Abstract:To investigate phenotypic and genotypic alterations before and after bone metastasis, we conducted genome-wide mRNA profiling and DNA exon sequencing of two cell lines (TMD and BMD) derived from a mouse xenograft model. TMD cells were harvested from the mammary fat pad after transfecting MDA-MB-231 breast cancer cells, while BMD cells were isolated from the metastasized bone. Compared to BMD cells, TMD cells exhibited higher cellular motility. In contrast, BMD cells formed a spheroid with a smoother and more c… Show more
“…These findings suggested that blocking S100A4 signaling reduces the formation of high proliferative active breast cancer hybrid cells also carrying metastatic properties. Supportive evidence is presented by recent work demonstrating reduced migration and metastases of breast cancer cells following down-modulation of S100A4 [ 50 ]. Fig.…”
BackgroundFusion of breast cancer cells with tumor-associated populations of the microenvironment including mesenchymal stroma/stem-like cells (MSC) represents a rare event in cell communication whereby the metastatic capacity of those hybrid cells remains unclear.MethodsFunctional changes were investigated in vitro and in vivo following spontaneous fusion and hybrid cell formation between primary human MSC and human MDA-MB-231 breast cancer cells. Thus, lentiviral eGFP-labeled MSC and breast cancer cells labeled with mcherry resulted in dual-fluorescing hybrid cells after co-culture.ResultsDouble FACS sorting and single cell cloning revealed two different aneuploid male hybrid populations (MDA-hyb1 and MDA-hyb2) with different STR profiles, pronounced telomerase activities, and enhanced proliferative capacities as compared to the parental cells. Microarray-based mRNA profiling demonstrated marked regulation of genes involved in epithelial-mesenchymal transition and increased expression of metastasis-associated genes including S100A4. In vivo studies following subcutaneous injection of the breast cancer and the two hybrid populations substantiated the in vitro findings by a significantly elevated tumor growth of the hybrid cells. Moreover, both hybrid populations developed various distant organ metastases in a much shorter period of time than the parental breast cancer cells.ConclusionTogether, these data demonstrate spontaneous development of new tumor cell populations exhibiting different parental properties after close interaction and subsequent fusion of MSC with breast cancer cells. This formation of tumor hybrids contributes to continuously increasing tumor heterogeneity and elevated metastatic capacities.Electronic supplementary materialThe online version of this article (10.1186/s12964-018-0215-4) contains supplementary material, which is available to authorized users.
“…These findings suggested that blocking S100A4 signaling reduces the formation of high proliferative active breast cancer hybrid cells also carrying metastatic properties. Supportive evidence is presented by recent work demonstrating reduced migration and metastases of breast cancer cells following down-modulation of S100A4 [ 50 ]. Fig.…”
BackgroundFusion of breast cancer cells with tumor-associated populations of the microenvironment including mesenchymal stroma/stem-like cells (MSC) represents a rare event in cell communication whereby the metastatic capacity of those hybrid cells remains unclear.MethodsFunctional changes were investigated in vitro and in vivo following spontaneous fusion and hybrid cell formation between primary human MSC and human MDA-MB-231 breast cancer cells. Thus, lentiviral eGFP-labeled MSC and breast cancer cells labeled with mcherry resulted in dual-fluorescing hybrid cells after co-culture.ResultsDouble FACS sorting and single cell cloning revealed two different aneuploid male hybrid populations (MDA-hyb1 and MDA-hyb2) with different STR profiles, pronounced telomerase activities, and enhanced proliferative capacities as compared to the parental cells. Microarray-based mRNA profiling demonstrated marked regulation of genes involved in epithelial-mesenchymal transition and increased expression of metastasis-associated genes including S100A4. In vivo studies following subcutaneous injection of the breast cancer and the two hybrid populations substantiated the in vitro findings by a significantly elevated tumor growth of the hybrid cells. Moreover, both hybrid populations developed various distant organ metastases in a much shorter period of time than the parental breast cancer cells.ConclusionTogether, these data demonstrate spontaneous development of new tumor cell populations exhibiting different parental properties after close interaction and subsequent fusion of MSC with breast cancer cells. This formation of tumor hybrids contributes to continuously increasing tumor heterogeneity and elevated metastatic capacities.Electronic supplementary materialThe online version of this article (10.1186/s12964-018-0215-4) contains supplementary material, which is available to authorized users.
“…The 4T1.2 cell line was isolated from metastasized bone, and represents a breast cancer model of efficient metastasis to bone ( 20 ). The responses to AZD7762 and PD407824 were examined in monolayer cell cultures, three-dimensional (3D) spheroids ( 21 ), and 3D bio-printed bone constructs ( 22 ). While PD407824 is able to inhibit tumor growth and osteoclastogenesis, its ability to promote osteoblast development is limited ( 11 ).…”
Chemotherapy for suppressing tumor growth and metastasis tends to induce various effects on other organs. Using AZD7762, an inhibitor of checkpoint kinase (Chk) 1 and 2, the present study examined its effect on mammary tumor cells in addition to bone cells (osteoclasts, osteoblasts and osteocytes), using monolayer cell cultures and three-dimensional (3D) cell spheroids. The results revealed that AZD7762 blocked the proliferation of 4T1.2 mammary tumor cells and suppressed the development of RAW264.7 pre-osteoclast cells by downregulating nuclear factor of activated T cells cytoplasmic 1. AZD7762 also promoted the mineralization of MC3T3 osteoblast-like cells and 3D bio-printed bone constructs of MLO-A5 osteocyte spheroids. While a Chk1 inhibitor, PD407824, suppressed the proliferation of tumor cells and the differentiation of pre-osteoclasts, its effect on gene expression in osteoblasts was markedly different compared with AZD7762. Western blotting indicated that the stimulating effect of AZD7762 on osteoblast development was associated with the inhibition of Chk2 and the downregulation of cellular tumor antigen p53. The results of the present study indicated that in addition to acting as a tumor suppressor, AZD7762 may prevent bone loss by inhibiting osteoclastogenesis and stimulating osteoblast mineralization.
“…TMD cells were isolated from tumors of injected MDA-MB-231 in a mouse, while BMD cells were isolated from bone metastasis after cardiac injection of TMD cells 9 . Our previous work showed that TMD cells exhibited more migratory characteristics in vitro than BMD cells 10 . Of note, primary human breast cancer cells are estrogen receptor-positive (ER+) 25 , while TMD and BMD cells are ER-negative.…”
Section: Discussionmentioning
confidence: 94%
“…TMD cells were isolated from the mammary tumor resulting from the injection of MDA-MB-231 cells to the mammary fat pad of NOD/SCHID mouse, while BMD cells were harvested from the metastasized bone 9 . Compared to BMD cells, it is reported that TMD cells exhibits higher cellular motility 10 .…”
Bone is a frequent site of metastasis from breast cancer. To understand the potential role of osteocytes in bone metastasis, we investigated tumor-osteocyte interactions using two cell lines derived from the MDA-MB-231 breast cancer cells, primary breast cancer cells, and MLO-A5/MLO-Y4 osteocyte cells. When three-dimensional (3D) tumor spheroids were grown with osteocyte spheroids, tumor spheroids fused with osteocyte spheroids and shrank. This size reduction was also observed when tumor spheroids were exposed to conditioned medium isolated from osteocyte cells. Mass spectrometry-based analysis predicted that several bone matrix proteins (e.g., collagen, biglycan) in conditioned medium could be responsible for tumor shrinkage. The osteocyte-driven shrinkage was mimicked by type I collagen, the most abundant organic component in bone, but not by hydroxyapatite, a major inorganic component in bone. RNA and protein expression analysis revealed that tumor-osteocyte interactions downregulated Snail, a transcription factor involved in epithelial-to-mesenchymal transition (EMT). An agarose bead assay showed that bone matrix proteins act as a tumor attractant. Collectively, the study herein demonstrates that osteocytes attract and compact migratory breast cancer cells through bone matrix proteins, suppress tumor migration, by Snail downregulation, and promote subsequent metastatic colonization.
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