Name: Snelle uitputting van niermacrofagen met behulp van menselijke CD59/Intermedilysin Cell Ablation Tool (Video) | JoVE Journal
Uploaded: 2025-05-09T14:10:00.000+00:00
Duration: 10 min 15 s
Description: Ontdek hoe niermacrofagen snel uitgeput en geregenereerd worden met behulp van het menselijke CD59/intermedilysin-ablatietool in muizen. Deze methode onthult chemokine-gemedieerde monocytrecrutering en herstel van immuunsurveillance na nierbeschadiging.

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5 minutes

<ol><li>Epelman, S., Lavine, K. J., Randolph, G. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Origin+and+functions+of+tissue+macrophages%5BTitle%5D)+AND+%22Immunity%22%5BJournal%5D)">Origin and functions of tissue macrophages.</a> Immunity. 41 (1), 21-35 (2014).</li>
<li>Nelson, P. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+renal+mononuclear+phagocytic+system%5BTitle%5D)+AND+%22J+Am+Soc+Nephrol%22%5BJournal%5D)">The renal mononuclear phagocytic system.</a> J Am Soc Nephrol. 23 (2), 194-203 (2012).</li>
<li>Wang, Y., Harris, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages+in+renal+disease%5BTitle%5D)+AND+%22J+Am+Soc+Nephrol%22%5BJournal%5D)">Macrophages in renal disease.</a> J Am Soc Nephrol. 22 (1), 21-27 (2011).</li>
<li>Islamuddin, M., Qin, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Renal+macrophages+and+NLRP3+inflammasomes+in+kidney+diseases+and+therapeutics%5BTitle%5D)+AND+%22Cell+Death+Discov%22%5BJournal%5D)">Renal macrophages and NLRP3 inflammasomes in kidney diseases and therapeutics.</a> Cell Death Discov. 10 (1), 229(2024).</li>
<li>Ricardo, S. D., van Goor, H., Eddy, A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophage+diversity+in+renal+injury+and+repair%5BTitle%5D)+AND+%22J+Clin+Invest%22%5BJournal%5D)">Macrophage diversity in renal injury and repair.</a> J Clin Invest. 118 (11), 3522-3530 (2008).</li>
<li>Mily, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Polarization+of+M1+and+M2+Human+Monocyte-Derived+Cells+and+Analysis+with+Flow+Cytometry+upon+Mycobacterium+tuberculosis+Infection%5BTitle%5D)+AND+%22J+Vis+Exp%22%5BJournal%5D)">Polarization of M1 and M2 Human Monocyte-Derived Cells and Analysis with Flow Cytometry upon Mycobacterium tuberculosis Infection.</a> J Vis Exp. (163), e61807(2020).</li>
<li>Huen, S. C., Cantley, L. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages+in+Renal+Injury+and+Repair%5BTitle%5D)+AND+%22Annu+Rev+Physiol%22%5BJournal%5D)">Macrophages in Renal Injury and Repair.</a> Annu Rev Physiol. 79, 449-469 (2017).</li>
<li>Tang, P. M., Nikolic-Paterson, D. J., Lan, H. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages%3A+versatile+players+in+renal+inflammation+and+fibrosis%5BTitle%5D)+AND+%22Nat+Rev+Nephrol%22%5BJournal%5D)">Macrophages: versatile players in renal inflammation and fibrosis.</a> Nat Rev Nephrol. 15 (3), 144-158 (2019).</li>
<li>Liu, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Distinct+fate%2C+dynamics+and+niches+of+renal+macrophages+of+bone+marrow+or+embryonic+origins%5BTitle%5D)+AND+%22Nat+Commun%22%5BJournal%5D)">Distinct fate, dynamics and niches of renal macrophages of bone marrow or embryonic origins.</a> Nat Commun. 11 (1), 2280(2020).</li>
<li>Chew, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Kidney+resident+macrophages+have+distinct+subsets+and+multifunctional+roles%5BTitle%5D)+AND+%22Matrix+Biol%22%5BJournal%5D)">Kidney resident macrophages have distinct subsets and multifunctional roles.</a> Matrix Biol. 127, 23-37 (2024).</li>
<li>Cheung, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Resident+macrophage+subpopulations+occupy+distinct+microenvironments+in+the+kidney%5BTitle%5D)+AND+%22JCI+Insight%22%5BJournal%5D)">Resident macrophage subpopulations occupy distinct microenvironments in the kidney.</a> JCI Insight. 7 (20), 161078(2022).</li>
<li>Epelman, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Embryonic+and+adult-derived+resident+cardiac+macrophages+are+maintained+through+distinct+mechanisms+at+steady+state+and+during+inflammation%5BTitle%5D)+AND+%22Immunity%22%5BJournal%5D)">Embryonic and adult-derived resident cardiac macrophages are maintained through distinct mechanisms at steady state and during inflammation.</a> Immunity. 40 (1), 91-104 (2014).</li>
<li>Feng, D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Cre-inducible+human+CD59+mediates+rapid+cell+ablation+after+intermedilysin+administration%5BTitle%5D)+AND+%22J+Clin+Invest%22%5BJournal%5D)">Cre-inducible human CD59 mediates rapid cell ablation after intermedilysin administration.</a> J Clin Invest. 126 (6), 2321-2333 (2016).</li>
<li>Hu, W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Rapid+conditional+targeted+ablation+of+cells+expressing+human+CD59+in+transgenic+mice+by+intermedilysin%5BTitle%5D)+AND+%22Nat+Med%22%5BJournal%5D)">Rapid conditional targeted ablation of cells expressing human CD59 in transgenic mice by intermedilysin.</a> Nat Med. 14 (1), 98-103 (2008).</li>
<li>Giddings, K. S., Zhao, J., Sims, P. J., Tweten, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Human+CD59+is+a+receptor+for+the+cholesterol-dependent+cytolysin+intermedilysin%5BTitle%5D)+AND+%22Nat+Struct+Mol+Biol%22%5BJournal%5D)">Human CD59 is a receptor for the cholesterol-dependent cytolysin intermedilysin.</a> Nat Struct Mol Biol. 11 (12), 1173-1178 (2004).</li>
<li>Parkhurst, C. N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Microglia+promote+learning-dependent+synapse+formation+through+brain-derived+neurotrophic+factor%5BTitle%5D)+AND+%22Cell%22%5BJournal%5D)">Microglia promote learning-dependent synapse formation through brain-derived neurotrophic factor.</a> Cell. 155 (7), 1596-1609 (2013).</li>
<li>Cao, Q., Harris, D. C., Wang, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages+in+kidney+injury%2C+inflammation%2C+and+fibrosis%5BTitle%5D)+AND+%22Physiology%22%5BJournal%5D)">Macrophages in kidney injury, inflammation, and fibrosis.</a> Physiology. 30 (3), 183-194 (2015).</li>
<li>Li, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((The+role+of+macrophages+in+fibrosis+of+chronic+kidney+disease%5BTitle%5D)+AND+%22Biomed+Pharmacother%22%5BJournal%5D)">The role of macrophages in fibrosis of chronic kidney disease.</a> Biomed Pharmacother. 177, 117079(2024).</li>
<li>Liu, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Versatile+cell+ablation+tools+and+their+applications+to+study+loss+of+cell+functions%5BTitle%5D)+AND+%22Cell+Mol+Life+Sci%22%5BJournal%5D)">Versatile cell ablation tools and their applications to study loss of cell functions.</a> Cell Mol Life Sci. 76 (23), 4725-4743 (2019).</li>
<li>Yashchenko, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Cx3cr1+kidney+resident+macrophage+heterogeneity%5BTitle%5D)+AND+%22Front+Immunol%22%5BJournal%5D)">Cx3cr1 kidney resident macrophage heterogeneity.</a> Front Immunol. 14, 1082078(2023).</li>
<li>Duffield, J. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages+and+immunologic+inflammation+of+the+kidney%5BTitle%5D)+AND+%22Semin+Nephrol%22%5BJournal%5D)">Macrophages and immunologic inflammation of the kidney.</a> Semin Nephrol. 30 (3), 234-254 (2010).</li>
<li>Lee, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Distinct+macrophage+phenotypes+contribute+to+kidney+injury+and+repair%5BTitle%5D)+AND+%22J+Am+Soc+Nephrol%22%5BJournal%5D)">Distinct macrophage phenotypes contribute to kidney injury and repair.</a> J Am Soc Nephrol. 22 (2), 317-326 (2011).</li>
<li>Wynn, T. A., Vannella, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Macrophages+in+Tissue+Repair%2C+Regeneration%2C+and+Fibrosis%5BTitle%5D)+AND+%22Immunity%22%5BJournal%5D)">Macrophages in Tissue Repair, Regeneration, and Fibrosis.</a> Immunity. 44 (3), 450-462 (2016).</li>
<li>Guo, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Crosstalk+between+proximal+tubular+epithelial+cells+and+other+interstitial+cells+in+tubulointerstitial+fibrosis+after+renal+injury%5BTitle%5D)+AND+%22Front+Endocrinol%22%5BJournal%5D)">Crosstalk between proximal tubular epithelial cells and other interstitial cells in tubulointerstitial fibrosis after renal injury.</a> Front Endocrinol. 14, 1256375(2023).</li>
<li>Guillot, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Bile+acid-activated+macrophages+promote+biliary+epithelial+cell+proliferation+through+integrin+%CE%B1v%CE%B26+upregulation+following+liver+injury%5BTitle%5D)+AND+%22J+Clin+Invest%22%5BJournal%5D)">Bile acid-activated macrophages promote biliary epithelial cell proliferation through integrin αvβ6 upregulation following liver injury.</a> J Clin Invest. 131 (9), 132305(2021).</li>
<li>Kim, S. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Adipocyte+Death+Preferentially+Induces+Liver+Injury+and+Inflammation+Through+the+Activation+of+Chemokine+%28C-C+Motif%29+Receptor+2-Positive+Macrophages+and+Lipolysis%5BTitle%5D)+AND+%22Hepatology%22%5BJournal%5D)">Adipocyte Death Preferentially Induces Liver Injury and Inflammation Through the Activation of Chemokine (C-C Motif) Receptor 2-Positive Macrophages and Lipolysis.</a> Hepatology. 69 (5), 1965-1982 (2019).</li>
<li>Saxena, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/pubmed?term=((Generation+of+Atp6v1g3-Cre+mice+for+investigation+of+intercalated+cells+and+the+collecting+duct%5BTitle%5D)+AND+%22Am+J+Physiol+Renal+Physiol%22%5BJournal%5D)">Generation of Atp6v1g3-Cre mice for investigation of intercalated cells and the collecting duct.</a> Am J Physiol Renal Physiol. 325 (6), F770-F778 (2023).</li>
</ol>

<table><tbody><tr><td>0.45 &micro;m syringe filter</td><td>Millipore</td><td>SLHVR33RS</td><td></td></tr><tr><td>4-15% TGX Stain-Free Protein Gels</td><td>Bio-Rad</td><td>4568084</td><td></td></tr><tr><td>6X Loading buffer</td><td>Fisher</td><td>50-103-6570</td><td></td></tr><tr><td>70% Ethanol</td><td>WWR Life Science</td><td>64-17-5</td><td></td></tr><tr><td>ACK (Ammonium-Chloride-Potassium) Lysing Buffer</td><td>Gibco</td><td>A1049201</td><td></td></tr><tr><td>Affinity resin beads</td><td>Millipore</td><td>69670</td><td></td></tr><tr><td>Ampicillin sodium solution</td><td>Zymo Research</td><td>A1001-5</td><td></td></tr><tr><td>Anti-CD11b-PE-Cy7 (Clone M1/70)</td><td>Invitrogen</td><td>25-0112-82</td><td></td></tr><tr><td>Anti-CD16/32 (Fc&gamma;RIII/II, Clone 93)</td><td>eBioscience</td><td>48-0161-80</td><td></td></tr><tr><td>Anti-CD45-e450 (Clone 30-F11)</td><td>eBioscience</td><td>48-0451-82</td><td></td></tr><tr><td>Anti-F4/80-BV605(Clone BM8)</td><td>BioLegend&nbsp;</td><td>123133</td><td></td></tr><tr><td>Anti-hCD59-PE (Clone OV9A2)</td><td>Invitrogen</td><td>12-0596-42</td><td></td></tr><tr><td>Aqua Live/Dead dye</td><td>Invitrogen</td><td>&nbsp;&nbsp;&nbsp; L34957A</td><td></td></tr><tr><td>Beads (resin)</td><td>Millipore</td><td>69670</td><td></td></tr><tr><td>Benzonase Nuclease</td><td>Millipore</td><td>70664-10KUN</td><td></td></tr><tr><td>BugBuster protein extraction reagent</td><td>Millipore</td><td>70584-4</td><td></td></tr><tr><td>Centrifuge for microtubes</td><td>Eppendorf</td><td>5424</td><td></td></tr><tr><td>Centrifuge for tubes</td><td>Thermo Scientific</td><td>75-001-241</td><td></td></tr><tr><td>Collagenase type IV</td><td>Worthington Biochemical Corporation</td><td>LS004188</td><td></td></tr><tr><td>Corn oil</td><td>Sigma Aldrich</td><td>C8267</td><td></td></tr><tr><td>Deoxyribonuclease (DNAse) I</td><td>Worthington Biochemical Corporation</td><td>LS002007</td><td></td></tr><tr><td>Detoxi-Gel resin column</td><td>Millipore</td><td>69670</td><td></td></tr><tr><td>DPBS (Dulbecco&rsquo;s phosphate-buffered saline) solution</td><td>Corning</td><td>21-031-CV</td><td></td></tr><tr><td>EDTA tubes</td><td>BD</td><td>365974</td><td></td></tr><tr><td>FBS (Fetal bovine serum)</td><td>Gibco</td><td>10082-139</td><td></td></tr><tr><td>Glycerol</td><td>Fisher</td><td>BP229-1</td><td></td></tr><tr><td>HBSS (Hank&rsquo;s Ballanced Salt Solution)</td><td>Gibco</td><td>24020117</td><td></td></tr><tr><td>IPTG (Isopropyl &beta;-D-1-thiogalactopyranoside)&nbsp;</td><td>Millipore-Sigma</td><td>206-703-0</td><td></td></tr><tr><td>Isoflurane</td><td>VET one</td><td>502017</td><td></td></tr><tr><td>LB media</td><td></td><td></td><td></td></tr><tr><td>LSRFortessa flow cytometer</td><td>BD Biosciences</td><td></td><td></td></tr><tr><td>MES [2-(N-morpholino) ethanesulfonic acid]</td><td>Fisher</td><td>50-488-796</td><td></td></tr><tr><td>MOPS [3-(N-morpholino) propanesulfonic acid]</td><td>Fisher</td><td>50-213-522</td><td></td></tr><tr><td>Percoll density gradient media</td><td>Cytiva</td><td>17089101</td><td></td></tr><tr><td>Peristaltic pump</td><td>Fisher Scientific</td><td></td><td>Discontinued now, use alternative</td></tr><tr><td>PFA (Paraformaldehyde)</td><td>Thermo Scientific</td><td>I28800</td><td></td></tr><tr><td>Purification column</td><td>Millipore</td><td>UFC900308</td><td></td></tr><tr><td>rLysozyme solution</td><td>Novagen</td><td>20C71110</td><td></td></tr><tr><td>Shaking water bath&nbsp;</td><td>Thermo Scientific</td><td>TSSB15</td><td></td></tr><tr><td>Slide-A-Lyzer dialysis kit</td><td>Thermo</td><td>66107</td><td></td></tr><tr><td>Sodium deoxycholate</td><td>Fisher</td><td>BP349-100</td><td>Fresh made or less than a month</td></tr><tr><td>Sterile cell strainer (40 &mu;m)</td><td>Fisher Scientific</td><td>22-363-547</td><td></td></tr><tr><td>Tamoxifen</td><td>Sigma Aldrich</td><td>6734</td><td></td></tr><tr><td>Ultra centrifugal filter</td><td>Millipore</td><td>UFC900308</td><td></td></tr><tr><td>Ultrapure water</td><td>Thermo</td><td>10977-015</td><td></td></tr><tr><td>Vortex mixer</td><td>Fisher Scientific</td><td>2215365</td><td></td></tr><tr><td>&beta;-ME (&beta;-mercaptoethanol)</td><td>Fisher</td><td>BP176-100</td><td></td></tr></tbody></table>

snelle uitputting van niermacrofagen met behulp van menselijke cd59/intermedilysin cell ablation tool

Rapid Depletion of Renal Macrophages Using Human CD59/Intermedilysin Cell Ablation Tool

Snelle uitputting van niermacrofagen met behulp van menselijke CD59/Intermedilysin Cell Ablation Tool

Renal macrophages (RMs) are essential for kidney health, orchestrating immune surveillance, tissue homeostasis, and responses to injury. Previously, we ...

rapid-depletion-renal-macrophages-using-human-cd59intermedilysin-cell

Research

JoVE Journal

Developmental Biology

Ontwikkelingsbiologie

Macrofagen spelen een centrale rol in homeostase en pathologie in vele weefsels. Het protocol hier gepresenteerde beschrijft methoden voor uitputting van...

Uitputting en reconstitutie van macrofagen in Muizen

Macrofagen zijn plastic cellen van het hematopoietische systeem die een cruciale rol spelen in beschermende immuniteit en homeostase. In dit rapport beschrijven we geoptimaliseerde in vitro technieken voor het fenotypisch en functioneel karakteriseren van transplantatie-infiltrerende regulerende macrofagen die zich in het getransplanteerde orgaan verzamelen onder tolerogene omstandigheden.

Functionele karakterisering van regulerende macrofagen die de transplantaatreactieve immuniteit verhinderen

Technieken die betrouwbaar en efficiënt zijn voor het isoleren van immuuncellen in de nieren zijn nodig voor downstream-toepassingen. Dit vereist oppervlakte-antilichaammarkering van een klein aantal immuuncellen in de nieren. Hierin beschrijven we een beknopte methode voor het isoleren van immuuncellen in de nieren die schijnbaar dit doel bereikt.

Betrouwbaar en High Efficiency Winning van Kidney immuuncellen

Dit manuscript beschrijft een gedetailleerd protocol voor fenotypische en kwantitatieve analyse van de aldaar gevestigde macrofagen van rat nieren door middel van flowcytometrie. De resulterende gekleurde cellen kunnen ook worden gebruikt voor andere toepassingen, zoals celsortering, genexpressie analyse of functionele studies, aldus verkregen in het experimentele modelinformatie verhogen.

Fenotypische karakterisering van macrofagen van Rat Kidney door flowcytometrie

Dit protocol kan een onderzoeker te isoleren en karakteriseren weefsel-ingezeten macrofagen in diverse hallmark ontstoken weefsels dieet-geïnduceerde modellen van metabolische wanorde is onttrokken.

Isolatie, karakterisering en zuivering van macrofagen uit weefsels beïnvloed door zwaarlijvigheidbetrekking hebbende ontsteking

Hier beschrijven we een microscopie-aanpak van lichte lakens om het cardiale CD45 + leukocyten infiltraat te visualiseren in een muizenmodel van aseptische fulminante myocarditis, die wordt geïnduceerd door de intratracheale difterie toxine behandeling van CD11c.DTR muizen.

Kwantitatieve visualisatie van leukocytinfectiet in een muizenmodel van Fulminant-myocarditis door middel van lichte microscopie

Dit protocol biedt een rendabele aanpak om te isoleren en karakteriseren van muis primaire renale tubulaire cellen die vervolgens sub kunnen worden gekweekt om te beoordelen renale biologische functies- ex vivo, met inbegrip van de mitochondriale Bioenergetica.

Isolatie, karakterisering en High Throughput extracellulaire Flux-analyse van muis primaire renale tubulaire epitheliale cellen

Hier presenteren we een protocol voor te bereiden apoptotic thymocyten en buikvlies macrofagen en analyseren van de efficiëntie van efferocytosis en de specifieke remmer-gemedieerde blokkering van apoptotic thymocyten overspoeling. Dit protocol heeft een brede toepassing in cel-gemedieerde ontruiming van andere deeltjes met inbegrip van kunstmatige kralen en bacteriën.

Experimentele analyse van apoptotic Thymocyte overspoeling door macrofagen

Hier presenteren we een mechanisch dissociatie protocol om macrofagen snel te isoleren van de dorsale wortel ganglion voor fenotypering en functionele analyse.

Snelle isolatie van dorsale wortel ganglion macrofagen

Gezien de beperkte xenotransplantaatmodellen die beschikbaar zijn om interacties tussen menselijke CART- en myeloïde cellen te bestuderen, hebben we in vitro en in vivo modellen opgesteld om de effecten van menselijke macrofagen op CART-cellen te begrijpen. Bevindingen kunnen mogelijk worden gegeneraliseerd om de rol van macrofagen in de micro-omgeving van de tumor te evalueren en macrofaaggerichte immunotherapieën te testen.

Onderzoek naar interacties tussen myeloïde cellen en CAR T-cellen in vitro en in vivo

Met een ex vivo explant-model worden kleine stukjes van het rostrale deel van visceraal wit vetweefsel van magere muizen 7–10 dagen gekweekt, waardoor ontsteking in vetweefsel tijdens vetrijke voeding kan worden geïnduceerd en bestudeerd.

Een ex vivo muismodel voor vetrijke vetachtige vetweefselontsteking

Ontdek hoe niermacrofagen snel uitgeput en geregenereerd worden met behulp van het menselijke CD59/intermedilysin-ablatietool in muizen. Deze methode onthult chemokine-gemedieerde monocytrecrutering en herstel van immuunsurveillance na nierbeschadiging.

Snelle uitputting van niermacrofagen met behulp van menselijke CD59/Intermedilysin Cell Ablation Tool

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