Microfluidic isolation of cancer-cell-derived microvesicles from hetergeneous extracellular shed vesicle populations

Extracellular shed vesicles, including exosomes and microvesicles, are disseminated throughout the body and represent an important conduit of cell communication. Cancer-cell-derived microvesicles have potential as a cancer biomarker as they help shape the tumor microenvironment to promote the growth of the primary tumor and prime the metastatic niche. It is likely that, in cancer cell cultures, the two constituent extracellular shed vesicle subpopulations, observed in dynamic light scattering, represent an exosome population and a cancer-cell-specific microvesicle population and that extracellular shed vesicle size provides information about provenance and cargo. We have designed and implemented a novel microfluidic technology that separates microvesicles, as a function of diameter, from heterogeneous populations of cancer-cell-derived extracellular shed vesicles. We measured cargo carried by the microvesicle subpopulation processed through this microfluidic platform. Such analyses could enable future investigations to more accurately and reliably determine provenance, functional activity, and mechanisms of transformation in cancer.

[1]  R. Johnstone,et al.  Vesicle formation during reticulocyte maturation. Association of plasma membrane activities with released vesicles (exosomes). , 1987, The Journal of biological chemistry.

[2]  Kevin Loutherback,et al.  Improved performance of deterministic lateral displacement arrays with triangular posts , 2010 .

[3]  Richard J Simpson,et al.  Comparison of ultracentrifugation, density gradient separation, and immunoaffinity capture methods for isolating human colon cancer cell line LIM1863-derived exosomes. , 2012, Methods.

[4]  S. Mathivanan,et al.  ExoCarta: A compendium of exosomal proteins and RNA , 2009, Proteomics.

[5]  Jared L. Johnson,et al.  Cancer cell-derived microvesicles induce transformation by transferring tissue transglutaminase and fibronectin to recipient cells , 2011, Proceedings of the National Academy of Sciences.

[6]  Hamid Cheshmi Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers , 2011 .

[7]  Jason P. Gleghorn,et al.  Transport and collision dynamics in periodic asymmetric obstacle arrays: rational design of microfluidic rare-cell immunocapture devices. , 2013, Physical review. E, Statistical, nonlinear, and soft matter physics.

[8]  D. Farber,et al.  Transfer of MicroRNAs by Embryonic Stem Cell Microvesicles , 2009, PloS one.

[9]  R. Cardigan,et al.  Microparticle sizing by dynamic light scattering in fresh‐frozen plasma , 2009, Vox sanguinis.

[10]  G. Taraboletti,et al.  Bioavailability of VEGF in tumor-shed vesicles depends on vesicle burst induced by acidic pH. , 2006, Neoplasia.

[11]  David W Inglis,et al.  Highly accurate deterministic lateral displacement device and its application to purification of fungal spores. , 2010, Biomicrofluidics.

[12]  Jason P. Gleghorn,et al.  Capture of circulating tumor cells from whole blood of prostate cancer patients using geometrically enhanced differential immunocapture (GEDI) and a prostate-specific antibody. , 2010, Lab on a chip.

[13]  Michael P Barrett,et al.  Separation of parasites from human blood using deterministic lateral displacement. , 2011, Lab on a chip.

[14]  Sanchita Bhatnagar,et al.  Exosome Function: From Tumor Immunology to Pathogen Biology , 2008, Traffic.

[15]  H. Bruus,et al.  A theoretical analysis of the resolution due to diffusion and size dispersion of particles in deterministic lateral displacement devices , 2007, 0711.0347.

[16]  Paul J. Harrison,et al.  Sizing and phenotyping of cellular vesicles using Nanoparticle Tracking Analysis , 2011, Nanomedicine : nanotechnology, biology, and medicine.

[17]  S. Kristensen,et al.  Extracellular Vesicle (EV) Array: microarray capturing of exosomes and other extracellular vesicles for multiplexed phenotyping , 2013, Journal of extracellular vesicles.

[18]  Clotilde Théry,et al.  Exosome Secretion: Molecular Mechanisms and Roles in Immune Responses , 2011, Traffic.

[19]  J. Slot,et al.  Proteomic and Biochemical Analyses of Human B Cell-derived Exosomes , 2003, The Journal of Biological Chemistry.

[20]  G. Parmiani,et al.  Tumor-released microvesicles as vehicles of immunosuppression. , 2007, Cancer research.

[21]  H. Schluesener,et al.  Role of exosomes in immune regulation , 2006, Journal of cellular and molecular medicine.

[22]  H. John Crabtree,et al.  Continuous dielectrophoretic cell separation microfluidic device. , 2007, Lab on a chip.

[23]  Milica Radisic,et al.  Deterministic lateral displacement as a means to enrich large cells for tissue engineering. , 2009, Analytical chemistry.

[24]  M. Rubin,et al.  Large oncosomes in human prostate cancer tissues and in the circulation of mice with metastatic disease. , 2012, The American journal of pathology.

[25]  Jason P. Gleghorn,et al.  Rare Cell Capture in Microfluidic Devices. , 2011, Chemical engineering science.

[26]  Richard J. Simpson,et al.  Proteomics Analysis of A33 Immunoaffinity-purified Exosomes Released from the Human Colon Tumor Cell Line LIM1215 Reveals a Tissue-specific Protein Signature* , 2009, Molecular & Cellular Proteomics.

[27]  Paul Harrison,et al.  Classification, Functions, and Clinical Relevance of Extracellular Vesicles , 2012, Pharmacological Reviews.

[28]  György Nagy,et al.  Membrane vesicles, current state-of-the-art: emerging role of extracellular vesicles , 2011, Cellular and Molecular Life Sciences.

[29]  A. Guha,et al.  Intercellular transfer of the oncogenic receptor EGFRvIII by microvesicles derived from tumour cells , 2008, Nature Cell Biology.

[30]  Michael T. Laub,et al.  Continuous Particle Separation Through Deterministic Lateral Displacement , 2004 .

[31]  Wenjie Wang,et al.  Peripheral blood microvesicles are potential biomarkers for hepatocellular carcinoma. , 2013, Cancer biomarkers : section A of Disease markers.

[32]  Thomas Hawighorst,et al.  Induction of tumor lymphangiogenesis by VEGF-C promotes breast cancer metastasis , 2001, Nature Medicine.

[33]  J. Rak,et al.  Microvesicles as mediators of intercellular communication in cancer—the emerging science of cellular ‘debris’ , 2011, Seminars in Immunopathology.

[34]  X. Breakefield,et al.  Prostate cancer-derived urine exosomes: a novel approach to biomarkers for prostate cancer , 2009, British Journal of Cancer.

[35]  E. Lechman,et al.  Exosomes derived from genetically modified DC expressing FasL are anti-inflammatory and immunosuppressive. , 2006, Molecular therapy : the journal of the American Society of Gene Therapy.

[36]  G. Raposo,et al.  Exosomes: a common pathway for a specialized function. , 2006, Journal of biochemistry.

[37]  James W. Clancy,et al.  Tumor-derived microvesicles: shedding light on novel microenvironment modulators and prospective cancer biomarkers. , 2012, Genes & development.

[38]  Crislyn D'Souza-Schorey,et al.  Microvesicles: mediators of extracellular communication during cancer progression , 2010, Journal of Cell Science.

[39]  M. Mizumoto,et al.  Predictive value of vascular endothelial growth factor (VEGF) in metastasis and prognosis of human colorectal cancer. , 1998, British Journal of Cancer.

[40]  G. Camussi,et al.  Microvesicles released from human renal cancer stem cells stimulate angiogenesis and formation of lung premetastatic niche. , 2011, Cancer research.

[41]  B. Kirby,et al.  Continuous-flow particle separation by 3D Insulative dielectrophoresis using coherently shaped, dc-biased, ac electric fields. , 2007, Analytical chemistry.

[42]  Robert L Moritz,et al.  Exosomes: proteomic insights and diagnostic potential , 2009, Expert review of proteomics.

[43]  H. Kwon,et al.  Proteomic analysis of microvesicles derived from human colorectal cancer cells. , 2007, Journal of proteome research.

[44]  Jacopo Meldolesi,et al.  Shedding microvesicles: artefacts no more. , 2009, Trends in cell biology.

[45]  Steven A. Stacker,et al.  VEGF-D promotes the metastatic spread of tumor cells via the lymphatics , 2001, Nature Medicine.

[46]  David W Inglis,et al.  Critical particle size for fractionation by deterministic lateral displacement. , 2006, Lab on a chip.

[47]  R. Cerione,et al.  Cancerous epithelial cell lines shed extracellular vesicles with a bimodal size distribution that is sensitive to glutamine inhibition , 2014, Physical biology.

[48]  D. Ott,et al.  CD45 immunoaffinity depletion of vesicles from Jurkat T cells demonstrates that exosomes contain CD45: no evidence for a distinct exosome/HIV-1 budding pathway , 2008, Retrovirology.

[49]  R. Cerione,et al.  R(h)oads to microvesicles , 2012, Small GTPases.

[50]  Nicole Pamme,et al.  Continuous flow separations in microfluidic devices. , 2007, Lab on a chip.

[51]  J. Rak,et al.  Microvesicles: Messengers and mediators of tumor progression , 2009, Cell cycle.

[52]  L. Mincheva-Nilsson,et al.  REVIEW ARTICLE: The Role of Placental Exosomes in Reproduction , 2010, American journal of reproductive immunology.

[53]  T G van Leeuwen,et al.  Optical and non‐optical methods for detection and characterization of microparticles and exosomes , 2010, Journal of thrombosis and haemostasis : JTH.

[54]  Brian J. Kirby,et al.  Microfluidic transport in microdevices for rare cell capture , 2012, Electrophoresis.

[55]  D. Lyden,et al.  The secreted factors responsible for pre-metastatic niche formation: old sayings and new thoughts. , 2011, Seminars in cancer biology.

[56]  Sascha Keller,et al.  Exosomes: from biogenesis and secretion to biological function. , 2006, Immunology letters.

[57]  S. Mathivanan,et al.  Exosomes: extracellular organelles important in intercellular communication. , 2010, Journal of proteomics.

[58]  Bo Li,et al.  RhoA triggers a specific signaling pathway that generates transforming microvesicles in cancer cells , 2012, Oncogene.