Name: a cre-lox p recombination approach for the detection of cell fusion in vivo
Uploaded: 2012-01-04T12:00:00
Description: Watch this Scientific Journal Video about A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo at JoVE.com

लेखकों के लिए उदारता से H1, MSCs और डॉ. टिम हैकर, डा. Gouqing गीत और विस्कॉन्सिन कार्डियोवास्कुलर फिजियोलॉजी के विश्वविद्यालय के सुश्री जिल कोच उपलब्ध कराने के लिए डा. Peiman Hemmati (मेडिसिन विभाग, मैडिसन विस्कॉन्सिन विश्वविद्यालय) का धन्यवाद करना चाहते हैं माउस सर्जरी प्रदर्शन करने के लिए कोर सुविधा. इस काम ब्रायन फ्रीमैन और NIH R21 HL089679 एक स्नातकोत्तर अनुसंधान फैलोशिप के माध्यम से राष्ट्रीय विज्ञान फाउंडेशन द्वारा समर्थित किया गया.

1. दाता सेल अभिकर्मक <ol><li> हार्वेस्ट mesenchymal स्टेम सेल (, MSCs, H1 भ्रूण स्टेम कोशिकाओं कृपया डॉ. Peiman Hematti द्वारा दान से प्राप्त वैकल्पिक रूप से, किसी भी vivo में फ्यूज धारणा प्रजातियों में से किसी भी सेल प्रकार नि?...

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J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+bioluminescence+imaging+of+cord+blood+derived+mesenchymal+stem+cell+transplantation+into+rat+myocardium.">In vivo bioluminescence imaging of cord blood derived mesenchymal stem cell transplantation into rat myocardium.</a> Ann. Nucl. Med. 20, 165-170 (2006).</li><li>Malstrom, S. E., Tornavaca, O., Meseguer, A., Purchio, A. F., West, D. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+characterization+and+hormonal+regulation+of+kidney+androgen-regulated+protein+(Kap)-luciferase+transgenic+mice.">The characterization and hormonal regulation of kidney androgen-regulated protein (Kap)-luciferase transgenic mice.</a> Toxicol. Sci. 79, 266-277 (2004).</li><li>Weir, L. 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विधि यहाँ की अनुमति देता है वर्णित है, पहली बार के लिए, छोटे जानवरों सहित जीवों में असतत पहचान और सेल संलयन अस्थायी विश्लेषण,. बाद biophotonic छवि विश्लेषण के साथ दृष्टिकोण Cre - LoxP पुनर्संयोजन को जोड़ती है. दृष्टि?...

<table border="1"><tbody><tr><td> अभिकर्मक के नाम </td><td> कंपनी </td><td> सूचीपत्र संख्या </td><td> टिप्पणियाँ </td></tr><tr><td> नियॉन अभिकर्मक सिस्टम </td><td> Invitrogen, Carlsbad, CA </td><td> MPK5000 </td><td></td></tr><tr><td> नियॉन 100 μL किट </td><td> Invitrogen, Carlsbad, CA </td><td> MPK10025 </td><td> अनुसंधान और ई बफर शामिल </td></tr><tr><td> एक सदस्य पाउडर </td><td> Invitrogen, Carlsbad, CA </td><td> 12000-022 </td><td></td></tr><tr><td> भ्रूण गोजातीय सीरम (FBS) </td><td> Hyclone, लोगान केन्द्र शासित प्रदेशों </td><td> SH30070.03 </td><td></td></tr><tr><td> B6.C - टीजी (सीएमवी - रचनात्मक) 1Cgn / जम्मू </td><td> जैक्सन प्रयोगशाला, बार हार्बर ME, </td><td> 006054 </td><td></td></tr><tr><td> 10X trypsin </td><td> फिशर साइंटिफिक, वन लॉन, NJ </tघ&gt; <td> मीट्रिक टन-25-054-सीएल </td><td></td></tr><tr><td> एल-Glutamine </td><td> फिशर साइंटिफिक, वन लॉन, NJ </td><td> 25030-081 </td><td></td></tr><tr><td> D-Luciferin </td><td> कैलिपर लाइफ साइंसेज, Hopkinton, एमए </td><td> 122796 </td><td></td></tr><tr><td> Xenogen Biophotonic इमेजिंग सिस्टम </td><td> कैलिपर लाइफ साइंसेज, Hopkinton, एमए </td><td> IVIS स्पेक्ट्रम </td><td></td></tr><tr><td> सोडियम Biocarbonate </td><td> सिग्मा Aldrich, सेंट लुइस, MO </td><td> S6014-500G </td><td></td></tr><tr><td> गैर आवश्यक एमिनो एसिड </td><td> Invitrogen, Carlsbad, CA </td><td> 11140-050 </td><td></td></tr></tbody></table>

a cre-lox p recombination approach for the detection of cell fusion in vivo

The ability of two or more cells of the same type to fuse has been utilized in metazoans throughout evolution to form many complex organs, including skeletal muscle, bone and placenta. Contemporary studies demonstrate fusion of cells of the same type confers enhanced function. For example, when the trophoblast cells of the placenta fuse to form the syncytiotrophoblast, the syncytiotrophoblast is better able to transport nutrients and hormones across the maternal-fetal barrier than unfused trophoblasts1-4. More recent studies demonstrate fusion of cells of different types can direct cell fate. The "reversion" or modification of cell fate by fusion was once thought to be limited to cell culture systems. But the advent of stem cell transplantation led to the discovery by us and others that stem cells can fuse with somatic cells in vivo and that fusion facilitates stem cell differentiation5-7. Thus, cell fusion is a regulated process capable of promoting cell survival and differentiation and thus could be of central importance for development, repair of tissues and even the pathogenesis of disease.
Limiting the study of cell fusion, is lack of appropriate technology to 1) accurately identify fusion products and to 2) track fusion products over time. Here we present a novel approach to address both limitations via induction of bioluminescence upon fusion (Figure 1); bioluminescence can be detected with high sensitivity in vivo8-15. We utilize a construct encoding the firefly luciferase (Photinus pyralis) gene placed adjacent to a stop codon flanked by LoxP sequences. When cells expressing this gene fuse with cells expressing the Cre recombinase protein, the LoxP sites are cleaved and the stop signal is excised allowing transcription of luciferase. Because the signal is inducible, the incidence of false-positive signals is very low. Unlike existing methods which utilize the Cre/LoxP system16, 17, we have incorporated a "living" detection signal and thereby afford for the first time the opportunity to track the kinetics of cell fusion in vivo.
To demonstrate the approach, mice ubiquitously expressing Cre recombinase served as recipients of stem cells transfected with a construct to express luciferase downstream of a floxed stop codon. Stem cells were transplanted via intramyocardial injection and after transplantation intravital image analysis was conducted to track the presence of fusion products in the heart and surrounding tissues over time. This approach could be adapted to analyze cell fusion in any tissue type at any stage of development, disease or adult tissue repair.

Watch this Scientific Journal Video about A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo at JoVE.com

A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo

एक समय पर रहने वाले जीवों में सेल संलयन ट्रैक विधि वर्णित है. दृष्टिकोण Cre - का इस्तेमाल<em&gt; LoxP</em&gt; पुनर्संयोजन सेल संलयन पर luciferase अभिव्यक्ति उत्पन्न करने के लिए. luminescent उत्पन्न संकेत रहने वाले ~ हज़ार कोशिकाओं की परिधि के ऊतकों में पता लगाने की संवेदनशीलता के साथ biophotonic इमेजिंग सिस्टम का उपयोग जीवों में पाया जा सकता है.

सेल फ्यूजन की जांच के लिए एक Cre Lox पी पुनर्संयोजन दृष्टिकोण Vivo में</em

The ability of two or more cells of the same type to fuse has been utilized in metazoans throughout evolution to form many complex organs, including ...

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A Cre-Lox P Recombination Approach for the Detection of Cell Fusion In Vivo

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