The immunomodulatory role of tumor Syndecan-1 (CD138) on ex vivo tumor microenvironmental CD4+ T cell polarization in inflammatory and non-inflammatory breast cancer patients.

Moshira Ezzat Saleh,Martin Götte,Ahmed Afifi,Ramy Gadalla,Sherif Abdelaziz Ibrahim,Mona Mostafa Mohamed,Hebatallah Hassan,Mohamed El-Shinawi,Alberto G Passi

doi:10.1371/journal.pone.0217550

Moshira Ezzat Saleh, Martin Götte + Show 7 more

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https://doi.org/10.1371/journal.pone.0217550

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Abstract

Herein, we aimed to identify the immunomodulatory role of tumor Syndecan-1 (CD138) in the polarization of CD4+ T helper (Th) subsets isolated from the tumor microenvironment of inflammatory breast cancer (IBC) and non-IBC patients. Lymphocytes and mononuclear cells isolated from the axillary tributaries of non-IBC and IBC patients during modified radical mastectomy were either stimulated with the secretome as indirect co-culture or directly co-cultured with control and Syndecan-1-silenced SUM-149 IBC cells. In addition, peripheral blood mononuclear cells (PBMCs) of normal subjects were used for the direct co-culture. Employing flow cytometry, we analyzed the expression of the intracellular IFN-γ, IL-4, IL-17, and Foxp3 markers as readout for basal and co-cultured Th1, Th2, Th17, and Treg CD4+ subsets, respectively. Our data revealed that IBC displayed a lower basal frequency of Th1 and Th2 subsets than non-IBC. Syndecan-1-silenced SUM-149 cells significantly upregulated only Treg subset polarization of normal subjects relative to controls. However, Syndecan-1 silencing significantly enhanced the polarization of Th17 and Treg subsets of non-IBC under both direct and indirect conditions and induced only Th1 subset polarization under indirect conditions compared to control. Interestingly, qPCR revealed that there was a negative correlation between Syndecan-1 and each of IL-4, IL-17, and Foxp3 mRNA expression in carcinoma tissues of IBC and that the correlation was reversed in non-IBC. Mechanistically, Syndecan-1 knockdown in SUM-149 cells promoted Th17 cell expansion via upregulation of IL-23 and the Notch ligand DLL4. Overall, this study indicates a low frequency of the circulating antitumor Th1 subset in IBC and suggests that tumor Syndecan-1 silencing enhances ex vivo polarization of CD4+ Th17 and Treg cells of non-IBC, whereby Th17 polarization is possibly mediated via upregulation of IL-23 and DLL4. These findings suggest the immunoregulatory role of tumor Syndecan-1 expression in Th cell polarization that may have therapeutic implications for breast cancer.

Highlights

Female breast cancer is the most broadly diagnosed cancer heading the list of life-threatening cancers in women all over the world and in Egypt [1, 2]
The Foxp3+ (Treg) cell subset ranked as the second highest proportion of CD4+ T cells in inflammatory breast cancer (IBC) patients representing 23.1%, whereas it ranked as the lowest proportion of CD4+ T cells in non-IBC patients representing 14.56%, there was no significant difference between non-IBC and IBC patients (Fig 1, Table 2)
Our data revealed that the Th17 (IL-17+CD4+) subset is the predominant subset among CD4+ T cells in both groups; non-IBC and IBC patients

Summary

Introduction

Female breast cancer is the most broadly diagnosed cancer heading the list of life-threatening cancers in women all over the world and in Egypt [1, 2]. Inflammatory breast cancer (IBC) is a deadly aggressive form of breast cancer that is featured by enrichment of cancer stemness, rapid invasion into the dermal lymphatic vasculature, increasing metastasis, and low survival rate in comparison to non-IBC [3, 4]. The Th1 subset typically produces IFN-γ to enhance the cytotoxic activity of tumor-specific CD8+ T cells [11]. The Th2 subset produces the classical anti-inflammatory cytokines IL-4, IL-13, and IL-5, to activate the M2 macrophage protumor phenotype and increase metastasis potential of breast carcinoma [12]. The Foxp3+ regulatory T cells (Treg) that create an immunosuppressive tumor microenvironment, allowing escape from immunosurveillance, and enhancing breast carcinoma progression [15, 16]. CD4+ T cells are highly plastic cells that can be inter-converted between different subsets according to cytokines and chemokines milieu of the environment [17]

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