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Mesenchymal stem cells and macrophages interact through IL-6 to promote inflammatory breast cancer in pre-clinical models.

ABSTRACT: Inflammatory breast cancer (IBC) is a unique and deadly disease with unknown drivers. We hypothesized the inflammatory environment contributes to the IBC phenotype. We used an in vitro co-culture system to investigate interactions between normal and polarized macrophages (RAW 264.7 cell line), bone-marrow derived mesenchymal stem cells (MSCs), and IBC cells (SUM 149 and MDA-IBC3). We used an in vivo model that reproduces the IBC phenotype by co-injecting IBC cells with MSCs into the mammary fat pad. Mice were then treated with a macrophage recruitment inhibitor, anti-CSF1. MSC and macrophages grown in co-culture produced higher levels of pro-tumor properties such as enhanced migration and elevated IL-6 secretion. IBC cells co-cultured with educated MSCs also displayed enhanced invasion and mammosphere formation and blocked by anti-IL-6 and statin treatment. The treatment of mice co-injected with IBC cells and MSCs with anti-CSF1 inhibited tumor associated macrophages and inhibited pSTAT3 expression in tumor cells. Anti-CSF1 treated mice also exhibited reduced tumor growth, skin invasion, and local recurrence. Herein we demonstrate reciprocal tumor interactions through IL-6 with cells found in the IBC microenvironment. Our results suggest IL-6 is a mediator of these tumor promoting influences and is important for the IBC induced migration of MSCs.

SUBMITTER: Wolfe AR

PROVIDER: S-EPMC5347707 | biostudies-literature | 2016 Dec

REPOSITORIES: biostudies-literature

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Mesenchymal stem cells and macrophages interact through IL-6 to promote inflammatory breast cancer in pre-clinical models.

Wolfe Adam R AR Trenton Nicholaus J NJ Debeb Bisrat G BG Larson Richard R Ruffell Brian B Chu Khoi K Hittelman Walter W Diehl Michael M Reuben Jim M JM Ueno Naoto T NT Woodward Wendy A WA

Oncotarget 20161201 50

Inflammatory breast cancer (IBC) is a unique and deadly disease with unknown drivers. We hypothesized the inflammatory environment contributes to the IBC phenotype. We used an in vitro co-culture system to investigate interactions between normal and polarized macrophages (RAW 264.7 cell line), bone-marrow derived mesenchymal stem cells (MSCs), and IBC cells (SUM 149 and MDA-IBC3). We used an in vivo model that reproduces the IBC phenotype by co-injecting IBC cells with MSCs into the mammary fat ...[more]

PMID: 27756885

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Project description:BackgroundMyocardial ischemia-reperfusion injury is a type of myocardial ischemia that has a significant impact on patients' health. We aimed to explore the protective effect of mesenchymal stem cell-derived exosomes (MSC-EXOs) on myocardial ischemia-reperfusion injury and their specific mechanism.MethodsThe effects of MSC-EXOs on myocardial ischemia-reperfusion injury were recorded. An enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of IL-6 and IL-10 in mouse myocardial tissue or culture supernatant. Co-cultured MSC-EXOs and RAW264.7 cells were used to study the effect of MSC-EXOs on the polarization of macrophages at the cellular level. The ratio of M1 and M2 macrophages were detected by flow cytometry, and RT-qPCR detected the mRNA expression levels of corresponding markers. After transfection with miR-21-5p inhibitors or mimics, flow cytometry and RT-qPCR experiments were performed to explore the specific role of MSC-EXOs in macrophage polarization.ResultsAfter injection of MSC-EXOs, the mRNA expression of M1 macrophage markers (iNOS, IL-1β, IL-6, and TNFα) in the myocardial tissue of model mice was significantly reduced (P<0.05), and the mRNA expression of M2 macrophage markers was significantly increased (P<0.05). The injection also reduced the inflammation response in the model mice (P<0.05). In the in vitro experiment, lipopolysaccharide (LPS) induced the inflammatory microenvironment. After MSC-EXOs were fixed in the cytoplasm of RAW264.7 cells, the level of IL-6 in the culture supernatant decreased (P<0.05), and the level of IL-10 increased (P<0.05). The addition of MSC-EXOs to LPS-induced RAW264.7 cells promoted their polarization toward the M2 phenotype and upregulated their marker expression levels (P<0.05). Following inhibition of miR-21-5p in MSC cells, the EXOs were collected, and it was found that MSC-EXOs that inhibited the expression of miR-21-5p promoted LPS-induced polarization of RAW264.7 cells to the M1 phenotype and upregulated inflammation in the culture supernatant. Furthermore, transfection with miR-21-5p mimics promoted the polarization of RAW264.7 cells to the M2 phenotype and reduced the level of inflammatory factors in the culture supernatant.ConclusionsMSC-EXOs promote the polarization of macrophages to the M2 phenotype via miR-21-5p, thereby reducing inflammation and promoting heart repair.

Project description:BACKGROUND:The mechanisms by which macrophage phenotype contributes to mesenchymal stem cells (MSC)-mediated bone repair remain unclear. In this work, we investigated the influence of factors released by human macrophages polarized to a pro-inflammatory or an anti-inflammatory phenotype on the ability of human MSC to attach, migrate, and differentiate toward the osteoblastic lineage. We focused on the role of TNF-? and IL-10, key pro-inflammatory and anti-inflammatory cytokines, respectively, in regulating MSC functions. METHODS:MSC were treated with media conditioned by pro-inflammatory or anti-inflammatory macrophages to study their influence in cell attachment, migration, and osteogenic differentiation. The involvement of TNF-? and IL-10 in the regulation of MSC functions was investigated using neutralizing antibodies and recombinant cytokines. RESULTS:Treatment of MSC with media conditioned by pro-inflammatory or anti-inflammatory macrophages promoted cell elongation and enhanced MSC ability to attach and migrate. These effects were more noticeable when MSC were treated with media from pro-inflammatory macrophages. Interestingly, MSC osteogenic activity was enhanced by factors released by anti-inflammatory macrophages, but not by pro-inflammatory macrophages. Significant IL-10 levels originated from anti-inflammatory macrophages enhanced MSC osteogenesis by increasing ALP activity and mineralization in MSC layers cultured under osteogenic conditions. Moreover, macrophage-derived IL-10 regulated the expression of the osteogenic markers RUNX2, COL1A1, and ALPL. Notably, low TNF-? levels secreted by anti-inflammatory macrophages increased ALP activity in differentiating MSC whereas high TNF-? levels produced by pro-inflammatory macrophages had no effects on osteogenesis. Experiments in which MSC were treated with cytokines revealed that IL-10 was more effective in promoting matrix maturation and mineralization than TNF-?. CONCLUSIONS:Factors secreted by pro-inflammatory macrophages substantially increased MSC attachment and migration whereas those released by anti-inflammatory macrophages enhanced MSC osteogenic activity as well as cell migration. IL-10 was identified as an important cytokine secreted by anti-inflammatory macrophages that potentiates MSC osteogenesis. Our findings provide novel insights into how environments provided by macrophages regulate MSC osteogenesis, which may be helpful to develop strategies to enhance bone regeneration.

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Mesenchymal stem cells and macrophages interact through IL-6 to promote inflammatory breast cancer in pre-clinical models.

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Mesenchymal stem cells and macrophages interact through IL-6 to promote inflammatory breast cancer in pre-clinical models.

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