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Endostatin inhibits the growth and migration of 4T1 mouse breast cancer cells by skewing macrophage polarity toward the M1 phenotype

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Abstract

The phenotypic diversity of tumor-associated macrophages (TAMs) increases with tumor development. One of the hallmarks of malignancy is the polarization of TAMs from a pro-immune (M1) phenotype to an immunosuppressive (M2) phenotype. However, the molecular basis of this process is still unclear. Endostatin is a powerful inhibitor of angiogenesis capable of suppressing tumor growth and metastasis. Here, we demonstrate that endostatin induces RAW264.7 cell polarization toward the M1 phenotype in vitro. Endostatin has no effect on TAM numbers in vivo, but results in an increased proportion of F4/80+Nos2+ cells and a decreased proportion of F4/80+CD206+ cells. Overexpression of endostatin in RAW264.7 cells resulted in a decrease in the phosphorylation of STAT3, an increase in expression of vascular endothelial growth factor A and placental growth factor, and an increase in the phosphorylation of STAT1, IκBα and p65 proteins compared with controls. These results indicate that endostatin regulates macrophage polarization, promoting the M1 phenotype by targeting NF-κB and STAT signaling.

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Abbreviations

APC:

Allophycocyanin

Arg1:

Arginase-1

BMDM:

Bone marrow-derived macrophage

CD206:

Mannose receptor 1

cDNA:

Complementary DNA

CFC:

Colony-forming cell

COX2:

Cyclooxygenase 2

DMSO:

Dimethylsulfoxide

ELISA:

Enzyme-linked immunosorbent assay

FBS:

Fetal bovine serum

FCM:

Flow cytometry

FITC:

Fluorescein isothiocyanate

IHC:

Immunohistochemistry

IL-6:

Interleukin 6

IL-10:

Interleukin 10

IL-12p40:

Interleukin 12p40

M1:

Classical activation macrophage

M2:

Alternative activation macrophage

MMP:

Matrix metalloproteinase

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-dimethyltetrazolium bromide

NF-κB:

Nuclear factor κB

NOS2:

Inducible nitric oxide synthase

PBS:

Phosphate-buffered saline

PE:

Phycoerythrin

PI:

Propidium iodide

PIGF:

Placenta growth factor

qRT-PCR:

Quantitative real-time reverse transcription polymerase chain reaction

SEM:

Scanning electron microscopy

STAT:

Signal transducer and activator of transcriptions

TAMs:

Tumor-associated macrophages

VEGF:

Vascular endothelial growth factor

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Acknowledgments

This work was supported by a Research Fund for the Scientific and Technological Development Plan Project in Jilin Province (20150414025GH). We are grateful to Shuqin Pan for assistance with the pathological technology applied in this study.

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Authors and Affiliations

  1. Department of Pathophysiology, Prostate Diseases Prevention and Treatment Research Center, School of Basic Medical Sciences, Jilin University, 126 Xinmin Street, Changchun, 130021, Jilin, People’s Republic of China

    Hua Guo, Yanan Liu, Yue Wang, Lianqin Liu, Ping Zhang & Yang Li

  2. Department of Pathology, Shandong Provincial Qianfoshan Hospital, Shandong University, Jinan, 250013, Shandong, People’s Republic of China

    Junlian Gu

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  1. Hua Guo
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  2. Yanan Liu
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  3. Junlian Gu
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  7. Yang Li
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Correspondence to Yang Li.

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Guo, H., Liu, Y., Gu, J. et al. Endostatin inhibits the growth and migration of 4T1 mouse breast cancer cells by skewing macrophage polarity toward the M1 phenotype. Cancer Immunol Immunother 65, 677–688 (2016). https://doi.org/10.1007/s00262-016-1824-7

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