Name: transfer of mammary gland-forming ability between mammary basal epithelial cells and mammary luminal cells via extracellular vesicles/exosomes
Uploaded: 2017-06-03T14:00:00
Description: Watch this Scientific Journal Video about Transfer of Mammary Gland-forming Ability Between Mammary Basal Epithelial Cells and Mammary Luminal Cells via Extracellular Vesicles/Exosomes at JoVE.com

Dette arbeidet ble støttet av tilskudd fra National Health Research Institutes (05A1-CSPP16-014, HJL) og fra departementet for vitenskap og teknologi (MOST 103-2320-B-400-015-MY3, HJL).

All forskning som involverte dyr fulgte protokoller godkjent av Institutt for dyrepleie. 1. Ekstracellulær vesikkel / exosomisolasjon og validering <ol><li> Kulturmammepitelbasale celler, NAMEC 4 , med friskt, serumfritt medium laget av 500 ml MCDB 170, pH 7,4 + 500 ml DMEM / F12 med natriumbikarbonat (0,2438%); EGF (5 ng / mL); Hydrokoriton (0,5 μg / ml); Insulin (5 μg / ml); Bovint hypofyse ekstrakt (BPE; 35 μg / ml); Og GW627368X (1 μg / mL) i 15 cm tallerke...

<ol>
	<li>Simons, M., Raposo, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes--vesicular+carriers+for+intercellular+communication.">Exosomes--vesicular carriers for intercellular communication.</a> Curr Opin Cell Biol. 21 (4), 575-581 (2009).</li><li>Th&#233;ry, C., Ostrowski, M., Segura, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+vesicles+as+conveyors+of+immune+responses.">Membrane vesicles as conveyors of immune responses.</a> Nat Rev Immunol. 9 (8), 581-593 (2009).</li><li>Boelens, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosome+Transfer+from+Stromal+to+Breast+Cancer+Cells+Regulates+Therapy+Resistance+Pathways.">Exosome Transfer from Stromal to Breast Cancer Cells Regulates Therapy Resistance Pathways.</a> Cell. 159 (3), 499-507 (2014).</li><li>Lin, M. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=PGE2+/EP4+Signaling+Controls+the+Transfer+of+the+Mammary+Stem+Cell+State+by+Lipid+Rafts+in+Extracellular+Vesicles.">PGE2 /EP4 Signaling Controls the Transfer of the Mammary Stem Cell State by Lipid Rafts in Extracellular Vesicles.</a> Stem Cells. , (2016).</li><li>Th&#233;ry, C., Amigorena, S., Raposo, G., Clayton, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+characterization+of+exosomes+from+cell+culture+supernatants+and+biological+fluids.">Isolation and characterization of exosomes from cell culture supernatants and biological fluids.</a> Curr Protoc Cell Biol. , (2006).</li><li>Gy&#246;rgy, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+vesicles,+current+state-of-the-art:+emerging+role+of+extracellular+vesicles.">Membrane vesicles, current state-of-the-art: emerging role of extracellular vesicles.</a> Cell Mol Life Sci. 68 (16), 2667-2688 (2011).</li><li>Olver, C., Vidal, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proteomic+analysis+of+secreted+exosomes.">Proteomic analysis of secreted exosomes.</a> Subcell Biochem. 43, 99-131 (2007).</li><li>Shackleton, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Generation+of+a+functional+mammary+gland+from+a+single+stem+cell.">Generation of a functional mammary gland from a single stem cell.</a> Nature. 439 (7072), 84-88 (2006).</li><li>Prater, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mammary+stem+cells+have+myoepithelial+cell+properties.">Mammary stem cells have myoepithelial cell properties.</a> Nat Cell Biol. 16 (10), 942-950 (2014).</li><li>Stingl, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Purification+and+unique+properties+of+mammary+epithelial+stem+cells.">Purification and unique properties of mammary epithelial stem cells.</a> Nature. 439 (7079), 993-997 (2006).</li><li>Gardiner, C., Ferreira, Y. J., Dragovic, R. A., Redman, C. W., Sargent, I. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+vesicle+sizing+and+enumeration+by+nanoparticle+tracking+analysis.">Extracellular vesicle sizing and enumeration by nanoparticle tracking analysis.</a> J Extracell Vesicles. 2, (2013).</li><li>Shapiro, A. L., Vi&#241;uela, E., Maizel, J. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+weight+estimation+of+polypeptide+chains+by+electrophoresis+in+SDS-polyacrylamide+gels.">Molecular weight estimation of polypeptide chains by electrophoresis in SDS-polyacrylamide gels.</a> Biochem Biophys Res Commun. 28 (5), 815-820 (1967).</li><li>Riches, A., Campbell, E., Borger, E., Powis, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+exosome+release+from+mammary+epithelial+and+breast+cancer+cells+-+a+new+regulatory+pathway.">Regulation of exosome release from mammary epithelial and breast cancer cells - a new regulatory pathway.</a> Eur J Cancer. 50 (5), 1025-1034 (2014).</li><li>Witwer, K. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardization+of+sample+collection,+isolation+and+analysis+methods+in+extracellular+vesicle+research.">Standardization of sample collection, isolation and analysis methods in extracellular vesicle research.</a> J Extracell Vesicles. 2, (2013).</li><li>van der Vlist, E. J., Nolte-'t Hoen, E. N., Stoorvogel, W., Arkesteijn, G. J., Wauben, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fluorescent+labeling+of+nano-sized+vesicles+released+by+cells+and+subsequent+quantitative+and+qualitative+analysis+by+high-resolution+flow+cytometry.">Fluorescent labeling of nano-sized vesicles released by cells and subsequent quantitative and qualitative analysis by high-resolution flow cytometry.</a> Nat Protoc. 7 (7), 1311-1326 (2012).</li><li>Kowal, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proteomic+comparison+defines+novel+markers+to+characterize+heterogeneous+populations+of+extracellular+vesicle+subtypes.">Proteomic comparison defines novel markers to characterize heterogeneous populations of extracellular vesicle subtypes.</a> Proc Natl Acad Sci U S A. 113 (8), E968-E977 (2016).</li><li>Li, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ADVANCED+IMAGING.+Extended-resolution+structured+illumination+imaging+of+endocytic+and+cytoskeletal+dynamics.">ADVANCED IMAGING. Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics.</a> Science. 349 (6251), (2015).</li><li>Outzen, H. C., Custer, R. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+of+human+normal+and+neoplastic+mammary+tissues+in+the+cleared+mammary+fat+pad+of+the+nude+mouse.">Growth of human normal and neoplastic mammary tissues in the cleared mammary fat pad of the nude mouse.</a> J Natl Cancer Inst. 55 (6), 1461-1466 (1975).</li><li>Sheffield, L. G., Welsch, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transplantation+of+human+breast+epithelia+to+mammary-gland-free+fat-pads+of+athymic+nude+mice:+influence+of+mammotrophic+hormones+on+growth+of+breast+epithelia.">Transplantation of human breast epithelia to mammary-gland-free fat-pads of athymic nude mice: influence of mammotrophic hormones on growth of breast epithelia.</a> Int J Cancer. 41 (5), 713-719 (1988).</li></ol>

Eksosomer har ofte egenskaper av cellene som frigjorde dem, og mengden av frigjorte eksosomer kan induseres av stimuli 4 . Kulturmediet fra celler kan samles og underkastes differensial ultracentrifugering for EV / eksosom samling ( Figur 1 ). Det er for øyeblikket ingen generell avtale om en ideell metode for å isolere EV / eksosomer. Den optimale fremgangsmåten som brukes her er blitt bestemt av nedstrømsapplikasjonen 14 . Ultracentrifug...

Eksosomer kan formidle cellulær kommunikasjon ved å overføre membran og cytosoliske proteiner, lipider og RNAer mellom celler 1 . Eksosomediert kommunikasjon har blitt vist å være involvert i mange fysiologiske og patologiske prosesser ( dvs. antigenpresentasjon, utvikling av toleranse 2 og tumorprogresjon 3 ). Eksosomer har ofte innhold som ligner på kildekildene som frigjør dem. Eksosomene kan således bære spesifikke celleegenskaper fra kildecellene og overføre disse egenskapene til cellene som inntar dem 4 . Eksosomer er 50- til 150 nm dobbeltlags membranvesikler og nåværende spesifikke markører ( f.eks. CD9, CD81, CD63, HSP70, Alix og TSG101). Eksosomer må derfor preges av ulike metoder for ulike aspekter. Overføringselektronmikroskopi kan brukes til å visualisere membranvesiklerSom eksosomer 4 , 5 . Nanopartikkelsporingsanalyse (NTA) og dynamisk lysspredningsanalyse (DLS) brukes til å måle størrelsen og antall rensede eksosomer 4 . Lipidmembraninnholdet i eksosomer kan verifiseres ved tetthetgradient. Eksosomale markører, slik som CD9, CD81, CD63, HSP70, Alix og TSG101 6 , 7 , kan måles ved Western blotting. Mammare basale celler har evnen til å generere brystkjertler når de implanteres i fettputer, mens luminale celler ikke kan 8 , 9 , 10 . Således refereres også basale celler i brystet til brystrepopulerende enheter. Ved bruk av modellen av basale og luminale celler i brom kan evnen til EV / eksosomer for å overføre celleegenskaper mellom forskjellige cellepopulasjoner undersøkes. Denne jobbenDemonstrerer metoden for å overføre kjerteldannende evne fra brystbasale epitelceller til mammale luminale epitelceller ved bruk av EVer / eksosomer avledet fra basale epithelialceller fra bryst. Luminale brystepitelceller kjøpte basalcelleegenskaper etter inntak av EV / eksosomer utskilt fra basale celler og kan deretter danne brystkjertler 4 .

<table>
	<tbody>
		<tr>
			<td>MCDB 170&#160;</td>
			<td>USBiological</td>
			<td>M2162</td>
			<td></td>
		</tr>
		<tr>
			<td>DMEM/F12</td>
			<td>Thermo</td>
			<td>1250062</td>
			<td></td>
		</tr>
		<tr>
			<td>Optima L-100K ultracentrifuge</td>
			<td>Beckman</td>
			<td>393253</td>
			<td></td>
		</tr>
		<tr>
			<td>SW28 Ti Rotor</td>
			<td>Beckman</td>
			<td>342204</td>
			<td></td>
		</tr>
		<tr>
			<td>SW41 Rotor</td>
			<td>Beckman</td>
			<td>331306</td>
			<td></td>
		</tr>
		<tr>
			<td>NANOSIGHT LM10</td>
			<td>Malvern</td>
			<td>NANOSIGHT LM10</td>
			<td>for nanoparticle tracking analysis (NTA)</td>
		</tr>
		<tr>
			<td>Optiprep&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>D1556</td>
			<td>60% (w/v) solution of iodixanol in water (sterile).</td>
		</tr>
		<tr>
			<td>CD81 antibody</td>
			<td>GeneTex</td>
			<td>GTX101766</td>
			<td>1:1000 in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween 20 at 4&#176;C, overnight&#160;</td>
		</tr>
		<tr>
			<td>CD9 antibody</td>
			<td>GeneTex</td>
			<td>GTX100912</td>
			<td>1:1000 in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween 20 at 4&#176;C, overnight&#160;</td>
		</tr>
		<tr>
			<td>CD63 antibody</td>
			<td>Abcam</td>
			<td>Ab59479</td>
			<td>1:1000 in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween 20 at 4&#176;C, overnight&#160;</td>
		</tr>
		<tr>
			<td>TSG101 antibody</td>
			<td>GeneTex</td>
			<td>GTX118736</td>
			<td>1:1000 in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween 20 at 4&#176;C, overnight&#160;</td>
		</tr>
		<tr>
			<td>GAPDH</td>
			<td>GeneTex</td>
			<td>GTX100118</td>
			<td>1:6000 in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween 20 at 4&#176;C, overnight&#160;</td>
		</tr>
		<tr>
			<td>CFSE (carboxyfluorescein succinimidyl diacetate ester)</td>
			<td>Thermo</td>
			<td>V12883</td>
			<td></td>
		</tr>
		<tr>
			<td>FACSCalibur</td>
			<td>BD Biosciences</td>
			<td></td>
			<td>fluorescence cell analyzer</td>
		</tr>
		<tr>
			<td>collagenase Type IV&#160;</td>
			<td>Thermo</td>
			<td>17104019</td>
			<td></td>
		</tr>
		<tr>
			<td>trypsin</td>
			<td>Thermo</td>
			<td>27250018</td>
			<td></td>
		</tr>
		<tr>
			<td>&#160;ITS</td>
			<td>Sigma-Aldrich</td>
			<td>I3146</td>
			<td>a mixture of recombinant human insulin, human transferrin, and sodium selenite</td>
		</tr>
		<tr>
			<td>accutase</td>
			<td>ebioscience</td>
			<td>00-4555-56</td>
			<td>a natural enzyme mixture with proteolytic and collagenolytic enzyme activity</td>
		</tr>
		<tr>
			<td>dispase&#160;</td>
			<td>STEMCELL</td>
			<td>7913</td>
			<td>5 mg/ml = 5 U/ml</td>
		</tr>
		<tr>
			<td>anti-CD49f antibody</td>
			<td>Biolegend</td>
			<td>313611</td>
			<td>1:50</td>
		</tr>
		<tr>
			<td>anti-EpCAM antibody</td>
			<td>Biolegend</td>
			<td>118213</td>
			<td>1:200</td>
		</tr>
		<tr>
			<td>FACSAria</td>
			<td>BD Biosciences</td>
			<td></td>
			<td>cell sorter</td>
		</tr>
		<tr>
			<td>carmine alum</td>
			<td>Sigma-Aldrich</td>
			<td>C1022</td>
			<td></td>
		</tr>
		<tr>
			<td>human mammary epithelial cells (HMLE cells, NAMECs)</td>
			<td></td>
			<td></td>
			<td>gifts from Dr. Robert Weinberg</td>
		</tr>
		<tr>
			<td>permount</td>
			<td>Thermo Fisher Scientific&#160;</td>
			<td>SP15-500</td>
			<td></td>
		</tr>
		<tr>
			<td>sodium bicarbonate</td>
			<td>Zymeset&#160;</td>
			<td>BSB101</td>
			<td></td>
		</tr>
		<tr>
			<td>EGF</td>
			<td>Peprotech</td>
			<td>AF-100-015</td>
			<td></td>
		</tr>
		<tr>
			<td>Hydrocoritisone</td>
			<td>Sigma-Aldrich</td>
			<td>SI-H0888</td>
			<td></td>
		</tr>
		<tr>
			<td>Insulin&#160;</td>
			<td>Sigma-Aldrich</td>
			<td>SI-I9278</td>
			<td></td>
		</tr>
		<tr>
			<td>BPE (bovine pituitary extract)</td>
			<td>Hammod Cell Tech&#160;</td>
			<td>1078-NZ</td>
			<td></td>
		</tr>
		<tr>
			<td>GW627368X&#160;</td>
			<td>Cayman</td>
			<td>10009162</td>
			<td></td>
		</tr>
		<tr>
			<td>15-cm culture dish</td>
			<td>Falcon&#160;</td>
			<td>353025</td>
			<td></td>
		</tr>
		<tr>
			<td>table-top centrifuge</td>
			<td>Eppendrof&#160;</td>
			<td>Centrifuge 3415R</td>
			<td></td>
		</tr>
		<tr>
			<td>ultracentrifuge tube</td>
			<td>Beckman</td>
			<td>344058</td>
			<td></td>
		</tr>
		<tr>
			<td>PBS (Phosphate-buffered saline)&#160;</td>
			<td>Corning</td>
			<td>46-013-CM</td>
			<td></td>
		</tr>
		<tr>
			<td>BCA Protein Assay</td>
			<td>Thermo Fisher Scientific&#160;</td>
			<td>23228</td>
			<td></td>
		</tr>
		<tr>
			<td>Transmission Electron Microscopy</td>
			<td>Hitachi</td>
			<td>HT7700</td>
			<td></td>
		</tr>
		<tr>
			<td>gelatin&#160;</td>
			<td>STEMCELL</td>
			<td>7903</td>
			<td></td>
		</tr>
		<tr>
			<td>10-cm culture dish</td>
			<td>Falcon&#160;</td>
			<td>353003</td>
			<td></td>
		</tr>
		<tr>
			<td>6-well culture dish</td>
			<td>Corning</td>
			<td>3516</td>
			<td></td>
		</tr>
		<tr>
			<td>female C57BL/6 mice</td>
			<td>NLAC (National Laboratory Animal Center</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>FBS (Fetal Bovine Serum)</td>
			<td>BioWest&#160;</td>
			<td>S01520</td>
			<td></td>
		</tr>
		<tr>
			<td>gentamycin</td>
			<td>Thermo Fisher Scientific&#160;</td>
			<td>15710072</td>
			<td></td>
		</tr>
		<tr>
			<td>Pen/Strep</td>
			<td>Corning</td>
			<td>30-002-Cl</td>
			<td></td>
		</tr>
		<tr>
			<td>DNase I</td>
			<td>5PRIMER</td>
			<td>2500120</td>
			<td></td>
		</tr>
		<tr>
			<td>isofluorane&#160;</td>
			<td>Halocarbon</td>
			<td>NPC12164-002-25</td>
			<td></td>
		</tr>
		<tr>
			<td>formaldehyde</td>
			<td>MACRON</td>
			<td>H121-08</td>
			<td></td>
		</tr>
		<tr>
			<td>EtOH (Ethanol)</td>
			<td>J.T. Baker</td>
			<td>800605</td>
			<td></td>
		</tr>
		<tr>
			<td>glacial acetic acid</td>
			<td>Panreac</td>
			<td>131008.1611</td>
			<td></td>
		</tr>
		<tr>
			<td>aluminum potassium sulfate</td>
			<td>Sigma-Aldrich</td>
			<td>12625</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylene&#160;</td>
			<td>Leica</td>
			<td>3803665</td>
			<td></td>
		</tr>
		<tr>
			<td>0.22 &#956;m membranes</td>
			<td>Merck Millipore</td>
			<td>Millex-GP</td>
			<td></td>
		</tr>
		<tr>
			<td>AUTOCLIP Wound Clips, 9 mm</td>
			<td>BD Biosciences</td>
			<td>427631</td>
			<td></td>
		</tr>
		<tr>
			<td>AUTOCLIP Wound Clip Applier</td>
			<td>BD Biosciences</td>
			<td>427630</td>
			<td></td>
		</tr>
		<tr>
			<td>CellMask&#8482; Deep Red</td>
			<td>Thermo Fisher Scientific&#160;</td>
			<td>C10046</td>
			<td>plasma membrane stain</td>
		</tr>
	</tbody>
</table>

transfer of mammary gland-forming ability between mammary basal epithelial cells and mammary luminal cells via extracellular vesicles/exosomes

Celler kan kommunisere via eksosomer, ~ 100 nm ekstracellulære vesikler (EVer) som inneholder proteiner, lipider og nukleinsyrer. Non-adherent / mesenchymal mammary epithelial cell (NAMEC) -deriverte ekstracellulære vesikler kan isoleres fra NAMEC medium via differensial ultracentrifugering. Basert på dens tetthet kan EVs renses via ultracentrifugering ved 110.000 x g. EV-preparatet fra ultracentrifugering kan separeres ytterligere under anvendelse av en kontinuerlig tetthetgradient for å forhindre forurensning med løselige proteiner. De rensede EV-stoffene kan deretter evalueres ytterligere ved hjelp av nanopartikkel-sporingsanalyse, som måler størrelsen og antall vesikler i preparatet. De ekstracellulære vesiklene med en størrelse som strekker seg fra 50 til 150 nm er eksosomer. De NAMEC-avledede EV / eksosomer kan inntas av brystepitelceller, som kan måles ved hjelp av flytcytometri og konfokal mikroskopi. Enkelte brystkreftegenskaper ( f.eks. Brystkreftdannende evne) kanOverføres fra stamme-lignende NAMECer til brystepitelceller via de NAMEC-avledede EV / eksosomer. Isolerte primære EpCAM hi / CD49f lo luminale brystepitelceller kan ikke danne brystkjertler etter transplantering i musfettputer, mens EpCAM lo / CD49f hi basale brystepitelceller danner brystkjertler etter transplantasjon. Opptak av NAMEC-avledede EV&#39;er / eksosomer av EpCAM hi / CD49f lo luminale brystepitelceller gjør det mulig for dem å generere brystkjertler etter transplantering i fettputer. EV-ene / eksosomene avledet fra stamlignende brystepitelceller overfører brystkjerteldannende evne til EpCAM hi / CD49f luminale brystepitelceller.

Siden det har blitt vist at blokkering av PGE 2 / EP 4- signalering utløser EV / eksosomfrigivelse fra basallignende stamceller 4 av brom, presenterer dette arbeid en metode for å isolere de induserte EVs / eksosomer fra brystepitelbasisk celle (NAMEC) -kultur. Siden NAMECs dyrkes i serumfritt medium, er det ingen eksisterende EV / eksosomer avledet fra serum 13 . For celler som dyrkes i serumholdig medium, må eksisterende eksos...

Watch this Scientific Journal Video about Transfer of Mammary Gland-forming Ability Between Mammary Basal Epithelial Cells and Mammary Luminal Cells via Extracellular Vesicles/Exosomes at JoVE.com

Transfer of Mammary Gland-forming Ability Between Mammary Basal Epithelial Cells and Mammary Luminal Cells via Extracellular Vesicles/Exosomes

Denne protokollen beskriver metoder for rensing, kvantifisering og karakterisering av ekstracellulære vesikler (EVs) / eksosomer fra ikke-adherent / mesenkymale brystepitelceller og for bruk av dem for å overføre brystkjerteldannende evne til luminale brystepitelceller. EVs / eksosomer avledet fra stamlignende brystepitelceller kan overføre denne celleegenskapen til celler som inntar EVs / eksosomer.

Overføring av Mammary Gland-forming Ability Mellom Mammary Basal Epithelial Cells og Mammary Luminal Cells Via Extracellular Vesicles / Exosomes

Cells can communicate via exosomes, ~100-nm extracellular vesicles (EVs) that contain proteins, lipids, and nucleic acids. Non-adherent/mesenchymal ...

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