Name: Caratterizzazione della secrezione di vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro (Video) | JoVE Journal
Uploaded: 2025-12-05T13:20:00.000+00:00
Duration: 10 min 40 s
Description: Scopri la caratterizzazione delle vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro. Questo metodo consente di studiare la comunicazione cellulare nel tessuto adiposo.

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<ol><li>Kershaw, E. E., Flier, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipose+tissue+as+an+endocrine+organ%5BTitle%5D)+AND+%22J+Clin+Endocrinol+Metab%22%5BJournal%5D)">Adipose tissue as an endocrine organ.</a> J Clin Endocrinol Metab. 89 (3), 2548-2556 (2004).</li>
<li>Wu, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Beige+adipocytes+are+a+distinct+type+of+thermogenic+fat+cell+in+mouse+and+human%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human.</a> Cell. 150 (2), 366-376 (2012).</li>
<li>Luo, L., Liu, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipose+tissue+in+control+of+metabolism%5BTitle%5D)+AND+%22J+Endocrinol%22%5BJournal%5D)">Adipose tissue in control of metabolism.</a> J Endocrinol. 231 (3), R77-R99 (2016).</li>
<li>Lynes, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+cold-induced+lipokine+12%2C13-diHOME+promotes+fatty+acid+transport+into+brown+adipose+tissue%5BTitle%5D)+AND+%22Nat+Med%22%5BJournal%5D)">The cold-induced lipokine 12,13-diHOME promotes fatty acid transport into brown adipose tissue.</a> Nat Med. 23, 631-637 (2017).</li>
<li>Sakers, A., De Siqueira, M. K., Seale, P., Villanueva, C. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipose-tissue+plasticity+in+health+and+disease%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Adipose-tissue plasticity in health and disease.</a> Cell. 185 (3), 419-446 (2022).</li>
<li>Santoro, A., McGraw, T. E., Kahn, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Insulin+action+in+adipocytes%2C+adipose+remodeling%2C+and+systemic+effects%5BTitle%5D)+AND+%22Cell+Metab%22%5BJournal%5D)">Insulin action in adipocytes, adipose remodeling, and systemic effects.</a> Cell Metab. 33 (4), 748-757 (2021).</li>
<li>Czech, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mechanisms+of+insulin+resistance+related+to+white%2C+beige%2C+and+brown+adipocytes%5BTitle%5D)+AND+%22Mol+Metab%22%5BJournal%5D)">Mechanisms of insulin resistance related to white, beige, and brown adipocytes.</a> Mol Metab. 34, 27-42 (2020).</li>
<li>Kahn, S. E., Hull, R. L., Utzschneider, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Mechanisms+linking+obesity+to+insulin+resistance+and+type+2+diabetes%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Mechanisms linking obesity to insulin resistance and type 2 diabetes.</a> Nature. 444, 840-846 (2006).</li>
<li>Feng, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((SNAP23+regulates+BAX-dependent+adipocyte+programmed+cell+death+independently+of+canonical+macroautophagy%5BTitle%5D)+AND+%22J+Clin+Invest%22%5BJournal%5D)">SNAP23 regulates BAX-dependent adipocyte programmed cell death independently of canonical macroautophagy.</a> J Clin Invest. 128 (9), 3941-3956 (2018).</li>
<li>Ouchi, N., Parker, J. L., Lugus, J. J., Walsh, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipokines+in+inflammation+and+metabolic+disease%5BTitle%5D)+AND+%22Nat+Rev+Immunol%22%5BJournal%5D)">Adipokines in inflammation and metabolic disease.</a> Nat Rev Immunol. 11, 85-97 (2011).</li>
<li>Zhang, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Positional+cloning+of+the+mouse+obese+gene+and+its+human+homologue%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Positional cloning of the mouse obese gene and its human homologue.</a> Nature. 372, 425-432 (1994).</li>
<li>Scherer, P. E., Williams, S., Fogliano, M., Baldini, G., Lodish, H. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+novel+serum+protein+similar+to+C1q%2C+produced+exclusively+in+adipocytes%5BTitle%5D)+AND+%22J+Biol+Chem%22%5BJournal%5D)">A novel serum protein similar to C1q, produced exclusively in adipocytes.</a> J Biol Chem. 270 (45), 26746-26749 (1995).</li>
<li>Considine, R. V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Serum+immunoreactive-leptin+concentrations+in+normal-weight+and+obese+humans%5BTitle%5D)+AND+%22N+Engl+J+Med%22%5BJournal%5D)">Serum immunoreactive-leptin concentrations in normal-weight and obese humans.</a> N Engl J Med. 334, 292-295 (1996).</li>
<li>Hotamisligil, G. S., Shargill, N. S., Spiegelman, B. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipose+expression+of+tumor+necrosis+factor-alpha%3A+Direct+role+in+obesity-linked+insulin+resistance%5BTitle%5D)+AND+%22Science%22%5BJournal%5D)">Adipose expression of tumor necrosis factor-alpha: Direct role in obesity-linked insulin resistance.</a> Science. 259 (5091), 87-91 (1993).</li>
<li>Steppan, C. M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+hormone+resistin+links+obesity+to+diabetes%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">The hormone resistin links obesity to diabetes.</a> Nature. 409, 307-312 (2001).</li>
<li>Yang, Q., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Serum+retinol+binding+protein+4+contributes+to+insulin+resistance+in+obesity+and+type+2+diabetes%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Serum retinol binding protein 4 contributes to insulin resistance in obesity and type 2 diabetes.</a> Nature. 436, 356-362 (2005).</li>
<li>Tilg, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipokines%3A+Masterminds+of+metabolic+inflammation%5BTitle%5D)+AND+%22Nat+Rev+Immunol%22%5BJournal%5D)">Adipokines: Masterminds of metabolic inflammation.</a> Nat Rev Immunol. 25, 250-265 (2025).</li>
<li>Clemente-Suarez, V. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+role+of+adipokines+in+health+and+disease%5BTitle%5D)+AND+%22Biomedicines%22%5BJournal%5D)">The role of adipokines in health and disease.</a> Biomedicines. 11 (5), 1290(2023).</li>
<li>Hallal, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Understanding+the+extracellular+vesicle+surface+for+clinical+molecular+biology%5BTitle%5D)+AND+%22J+Extracell+Vesicles%22%5BJournal%5D)">Understanding the extracellular vesicle surface for clinical molecular biology.</a> J Extracell Vesicles. 11 (10), e12260(2022).</li>
<li>Welsh, J. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Minimal+information+for+studies+of+extracellular+vesicles+%28MISEV2023%29%3A+From+basic+to+advanced+approaches%5BTitle%5D)+AND+%22J+Extracell+Vesicles%22%5BJournal%5D)">Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches.</a> J Extracell Vesicles. 13 (2), e12404(2024).</li>
<li>Le Lay, S., Scherer, P. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Exploring+adipose+tissue-derived+extracellular+vesicles+in+inter-organ+crosstalk%3A+Implications+for+metabolic+regulation+and+adipose+tissue+function%5BTitle%5D)+AND+%22Cell+Rep%22%5BJournal%5D)">Exploring adipose tissue-derived extracellular vesicles in inter-organ crosstalk: Implications for metabolic regulation and adipose tissue function.</a> Cell Rep. 44 (6), 115732(2025).</li>
<li>Shimoda, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Exosome+surface+glycans+reflect+osteogenic+differentiation+of+mesenchymal+stem+cells%3A+Profiling+by+an+evanescent+field+fluorescence-assisted+lectin+array+system%5BTitle%5D)+AND+%22Sci+Rep%22%5BJournal%5D)">Exosome surface glycans reflect osteogenic differentiation of mesenchymal stem cells: Profiling by an evanescent field fluorescence-assisted lectin array system.</a> Sci Rep. 9, 11497(2019).</li>
<li>Albuquerque, P. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+Paracoccidioides+brasiliensis+glycan+shares+serologic+and+functional+properties+with+cryptococcal+glucuronoxylomannan%5BTitle%5D)+AND+%22Fungal+Genet+Biol%22%5BJournal%5D)">A Paracoccidioides brasiliensis glycan shares serologic and functional properties with cryptococcal glucuronoxylomannan.</a> Fungal Genet Biol. 49 (11), 943-954 (2012).</li>
<li>Li, Y., Tang, X., Gu, Y., Zhou, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipocyte-derived+extracellular+vesicles%3A+Small+vesicles+with+big+impact%5BTitle%5D)+AND+%22Front+Biosci+%28Landmark+Ed%29%22%5BJournal%5D)">Adipocyte-derived extracellular vesicles: Small vesicles with big impact.</a> Front Biosci (Landmark Ed). 28 (7), 149(2023).</li>
<li>Thomou, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipose-derived+circulating+miRNAs+regulate+gene+expression+in+other+tissues%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">Adipose-derived circulating miRNAs regulate gene expression in other tissues.</a> Nature. 542, 450-455 (2017).</li>
<li>Crewe, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((An+endothelial-to-adipocyte+extracellular+vesicle+axis+governed+by+metabolic+state%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">An endothelial-to-adipocyte extracellular vesicle axis governed by metabolic state.</a> Cell. 175 (3), 695-708.e13 (2018).</li>
<li>Flaherty, S. E. 3rd, et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+lipase-independent+pathway+of+lipid+release+and+immune+modulation+by+adipocytes%5BTitle%5D)+AND+%22Science%22%5BJournal%5D)">A lipase-independent pathway of lipid release and immune modulation by adipocytes.</a> Science. 363 (6430), 989-993 (2019).</li>
<li>Clement, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipocyte+extracellular+vesicles+carry+enzymes+and+fatty+acids+that+stimulate+mitochondrial+metabolism+and+remodeling+in+tumor+cells%5BTitle%5D)+AND+%22EMBO+J%22%5BJournal%5D)">Adipocyte extracellular vesicles carry enzymes and fatty acids that stimulate mitochondrial metabolism and remodeling in tumor cells.</a> EMBO J. 39, e102525(2020).</li>
<li>Kulaj, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipocyte-derived+extracellular+vesicles+increase+insulin+secretion+through+transport+of+insulinotropic+protein+cargo%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Adipocyte-derived extracellular vesicles increase insulin secretion through transport of insulinotropic protein cargo.</a> Nat Commun. 14, 709(2023).</li>
<li>McCann, J. V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Reporter+mice+for+isolating+and+auditing+cell+type-specific+extracellular+vesicles+in+vivo%5BTitle%5D)+AND+%22Genesis%22%5BJournal%5D)">Reporter mice for isolating and auditing cell type-specific extracellular vesicles in vivo.</a> Genesis. 58 (7), e23369(2020).</li>
<li>Men, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Exosome+reporter+mice+reveal+the+involvement+of+exosomes+in+mediating+neuron+to+astroglia+communication+in+the+CNS%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Exosome reporter mice reveal the involvement of exosomes in mediating neuron to astroglia communication in the CNS.</a> Nat Commun. 10, 4136(2019).</li>
<li>Jeffery, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Characterization+of+Cre+recombinase+models+for+the+study+of+adipose+tissue%5BTitle%5D)+AND+%22Adipocyte%22%5BJournal%5D)">Characterization of Cre recombinase models for the study of adipose tissue.</a> Adipocyte. 3 (3), 206-211 (2014).</li>
<li>Emont, M. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((A+single-cell+atlas+of+human+and+mouse+white+adipose+tissue%5BTitle%5D)+AND+%22Nature%22%5BJournal%5D)">A single-cell atlas of human and mouse white adipose tissue.</a> Nature. 603, 926-933 (2022).</li>
<li>Krylova, S. V., Feng, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+machinery+of+exosomes%3A+Biogenesis%2C+release%2C+and+uptake%5BTitle%5D)+AND+%22Int+J+Mol+Sci%22%5BJournal%5D)">The machinery of exosomes: Biogenesis, release, and uptake.</a> Int J Mol Sci. 24 (2), (2023).</li>
<li>Crewe, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+challenges+of+interrogating+adipose+tissue+extracellular+vesicle+functions+in+physiology%5BTitle%5D)+AND+%22Commun+Biol%22%5BJournal%5D)">The challenges of interrogating adipose tissue extracellular vesicle functions in physiology.</a> Commun Biol. 5, 581(2022).</li>
<li>Blandin, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Lipidomic+analysis+of+adipose-derived+extracellular+vesicles+reveals+specific+EV+lipid+sorting+informative+of+the+obesity+metabolic+state%5BTitle%5D)+AND+%22Cell+Rep%22%5BJournal%5D)">Lipidomic analysis of adipose-derived extracellular vesicles reveals specific EV lipid sorting informative of the obesity metabolic state.</a> Cell Rep. 42 (3), 112169(2023).</li>
<li>Matilainen, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Increased+secretion+of+adipocyte-derived+extracellular+vesicles+is+associated+with+adipose+tissue+inflammation+and+the+mobilization+of+excess+lipid+in+human+obesity%5BTitle%5D)+AND+%22J+Transl+Med%22%5BJournal%5D)">Increased secretion of adipocyte-derived extracellular vesicles is associated with adipose tissue inflammation and the mobilization of excess lipid in human obesity.</a> J Transl Med. 22, 623(2024).</li>
<li>Thery, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Minimal+information+for+studies+of+extracellular+vesicles+2018+%28MISEV2018%29%3A+A+position+statement+of+the+International+Society+for+Extracellular+Vesicles+and+update+of+the+MISEV2014+guidelines%5BTitle%5D)+AND+%22J+Extracell+Vesicles%22%5BJournal%5D)">Minimal information for studies of extracellular vesicles 2018 (MISEV2018): A position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines.</a> J Extracell Vesicles. 7 (1), 1535750(2018).</li>
<li>Olson, A. L., Knight, J. B., Pessin, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Syntaxin+4%2C+VAMP2%2C+and%2For+VAMP3%2Fcellubrevin+are+functional+target+membrane+and+vesicle+SNAP+receptors+for+insulin-stimulated+GLUT4+translocation+in+adipocytes%5BTitle%5D)+AND+%22Mol+Cell+Biol%22%5BJournal%5D)">Syntaxin 4, VAMP2, and/or VAMP3/cellubrevin are functional target membrane and vesicle SNAP receptors for insulin-stimulated GLUT4 translocation in adipocytes.</a> Mol Cell Biol. 17 (5), 2425-2435 (1997).</li>
<li>Oh, E., Miller, R. A., Thurmond, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Syntaxin+4+overexpression+ameliorates+effects+of+aging+and+high-fat+diet+on+glucose+control+and+extends+lifespan%5BTitle%5D)+AND+%22Cell+Metab%22%5BJournal%5D)">Syntaxin 4 overexpression ameliorates effects of aging and high-fat diet on glucose control and extends lifespan.</a> Cell Metab. 22 (3), 499-507 (2015).</li>
<li>Merz, K. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Enrichment+of+the+exocytosis+protein+STX4+in+skeletal+muscle+remediates+peripheral+insulin+resistance+and+alters+mitochondrial+dynamics+via+Drp1%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Enrichment of the exocytosis protein STX4 in skeletal muscle remediates peripheral insulin resistance and alters mitochondrial dynamics via Drp1.</a> Nat Commun. 13, 424(2022).</li>
<li>Yu, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Involution+of+brown+adipose+tissue+through+a+syntaxin+4+dependent+pyroptosis+pathway%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Involution of brown adipose tissue through a syntaxin 4 dependent pyroptosis pathway.</a> Nat Commun. 15, 2856(2024).</li>
<li>Verweij, F. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Quantifying+exosome+secretion+from+single+cells+reveals+a+modulatory+role+for+GPCR+signaling%5BTitle%5D)+AND+%22J+Cell+Biol%22%5BJournal%5D)">Quantifying exosome secretion from single cells reveals a modulatory role for GPCR signaling.</a> J Cell Biol. 217 (3), 1129-1142 (2018).</li>
<li>Arora, A., Sharma, V., Gupta, R., Aggarwal, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Isolation+and+characterization+of+extracellular+vesicles+derived+from+ex+vivo+culture+of+visceral+adipose+tissue%5BTitle%5D)+AND+%22Bio+Protoc%22%5BJournal%5D)">Isolation and characterization of extracellular vesicles derived from ex vivo culture of visceral adipose tissue.</a> Bio Protoc. 14 (11), e5011(2024).</li>
<li>Benedikter, B. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ultrafiltration+combined+with+size+exclusion+chromatography+efficiently+isolates+extracellular+vesicles+from+cell+culture+media+for+compositional+and+functional+studies%5BTitle%5D)+AND+%22Sci+Rep%22%5BJournal%5D)">Ultrafiltration combined with size exclusion chromatography efficiently isolates extracellular vesicles from cell culture media for compositional and functional studies.</a> Sci Rep. 7, 15297(2017).</li>
<li>Nordin, J. Z., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Ultrafiltration+with+size-exclusion+liquid+chromatography+for+high+yield+isolation+of+extracellular+vesicles+preserving+intact+biophysical+and+functional+properties%5BTitle%5D)+AND+%22Nanomedicine%22%5BJournal%5D)">Ultrafiltration with size-exclusion liquid chromatography for high yield isolation of extracellular vesicles preserving intact biophysical and functional properties.</a> Nanomedicine. 11 (4), 879-883 (2015).</li>
</ol>

<table><tbody><tr><td>0,5 M EDTA, pH 8,0</td><td>Fisher</td><td>AM9260G</td><td>Diluizione 500x</td></tr><tr><td>10x PBS</td><td>Fisher Scientific</td><td>70-013-032</td><td>Diluizione 1000x con acqua</td></tr><tr><td>1 M HEPES</td><td>Gibco</td><td>15630080</td><td>Diluizione 1000x</td></tr><tr><td>Kit di isolamento del progenitore del tessuto adiposo, topo</td><td>Miltenyi biotec</td><td>30-106-639</td><td>Pronto all'uso</td></tr><tr><td>Filtro centrifugo Amicon Ultra, MWCO da 100 kDa</td><td>Sigma</td><td>UFC9100</td><td>Volume di campione di 15 mL</td></tr><tr><td>Filtro centrifugo Amicon Ultra, MWCO da 100 kDa</td><td>Sigma</td><td>UFC9100</td><td>Volume di campione di 4 mL</td></tr><tr><td>BSA</td><td>Sigma</td><td>A6003</td><td>Pronto all'uso</td></tr><tr><td>Anticorpo di Calnexina</td><td>Calnexina</td><td>ab22595</td><td>Diluizione 1000x</td></tr><tr><td>Anticorpo CD63</td><td>abcam</td><td>ab217345</td><td>Diluizione 500x</td></tr><tr><td>CD81</td><td>Tecnologia di segnalazione cellulare</td><td>10037s</td><td>Diluizione 500x</td></tr><tr><td>Corning&nbsp; BioCoat&nbsp; Diapositiva della Cultura Collagen I</td><td>Sigma</td><td>CLS354630</td><td>Pronto all'uso</td></tr><tr><td>Corning&nbsp; Filtro cellulare</td><td>Sigma</td><td>CLS431750</td><td>40 & mu; m dimensione dei pori</td></tr><tr><td>Corning&nbsp; Filtro cellulare</td><td>Sigma</td><td>CLS431752</td><td>100 & mu; m dimensione dei pori</td></tr><tr><td>Anticorpo COX IV</td><td>Tecnologia di segnalazione cellulare</td><td>4844</td><td>Diluizione 1000x</td></tr><tr><td>DMEM</td><td>Gibco</td><td>11965092</td><td>Pronto all'uso</td></tr><tr><td>DMEM/F-12, integratore GlutaMAX</td><td>Thermofisher</td><td>10565018</td><td>Pronto all'uso</td></tr><tr><td>Dnase 1</td><td>Sigma</td><td>D4513</td><td>Stock 5 KU/mL a -20 &deg; C</td></tr><tr><td>Integratore GlutaMAX</td><td>Gibco</td><td>35050061</td><td>Diluizione 100x</td></tr><tr><td>Halt & trade; Cocktail di inibitori di proteasi e fosfatasi (100X)</td><td>Thermofisher</td><td>78444</td><td>Pronto all'uso</td></tr><tr><td>Anticorpo HSP70</td><td>abcam</td><td>ab181606</td><td>Diluizione 1000x</td></tr><tr><td>Invitrogen eBioscience 10X RBC Lysis Buffer (Multi-specie)</td><td>Fisher Scientific</td><td>50-112-9743</td><td>Diluizione 10x in acqua</td></tr><tr><td>Izon Science Usa Ltd QEVORIGINAL 70NMGEN2 5/PK SDP</td><td>Fisher Scientific</td><td>NC2042225</td><td>Pronto all'uso</td></tr><tr><td>Microscopio confocale LEICA STELLARIS 8</td><td>Microscopi Leica</td><td>N/A</td><td></td></tr><tr><td>Liberase TM Grado di Ricerca</td><td>Sigma</td><td>LIBTM-RO</td><td>Scorta 5 mg/mL a -20 &deg; C</td></tr><tr><td>Filtro a siringa Millex MCE</td><td>Sigma</td><td>SLAA025</td><td>0,8 &mu; m dimensione dei pori</td></tr><tr><td>NanoSight Pro</td><td>Malvern Panalytical</td><td>N/A</td><td></td></tr><tr><td>Novex Tris-Glycine Mini Protein Gels, 4&ndash; 12%,</td><td>Thermofisher</td><td>XP04120BOX</td><td>Pronto all'uso</td></tr><tr><td>Tampone campione Novex Tris-Glicina SDS (2x)</td><td>Thermofisher</td><td>LC2676</td><td>Pronto all'uso</td></tr><tr><td>Anticorpo anti-topo Ly-6A/E (Sca-1) Blu del Pacifico</td><td>BioLegend</td><td>108119</td><td>Diluizione 25x</td></tr><tr><td>Soluzione di paraformaldeide (16%)</td><td>Fisher Scientific</td><td>50-980-487</td><td>diluire al 4% con PBS</td></tr><tr><td>pluriStrainer S / 30 & micro; m (Filtro cellulare)</td><td>Fisher Scientific</td><td>NC0922459</td><td>30 & mu; m dimensione dei pori</td></tr><tr><td>Soluzione Ponceau S</td><td>Sigma</td><td>P7170-1L</td><td>Pronto all'uso</td></tr><tr><td>Primocin</td><td>Invivogen</td><td>NC9141851</td><td>Diluizione 500x</td></tr><tr><td>Reagente Antifade ProLong Gold con DAPI</td><td>Tecnologia di segnalazione cellulare</td><td>8961S</td><td>Pronto all'uso</td></tr><tr><td>Anticorpo anti-topo purificato CD16/32</td><td>BioLegend</td><td>101301</td><td>Diluizione 100x</td></tr><tr><td>RIPA buffer&nbsp;</td><td>Fisher Scientific</td><td>PI89900</td><td>Diluizione 10x in acqua</td></tr><tr><td>Citometro a flusso spettrale (Aurora 3)</td><td>Cytek Biosciences</td><td>N/A</td><td></td></tr><tr><td>Anticorpo sintassina 4</td><td>Sistemi sinaptici</td><td>110042</td><td>Diluizione 5000x</td></tr><tr><td>Tecnai 20 (Microscopio Elettronico a Trasmissione)</td><td>FEI</td><td>N/A</td><td></td></tr><tr><td>Tergeo-EM</td><td>PIE Scientific LLC</td><td>N/A</td><td></td></tr><tr><td>Anticorpo Vinculina</td><td>abcam</td><td>circa 18508</td><td>Diluizione 5000x</td></tr></tbody></table>

caratterizzazione della secrezione di vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter cd63-gfp in vivo e in vitro

Characterization of Adipocyte-Derived Extracellular Vesicle Secretion Using a CD63-GFP Reporter Mouse Model In Vivo and In Vitro

Utilizzando questo protocollo di rilevamento degli AdEV tramite l'assorbimento di APC tramite microscopia confocale, abbiamo isolato le APC dai tessuti adiposi inguinali di topi AdipCD63-GFP e Stopfl/fl/CD63-GFP e le abbiamo colorate con DAPI per la visualizzazione dei nuclei. Esaminate con microscopia confocale, le APC di topi Stopfl/fl/CD63-GFP non hanno mostrato punti GFP rilevabili (Figura 1A). Al contrario, le APC isolate da topi AdipCD63-GFP hanno mostrato numeros...

Caratterizzazione della secrezione di vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro

Adipocytes are increasingly recognized as active endocrine cells that secrete substantial quantities of EVs enriched in proteins, lipids, and nucleic ...

characterization-adipocyte-derived-extracellular-vesicle-secretion

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JoVE Journal

Biology

Biologia

Il glicocalice endoteliale / endoteliale strato superficiale è perfettamente studiata usando la microscopia intravitale. Microscopia intravitale è tecnicamente difficile in un organo in movimento come il polmone. Si dimostra come brightfield simultanea e microscopia a fluorescenza possono essere utilizzate per stimare endoteliale spessore dello strato di superficie in un movimento...

Nella misura in vivo della superficie polmonare strato endoteliale mouse

Questo protocollo descrive metodi per purificare, quantificare e caratterizzare vescicole extracellulari (EVs) / esosomi da cellule epiteliali mammarie non aderenti / mesenchimali e per usarle per trasferire la capacità di formazione delle ghiandole mammarie alle cellule epiteliali mammarie luminali. EVs / esosomi derivati ​​da cellule epiteliali mammarie simili a stelo possono trasferire questa proprietà cellulare alle cellule che ingeriscono gli EV / esosomi.

Trasferimento di abilità di formazione del ghiandale mammario tra le cellule epiteliali basali del mammifero e le cellule luminescenti mammarie attraverso le vescicole / esosomi extracellulari

La differenziazione degli adipociti bianchi e beige da progenitori vascolari del tessuto adiposo porta il potenziale di miglioramento metabolico nell'obesità. Descriviamo i protocolli per CD34 + CD31 + isolamento di cellule endoteliali da grasso umano e per una successiva in vitro espansione e differenziazione in adipociti bianchi e beige. Diverse applicazioni a valle sono discussi.

Isolamento, espansione e adipogenico induzione di CD34 + CD31 + cellule endoteliali dal tessuto adiposo umano omentale e sottocutaneo

In questo protocollo, descriviamo il flusso di lavoro completo per rapido isolamento delle vescicole extracellulari da sangue umano intero e la caratterizzazione di marcatori specifici di analisi di fluorescenza-basata nanoparticle tracking. I risultati presentati mostrano un elevato livello di riproducibilità e possono essere regolati per i surnatanti di coltura cellulare.

Rapida caratterizzazione basata sulla fluorescenza delle singole vescicole extracellulari nel sangue umano con analisi Nanoparticle tracking

Descriviamo un sistema 3D di coltura extracellulare extracellulare umana-adipocito in vitro che permette la dissezione dei ruoli della matrice e adipociti nel contribuire al fenotipo metabolico del tessuto adiposo.

Un modello di coltura a matrice extracellulare extracellulare -adipocyte umano per lo studio del crosstalk metabolico a matrice

L'ultima generazione di strumenti di caratterizzazione dei veicoli elettrici è in grado di eseguire l'analisi di singoli veicoli elettrici su più parametri contemporaneamente. La citometria a nano-flusso misura tutte le particelle biologiche più grandi di 45 nm senza etichettatura e identifica le caratteristiche specifiche delle sottopopolazioni mediante una varietà di tecniche di etichettatura fluorescente.

Caratterizzazione di proteine transmembrana di vescicole extracellulari singole mediante citometria a nano-flusso

Il presente protocollo descrive la centrifugazione differenziale per isolare e caratterizzare EV rappresentative (esosomi e microvescicole) da MSC umane in coltura. Ulteriori applicazioni di questi veicoli elettrici sono spiegate anche in questo articolo.

Isolamento, caratterizzazione e applicazione terapeutica di vescicole extracellulari da cellule staminali mesenchimali umane in coltura

Questo protocollo descrive tutte le procedure, dalla coltura di cellule staminali mesenchimali di derivazione adiposa umana (ADSC) e la raccolta del surnatante all'estrazione delle vescicole extracellulari (EV) mediante ultracentrifugazione.

Purificazione e caratterizzazione di vescicole extracellulari da cellule staminali mesenchimali di derivazione adiposa umana

Qui, descriviamo un protocollo per isolare, coltivare e fenotipizzare le cellule endoteliali microvascolari dal tessuto adiposo sottocutaneo umano (hSATMVECs). Inoltre, descriviamo un modello sperimentale di cross-talk hSATMVEC-adipociti.

Studio del cross-talk tra cellule endoteliali adipose e adipociti nel tessuto adiposo sottocutaneo umano

Utilizzando un modello ex vivo di espianti, piccoli pezzi della porzione rostrale del tessuto adiposo bianco viscerale di topi magri vengono coltivati per 7–10 giorni, permettendo l'induzione e lo studio dell'infiammazione del tessuto adiposo durante l'alimentazione ricca di grassi.

Un modello topino ex vivo per l'infiammazione del tessuto adiposo ad alto contenuto di grassi

Scopri la caratterizzazione delle vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro. Questo metodo consente di studiare la comunicazione cellulare nel tessuto adiposo.

Caratterizzazione della secrezione di vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro (Video) | JoVE Journal

Caratterizzazione della secrezione di vescicole extracellulari derivate da adipociti utilizzando un modello murino con reporter CD63-GFP in vivo e in vitro

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