Src-mediated morphology transition of lung cancer cells in three-dimensional organotypic culture

Cancer Cell International

Bin Shan¹  Ying Zhuo¹  Zongbing You³  Jay K Kolls²  Steven P Kantrow⁴  Lichun Sun¹  Yan Zhuang¹  Hong T Nguyen¹ 

[1] Department of Medicine, Tulane University School of Medicine, 1430 Tulane Avenue, New Orleans, LA 70112, USA;Department of Pediatrics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15224, USA;Department of Structural and Cellular Biology, Tulane University School of Medicine, 1430 Tulane Avenue, New Orleans, LA 70112, USA;Department of Medicine, Louisiana State University Health Sciences Center, 1901 Perdido Street, New Orleans, LA 70112, USA

关键词: Extracellular matrix;    Three-dimensional culture;    Type I collagen;    Src;    TGF-β1;   

Others  :  794333
DOI  :  10.1186/1475-2867-13-16

 received in 2012-10-03, accepted in 2013-02-11,  发布年份 2013

【 摘 要 】

A fribotic tumor microenvironment promotes progression of cancer. In this study, we utilize a reconstituted basement membrane mimics Matrigel based three-dimensional organotypic culture (rBM 3-D) to investigate the mechanisms that mediate the tumor promoting effects of the fibrogenic mediators TGF-β1 and type I collagen (Col-1) on lung adenocarcinoma cells. Similar to normal alveolar epithelial cells, the well-differentiated lung adenocarcinoma cells in rBM 3-D culture undergo acinar morphogeneis that features polarized epithelial cell spheres with a single central lumen. Either TGF-β1 or Col-1 modestly distorts acinar morphogenesis. On the other hand, TGF-β1 and Col-1 synergistically induce a transition from acinar morphology into stellate morphology that is characteristic of invasive and metastatic cancer cells. Inhibition of the Src kinase activity abrogates induction of stellate morphology, activation of Akt and mTOR, and the expression of tumor promoting genes by TGF-β1 and Col-1. To a similar extent, pharmacological inhibition of mTOR abrogates the cellular responses to TGF-β1 and Col-1. In summary, we demonstrate that TGF-β1 and Col-1 promote stellate morphogenesis of lung cancer cells. Our findings further suggest that the Src-Akt-mTOR axis mediates stellate morphogenesis. These findings also indicate that rBM 3-D culture can serve as an ideal platform for swift and cost-effective screening of therapeutic candidates at the interface of the tumor and its microenvironment.

【 授权许可】

   
2013 Nguyen et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Paszek MJ, Zahir N, Johnson KR, Lakins JN, Rozenberg GI, Gefen A, Reinhart-King CA, Margulies SS, Dembo M, Boettiger D: Tensional homeostasis and the malignant phenotype. Cancer Cell 2005, 8(3):241-254.
• [2]Bian YF, Yang HY, Yang ZM, Gao F, Zhang NN, Xiao CS: Amlodipine treatment prevents angiotensin II-induced human umbilical vein endothelial cell apoptosis. Arch Med Res 2011, 42(1):22-27.
• [3]Kalluri R, Zeisberg M: Fibroblasts in cancer. Nat Rev Cancer 2006, 6(5):392-401.
• [4]Provenzano PP, Eliceiri KW, Campbell JM, Inman DR, White JG, Keely PJ: Collagen reorganization at the tumor-stromal interface facilitates local invasion. BMC Med 2006, 4(1):38. BioMed Central Full Text
• [5]Levental KR, Yu H, Kass L, Lakins JN, Egeblad M, Erler JT, Fong SF, Csiszar K, Giaccia A, Weninger W: Matrix crosslinking forces tumor progression by enhancing integrin signaling. Cell 2009, 139(5):891-906.
• [6]Borczuk AC, Papanikolaou N, Toonkel RL, Sole M, Gorenstein LA, Ginsburg ME, Sonett JR, Friedman RA, Powell CA: Lung adenocarcinoma invasion in TGFbetaRII-deficient cells is mediated by CCL5/RANTES. Oncogene 2008, 27(4):557-564.
• [7]Han S, Khuri FR, Roman J: Fibronectin stimulates non-small cell lung carcinoma cell growth through activation of Akt/mammalian target of rapamycin/S6 kinase and inactivation of LKB1/AMP-activated protein kinase signal pathways. Cancer Res 2006, 66(1):315-323.
• [8]Soltermann A, Tischler V, Arbogast S, Braun J, Probst-Hensch N, Weder W, Moch H, Kristiansen G: Prognostic significance of epithelial-mesenchymal and mesenchymal-epithelial transition protein expression in non-small cell lung cancer. Clin Cancer Res 2008, 14(22):7430-7437.
• [9]Borczuk AC, Sole M, Lu P, Chen J, Wilgus ML, Friedman RA, Albelda SM, Powell CA: Progression of human bronchioloalveolar carcinoma to invasive adenocarcinoma is modeled in a transgenic mouse model of K-ras-induced lung cancer by loss of the TGF-beta type II receptor. Cancer Res 2011, 71(21):6665-6675.
• [10]Gao Y, Xiao Q, Ma H, Li L, Liu J, Feng Y, Fang Z, Wu J, Han X, Zhang J: LKB1 inhibits lung cancer progression through lysyl oxidase and extracellular matrix remodeling. Proc Natl Acad Sci USA 2010, 107(44):18892-18897.
• [11]Debnath J, Muthuswamy SK, Brugge JS: Morphogenesis and oncogenesis of MCF-10A mammary epithelial acini grown in three-dimensional basement membrane cultures. Methods 2003, 30(3):256-268.
• [12]Hebner C, Weaver VM, Debnath J: Modeling morphogenesis and oncogenesis in three-dimensional breast epithelial cultures. Annu Rev Pathol 2008, 3:313-339.
• [13]Lee GY, Kenny PA, Lee EH, Bissell MJ: Three-dimensional culture models of normal and malignant breast epithelial cells. Nat Methods 2007, 4(4):359-365.
• [14]Martin KJ, Patrick DR, Bissell MJ, Fournier MV: Prognostic breast cancer signature identified from 3D culture model accurately predicts clinical outcome across independent datasets. PLoS One 2008, 3(8):e2994.
• [15]Kenny PA, Lee GY, Myers CA, Neve RM, Semeiks JR, Spellman PT, Lorenz K, Lee EH, Barcellos-Hoff MH, Petersen OW: The morphologies of breast cancer cell lines in three-dimensional assays correlate with their profiles of gene expression. Mol Oncol 2007, 1(1):84-96.
• [16]Yu W, Fang X, Ewald A, Wong K, Hunt CA, Werb Z, Matthay MA, Mostov K: Formation of cysts by alveolar type II cells in three-dimensional culture reveals a novel mechanism for epithelial morphogenesis. Mol Biol Cell 2007, 18(5):1693-1700.
• [17]Bren-Mattison Y, Van Putten V, Chan D, Winn R, Geraci MW, Nemenoff RA: Peroxisome proliferator-activated receptor-gamma (PPAR(gamma)) inhibits tumorigenesis by reversing the undifferentiated phenotype of metastatic non-small-cell lung cancer cells (NSCLC). Oncogene 2005, 24(8):1412-1422.
• [18]De Palma M, Hanahan D: The biology of personalized cancer medicine: Facing individual complexities underlying hallmark capabilities. Mol Oncol 2012, 6(2):111-127.
• [19]Wang D, Haviland DL, Burns AR, Zsigmond E, Wetsel RA: A pure population of lung alveolar epithelial type II cells derived from human embryonic stem cells. Proc Natl Acad Sci USA 2007, 104(11):4449-4454.
• [20]Johnson L, Mercer K, Greenbaum D, Bronson RT, Crowley D, Tuveson DA, Jacks T: Somatic activation of the K-ras oncogene causes early onset lung cancer in mice. Nature 2001, 410(6832):1111-1116.
• [21]Playford MP, Schaller MD: The interplay between Src and integrins in normal and tumor biology. Oncogene 2004, 23(48):7928-7946.
• [22]Lee MY, Lee SH, Park JH, Han HJ: Interaction of galectin-1 with caveolae induces mouse embryonic stem cell proliferation through the Src, ERas, Akt and mTOR signaling pathways. Cell Mol Life Sci 2009, 66(8):1467-1478.
• [23]Anand S, Muthusamy VS, Sujatha S, Sangeetha KN, Bharathi Raja R, Sudhagar S, Poornima Devi N, Lakshmi BS: Aloe emodin glycosides stimulates glucose transport and glycogen storage through PI3K dependent mechanism in L6 myotubes and inhibits adipocyte differentiation in 3T3L1 adipocytes. FEBS Lett 2010, 584(14):3170-3178.
• [24]Calalb MB, Zhang X, Polte TR, Hanks SK: Focal adhesion kinase tyrosine-861 is a major site of phosphorylation by Src. Biochem Biophys Res Commun 1996, 228(3):662-668.
• [25]Thoreen CC, Kang SA, Chang JW, Liu Q, Zhang J, Gao Y, Reichling LJ, Sim T, Sabatini DM, Gray NS: An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem 2009, 284(12):8023-8032.
• [26]Affolter M, Zeller R, Caussinus E: Tissue remodelling through branching morphogenesis. Nat Rev Mol Cell Biol 2009, 10(12):831-842.
• [27]Kim KK, Kugler MC, Wolters PJ, Robillard L, Galvez MG, Brumwell AN, Sheppard D, Chapman HA: Alveolar epithelial cell mesenchymal transition develops in vivo during pulmonary fibrosis and is regulated by the extracellular matrix. Proc Natl Acad Sci USA 2006, 103(35):13180-13185.
• [28]Moreno-Bueno G, Peinado H, Molina P, Olmeda D, Cubillo E, Santos V, Palacios J, Portillo F, Cano A: The morphological and molecular features of the epithelial-to-mesenchymal transition. Nat Protoc 2009, 4(11):1591-1613.
• [29]Li C, Nguyen HT, Zhuang Y, Lin Y, Flemington EK, Guo W, Guenther J, Burow ME, Morris GF, Sullivan D: Post-transcriptional up-regulation of miR-21 by type I collagen. Mol Carcinog 2011, 50(7):563-570.
• [30]Wrana JL, Attisano L, Carcamo J, Zentella A, Doody J, Laiho M, Wang XF, Massague J: TGF beta signals through a heteromeric protein kinase receptor complex. Cell 1992, 71(6):1003-1014.
• [31]Dy GK, Adjei AA: Emerging therapeutic targets in non-small cell lung cancer. Proc Am Thorac Soc 2009, 6(2):218-223.
• [32]Nagashima T, Maruyama T, Uchida H, Kajitani T, Arase T, Ono M, Oda H, Kagami M, Masuda H, Nishikawa S: Activation of SRC kinase and phosphorylation of signal transducer and activator of transcription-5 are required for decidual transformation of human endometrial stromal cells. Endocrinology 2008, 149(3):1227-1234.
• [33]Thomas JE, Soriano P, Brugge JS: Phosphorylation of c-Src on tyrosine 527 by another protein tyrosine kinase. Science 1991, 254(5031):568-571.
• [34]Shan B, Zhuo Y, Chin D, Morris CA, Morris GF, Lasky JA: Cyclin-dependent kinase 9 is required for tumor necrosis factor-alpha-stimulated matrix metalloproteinase-9 expression in human lung adenocarcinoma cells. J Biol Chem 2005, 280(2):1103-1111.
• [35]Zhuang Y, Nguyen HT, Lasky JA, Cao S, Li C, Hu J, Guo X, Burow ME, Shan B: Requirement of a novel splicing variant of human histone deacetylase 6 for TGF-beta1-mediated gene activation. Biochem Biophys Res Commun 2010, 392(4):608-613.
• [36]Bao L, Jaligam V, Zhang XY, Kutner RH, Kantrow SP, Reiser J: Stable transgene expression in tumors and metastases after transduction with lentiviral vectors based on human immunodeficiency virus type 1. Hum Gene Ther 2004, 15(5):445-456.
• [37]Li C, Nguyen HT, Zhuang Y, Lin Z, Flemington EK, Zhuo Y, Kantrow SP, Morris GF, Sullivan DE, Shan B: Comparative profiling of miRNA expression of lung adenocarcinoma cells in two-dimensional and three-dimensional cultures. Gene 2012, 511(2):143-150.
• [38]Shan B, Morris CA, Zhuo Y, Shelby BD, Levy DR, Lasky JA: Activation of proMMP-2 and Src by HHV8 vGPCR in human pulmonary arterial endothelial cells. J Mol Cell Cardiol 2007, 42(3):517-525.
• [39]Nguyen HT, Li C, Lin Z, Zhuang Y, Flemington EK, Burow ME, Lin Y, Shan B: The microRNA expression associated with morphogenesis of breast cancer cells in three-dimensional organotypic culture. Oncol Rep 2012, 28(1):117-126.
• [40]Shan B, Hagood JS, Zhuo Y, Nguyen HT, MacEwen M, Morris GF, Lasky JA: Thy-1 attenuates TNF-alpha-activated gene expression in mouse embryonic fibroblasts via Src family kinase. PLoS One 2010, 5(7):e11662.
• [41]Sun L, Vasilevich NI, Fuselier JA, Coy DH: Abilities of 3,4-diarylfuran-2-one analogs of combretastatin A-4 to inhibit both proliferation of tumor cell lines and growth of relevant tumors in nude mice. Anticancer Res 2004, 24(1):179-186.