Associations of chemo- and radio-resistant phenotypes with the gap junction, adhesion and extracellular matrix in a three-dimensional culture model of soft sarcoma

Abstract

BackgroundThree-dimensional (3D) culture models are considered to recapitulate the cell microenvironment in solid tumors, including the extracellular matrix (ECM), cell-cell interactions, and signal transduction. These functions are highly correlated with cellular behaviors and contribute to resistances against chemo- and radio-therapies. However, the biochemical effects and mechanisms remain unknown in soft sarcoma. Therefore, we developed an in vitro 3D model of sarcoma to analyze the reasons of the chemo- and radio-resistance in therapies.MethodsFour soft sarcoma cell lines, HT1080, RD, SW872, and human osteosarcoma cell line 1 (HOSS1), a cell line established from a patient-derived xenograft, were applied to 3D culture and treated with growth factors in methylcellulose-containing medium. Spheroids were examined morphologically and by western blotting, RT-qPCR, and immunofluorescence staining to analyze cell adhesion, gap junctions, ECM genes, and related factors. Proliferation and colony formation assays were performed to assess chemo- and radio-resistances between 3D and two-dimensional (2D) cell cultures. Annexin V and Propidium Iodide staining was used to detect early apoptotic sarcoma cells treated with Doxorubicin, Gemcitabine, and Docetaxel in the 3D model.ResultsThe four soft sarcoma cell lines formed spheres in vitro by culture in modified condition medium. Compared with 2D cell culture, expression of ECM genes and proteins, including COL1A1, LOX, SED1, FN1, and LAMA4, was significantly increased in 3D culture. Analysis of cadherin and gap junction molecules showed significant changes in the gene and protein expression profiles under 3D conditions. These changes affected cell–cell communication and were mainly associated with biological processes such as cell proliferation and apoptosis related to chemo- and radio-resistances.ConclusionsOur findings revealed significant differences between 3D and 2D cell culture systems, and indicated that cellular responsiveness to external stress such as radiation and chemotherapeutics is influenced by differential expression of genes and proteins involved in regulation of the ECM, cell adhesion, and gap junction signaling.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-015-0175-0) contains supplementary material, which is available to authorized users.

3D tumor spheroids as in vitro models to mimic in vivo human solid tumors resistance to therapeutic drugs

“…also demonstrated that collagen type IV expression is upregulated in BT4C glioma tumor spheroids, whereas no significant expression was observed in monolayer cultures of BT4C cells.Further studies showed that the increased deposition of ECM proteins in spheroids can influence the response of the cells to the therapeutics Bai et al (2015). verified that the increased expression of ECM proteins (e.g., collagen and fibronectin-1) in 3D cultures of soft sarcoma (HT1080, RD, and SW872) and osteosarcoma (HOSS1) cell lines contributed for the establishment of a chemoresistant environment to doxorubicin, gemcitabine, and docetaxel.…”

“…cells were superior in spheroids(4.61, 23.55, and 103.2 µM, respectively) than in 2D cell cultures (0.078, 6.23, and 6.72 µM, respectively;Bai et al, 2015).To reduce the cell-ECM proteins interactions,Cheng, van Zandwijk, and Reid (2014) used an integrin-targeted chemotherapeutic drug (Cilengitide-synthetic peptide inhibitor of integrin αv heterodimer with high specificity for αvβ3 and αvβ5) to reduce the malignance behavior of malignant pleural mesothelioma cells (e.g., H28) spheroids. Their results demonstrated that H28 cells viability was significantly reduced because these cells display a high expression of Cilengitide target integrins.…”

“…Nederman, Norling, Glimelius, Carlsson, and Brunk (1984) studied the presence and expression of ECM proteins in U-118 MG glioma and HTH-7 thyroid cancer spheroids and they noticed that spheroids were able to produce ECM components such as collagens (Type I, III, and V), fibronectin and laminin. Bjerkvig, Laerum, and Rucklidge (1989) in HOSS1 cells were two-and fourfolds higher in the spheroids than in 2D cell cultures, respectively (Bai et al, 2015). Bjerkvig, Laerum, and Rucklidge (1989) in HOSS1 cells were two-and fourfolds higher in the spheroids than in 2D cell cultures, respectively (Bai et al, 2015).…”

Leveraging Naked Mole Rat (Heterocephalus glaber) Comparative Genomics to Identify Canine Genes Modulating Susceptibility to Tumorigenesis and Cancer Phenotypes

“…A genetic association study published in 2012 identified a gene within one of the most conserved tumorigenesis orthologs, FN1, as associated with elbow dysplasia in Bernese Mountain Dogs [67]. Not surprisingly, the FN1 gene is expressed in soft sarcoma and osteosarcoma cells [68].…”

Three‐Dimensional Osteosarcoma Models for Advancing Drug Discovery and Development

“…[60] Although the cytotoxicity is confirmed by ATP blocking, the maintenance of spheroid structure demonstrates that chemotherapeutic treatments have a crucial effect in ECM-related gene and protein expression, which modulates cell-cell interactions and tumor networks. [61] In fact, a broad proteomic analysis displayed a high number of species associated with doxorubicin resistance, with a focus on cathepsin D, lately implicated in several malignant phenotypes. [61] In fact, a broad proteomic analysis displayed a high number of species associated with doxorubicin resistance, with a focus on cathepsin D, lately implicated in several malignant phenotypes.…”