Name: rotating cell culture systems for human cell culture: human trophoblast cells as a model
Uploaded: 2012-01-18T12:00:00
Description: Watch this Scientific Journal Video about Rotating Cell Culture Systems for Human Cell Culture: Human Trophoblast Cells as a Model at JoVE.com

وأيد هذا العمل من قبل المعاهد الوطنية الأميركية للصحة منح HD051998 # المعاهد الوطنية للصحة / معاهد الصحة القومية (لCAM).

1. الكولاجين الخرزة التحضير <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> قبل EVTs التحميل لمدة 3 - D خلية ثقافة ، ويحتاج المرء لتحضير حبات microcarrier Cytodex - 3 : <ol style=";text-align:right;direction:rtl"><li style=";text-align:right;direction:rtl"> تزن خارج القدر المناسب من Cytodex - 3 حبات المطلوبة لهذه التجربة. ويت?...

<ol>
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Immun. 73, 1129-1129 (2005).</li><li>Myers, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Closing+the+phenotypic+gap+between+transformed+neuronal+cell+lines+in+culture+and+untransformed+neurons.">Closing the phenotypic gap between transformed neuronal cell lines in culture and untransformed neurons.</a> J. Neurosci. Methods. 174, 31-31 (2008).</li><li>Hjelm, B. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+and+characterization+of+a+three-dimensional+organotypic+human+vaginal+epithelial+cell+model.">Development and characterization of a three-dimensional organotypic human vaginal epithelial cell model.</a> Biol. Reprod. 82, 617-617 (2010).</li><li>Straub, T. 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D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+three-dimensional+tissue+culture+model+for+the+study+of+attach+and+efface+lesion+formation+by+enteropathogenic+and+enterohaemorrhagic+Escherichia+coli.">A three-dimensional tissue culture model for the study of attach and efface lesion formation by enteropathogenic and enterohaemorrhagic Escherichia coli.</a> Cell. Microbiol. 7, 1771-1771 (2005).</li><li>Sainz, B., TenCate, V., Uprichard, S. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Three-dimensional+Huh7+cell+culture+system+for+the+study+of+Hepatitis+C+virus+infection.">Three-dimensional Huh7 cell culture system for the study of Hepatitis C virus infection.</a> Virol. J. 6, 103-103 (2009).</li><li>Lelkes, P. I., Ramos, E., Nikolaychik, V. V., Wankowski, D. M., Unsworth, B. R., Goodwin, T. 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ثقافة تقنية المعروضة هنا يقدم المحققون مع الخلايا EVT مثل الغازية للغاية. وقد تم الآن من المسلم به أن فقدان التمايز يحدث في monolayers بسبب تثبيط الاستجابات الخلوية لمنبهات الكيميائية والجزيئية في ثلاثة أبعاد (سطوح الخلايا القمي ، القاعدية ، والجانبية) 10،13. هذا الأسل...

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><td> اسم الكاشف </td><td> شركة </td><td> فهرس العدد </td><td> تعليقات </td></tr><tr><td> Cytodex الخرز microcarrier </td><td> سيغما الدريخ </td><td> C3275 </td><td></td></tr><tr><td> نظام التناوب الثقافة الخلية (المراكز المناخية الإقليمية) </td><td> Synthecon </td><td> المراكز المناخية الإقليمية - D </td><td> ويشمل العرض الدوار قاعدة السلطة ، و 4 وحدات للتصرف المراكز المناخية الإقليمية </td></tr><tr><td> وحدات المراكز المناخية الإقليمية يعاد </td><td> Synthecon </td><td> الاتصال Synthecon </td><td></td></tr><tr><td> 3ml حقنة غيض Luer لوك </td><td> دينار بحريني </td><td> 309585 </td><td></td></tr><tr><td> 10ML اسعة طرف المصلية ماصة </td><td> دينار بحريني </td><td> 357504 </td><td></td></tr><tr><td> MEM ألفا </td><td> Invitrogen </td><td> 12561-072 </td><td></td></tr><tr><td> L - 15 ليبوفيتش المتوسطة ، ومسحوق </td><td> Invitrogen </td><td> 41300-039 </td><td></td></tr><tr><td> H 2 O ، الذيفان الداخلي مجانا </td><td> الصياد </td><td> MT - 25 - 055 - CM </td><td></td></tr><tr><td> بيكربونات الصوديوم </td><td> سيغما الدريخ </td><td> S - 7795 </td><td></td></tr><tr><td> الببتون مادة تنشأ عن البروتينات </td><td> فيشر العلمية </td><td> BP1420 - 100 </td><td></td></tr><tr><td> سكر الفاكهة </td><td> سيغما الدريخ </td><td> F3510 - 100 </td><td></td></tr><tr><td> الجالاكتوز </td><td> سيغما الدريخ </td><td> G5388 - 100 </td><td></td></tr><tr><td> جلوكوز </td><td> سيغما الدريخ </td><td> G7528 - 250 </td><td></td></tr><tr><td> HEPES </td><td> Invitrogen </td><td> 15630-080 </td><td></td></tr><tr><td> L - الجلوتامين </td><td> Invitrogen </td><td> 25030 </td><td></td></tr><tr><td> الانسولين ترانسفيرين - زيلونيت الصوديوم (ITS) </td><td> سيغما الدريخ </td><td> I1884 </td><td></td></tr><tr><td> FBS </td><td> Invitrogen </td><td> 10437 </td><td></td></tr><tr><td> البنسلين الستربتوميسين </td><td> Invitrogen </td><td> 15140 </td><td></td></tr></tbody></table>

rotating cell culture systems for human cell culture: human trophoblast cells as a model

The field of human trophoblast research aids in understanding the complex environment established during placentation. Due to the nature of these studies, human in vivo experimentation is impossible. A combination of primary cultures, explant cultures and trophoblast cell lines1 support our understanding of invasion of the uterine wall2 and remodeling of uterine spiral arteries3,4 by extravillous trophoblast cells (EVTs), which is required for successful establishment of pregnancy. Despite the wealth of knowledge gleaned from such models, it is accepted that in vitro cell culture models using EVT-like cell lines display altered cellular properties when compared to their in vivo counterparts5,6. Cells cultured in the rotating cell culture system (RCCS) display morphological, phenotypic, and functional properties of EVT-like cell lines that more closely mimic differentiating in utero EVTs, with increased expression of genes mediating invasion (e.g. matrix metalloproteinases (MMPs)) and trophoblast differentiation7,8,9. The Saint Georges Hospital Placental cell Line-4 (SGHPL-4) (kindly donated by Dr. Guy Whitley and Dr. Judith Cartwright) is an EVT-like cell line that was used for testing in the RCCS.
The design of the RCCS culture vessel is based on the principle that organs and tissues function in a three-dimensional (3-D) environment. Due to the dynamic culture conditions in the vessel, including conditions of physiologically relevant shear, cells grown in three dimensions form aggregates based on natural cellular affinities and differentiate into organotypic tissue-like assemblies10,11,12 . The maintenance of a fluid orbit provides a low-shear, low-turbulence environment similar to conditions found in vivo. Sedimentation of the cultured cells is countered by adjusting the rotation speed of the RCCS to ensure a constant free-fall of cells. Gas exchange occurs through a permeable hydrophobic membrane located on the back of the bioreactor. Like their parental tissue in vivo, RCCS-grown cells are able to respond to chemical and molecular gradients in three dimensions (i.e. at their apical, basal, and lateral surfaces) because they are cultured on the surface of porous microcarrier beads. When grown as two-dimensional monolayers on impermeable surfaces like plastic, cells are deprived of this important communication at their basal surface. Consequently, the spatial constraints imposed by the environment profoundly affect how cells sense and decode signals from the surrounding microenvironment, thus implying an important role for the 3-D milieu13.
We have used the RCCS to engineer biologically meaningful 3-D models of various human epithelial tissues7,14,15,16. Indeed, many previous reports have demonstrated that cells cultured in the RCCS can assume physiologically relevant phenotypes that have not been possible with other models10,17-21. In summary, culture in the RCCS represents an easy, reproducible, high-throughput platform that provides large numbers of differentiated cells that are amenable to a variety of experimental manipulations. 
In the following protocol, using EVTs as an example, we clearly describe the steps required to three-dimensionally culture adherent cells in the RCCS.

Watch this Scientific Journal Video about Rotating Cell Culture Systems for Human Cell Culture: Human Trophoblast Cells as a Model at JoVE.com

Rotating Cell Culture Systems for Human Cell Culture: Human Trophoblast Cells as a Model

التقليدية ، واثنين من خلية ثقافة تقنيات الأبعاد غالبا ما تؤدي إلى تغيير الخصائص فيما يتعلق علامات التمايز ، السيتوكينات وعوامل النمو. ثلاثي الأبعاد ثقافة خلية في نظام خلية ثقافة الدورية (المراكز المناخية الإقليمية) استعادت التعبير عن كثير من هذه العوامل كما هو موضح هنا مع خط الأرومة الغاذية extravillous الخلية.

نظم الدورية الثقافة خلية خلية للثقافة الإنسان : خلايا الأرومة الغاذية الإنسان نموذجا

The field of human trophoblast research aids in  understanding the complex environment established during placentation. Due to the  nature of these ...

Research

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Bioengineering

Microfabricated Platforms for Mechanically Dynamic Cell Culture

Microfabricated المنصات الحيوي للثقافة الخلية ميكانيكيا

The ability to systematically probe in vitro cellular response to combinations of mechanobiological stimuli for tissue engineering, drug discovery or ...

Shape Memory Polymers for Active Cell Culture

البوليمرات الذاكرة شكل خلية للثقافة أحدث

Shape memory polymers (SMPs) are a class of "smart" materials that have the ability to change from a fixed, temporary shape to a pre-determined permanent ...

Alginate Microcapsule as a 3D Platform for Propagation and Differentiation of Human Embryonic Stem Cells (hESC) to Different Lineages

Alginate Microcapsule as a 3D Platform for Propagation and Differentiation of Human Embryonic Stem Cells hESC to Different Lineages

Microcapsule الجينات كمنصة 3D لإكثار وتمايز الخلايا الجذعية الجنينية البشرية (hESC) إلى أنساب مختلفة

Human embryonic stem cells (hESC) are emerging as an attractive alternative source for cell replacement therapy since they can be expanded in culture ...

Three-dimensional Cell Culture Model for Measuring the Effects of Interstitial Fluid Flow on Tumor Cell Invasion

نموذج ثلاثي الأبعاد للخلية ثقافة قياس آثار تدفق السائل الخلالي على غزو الخلايا السرطانية

The growth and progression of most solid tumors depend on the initial transformation of the cancer cells and their response to stroma-associated signaling ...

Cell Co-culture Patterning Using Aqueous Two-phase Systems

خلية المشارك الثقافة الزخرفة باستخدام مائي مرحلتين أنظمة

Cell patterning  technologies that are fast, easy to use and affordable will be required for the  future development of high throughput cell assays, ...

In vitro Cell Culture Model for Toxic Inhaled Chemical Testing

In vitro Cell Culture Model for Toxic Inhaled Chemical Testing

في المختبر خلية الثقافة نموذج لاستنشاق الفحوصات الكيميائية السامة

Cell cultures are indispensable to develop and study efficacy of therapeutic agents, prior to their use in animal models. We have the unique ability to ...

A Hormone-responsive 3D Culture Model of the Human Mammary Gland Epithelium

نموذج الثقافة 3D-الهرمون تستجيب للالغدة الثديية الظهارة الإنسان

The process of mammary epithelial morphogenesis is influenced by hormones. The study of hormone action on the breast epithelium using 2D cultures is ...

Tunable Hydrogels from Pulmonary Extracellular Matrix for 3D Cell Culture

الانضباطي الهلاميات المائية من الرئوي خارج الخلية مصفوفة للثقافة خلية 3D

Here we present a method for establishing multiple component cell culture hydrogels for in vitro lung cell culture. Beginning with healthy en bloc lung ...

Ultrathin Porated Elastic Hydrogels As a Biomimetic Basement Membrane for Dual Cell Culture

سامسونج بوراتيد مطاطا الهلاميات المائية كغشاء الطابق السفلي المحاكاة البيولوجية لثنائي خلية ثقافة

The basement membrane is a critical component of cellular bilayers that can vary in stiffness, composition, architecture, and porosity. In vitro studies ...

Human Pluripotent Stem Cell Culture on Polyvinyl Alcohol-Co-Itaconic Acid Hydrogels with Varying Stiffness Under Xeno-Free Conditions

ثقافة الخلايا الجذعية Pluripotent البشرية على حمض البولي فينيل الكحول-Co-إيتاكونيك الهلاميات المائية مع صلابة متفاوتة تحت ظروف خالية من كرة

The effect of physical cues, such as the stiffness of biomaterials on the proliferation and differentiation of stem cells, has been investigated by ...