9 cooperatively determine the mammary stem cell state
Cell
Lin28: primal regulator of growth and metabolism in stem cells
Cell Stem Cell
Lin28A and Lin28B inhibit let-7 microRNA biogenesis by distinct mechanisms
Cell
let-7 regulates self renewal and tumorigenicity of breast cancer cells
Cell
Hmga2 promotes neural stem cell self-renewal in young but not old mice by reducing p16Ink4a and p19Arf Expression
Cell
Epigenetic regulation of the miR142-3p/interleukin-6 circuit in glioblastoma
Mol. Cell
Coordinated regulation of myeloid cells by tumours
Nat. Rev. Immunol.
Human breast cancer cells educate macrophages toward the M2 activation status
Breast Cancer Res.
A high M1/M2 ratio of tumor-associated macrophages is associated with extended survival in ovarian cancer patients
J. Ovarian Res.
The distribution of macrophages with a M1 or M2 phenotype in relation to prognosis and the molecular characteristics of colorectal cancer
PLoS One
Opposite effects of M1 and M2 macrophage subtypes on lung cancer progression
Sci. Rep.
The role of macrophages polarization in predicting prognosis of radically resected gastric cancer patients
J. Cell Mol. Med.
Targeting tumor-infiltrating macrophages to combat cancer
Immunotherapy
The promise of targeting macrophages in cancer therapy
Clin. Cancer Res.
Reprogramming of tumor-associated macrophages with anticancer therapies: radiotherapy versus chemo- and immunotherapies
Front. Immunol.
The anti-tumor effects of M1 macrophage-loaded poly (ethylene glycol) and gelatin-based hydrogels on hepatocellular carcinoma
Theranostics
Using macrophage activation to augment immunotherapy of established tumours
Br. J. Canc.
In vivo imaging reveals a tumor-associated macrophage-mediated resistance pathway in anti-PD-1 therapy
Sci. Transl. Med.
Tumor-associated macrophages exhibit pro- and anti-inflammatory properties by which they impact on pancreatic tumorigenesis
Int. J. Cancer
Comparative characterization of stroma cells and ductal epithelium in chronic pancreatitis and pancreatic ductal adenocarcinoma
PLoS One
Macrophages in T cell/histiocyte rich large B cell lymphoma strongly express metal-binding proteins and show a bi-activated phenotype
Int. J. Cancer
Mixed-polarization phenotype of ascites-associated macrophages in human ovarian carcinoma: correlation of CD163 expression, cytokine levels and early relapse
Int. J. Cancer
Pro-inflammatory M1 macrophage enhances metastatic potential of ovarian cancer cells through NF-kappaB activation
Mol. Carcinog.
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Contributed equally.