{"id":22563,"date":"2019-11-29T16:14:15","date_gmt":"2019-11-29T08:14:15","guid":{"rendered":"http:\/\/biopioneer.com.tw\/?p=22563"},"modified":"2025-03-21T14:28:03","modified_gmt":"2025-03-21T06:28:03","slug":"pulmonary-microvascular-endothelial-cells-pmec-%e8%b2%a8%e8%99%9ft0491","status":"publish","type":"post","link":"http:\/\/biopioneer.com.tw\/?p=22563","title":{"rendered":"\u80ba\u5fae\u8840\u7ba1\u5167\u76ae\u7d30\u80de | pulmonary microvascular endothelial cells (PMEC) \u8ca8\u865fT0491"},"content":{"rendered":"

A<\/strong>bmgood<\/strong>\u85e5\u7269\u7be9\u9078\u5e73\u53f0<\/strong> | <\/strong>\u7d30\u80de\u7cfb\u5217\u5546\u54c1<\/strong> (immortalized cell)<\/strong><\/p>\n

pulmonary microvascular endothelial cells (PMEC)\u00a0 | \u8ca8\u865fT0491\u00a0<\/a> <\/span>\u7d30\u80de\u6578\u91cf1×106<\/sup> cells \/ 1.0 ml<\/p>\n

\u7522\u54c1\u4ecb\u7d39: \u80ba\u5fae\u8840\u7ba1\u5167\u76ae\u7d30\u80de\uff08pulmonary microvascular endothelial cells , PMEC\uff09\u5206\u96e2\u81ea\u4eba\u80ba\u52d5\u8108 (pulmonary arteries)\u3002 PMEC\u5f62\u6210\u534a\u9078\u64c7\u5c4f\u969c(semiselective barrier)\uff0c\u9019\u5c0d\u65bc\u80ba\u6c23\u9ad4\u4ea4\u63db\u4ee5\u53ca\u8abf\u7bc0\u8840\u6db2\u8207\u80ba\u9593\u8cea\u8154\u5ba4\u4e4b\u9593\u7684\u6d41\u9ad4\u548c\u6eb6\u8cea\u901a\u904e\u81f3\u95dc\u91cd\u8981\u3002 HPAEC\u5df2\u7528\u65bc\u7814\u7a76\u8840\u7ba1\u901a\u900f\u6027(vascular permeability)\u548c\u708e\u75c7\u53cd\u61c9(inflammatory responses)\u3002 \u8207HPASMC\u5171\u57f9\u990a\u7684HPAEC\u5df2\u88ab\u7528\u4f5c\u80ba\u8840\u7ba1\u75be\u75c5\u7684\u6a21\u578b\u3002<\/p>\n

\"\"<\/p>\n

pulmonary microvascular endothelial cells (PMEC)<\/p>\n\n\n\n\n\n\n\n\n\n\n\n\n
Organism<\/strong><\/td>\nHomo sapiens<\/td>\n<\/tr>\n
Source Organ<\/strong><\/td>\nLung<\/td>\n<\/tr>\n
BioSafety Level<\/strong><\/td>\nII<\/td>\n<\/tr>\n
Growth Properties<\/strong><\/td>\nAdherent<\/td>\n<\/tr>\n
Morphology<\/strong><\/td>\nPolygonal<\/td>\n<\/tr>\n
Recommended Seeding Density<\/strong><\/td>\n50,000 cells\/cm2<\/sup><\/td>\n<\/tr>\n
Population Doubling<\/strong><\/td>\n25-35 hours<\/td>\n<\/tr>\n
Immortalization Method<\/strong><\/td>\nSerial passaging and transduction with recombinant lentiviruses carrying SV40 Large T antigen<\/td>\n<\/tr>\n
Description<\/strong><\/td>\nThe pulmonary microvascular endothelial cells (PMEC) are isolated from human pulmonary arteries. The PMECs form a semiselective barrier that is critical for lung gas exchange and regulation of fluid and solute passage between the blood and interstitial compartments in the lung. HPAEC have been used for the study of vascular permeability and inflammatory responses. HPAEC in co-culture with HPASMC have been used as a model for pulmonary angiopathy.<\/td>\n<\/tr>\n
Applications<\/strong><\/td>\nFor Research Use Only<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n

pulmonary microvascular endothelial cells (PMEC)\u00a0\u7d30\u80de\u682a | \u4e0d\u6747\u65b9\u6cd5 : Serial passaging and transduction with recombinant lentiviruses carrying SV40 Large T antigen | \u5668\u5b98\u4f86\u6e90Lung | \u7d30\u80de\u57f9\u990a\u689d\u4ef6: \u4f7f\u7528PriCoat\u2122T25\u57f9\u990a\u74f6 (\u8ca8\u865fG299<\/a><\/span>)\uff0c\u4ee5\u6700\u4f73\u5316\u521d\u4ee3\u548c\u4e0d\u673d\u7d30\u80de\u7684\u751f\u9577\u3001\u589e\u6b96\u548c\u64f4\u589e\u3002\u6216\u5857\u5c64\u7d30\u80de\u5916\u57fa\u8cea(Applied Cell Extracellular Matrix) \u589e\u5f37\u7d30\u80de\u8cbc\u9644 (\u8ca8\u865fG422<\/a><\/span>)\u3002Grow cells in ECM-coated culture vessels with the following conditions. \u57f9\u990a\u6db2PriGrow I (<\/u>\u8ca8\u865f<\/u>\u00a0TM001<\/u><\/a><\/u><\/span>)<\/u>\u3002\u70ba\u4e86\u88fd\u5099\u5b8c\u6574\u7684\u751f\u9577\u57f9\u990a\u57fa\uff0c\u5c07\u4e0b\u5217\u7d44\u5206\u6dfb\u52a0\u5230\u57fa\u790e\u57f9\u990a\u57fa\u4e2d\uff1a\u80ce\u725b\u8840\u6e05\uff08TM999<\/span>\uff09\u81f3\u6700\u7d42\u6fc3\u5ea610\uff05\u548cPenicillin\/Streptomycin\u9752\u9ef4\u7d20\/\u93c8\u9ef4\u7d20\u6eb6\u6db2\uff08G255\uff09\u81f3\u6700\u7d42\u6fc3\u5ea6\u70ba1\uff05\u3002\u4e8c\u6c27\u5316\u78b3\uff08CO2\uff09\uff1a5\uff05\uff0c\u6eab\u5ea6\uff1a37.0\u2103\u3002<\/p>\n

\u9ef4\u6f3f\u83ccPCR\u5075\u6e2c<\/a>\u00a0|<\/strong>\u00a0DAPI\u67d3\u6838<\/a>\u00a0|<\/strong>\u00a0\u7d30\u80de\u5167\u9ef4\u6f3f\u83cc\u53bb\u9664\u5291<\/a>\u00a0|<\/strong>\u00a0\u6c34\u6d74\u69fd\u6291\u83cc\u5291<\/a>\u00a0|<\/strong>\u00a0\u5674\u9727\u5f0f\u53bb\u9ef4\u5291 \u7b49\u7cfb\u5217\u5546\u54c1<\/a><\/span><\/p>\n

Abmgood<\/strong>\u4e0d\u673d\u5316\u7cfb\u5217\u5546\u54c1<\/strong>\u00a0(immortalized reagenet)<\/strong>\u00a0Ready-to-use<\/p>\n

Abmgood\u516c\u53f8\u65bc\u4eba\u985e\u8207\u8001\u9f20\u4e0d\u673d\u4e4b\u7d30\u80de\u682a\u4e0a\uff0c\u5177\u6709\u5e7e\u5341\u5e74\u7684\u5be6\u9a57\u7d93\u9a57\uff1b\u63d0\u4f9b\u4e00\u7cfb\u5217\u4e0d\u673d (immortalized)\u5316\u4e4b\u7d30\u80de\u682a\u3001\u53ca\u4e0d\u673d\u5316\u4e4b\u8a66\u5291\u5546\u54c1\u3002ready-to-use\u5b9a\u91cf\u597d\u4e4bretrovirus\u3001lentiviru\u3001 adenovirus\uff0c\u53ef\u76f4\u63a5\u52a0\u65bc\u7d30\u80de\u4e2d\u9032\u884c\u611f\u67d3\u8868\u73fe\uff1b\u75c5\u6bd2\u6db2\u5305\u62ec\u6709SV40\u91cd\u7d44\u75c5\u6bd2\u3001EBV\u91cd\u7d44\u75c5\u6bd2\u3001hTERT\u75c5\u6bd2\u6db2\u3001Myc\u57fa\u56e0\u75c5\u6bd2\u6db2\u3001p53\u57fa\u56e0\u75c5\u6bd2\u6db2\u3001Rb\u57fa\u56e0\u75c5\u6bd2\u6db2\u3001Ras\u57fa\u56e0\u75c5\u6bd2\u6db2\u3002\u63d0\u4f9b106\u53ca109\u5169\u7a2e\u5305\u88dd\u529b\u50f9\u4e4b\u75c5\u6bd2\u6db2\uff1b\u4ea6\u63d0\u4f9b\u4e00\u7cfb\u5217\u75c5\u6bd2\u529b\u50f9\u6fc3\u7e2e\u8207\u5b9a\u91cf\u8a66\u5291\u5546\u54c1\u3002<\/p>\n

\u7d30\u80de\u4e0d\u673d\u5316\u7684\u5de5\u5177\u4e2d\uff0cSV40 T antigen\u662f\u6700\u7c21\u55ae\u4e14\u6700\u5e38\u7528\u4f86\u9020\u6210\u4e0d\u673d\u5316\u7d30\u80de\u751f\u6210\u7684\u65b9\u6cd5\u3002\u591a\u6578\u7684\u75c5\u6bd2\u57fa\u56e0\u548c\u6291\u764c\u57fa\u56e0\uff08p53\u3001Rb\u7b49\uff09\u7684\u4e0d\u6d3b\u5316\u76f8\u95dc\uff0c\u6703\u8a98\u5c0e\u7d30\u80de\u8655\u65bcreplicative senescence\u7684\u968e\u6bb5\u3002\u76ee\u524d\u4e5f\u6709\u7814\u7a76\u986f\u793a\u88ab\u611f\u67d3\u7684\u7d30\u80de\uff0cSV40 T anbtigen\u6703\u8a98\u5c0e\u7aef\u7c92\u9176\uff08Telomerase\uff09\u7684\u6d3b\u6027\u3002\u8207\u7aef\u7c92\uff08telomere\uff09\u9577\u5ea6\u76f8\u95dc\u7684Telomerase Reverse Transcriptase protein (TERT)\u662f\u4e00\u7a2e\u4e0d\u673d\u7d30\u80de\u7684\u7814\u7a76\u61c9\u7528\u3002\u5e73\u5e38\u7d30\u80de\u4e2d\uff0chTERT\u662f\u8655\u65bc\u4e0d\u6d3b\u5316\u614b\uff0c\u5fc5\u8981\u6642\u6703\u9032\u884c\u6d3b\u5316\u4fdd\u6301telomere\u7684\u9577\u5ea6\uff0c\u907f\u514d\u7d30\u80de\u767c\u751freplicative senescence\u7684\u73fe\u8c61\u3002\u67d0\u4e9b\u7d30\u80dehTERT \u904e\u5ea6\u8868\u73fe\uff08over-expression\uff09\u5c0d\u65bc\u8a98\u767c\u7d30\u80de\u4e0d\u673d\u5316\u662f\u7121\u6548\u7684\uff08\u5982\u521d\u4ee3\u4e0a\u76ae\u7d30\u80de\uff09\uff0c\u53cd\u800c\u6703\u9020\u6210\u7d30\u80de\u6b7b\u4ea1\uff1b\u540c\u6642\u8868\u73fehTERT catalytic subunit\u4ee5\u53cap53\u6216RB siRNA\u5728\u4eba\u985e\u521d\u4ee3\u5375\u5de2\u4e0a\u76ae\u7d30\u80de\u6703\u9020\u6210\u7d30\u80de\u4e0d\u673d\u5316\u3002\u904e\u5ea6\u8868\u73feRas\u6216Myc T58A mutants\u4e5f\u6709\u521d\u4ee3\u7d30\u80de\u4e0d\u673d\u5316\u7684\u73fe\u8c61\u7522\u751f\u3002<\/p>\n

Decomplement<\/strong>\u52a0\u71b1\u975e\u6d3b\u5316<\/strong>FBS<\/strong>\uff1a<\/strong><\/p>\n

1.\u8b93FBS\u74f6\u57284\u00b0C\u4e0b\u5b8c\u5168\u89e3\u51cd\u904e\u591c\u3002<\/p>\n

2.\u5c07FBS\u74f6\u7f6e\u65bc56\u00b0C\u7684\u6c34\u6d74\u4e2d30\u5206\u9418\u3002<\/p>\n

Decomplemented FBS\u53ef\u4ee5\u5728-20\u00b0C\u4e0b\u9577\u671f\u4fdd\u5b58\u3002\u907f\u514d\u983b\u7e41\u7684\u51cd\u878d\u5faa\u74b0\u3002\u6bcf2-3\u5929\u66f4\u63db\u4e00\u6b21\u5b8c\u6574\u7684\u5a92\u9ad4\u3002\u4e0d\u8981\u8b93\u4ecb\u8cea\u984f\u8272\u8b8a\u70ba\u9ec3\u8272\u3002<\/p>\n

Thawing<\/strong>\u7d30\u80de\u89e3\u51cd\u65b9\u6cd5\uff1a<\/strong>(<\/strong>\u6bcf\u7a2e\u7d30\u80de\u5efa\u8b70\u689d\u4ef6\u53ef\u80fd\u6709\u8a31\u4e0d\u540c\uff0c\u4f9d\u7167\u539f\u5ee0\u5efa\u8b70\u64cd\u4f5c\uff0c\u4ee5\u4e0b\u70ba\u901a\u7528\u65b9\u5f0f<\/strong>)<\/strong><\/p>\n

1.\u572837\u2103\u6c34\u6d74\u4e2d\u89e3\u51cd\u5c0f\u74f6\uff0c\u76f4\u5230\u53ea\u6709\u4e00\u5c0f\u584a\u51b0\u584a\u3002<\/p>\n

2.\u5c07\u7d30\u80de\u8f49\u79fb\u5230\u542b\u67095ml pre-warmed complete media \u4e4b15ml\u7ba1\u4e2d\u3002<\/p>\n

3.\u5c07\u7d30\u80de\u4ee5290xg\u96e2\u5fc35\u5206\u9418\u3002<\/p>\n

4.\u5c0f\u5fc3\u4e1f\u68c4\u4e0a\u6e05\u6db2\uff0c\u4e0d\u8981\u6253\u64fe\u7d30\u80de\u6c89\u6fb1\uff0c\u8f15\u8f15\u5730\u4e0a\u4e0b\u79fb\u6db2\uff0c\u5c07\u7d30\u80de\u91cd\u65b0\u61f8\u6d6e\u57281ml\u5b8c\u5168\u57f9\u990a\u57fa\u4e2d\u3002<\/p>\n

5.\u5c07\u7d30\u80de\u57f9\u990a\u5728150,000-200,000\u500b\u7d30\u80de\/ cm2\u3002<\/p>\n

Subculture<\/strong>\u7d30\u80de\u7e7c\u4ee3\u65b9\u6cd5<\/strong>:<\/strong>\u00a0(<\/strong>\u6bcf\u7a2e\u7d30\u80de\u5efa\u8b70\u689d\u4ef6\u53ef\u80fd\u6709\u8a31\u4e0d\u540c\uff0c\u4f9d\u7167\u539f\u5ee0\u5efa\u8b70\u64cd\u4f5c\uff0c\u4ee5\u4e0b\u70ba\u901a\u7528\u65b9\u5f0f<\/strong>)<\/strong><\/p>\n

1.\u7576\u7d30\u80de70-90\uff05 confluency\u6642\uff0c\u5efa\u8b70\u7e7c\u4ee3\u57f9\u990a\u3002<\/p>\n

    \n
  1. Aspirate old media\u3002\u61f8\u6d6e\u6db2\u4e2d\u7684\u4e00\u4e9b\u7d30\u80de\u4ecd\u7136\u662f\u6d3b\u7d30\u80de\u3002\u6216\u8005\uff0c\u4e5f\u53ef\u4ee5\u6536\u96c6\u542b\u6709\u61f8\u6d6e\u7d30\u80de\u7684\u4e0a\u6e05\u6db2\u4e26\u96e2\u5fc3\uff08Skip to Step 6 in protocol\uff09\u3002<\/li>\n<\/ol>\n

    3.\u52a0\u51651\uff1a1\u6bd4\u4f8b\u76841X\u7121\u83ccPBS\u548c0.25\uff05\u80f0\u86cb\u767d\u9176-EDTA\uff08TM051\uff09\u3002<\/p>\n

    4.\u5728\u5ba4\u6eab\u4e0b\u5b75\u80b2\u7d30\u80de3-5\u5206\u9418\u4e26\u652a\u62cc\u57f9\u990a\u5bb9\u5668\u76f4\u81f390\uff05\u7684\u7d30\u80de\u812b\u843d\u3002<\/p>\n

      \n
    1. \u4e2d\u548c\u80f0\u86cb\u767d\u9176:\u6dfb\u52a0\u7b49\u91cf\u4e4btrypsin + PBS\u3002<\/li>\n<\/ol>\n

      6.\u6536\u96c6\u7d30\u80de\u4e26\u4ee5290xg\u96e2\u5fc35\u5206\u9418\u3002<\/p>\n

      7.\u4e1f\u68c4\u4e0a\u6e05\u6db2\uff0c\u8f15\u8f15\u5730\u4e0a\u4e0b\u79fb\u6db2\uff0c\u5c07\u7d30\u80de\u91cd\u65b0\u61f8\u6d6e\u5728\u5b8c\u5168\u57f9\u990a\u57fa\u4e2d\u3002<\/p>\n

      8.\u5efa\u8b70split ratio\u6bd4\u4e0d\u8d85\u904e1\uff1a3\u3002<\/p>\n

      Freezing<\/strong>\u7d30\u80de\u51cd\u7d50\uff1a<\/strong>(<\/strong>\u6bcf\u7a2e\u7d30\u80de\u5efa\u8b70\u689d\u4ef6\u53ef\u80fd\u6709\u8a31\u4e0d\u540c\uff0c\u4f9d\u7167\u539f\u5ee0\u5efa\u8b70\u64cd\u4f5c\uff0c\u4ee5\u4e0b\u70ba\u901a\u7528\u65b9\u5f0f<\/strong>)<\/strong><\/p>\n

      Complete growth medium with decomplemented FBS to a final concentration of 50% from PAN Biotech (P40-37500) or from Nucleus Biologics (FBS1824-001) and 5% DMSO.<\/p>\n

      \u5132\u5b58\u6eab\u5ea6\uff1a\u6db2\u6c2e\u6c23\u76f8\u3002\u91cd\u8981\u63d0\u793a\uff1a\u6b64\u7d30\u80de\u4fc2\u5c0dFBS\u6279\u6b21\u975e\u5e38\u654f\u611f\u3002\u6211\u5011\u5f37\u70c8\u5efa\u8b70\u6700\u7d42\u7528\u6236\u4f7f\u7528\u5f9ePAN biotech\u6216Nucleus Biologics \u4e4bFBS\u3002<\/p>\n

      \u6162\u75c5\u6bd2\u5305\u88dd\u7cfb\u5217\u7522\u54c1\u00a0Lentivirus<\/u>\u6162\u75c5\u6bd2\u4f4e\u901f\u7d14\u5316<\/u>(\u7d14\u5316)\u00a0<\/a>| Lentivirus <\/u>\u6162\u75c5\u6bd2\u9ad8\u6fc3\u5ea6\u7d14\u5316<\/u>(\u7d14\u5316)\u00a0<\/a>|\u00a0Lentivirus<\/u>\u75c5\u6bd2<\/u>RT-QPCR<\/u>\u5b9a\u91cf<\/u>(\u5b9a\u91cf)<\/a>\u00a0<\/span><\/p>\n","protected":false},"excerpt":{"rendered":"

      Abmgood\u85e5\u7269\u7be9\u9078\u5e73\u53f0 | \u7d30\u80de\u7cfb\u5217\u5546\u54c1 (immortalized cell) pulmonary mi […]<\/p>\n","protected":false},"author":3,"featured_media":11653,"comment_status":"closed","ping_status":"closed","sticky":false,"template":"","format":"standard","meta":[],"categories":[453],"tags":[1807],"_links":{"self":[{"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/posts\/22563"}],"collection":[{"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/posts"}],"about":[{"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/users\/3"}],"replies":[{"embeddable":true,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=%2Fwp%2Fv2%2Fcomments&post=22563"}],"version-history":[{"count":4,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/posts\/22563\/revisions"}],"predecessor-version":[{"id":50798,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/posts\/22563\/revisions\/50798"}],"wp:featuredmedia":[{"embeddable":true,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=\/wp\/v2\/media\/11653"}],"wp:attachment":[{"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=%2Fwp%2Fv2%2Fmedia&parent=22563"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=%2Fwp%2Fv2%2Fcategories&post=22563"},{"taxonomy":"post_tag","embeddable":true,"href":"http:\/\/biopioneer.com.tw\/index.php?rest_route=%2Fwp%2Fv2%2Ftags&post=22563"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}