Co-cultivation of murine BMDCs with 67NR mouse mammary carcinoma cells give rise to highly drug resistant cells

Christa Nagler,Thomas Dittmar,Kurt S Zänker,Cornelia Hardt

doi:10.1186/1475-2867-11-21

Abstract

BackgroundTumor tissue resembles chronically inflamed tissue. Since chronic inflammatory conditions are a strong stimulus for bone marrow-derived cells (BMDCs) it can be assumed that recruitment of BMDCs into cancer tissue should be a common phenomenon. Several data have outlined that BMDC can influence tumor growth and metastasis, e.g., by inducing a paracrine acting feedback loop in tumor cells. Likewise, cell fusion and horizontal gene transfer are further mechanisms how BMDCs can trigger tumor progression.ResultsHygromycin resistant murine 67NR-Hyg mammary carcinoma cells were co-cultivated with puromycin resistant murine BMDCs from Tg(GFPU)5Nagy/J mice. Isolation of hygromycin/puromycin resistant mBMDC/67NR-Hyg cell clones was performed by a dual drug selection procedure. PCR analysis revealed an overlap of parental markers in mBMDC/67NR-Hyg cell clones, suggesting that dual resistant cells originated by cell fusion. By contrast, both STR and SNP data analysis indicated that only parental 67NR-Hyg alleles were found in mBMDC/67NR-Hyg cell clones favoring horizontal gene transfer as the mode of origin. RealTime-PCR-array analysis showed a marked up-regulation of Abcb1a and Abcb1b ABC multidrug transporters in mBMDC/67NR-Hyg clones, which was verified by Western Blot analysis. Moreover, the markedly increased Abcb1a/Abcb1b expression was correlated to an efficient Rhodamine 123 efflux, which was completely inhibited by verapamil, a well-known Abcb1a/Abcb1b inhibitor. Likewise, mBMDCs/67NR-Hyg clones revealed a marked resistance towards chemotherapeutic drugs including 17-DMAG, doxorubicin, etoposide and paclitaxel. In accordance to Rhodamine 123 efflux data, chemotherapeutic drug resistance of mBMDC/67NR-Hyg cells was impaired by verapamil mediated blockage of Abc1a/Abcb1b multidrug transporter function.ConclusionCo-cultivation of mBMDCs and mouse 67NR-Hyg mammary carcinoma cells gave rise to highly drug resistant cells. Even though it remains unknown whether mBMDC/67NR-Hyg clones originated by cell fusion or horizontal gene transfer, our data indicate that the exchange of genetic information between two cellular entities is crucial for the origin of highly drug resistant cancer (hybrid) cells, which might be capable to survive chemotherapy.

Highlights

Gene expression analysis revealed a nuclear reprogramming in hybrid cells [11]. Even though both studies did not investigate whether bone marrow-derived cells (BMDCs)/tumor cell hybrids do contribute to tumor progression at all, several data of the past years indicated that cell fusion is a common phenomenon in cancer and that hybrid cells possess novel characteristics, which can be definitely linked to tumor progression
Co-cultivation of Murine bone marrow-derived cells (mBMDCs) and murine 67NR mammary carcinoma cells resulted in hygromycin and puromycin double resistant cells In the present study we investigated the co-cultivation of mBMDCs and murine mammary carcinoma cells (67NR-Hyg)
Both STR and single nucleotide polymorphism (SNP) analyses revealed that only parental 67NR-Hyg alleles were present in mBMDC/67NR-Hyg clones, facing the question whether mBMDC/ 67NR-Hyg clones rather originated by horizontal gene transfer (HGT). 67NR-Hyg cells might have uptaken DNA containing microvesicles or apoptotic bodies shedded from mBMDCs

Summary

Introduction

It is assumed that BMDCs can promote a proglycolytic phenotype in tumor cells, giving them a survival advantage in hypoxic and inflammatory conditions [9], or promote tumor cell survival through the activation of the integrin-linked kinase (ILK), thereby activating the prosurvival AKT signaling pathway [10] Another mechanism has been described by Karnoub and colleagues by demonstrating that breast cancer cells stimulated the de novo secretion of the chemokine CCL5 ( named RANTES) from MSCs, which acted in a paracrine fashion on the cancer cells to enhance their motility, invasion and metastasis [7]. Cell fusion has been suggested as one mechanism how cancer stem cells might originate [13,19,20]

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