Name: enrichment for chemoresistant ovarian cancer stem cells from human cell lines
Uploaded: 2014-09-10T15:00:00
Description: Watch this Scientific Journal Video about Enrichment for Chemoresistant Ovarian Cancer Stem Cells from Human Cell Lines at JoVE.com

Serene Samyesudhas and Dr. Lynn Roy assisted in preparing samples for filming.

1. خلية الثقافة والفلورسنت وصفها من سرطان المبيض خطوط الخلايا <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> إعداد SKOV3 وسائل الإعلام: وسائل الإعلام مكوي تستكمل مع 10٪ مصل بقري جنيني (FBS)، 0.1 مم L-الجلوتامين، 50 U / البنسلين مل، و 50 ميكروغرام / مل الستربتومايسين. الحفاظ عل?...

<ol>
	<li>Pennington, K., Pulaski, H., Pennington, M., Liu, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Too+Much+of+a+Good+Thing:+Suicide+Prevention+Promotes+Chemoresistance+in+Ovarian+Carcinoma.">Too Much of a Good Thing: Suicide Prevention Promotes Chemoresistance in Ovarian Carcinoma.</a> Curr Cancer Drug Targets. 10, 575-583 (2010).</li><li>Thigpen, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+rational+approach+to+the+management+of+recurrent+or+persistent+ovarian+carcinoma.">A rational approach to the management of recurrent or persistent ovarian carcinoma.</a> Clin Obstet Gynecol. 55, 114-130 (2012).</li><li>Burgos-Ojeda, D., Rueda, B. R., Buckanovich, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ovarian+cancer+stem+cell+markers:+prognostic+and+therapeutic+implications.">Ovarian cancer stem cell markers: prognostic and therapeutic implications.</a> Cancer letters. 322, 1-7 (2012).</li><li>Ma, L., Lai, D., Liu, T., Cheng, W., Guo, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer+stem-like+cells+can+be+isolated+with+drug+selection+in+human+ovarian+cancer+cell+line+SKOV3.">Cancer stem-like cells can be isolated with drug selection in human ovarian cancer cell line SKOV3.</a> Acta Biochim Biophys Sin (Shanghai). 42, 593-602 (2010).</li><li>Bonnet, D., Dick, J. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+acute+myeloid+leukemia+is+organized+as+a+hierarchy+that+originates+from+a+primitive+hematopoietic+cell.">Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell.</a> Nature Medicine. 3, 730-737 (1997).</li><li>Alison, M. R., Lim, S. M., Nicholson, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer+stem+cells:+problems+for+therapy.">Cancer stem cells: problems for therapy.</a> The Journal of pathology. 223, 147-161 (2011).</li><li>Luo, X., Dong, Z., Chen, Y., Yang, L., Lai, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enrichment+of+ovarian+cancer+stem-like+cells+is+associated+with+epithelial+to+mesenchymal+transition+through+an+miRNA-activated+AKT+pathway.">Enrichment of ovarian cancer stem-like cells is associated with epithelial to mesenchymal transition through an miRNA-activated AKT pathway.</a> Cell proliferation. 46, 436-446 (2013).</li><li>Wang, L., Mezencev, R., Bowen, N. J., Matyunina, L. V., McDonald, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+characterization+of+stem-like+cells+from+a+human+ovarian+cancer+cell+line.">Isolation and characterization of stem-like cells from a human ovarian cancer cell line.</a> Molecular and cellular biochemistry. 363, 257-268 (2012).</li><li>Abubaker, K., 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=Short-term+single+treatment+of+chemotherapy+results+in+the+enrichment+of+ovarian+cancer+stem+cell-like+cells+leading+to+an+increased+tumor+burden.">Short-term single treatment of chemotherapy results in the enrichment of ovarian cancer stem cell-like cells leading to an increased tumor burden.</a> Molecular cancer. 12, 24 (2013).</li><li>Conic, I., Dimov, I., Tasic-Dimov, D., Djordjevic, B., Stefanovic, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ovarian+epithelial+cancer+stem+cells.">Ovarian epithelial cancer stem cells.</a> ScientificWorldJournal. 11, 1243-1269 (2011).</li><li>Liu, K. C., 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=Ovarian+cancer+stem-like+cells+show+induced+translineage-differentiation+capacity+and+are+suppressed+by+alkaline+phosphatase+inhibitor.">Ovarian cancer stem-like cells show induced translineage-differentiation capacity and are suppressed by alkaline phosphatase inhibitor.</a> Oncotarget. 4 (12), 2366-2382 (2013).</li><li>Liu, T., Cheng, W., Lai, D., Huang, Y., Guo, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+primary+ovarian+cancer+cells+in+different+culture+systems.">Characterization of primary ovarian cancer cells in different culture systems.</a> Oncology reports. 23, 1277-1284 (2010).</li></ol>

الخلايا الجذعية السرطانية التي تقاوم العلاج قد يكون مسؤولا عن الانتكاس بعد العلاج من الورم الرئيسي. توصيف الخلايا الجذعية السرطانية قد يؤدي إلى تحسن العلاجات لسرطان المبيض. المعلمات الحرجة في تأسيس الخلايا الجذعية السرطانية chemoresistant باستخدام بروتوكول أعلاه وتوقيت ...

Resistance to chemotherapy is a major impediment to the treatment and cure of cancer. Ovarian cancer is the 5th leading cause of cancer-related deaths among women in the United States (American Cancer Society Facts and Figures 2013). Patients initially respond well to chemotherapy, but most patients relapse1. After relapse patients are treated with a variety of additional chemotherapy agents with very little benefit2. General properties of CSCs include unlimited proliferative capabilities, retention of an undifferentiated state, resistance to drug treatment, efficient DNA repair, and ability to generate malignant tumor cells with different phenotypes3. CSCs frequently exhibit expression of stem cell genes such as Nanog, Oct4, Sox2, Nestin, CD133, and CD117. CSCs often express elevated levels of ABCG2, and ALDH genes that may contribute to drug efflux and metabolism3,4. 
The first definitive evidence for CSCs was demonstrated by isolating acute myeloid leukemia-initiating cells that were capable of self-renewal and tumor generation5. These leukemic stem cells expressed surface CD34 and generated leukemia in NOD/SCID (immunocompromised) mice5. Since then CSCs have been identified in many cancer types including leukemias/lymphomas, breast, bladder, colorectal, endometrial, sarcomas, hepatocellular carcinoma, melanomas, gliomas, ovarian, pancreatic, prostate, squamous cell carcinoma, and lung6. Therefore, being able to study this subtype of cancer cell is advantageous.
The goal of this study is to create a protocol for the selection and isolation of chemoresistant CSCs. Several methods have been reported for the isolation of CSCs from ovarian cancer cell lines. Non-adherent spheroids isolated from OVCAR-3, SKOV3, or HO8910 cultures demonstrate stem-like properties7,8. Isolation of CD133+ cells from OVCAR-3 cultures also yields CSCs. CSCs have also been selected in culture by treatment with chemotherapeutic agents. Treating tumor cell lines (OVCA433, Hey, and SKOV3 cells) with cisplatin and paclitaxel allows for the expansion and isolation of CSCs4,9. While culture of some cell types in CSC media leads to isolation of CSCs, SKOV3 cells did not survive culture in serum-free media or form sphere cells4. Therefore, treatment of cells with cisplatin and paclitaxel aided the expansion or isolation of this population4.
Using a modification of the procedure presented by Ma and colleagues4 we developed a method to isolate CSCs from the ovarian cancer cell lines. Our protocol is advantageous as it yields more viable cells while using less toxic chemotherapeutic agents. Cells are treated with cisplatin and subsequently grown in serum-free media supplemented with growth factors (stem cell media). We isolate the resulting non-adherent sphere cells and assay them for their expression of stem cell markers. This model enables the study of CSC properties and response to drug therapy.

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enrichment for chemoresistant ovarian cancer stem cells from human cell lines

وتعرف الخلايا الجذعية السرطانية (الخلايا الجذعية السرطانية)، ومجموعة فرعية من الدراجات بطيئة والخلايا غير المتمايزة التي تقسم غير متماثلة لتوليد التكاثري للغاية، الغازية، والخلايا السرطانية chemoresistant. ولذلك، الخلايا الجذعية السرطانية هي الخلايا السكان جذابة لاستهداف علاجيا. ومن المتوقع الخلايا الجذعية السرطانية إلى المساهمة في عدد من أنواع الأورام الخبيثة بما في ذلك تلك الموجودة في الدم والدماغ والرئة والجهاز الهضمي والبروستاتا والمبيض. عزل وإثراء ويبلغ عدد سكانها الخلايا السرطانية عن الخلايا الجذعية السرطانية تمكين الباحثين من دراسة خصائص وعلم الوراثة، والاستجابة العلاجية الخلايا الجذعية السرطانية. ولدت لدينا البروتوكول الذي يثري بتكاثر الخلايا الجذعية السرطانية لسرطان المبيض المبيض من خطوط الخلايا السرطانية (SKOV3 وOVCA429). يتم التعامل مع خطوط الخلايا مع 20 ميكرومتر سيسبلاتين لمدة 3 أيام. الخلايا على قيد الحياة هي معزولة ومثقف في وسائل الاعلام الخلايا الجذعية خالية من مصل يحتوي على السيتوكينات وعوامل النمو. علينا أن نبرهن على إثراء هذه الخلايا الجذعية السرطانية أكثر نظافة من خلال تحليل خلايا منعزلة لكnown وقف علامات خلية Oct4، NANOG، وProm1 (CD133) والتعبير سطح الخلية من CD177 وCD133. زاد المعرض الخلايا الجذعية السرطانية مقاومة كيميائية. هذه الطريقة لعزل الخلايا الجذعية السرطانية هي أداة مفيدة لدراسة دور الخلايا الجذعية السرطانية في الورم مقاومة كيميائية والانتكاس.

لإثبات أننا عزل الخلايا الجذعية السرطانية من المبيض خطوط الخلايا السرطانية الظهارية باستخدام العلاج سيسبلاتين، حصلنا على أول صور للخطوط الخلايا قبل العلاج وبعد الاختيار. استخدمنا المجهر الضوئي لالتقاط الصور من ملتصقة (غير المعالجة) SKOV3 وOVCA429 الخلايا وSKOV3 وOVCA429 الخل?...

Watch this Scientific Journal Video about Enrichment for Chemoresistant Ovarian Cancer Stem Cells from Human Cell Lines at JoVE.com

Enrichment for Chemoresistant Ovarian Cancer Stem Cells from Human Cell Lines

وتورط خلايا الجذعية السرطانية (الخلايا الجذعية السرطانية) في ورم الانتكاس بسبب مقاومة كيميائية. لقد الأمثل بروتوكول للاختيار والتوسع في الخلايا الجذعية السرطانية من خطوط الخلايا السرطانية في المبيض. عن طريق علاج الخلايا مع سيسبلاتين العلاج الكيميائي وزرع الخلايا الجذعية في تعزيز وسائل الإعلام أننا إثراء للثقافات CSC غير ملتصقة.

لتخصيب Chemoresistant سرطان المبيض الخلايا الجذعية من خطوط الخلايا البشرية

Cancer stem cells (CSCs) are defined as a subset of slow cycling and undifferentiated cells that divide asymmetrically to generate highly proliferative, ...

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Time-lapse Imaging of Primary Preneoplastic Mammary Epithelial Cells Derived from Genetically Engineered Mouse Models of Breast Cancer

الوقت الفاصل بين التصوير من الخلايا الثديية سابق للورم ظهاري الأولية المستمدة من نماذج الماوس المعدلة وراثيا لسرطان الثدي

Time-lapse  imaging can be used to compare behavior of cultured primary preneoplastic  mammary epithelial cells derived from different genetically ...

Patient Derived Cell Culture and Isolation of CD133+ Putative Cancer Stem Cells from Melanoma

Patient Derived Cell Culture and Isolation of CD133+ Putative Cancer Stem Cells from Melanoma

زراعة الخلايا المشتقة المريض وعزل CD133<sup&gt; +</sup&gt; خلايا الجذعية السرطانية المزعومين من الميلانوما

Despite improved treatments options for melanoma available today, patients with advanced malignant melanoma still have a poor prognosis for ...

Isolation of Cancer Stem Cells From Human Prostate Cancer Samples

عزل الخلايا الجذعية للسرطان من العينات البشرية سرطان البروستاتا

The cancer stem cell (CSC) model has been considerably revisited over the last two decades. During this time CSCs have been identified and directly ...

Method for Obtaining Primary Ovarian Cancer Cells From Solid Specimens

طريقة الحصول على خلايا سرطان المبيض الابتدائية من العينات الصلبة

Reliable tools for investigating ovarian cancer initiation and progression are urgently needed. While the use of ovarian cancer cell lines remains a ...

Evaluation of Stem Cell Properties in Human Ovarian Carcinoma Cells Using Multi and Single Cell-based Spheres Assays

تقييم خصائص الخلايا الجذعية في الإنسان خلايا سرطان المبيض عن طريق متعدد واحدة على أساس خلية المجالات فحوصات

Years of research indicates that ovarian cancers harbor a heterogeneous mixture of cells including a subpopulation of so-called &#8220;cancer stem ...

In vitro Enrichment of Ovarian Cancer Tumor-initiating Cells

In vitro Enrichment of Ovarian Cancer Tumor-initiating Cells

في التخصيب في المختبر من خلايا سرطان المبيض-الشروع ورم

Evidence suggests that small subpopulations of tumor cells maintain a unique self-renewing and differentiation capacity and may be responsible for tumor ...