<meta charset="utf8"></head><body>ヒトSATMVECs:単離と脂肪細胞クロストーク研究

<ol>
	<li>Bierhansl, L., 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=Central+role+of+metabolism+in+endothelial+cell+function+and+vascular+disease.">Central role of metabolism in endothelial cell function and vascular disease.</a> Physiology. 32 (2), 126-140 (2017).</li><li>Aird, W. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endothelial+cell+heterogeneity.">Endothelial cell heterogeneity.</a> Cold Spring Harb Perspect Med. 2 (1), a006429 (2012).</li><li>Rajendran, P., 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=The+vascular+endothelium+and+human+diseases.">The vascular endothelium and human diseases.</a> Int J Biol Sci. 9 (10), 1057-1069 (2013).</li><li>Brown, O. I., Bridge, K., Kearney, M. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Nicotinamide+adenine+dinucleotide+phosphate+oxidases+in+glucose+homeostasis+and+diabetes-related+endothelial+cell+dysfunction.">Nicotinamide adenine dinucleotide phosphate oxidases in glucose homeostasis and diabetes-related endothelial cell dysfunction.</a> Cells. 10 (9), 2315 (2021).</li><li>Luk, C., Haywood, N. J., Bridge, K. I., Kearney, M. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Paracrine+role+of+the+endothelium+in+metabolic+homeostasis+in+health+and+nutrient+excess.">Paracrine role of the endothelium in metabolic homeostasis in health and nutrient excess.</a> Front Cardiovasc Med. 26 (9), 882923 (2022).</li><li>. Click-iT EdU Imaging Kits Available from: <a target="_blank" href="https://www.thermofisher.com/document-connect/document-connect.html?url=https://assets.thermofisher.com/TFS-Assets%2FLSG%2Fmanuals%2Fmp10338.pdf">https://www.thermofisher.com/document-connect/document-connect.html?url=https://assets.thermofisher.com/TFS-Assets%2FLSG%2Fmanuals%2Fmp10338.pdf</a> (2024)</li><li>Bou&#239;s, 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=Endothelium+in+vitro:+A+review+of+human+vascular+endothelial+cell+lines+for+blood+vessel-related+research.">Endothelium in vitro: A review of human vascular endothelial cell lines for blood vessel-related research.</a> Angiogenesis. 4 (2), 91-102 (2001).</li><li>Ades, E. 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=HMEC-1:+establishment+of+an+immortalized+human+microvascular+endothelial+cell+line.">HMEC-1: establishment of an immortalized human microvascular endothelial cell line.</a> J Invest Dermatol. 99 (6), 683-690 (1992).</li><li>Chi, J. T., 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=Endothelial+cell+diversity+revealed+by+global+expression+profiling.">Endothelial cell diversity revealed by global expression profiling.</a> Proc Natl Acad Sci U S A. 100 (19), 10623-10628 (2003).</li><li>Brignole, 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=213+ESC+Guidelines+on+cardiac+pacing+and+cardiac+resynchronization+therapy.">213 ESC Guidelines on cardiac pacing and cardiac resynchronization therapy.</a> Eur Heart J. 34 (29), 2281-2329 (2013).</li><li>Greenspon, A. 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=Trends+in+permanent+pacemaker+implantation+in+the+United+States+from+1993+to+2009:+Increasing+complexity+of+patients+and+procedures.">Trends in permanent pacemaker implantation in the United States from 1993 to 2009: Increasing complexity of patients and procedures.</a> J Am Coll Cardiol. 60 (16), 1540-1545 (2012).</li><li>Mendis, S. A., Alwan, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Prioritized Research Agenda for Prevention and Control of Noncommunicable Diseases. , (2011).</li><li>Brown, O. I., 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=Relationship+among+diabetes,+obesity,+and+cardiovascular+disease+phenotypes:+A+UK+biobank+cohort+study.">Relationship among diabetes, obesity, and cardiovascular disease phenotypes: A UK biobank cohort study.</a> Diabetes Care. 46 (8), 1531-1540 (2023).</li></ol>

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>170L CO2 Incubator</td>
			<td>GS Biotech</td>
			<td>170G-014</td>
			<td></td>
		</tr>
		<tr>
			<td>2-NBDG (2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose)</td>
			<td>Invitrogen</td>
			<td>N13195</td>
			<td></td>
		</tr>
		<tr>
			<td>4% PARAFIX buffered histological fixative</td>
			<td>VWR Chemicals</td>
			<td>PRC/R/38/1</td>
			<td></td>
		</tr>
		<tr>
			<td>Akt (tAkt) Rabbit&#160; 1:1000</td>
			<td>Cell Signalling Technology</td>
			<td>9272</td>
			<td></td>
		</tr>
		<tr>
			<td>BD Venflon Pro Safety 14 g x 45 mm, Orange, 50/pk</td>
			<td>Medisave</td>
			<td>393230</td>
			<td></td>
		</tr>
		<tr>
			<td>Bovine Serum Albumin solution, 7.5%</td>
			<td>Merck</td>
			<td>A8412-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>CD144&#160;(VE-Cadherin) Antibody, anti-human, PE, REAfinity</td>
			<td>Miltenyi Biotec</td>
			<td>130-118-495</td>
			<td></td>
		</tr>
		<tr>
			<td>CD31 Antibody, anti-human, PerCP-Vio 700, REAfinity</td>
			<td>Miltenyi Biotec</td>
			<td>130-110-811</td>
			<td></td>
		</tr>
		<tr>
			<td>CD31 MicroBead Kit, human, 1 kit</td>
			<td>Miltenyi Biotec</td>
			<td>130-091-935</td>
			<td></td>
		</tr>
		<tr>
			<td>CD45 Antibody, anti-human, FITC, REAfinity</td>
			<td>Miltenyi Biotec</td>
			<td>130-110-769</td>
			<td></td>
		</tr>
		<tr>
			<td>Cell Extraction Buffer</td>
			<td>Invitrogen</td>
			<td>FNN0011</td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge 5804 R</td>
			<td>Eppendorf</td>
			<td>5805000060</td>
			<td></td>
		</tr>
		<tr>
			<td>Click-iT EdU Cell Proliferation Kit for Imaging, Alexa Fluor 488 dye</td>
			<td>Invitrogen</td>
			<td>C10337</td>
			<td>Cell proliferation imaging kit</td>
		</tr>
		<tr>
			<td>Collagenase/Dispase, 500 mg</td>
			<td>Roche/Merck</td>
			<td>11097113001</td>
			<td></td>
		</tr>
		<tr>
			<td>Corning Matrigel Growth Factor Reduced (GFR) Basement Membrane Matrix</td>
			<td>Corning&#160;</td>
			<td>354230</td>
			<td>Basement Membrane Matrix</td>
		</tr>
		<tr>
			<td>Corning 100 mm TC-treated Culture Dish</td>
			<td>Corning&#160;</td>
			<td>430167</td>
			<td></td>
		</tr>
		<tr>
			<td>Costar 24-well Clear TC-treated Multiple Well Plates, Individually Wrapped, Sterile</td>
			<td>Corning&#160;</td>
			<td>3526</td>
			<td></td>
		</tr>
		<tr>
			<td>Costar 6-well Clear TC-treated Multiple Well Plates, Individually Wrapped, Sterile</td>
			<td>Corning&#160;</td>
			<td>3516</td>
			<td></td>
		</tr>
		<tr>
			<td>CytoFLEX S &#8211; 4 laser flow Cytometer</td>
			<td>Beckman</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Dead Cell Removal Kit</td>
			<td>Miltenyi Biotec</td>
			<td>130-090-101</td>
			<td></td>
		</tr>
		<tr>
			<td>Dulbecco&#8242;s Phosphate Buffered Saline</td>
			<td>Merck</td>
			<td>D8537-500ML</td>
			<td></td>
		</tr>
		<tr>
			<td>EASYstrainer Cell sieve for 50 mL tubes, 70 &#181;m mesh, Blue, sterile, 50/pk</td>
			<td>Greiner Bio-One</td>
			<td>542070</td>
			<td></td>
		</tr>
		<tr>
			<td>Endothelial Cell Growth Medium MV</td>
			<td>PromoCell</td>
			<td>C-22020</td>
			<td></td>
		</tr>
		<tr>
			<td>Eppendorf Safe-Lock micro test tubes</td>
			<td>Merck</td>
			<td>EP0030120094</td>
			<td></td>
		</tr>
		<tr>
			<td>Ethylenediaminetetraacetic acid solution</td>
			<td>Merck</td>
			<td>E8008-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Falcon 24 Well TC-Treated Cell Polystyrene Permeable Support Companion Plate, with Lid, Sterile, 1/Pack, 50/Case</td>
			<td>Appleton Woods</td>
			<td>CF537</td>
			<td></td>
		</tr>
		<tr>
			<td>Falcon Permeable Support for 24 Well Plate with 0.4 &#181;m Transparent PET Membrane, Sterile, 1/Pack, 48/Case</td>
			<td>Appleton Woods</td>
			<td>CF521</td>
			<td></td>
		</tr>
		<tr>
			<td>Freezing Medium Cryo-SFM</td>
			<td>PromoCell</td>
			<td>C-29912</td>
			<td></td>
		</tr>
		<tr>
			<td>Gelatin solution, 2% in water</td>
			<td>Merck</td>
			<td>G1393-100ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Gibco Antibiotic-Antimycotic (100x), 100mL</td>
			<td>Fisher Scientific</td>
			<td>11570486</td>
			<td></td>
		</tr>
		<tr>
			<td>Gibco TrypLE Select Enzyme (1x), no phenol red</td>
			<td>Fisher Scientific</td>
			<td>12563029</td>
			<td></td>
		</tr>
		<tr>
			<td>Hanks&#8242; Balanced Salt solution</td>
			<td>Merck</td>
			<td>H6648-500ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Human Subcutaneous Preadipocyte Cells</td>
			<td>Lonza</td>
			<td>PT-5020</td>
			<td></td>
		</tr>
		<tr>
			<td>Incucyte ZOOM</td>
			<td>Essen BioScience</td>
			<td></td>
			<td>Live-cell analysis system</td>
		</tr>
		<tr>
			<td>Insulin solution human</td>
			<td>Merck</td>
			<td>I9278-5ML</td>
			<td></td>
		</tr>
		<tr>
			<td>LS Columns, 25/pk</td>
			<td>Miltenyi Biotec</td>
			<td>130-042-401</td>
			<td></td>
		</tr>
		<tr>
			<td>MACS MultiStand</td>
			<td>Miltenyi Biotec</td>
			<td>130-042-303</td>
			<td></td>
		</tr>
		<tr>
			<td>MACS Tissue Storage Solution</td>
			<td>Miltenyi Biotec</td>
			<td>130-100-008</td>
			<td></td>
		</tr>
		<tr>
			<td>MACSmix Tube Rotator</td>
			<td>Miltenyi Biotec</td>
			<td>130-090-753</td>
			<td>Tube Rotator</td>
		</tr>
		<tr>
			<td>MS Columns, 25/pk</td>
			<td>Miltenyi Biotec</td>
			<td>130-042-201</td>
			<td></td>
		</tr>
		<tr>
			<td>NuPAGE 4&#8211;12% Bis-Tris Gel</td>
			<td>Invitrogen</td>
			<td>NP0322BOX</td>
			<td></td>
		</tr>
		<tr>
			<td>OctoMACS Separator</td>
			<td>Miltenyi Biotec</td>
			<td>130-042-109</td>
			<td></td>
		</tr>
		<tr>
			<td>PGM-2 Preadipocyte Growth Medium-2 BulletKit</td>
			<td>Lonza</td>
			<td>PT-8002</td>
			<td></td>
		</tr>
		<tr>
			<td>Phospho (Ser1177) eNOS Rabbit 1:1000</td>
			<td>Cell Signalling Technology</td>
			<td>9570</td>
			<td></td>
		</tr>
		<tr>
			<td>Phospho-Akt (Ser473) Rabbit&#160; 1:1000</td>
			<td>Cell Signalling Technology</td>
			<td>4060</td>
			<td></td>
		</tr>
		<tr>
			<td>Pre-Separation Filters 30 &#181;m, 50/pk</td>
			<td>Miltenyi Biotec</td>
			<td>130-041-407</td>
			<td></td>
		</tr>
		<tr>
			<td>Propidium Iodide (PI)/RNase Staining Solution</td>
			<td>Cell Signalling Technology</td>
			<td>4087</td>
			<td></td>
		</tr>
		<tr>
			<td>QuadroMACS Separator</td>
			<td>Miltenyi Biotec</td>
			<td>130-090-976</td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel Disposable Sterile Style 10</td>
			<td>VWRI</td>
			<td>501</td>
			<td></td>
		</tr>
		<tr>
			<td>Screw cap tube, 15 ml, (Lx&#216;): 120 x 17 mm, PP, with print</td>
			<td>Sarstetd</td>
			<td>62.554.502</td>
			<td></td>
		</tr>
		<tr>
			<td>Screw cap tube, 50 ml, (Lx&#216;): 114 x 28 mm, PP, with print</td>
			<td>Sarstedt</td>
			<td>62.547.254</td>
			<td></td>
		</tr>
		<tr>
			<td>Screw cap tube, 50 ml, (Lx&#216;): 114 x 28 mm, PP, with print</td>
			<td>Sarstetd</td>
			<td>62.547.254</td>
			<td></td>
		</tr>
		<tr>
			<td>Total eNOS Mouse 1:1000</td>
			<td>BD Biosciences</td>
			<td>610297</td>
			<td></td>
		</tr>
		<tr>
			<td>Triton X-100, BioUltra, for molecular biology</td>
			<td>Merck</td>
			<td>93443-500ML</td>
			<td></td>
		</tr>
		<tr>
			<td>&#946;-Actin (C4) Mouse 1:5000</td>
			<td>Santa Cruz Biotechnology</td>
			<td>&#160;Sc-47778</td>
			<td></td>
		</tr>
	</tbody>
</table>

studying adipose endothelial cell/adipocyte cross-talk in human subcutaneous adipose tissue

Endothelial cells (ECs) are squamous cells that line the inner surface of the blood vessel wall as a monolayer. They have many essential roles, including control of vascular tone, regulation of thrombosis, modulating the inflammatory response, and contributing to angiogenesis1. Given the importance of endothelial cells in cardiometabolic physiology, they are frequently used experimentally to further the understanding of pathophysiology and to examine new pharmacological treatments for cardiometabolic disease.
However, there is enormous heterogeneity in endothelial cell morphology, function, gene expression, and antigen composition depending on the origin of their vascular bed2. While endothelial cells from large arteries are best suited to atherosclerosis studies, endothelial cells from small vessels, known as microvascular endothelial cells (MVECs), are more suitable for angiogenesis studies2. Understanding the molecular basis for endothelial heterogeneity may provide valuable insights into vascular bed-specific therapies. Microvascular endothelial function also significantly differs in numerous diseases, including diabetes, cardiovascular disease, and systemic infection3,4. Therefore, the ability to isolate endothelial cells from defined patient groups allows direct comparison of their endothelial cell function and cellular cross-talk5. 
In this paper, we describe a novel method of isolating human MVECs from subcutaneous adipose tissue (hSATMVEC) taken at the time of cardiac implantable electronic device (CIED) insertion. hSATMVEC isolated following enzymatic digestion of subcutaneous adipose tissue (SAT) were grown and passaged on gelatin-coated plates. We then describe a range of phenotyping assays that have been successfully applied to hSATMVECs in order to validate their phenotype and demonstrate use in routine endothelial cell assays. Finally, we describe an application of hSATMVECs in an experimental model of hSATMVECs-adipocyte cross-talk.

<meta charset="utf8"></head><body>著者スポットライト:皮下脂肪組織微小血管系研究による心血管代謝疾患への洞察

ヒト皮下脂肪組織における脂肪内皮細胞/脂肪細胞クロストークの研究

We want to understand the role of the subcutaneous adipose tissue microvasculature in people with and without cardiometabolic disease. To do this, we've been isolating subcutaneous adipose tissue microvascular endothelial cells from patients undergoing cardiac implantable electronic devices. This is to study the endothelial cell phenotype and cell-cell crosstalk with adipocytes.Historically, we have used immortalized human cell lines or animal models to study cardiometabolic disease in vitro. However, these are not representative of diseases in human subjects. Our technique overcomes this by allowing us to understand the cellular basis of cardiometabolic disease using tissue from real-world patients.Our group is currently investigating the mechanism of dysregulated subcutaneous adipose tissue endothelial cell adipocyte crosstalk in people with diabetes, in particular, the impact of diabetic endothelium on adipocyte function and how this can be targeted to improve cardiometabolic health.

Read this Scientific Article Protocol about Studying Adipose Endothelial Cell/Adipocyte Cross-Talk in Human Subcutaneous Adipose Tissue at JoVE.com

Microvascular endothelial cells (MVECs) have many critical roles, including control of vascular tone, regulation of thrombosis, and angiogenesis. ...

studying-adipose-endothelial-celladipocyte-cross-talk-human

Research

JoVE Journal

Medicine

Studying Adipose Endothelial Cell/Adipocyte Cross-Talk in Human Subcutaneous Adipose Tissue

477 ビュー.  University of Leeds. 私たちは、心血管代謝疾患のある人とない人における皮下脂肪組織微小血管系の役割を理解したいと考えています。そのために、私たちは心臓埋め込み型電子機器を装着している患者さんから皮下脂肪組織微小血管内皮細胞を単離しています。これは、内皮細胞の表現型と脂肪細胞との細胞間クロストークを研究するためです。歴史的に、...