Name: planarian immobilization, partial irradiation, and tissue transplantation
Uploaded: 2012-08-06T12:00:00
Description: Watch this Scientific Journal Video about Planarian Immobilization, Partial Irradiation, and Tissue Transplantation at JoVE.com

Авторы хотели бы поблагодарить Chiyoko Кобаяси и Kiyokazu Агата полезные советы по трансплантации планарий, а также прошлых и настоящих членов Санчес лаборатории за неоценимую обсуждения в ходе развития этих технологий. Эта работа была поддержана NIH обучения Грант (5T32 HD0791), чтобы КГК и NIH R37GM057260 в ASA. ASA является Медицинского института Говарда Хьюза следователь.

Примечание: этот протокол предполагает использование потенциально опасных материалов (свинец и chloretone). Приобретать, читать и следовать MSDS для всех потенциально опасных материалов. 1. Животное культуры, селекция и подготовка <ol><li> Для культуры животных и обраще...

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	<li>Morgan, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experimental+studies+of+the+regeneration+of+Planaria+maculata.">Experimental studies of the regeneration of Planaria maculata.</a> Arch. Entw. Mech. Org. 7, 364-397 .</li><li>Reddien, P. W., Sanchez Alvarado, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fundamentals+of+planarian+regeneration.">Fundamentals of planarian regeneration.</a> Annu Rev Cell Dev Biol. 20, 725-757 (2004).</li><li>Randolph, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+regeneration+of+the+tail+in+lumbriculus.">The regeneration of the tail in lumbriculus.</a> J. Morphol. 7, 317-344 .</li><li>Wagner, D. E., Wang, I. E., Reddien, P. 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A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Planarian+homologs+of+netrin+and+netrin+receptor+are+required+for+proper+regeneration+of+the+central+nervous+system+and+the+maintenance+of+nervous+system+architecture.">Planarian homologs of netrin and netrin receptor are required for proper regeneration of the central nervous system and the maintenance of nervous system architecture.</a> Development. , 132-3691 (2005).</li><li>Newmark, P. A., S&#225;nchez Alvarado, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bromodeoxyuridine+specifically+labels+the+regenerative+stem+cells+of+planarians.">Bromodeoxyuridine specifically labels the regenerative stem cells of planarians.</a> Dev. Biol. 220, 142-153 (2000).</li><li>Miller, W., Kennedy, R. 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Dyn. 238, 443-450 (2009).</li><li>Hayashi, T., Asami, M., Higuchi, S., Shibata, N., Agata, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+of+planarian+X-ray-sensitive+stem+cells+by+fluorescence-activated+cell+sorting.">Isolation of planarian X-ray-sensitive stem cells by fluorescence-activated cell sorting.</a> Dev. Growth Differ. 48, 371-380 (2006).</li><li>Reddien, P. W., Oviedo, N. J., Jennings, J. R., Jenkin, J. C., S&#225;nchez Alvarado, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SMEDWI-2+is+a+PIWI-like+protein+that+regulates+planarian+stem+cells.">SMEDWI-2 is a PIWI-like protein that regulates planarian stem cells.</a> Science. 310, 1327-1330 (2005).</li><li>St&#233;phan-Dubois, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Les+cellules+de+r&#233;g&#233;n&#233;ration+chez+la+planaire+Dendrocoleum+lacteum.">Les cellules de r&#233;g&#233;n&#233;ration chez la planaire Dendrocoleum lacteum.</a> Bulletin de la Société Zooologique de France. 86, 172-185 (1961).</li><li>Gurley, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Expression+of+secreted+Wnt+pathway+components+reveals+unexpected+complexity+of+the+planarian+amputation+response.">Expression of secreted Wnt pathway components reveals unexpected complexity of the planarian amputation response.</a> Dev. Biol. 347, 24-39 (2010).</li><li>Salvetti, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adult+stem+cell+plasticity:+neoblast+repopulation+in+non-lethally+irradiated+planarians.">Adult stem cell plasticity: neoblast repopulation in non-lethally irradiated planarians.</a> Dev. Biol. 328, 305-314 (2009).</li></ol>

Важность иммобилизации Иммобилизация на сегодняшний день является наиболее важным шагом для успешного завершения любой из этих процессов. Если планарии неправильно иммобилизованных до частичного облучения, они могут двигаться под свинцовый экран, производя непоследо?...

<table border="1"><tbody><tr><td> Название реагента </td><td> Компания </td><td> Номер по каталогу </td></tr><tr><td> General Purpose передачи пипетки </td><td> Samco </td><td> 691 </td></tr><tr><td> Капиллярные трубки (ID 0,75 мм) </td><td> FHC </td><td> 30-30-0 </td></tr><tr><td> Капиллярные трубы (диаметр 0,7 мм) </td><td> FHC </td><td> 30-50-08 </td></tr><tr><td> Парафильмом M </td><td> VWR </td><td> 52858-076 </td></tr><tr><td> Kimwipes 34155 </td><td> VWR </td><td> 500029-891 </td></tr><tr><td> Черный фильтровальная бумага </td><td> Шлейхер и Schuell </td><td> 10310809 </td></tr><tr><td> Ватман № 2 фильтровальной бумаги 1002-055 </td><td> Fisher Scientific </td><td> 09-810B </td></tr><tr><td> Ватман № 3 фильтровальной бумаги 1003-185 </td><td> Fisher Scientific </td><td> 09-820E </td></tr><tr><td> Сигареты прокатки бумаги </td><td> Zig-Zag, оригинальные </td><td> Не Доступно </td></tr><tr><td> Петрипосуда </td><td> VWR </td><td> 82050-544 </td></tr><tr><td> Пинцет Дюмон, INOX # 5 </td><td> FST </td><td> 11251-20 </td></tr><tr><td> Chloretone </td><td> Sigma Aldrich </td><td> 112054 </td></tr><tr><td> Казеин </td><td> Sigma Aldrich </td><td> C3400 </td></tr><tr><td> Ведущий Шилдс </td><td> Альфа-системы корпорации, Bluffdale, UT </td><td> Нестандартная конструкция </td></tr><tr><td> XRAD-320 биологических облучатель </td><td> Precision X-Ray, Северном Брэнфорд, CT </td><td> Не Доступно </td></tr></tbody></table>

planarian immobilization, partial irradiation, and tissue transplantation

The planarian, a freshwater flatworm, has proven to be a powerful system for dissecting metazoan regeneration and stem cell biology1,2. Planarian regeneration of any missing or damaged tissues is made possible by adult stem cells termed neoblasts3. Although these stem cells have been definitively shown to be pluripotent and singularly capable of reconstituting an entire animal4, the heterogeneity within the stem cell population and the dynamics of their cellular behaviors remain largely unresolved. Due to the large number and wide distribution of stem cells throughout the planarian body plan, advanced methods for manipulating subpopulations of stem cells for molecular and functional study in vivo are needed.
Tissue transplantation and partial irradiation are two methods by which a subpopulation of planarian stem cells can be isolated for further study. Each technique has distinct advantages. Tissue transplantation allows for the introduction of stem cells, into a na&iuml;ve host, that are either inherently genetically distinct or have been previously treated pharmacologically. Alternatively, partial irradiation allows for the isolation of stem cells within a host, juxtaposed to tissue devoid of stem cells, without the introduction of a wound or any breech in tissue integrity. Using these two methods, one can investigate the cell autonomous and non-autonomous factors that control stem cell functions, such as proliferation, differentiation, and migration.
Both tissue transplantation5,6 and partial irradiation7 have been used historically in defining many of the questions about planarian regeneration that remain under study today. However, these techniques have remained underused due to the laborious and inconsistent nature of previous methods. The protocols presented here represent a large step forward in decreasing the time and effort necessary to reproducibly generate large numbers of grafted or partially irradiated animals with efficacies approaching 100 percent. We cover the culture of large animals, immobilization, preparation for partial irradiation, tissue transplantation, and the optimization of animal recovery. Furthermore, the work described here demonstrates the first application of the partial irradiation method for use with the most widely studied planarian, Schmidtea mediterranea. Additionally, efficient tissue grafting in planaria opens the door for the functional testing of subpopulations of na&iuml;ve or treated stem cells in repopulation assays, which has long been the gold-standard method of assaying adult stem cell potential in mammals8. Broad adoption of these techniques will no doubt lead to a better understanding of the cellular behaviors of adult stem cells during tissue homeostasis and regeneration.

Watch this Scientific Journal Video about Planarian Immobilization, Partial Irradiation, and Tissue Transplantation at JoVE.com

Planarian Immobilization, Partial Irradiation, and Tissue Transplantation

Эффективный метод для пересадки ткани определенной и последовательной размер между планарии описано. Также имеется описание того, как иммобилизация метод, используемый для трансплантации могут быть адаптированы в сочетании с ведущим щиты, для частичного облучения животных.

Планарии иммобилизации, частичное облучение, и тканей

The planarian, a freshwater flatworm, has proven to be a powerful system for dissecting metazoan regeneration and stem cell biology1,2. Planarian ...

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